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IOBC / WPRS

Study Group “Integrated Control of Plant-Feeding Mites”

OILB / SROP

Groupe d’étude “Lutte Intégrée Contre les Acariens Phytophages”

Proceedings of the Study Group Meeting

at

Jerusalem, Israel

12 – 14 March 2007

Editor:

Phyllis G. Weintraub

IOBC wprs Bulletin Bulletin OILB srop Vol. 30 (5) 2007

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The IOBC/WPRS Bulletin is published by the International Organization for Biological and Integrated Control of Noxious Animals and Plants, West Palearctic Regional Seciton (IOBC/WPRS) Le Bulletin OILB/SROP est publié par l’organisation Internationale de Lutte Biologique et Intégrée contre les Animaux et les Plantes Nuisibles, section Régionale Ouest Paléarctique (OILB/SROP) Copyright : IOBC/WPRS 2007 The Publication Commission: Dr. Horst Bathon Prof. Dr. Luc Tirry Federal Biological Research Center Ghent University For Agriculture and Forestry (BBA) Laboratory of Agrozoology Institute for Biological Control Department of Crop Protection Heinrichstrasse 243 Coupure Links 653 D-64287 Darmstadt (Germany) B-9000 Gent (Belgium) Tel +49 6151 407-225, Fax+49 6151 407-290 Tel. +32 9 2646152, Fax +32 2646239 e-mail: [email protected] e-mail: [email protected] Address General Secretariat IOBC/WPRS Dr. Philippe C. Nicot INRA – Unité de Pathologie Végétale Domaine St. Maurice – B.P. 94 F-84143 Montfavet Cedex France ISBN 92-9067-200-3 web: http://www.iobc-wprs.org

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The first meeting of the Study Group: IPM of Plant-Feeding Mites was supported (in alphabetical order) by:

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Subject Index

Plant Pest Subject First Author Acaricides Cotton Tetranychus urticae Thamethoxam, Imidacloprid, Aldicarb Troxclair Microbial Control Cucumber, strawberry

Tetranychus urticae Fungus, Neozygites floridana Klingen

Neoseiulus californicus, Rhizoglyphus robini

Symbionts, Spiroplasma, Defluvibacter Zchori-Fein

Plant-pathogen Mite interactions Mango Aceria mangiferae Fusarium mangiferae Gamliel-Atinsky Papaya Calacarus flagelliseta Oidium caricae Rosenheim Onion Rhizoglyphus robini Rhizoctonia, Fusarium Hanuny Predators Avacado Oligonychus perseae Euseius scutalis, Neoseiulus

californicus Maoz

Citrus Phyllocoptruta oleivora Amblyseius swirskii Iphiseius degenerans

Argov

Coconut Aceria guerreronis Phytoseiidae, Ascidae, Cheyletidae Lawson-Balagbo Grape Eotetranychus carpini Typhlodromus exhilaratus Liguori Papaya Tetranychus cinnabarinus Phytoseiulus macropilis Rosenheim Amaryllis Steneotarsonemus laticeps Neoseiulus barkeri, Amblyseius

andersoni Messelink

Pepper Polyphagotarsonemus latus Amblyseius swirskii Tal Tomato Tetranychus evansi Phytoseiulus longipes Knapp Solanaceae Tetranychus evansi Phytoseiulus longipes Ferrero Predator Interactions Papaya Tetranychus cinnabarinus Phytoseiulus macropilis, Sten Rosenheim Bean Tetranychus urticae Phytoseiulus persimilis, Neoseiulus

californicus Walzer

Pepper Tetranychus urticae Phytoseiulus persimilis, Neoseiulus californicus, Amblyseius swirskii, A. andersoni

Van Houten

Strawberry Neoseiulus cucumeris, Orius laevigatus Coll Resistant Plants Bean Tetranychus cinnabarinus Antibiosis Vasquez Transgenic plants Zemek Cucumber tomato

Polyphagotarsonemus latus Host-plant selection Alagarmalai

Various Grape Tydeids, Tetranychids Management practices Simoni Tetranychus spp. Systematics, Genetic Variability Ben David Quality Control Steinberg Tribute to Professor E. Swirski Gerson History of Israeli Phytoseiid research Rubin

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List of Participants Jeyasankar Alagarmalai Agricultural Research Organization, Department of Entomology, P.O. Box 6, Bet Dagan, 50250, ISRAEL e-mail: [email protected] Yael Argov

Israel Cohen Institute for Biological Control, Plant Production and Marketing Board, Citrus Division, POB 80 Bet Dagan, 50250, ISRAEL e-mail: [email protected] Yves Arijs Biobest N.V., Ilse Velden 18, B-2260 Westerlo, BELGIUM e-mail: [email protected] Tselila Ben David Hebrew University of Jerusalem, Faculty of Agricultural, Food and Environmental Quality Sciences, P.O Box 12, Rehovot 76100, ISRAEL e-mail: [email protected] Karel J.F. Bolckmans Koppert Biological Systems, P.O. Box 155, 2650 AD Berkel en Rodenrijs, THE NETHERLANDS e-mail: [email protected] Moshe Cohen Bio-Bee Biological Systems, Kibbutz Sde Eliyahu, Beit Shean Valley 10810, ISRAEL Moshe Coll Hebrew University of Jerusalem, Faculty of Agricultural, Food and Environmental Quality Sciences, P.O Box 12, Rehovot 76100, ISRAEL e-mail: [email protected]

Myriam Freund Bio-Bee Sde Eliyahu Ltd, Dept of Research & Development, Kibbutz Sde Eliyahu 10810, ISRAEL e-mail: [email protected] Shira Gal

Agricultural Research Organization (ARO), Newe-Ya’ar Research Center, Ramat Yishay, 30095, ISRAEL e-mail: [email protected] Efrat Gamliel-Atinsky Agricultural Research Organization (ARO), Dept. of Plant Pathology, The Volcani Center, Bet Dagan 50250, ISRAEL e-mail: [email protected] Uri Gerson Dept. of Entomology, Faculty of Agriculture, Hebrew University of Jerusalem, Rehovot 76000, ISRAEL e-mail: [email protected] Danny Gouldman Bio-Bee Biological Systems, Kibbutz Sde Eliyahu, Beit Shean Valley 10810, ISRAEL e-mail: [email protected] Richard GreatRex Syngenta BioLine, Telstar Nursery, Holland Road, Little Clacton, Essex CO16 9QG, U K [email protected] Mor Grinberg Department of Life Sciences, Bar Ilan University, Ramat Gan, ISRAEL e-mail: [email protected] Tal Hanuny

Agricultural Research Organization (ARO), Newe-Ya’ar Research Center, Ramat Yishay, 30095, ISRAEL e-mail: [email protected]

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Nina Svae Johansen Norwegian Crop Protection institute, Plant Protection Centre, Dept of Entomologyand Nematology, Fellesbygget, NORWAY e-mail: [email protected] Sophia Kleitman ARO, Gilat Research Center, Dept of Entomology, D.N. Negev, 85280, ISRAEL Markus Knapp Icipe – African Insect Science for Food and Health, P.O. Box 30772, 00100 Nairobi, KENYA e-mail: [email protected] Serge Kreiter Département d'Enseignement Ecologie et Santé des Plantes, 2 Place Pierre Viala - 34060 Montpellier cedex 01, FRANCE e-mail: [email protected] Marialivia Liguori Istituto Sperimentale per la Zoologia Agraria, via Lanciola 12/A, 50125 Firenze, ITALY e-mail: [email protected] Alon Lotan

Agricultural Research Organization (ARO), Newe-Ya’ar Research Center, Ramat Yishay, 30095, ISRAEL e-mail: [email protected] Anna Luczynski Bio-Bugs Consulting Ltd, Surrey, British Columbia, V4P 2X7, CANADA e-mail: [email protected] Yonattan Maoz Agricultural Research Organization (ARO), Newe-Ya’ar Research Center, Ramat Yishay, 30095, ISRAEL

Gerben J. Messelink Wageningen UR Greenhouse Horticulture, P.O. Box 20, 2265 ZG Bleiswijk, THE NETHERLANDS e-mail: [email protected] Sandra Mulder Koppert Biological Systems, P.O. Box 155, 2650 AD Berkel en Rodenrijs, THE NETHERLANDS e-mail: [email protected] Eric Palevsky

Agricultural Research Organization, Department of Entomology, Newe-Ya’ar Research Center, Ramat Yishay, 30095, ISRAEL e-mail: [email protected] Yeshurum Plesser Bio-Bee Biological Systems, Kibbutz Sde Eliyahu, Beit Shean Valley 10810, ISRAEL e-mail: [email protected] Salvatore Ragusa Dipartimento S. En. Fi. Mi. Zo., Viale delle Scienze, 90128 Palermo, ITALY e-mail: [email protected] Jay Rosenheim Department of Entomology and Center for Population Biology, University of California, Davis, 95616, USA e-mail: [email protected] Amos Rubin Biological Control laboratory, IPM consulting Service, 14 Rahavat Ilan, Givat Shmuel, 54056, ISRAEL e-mail: [email protected] Nurit Shapira Arava Research & Development, D.N. Arava 86825, ISRAEL e-mail: [email protected]

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Sauro Simoni C.R.A., Istituto Sperimentale per la Zoologia Agraria, lab. Acarology, via di Lanciola 12/a, 50125 Florence ITALY e-mail: [email protected] Peter Smytheman Biological Crop Protection Ltd, Occupation Road, Ashford, TN25 5EN, UK; e-mail: [email protected] Victoria Soroker Agricultural Research Organization, Department of Entomology, P.O. Box 6, Bet Dagan, 50250, ISRAEL e-mail: [email protected] Shimon Steinberg Bio-Bee Biological Systems, Kibbutz Sde Eliyahu, Beit Shean Valley 10810, Israel; e-mail: [email protected] Carmit Tal Hebrew University of Jerusalem, Faculty of Agricultural, Food and Environmental Quality Sciences, Rehovot 76100, ISRAEL e-mail: [email protected] Mary Troxclair P.O. Box 1849 Uvalde, Texas 78802-1849, USA Noel Troxclair Texas Cooperative Extension, Texas A&M Research and Extension Center, P.O. Box 1849, Uvalde, Texas 78802-1849 USA e-mail: [email protected] Yvonne M. van Houten Koppert Biological Systems, P.O. Box 155, 2650 AD Berkel en Rodenrijs, THE NETHERLANDS e-mail: [email protected]

Yehudit Vardi Ministry of Agriculture, Extension Service, Bet Dagan, ISRAEL e-mail: [email protected] Carlos Vásquez Universidad Centroccidental Lisandro Alvarado, Decanato de Agronomía Municipio Palavecino, Estado Lara, VENEZUELA e-mail: [email protected] Andreas Walzer Institute of Plant Protection, Department of Applied Plant Sciences and Plant Biotechnology, University of Natural Resources and Applied Life Sciences, Peter Jordanstrasse 82, Vienna, AUSTRIA e-mail: [email protected] Dan Weil Pollination Services Yad-Mordechai, Kibbutz Yad-Mordechai 79145 ISRAEL e-mail: [email protected] Phyllis G. Weintraub Agricultural Research Organization, Gilat Research Center, DN Negev, 85280, ISRAEL e-mail: [email protected] Shaul Zaban Oved, Gubi & Co., Kfar Saba, ISRAEL e-mail: [email protected] Einat Zchori-Fein Agricultural Research Organization, Newe-Ya’ar Research Center, Ramat Yishay, 30095, ISRAEL e-mail: [email protected] Rostislav Zemek Institute of Entomology, Academy of Sciences, Branišovaká 31, České Budějovice, CZ-370 05 Czech Republic e-mail: [email protected]

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Hillel Zuntz Bio-Bee Sde Eliyahu Ltd, Kibbutz Sde Eliyahu 10818 ISRAEL e-mail: [email protected]

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Preface This Bulletin contains the contributions to the first meeting of the IOBC/wprs Study Group “Integrated Control of Plant-Feeding Mites” held in Jerusalem, Israel, 12-14 March, 2007. The Bulletin contains 28 contributions authored by 48 people from 13 countries in Africa, North and South America, Europe, and the Middle East. Topics in this Bulletin include biological control of tetranychid and non-tetranychid mites, chemical control, microbial control, predator-prey interactions, mite-fungal interactions, host plant-mite interactions and the role of symbionts in pest control. Mites are small arthropods that often “get lost” among their larger insect cousins. The aim of this Study Group is to bring together students, researchers and company representatives and focus on all aspects of integrated and biological control of plant feeding mites, in order to discuss approaches that can be taken to increase control efficacy. I would like to thank the scientific committee, Uri Gerson and Eric Palevsky, for selecting the keynote speakers and all of the private sponsors that generously contributed to finance this meeting. Furthermore, I would like to thank the participants for their enthusiasm in preparing their contributions and for their collaboration in the success of the meeting. Phyllis G. Weintraub Convenor of the Study Group March 2007

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Contents Subject Index .................................................................................................................................. v List of Participants .........................................................................................................................vii Preface ............................................................................................................................................xi Contents........................................................................................................................................ xiii Host selection behavior of the broad mite, Polyphagotarsonemus latus (Acari: Tarsonemidae) ................................................................................................................................ 1

Jeyasankar Alagarmalai , Mor Grinberg, Saadia Reneh, Lital Sharon, Rafael Perl-Treves , Victoria Soroker

Augmentation and conservation of indigenous generalist acarine predators for the control of citrus rust mite, Phyllocoptruta oleivora, in Israel ........................................................ 9

Yael Argov, Sylvi Domeratzky, Uri Gerson, Shimon Steinberg, Eric Palevsky

The genetic variability of the spider mites of Israel ..................................................................... 17 Tselila Ben-David, Sarah Melamed, Uri Gerson, Shai Morin Interplay between omnivory and intraguild predation: thrips spatial dynamics and damage to strawberry ............................................................................................................ 19 Moshe Coll, Sulochana Shakya, Phyllis Weintraub Tetranychus evasi Baker & Pritchard control in European solanaceous greenhouses: facts and perspectives ................................................................................................................... 21 Maxine Ferrero, Marie-Stephane Tixier, Karel Bolckmans, Serge Kreiter Interactions of the mango bud mite, Aceria mangiferae, with Fusarium mangiferae, the causal agent of mango malformation disease ......................................................................... 23 Efrat Gamliel-Atinsky, Stanley Freeman, Abraham Sztejnberg, Marcel Maymon, Eduard

Belausov, Eric Palevsky A Tribute to the late Professor Eliahu Swirski, our foremost agricultural acarologist ................ 29

Uri Gerson Acaricides and the integrated control of plant feeding mites ....................................................... 33 Richard GreatRex The interaction between Rhizolyphus robini and plant pathogens on onion ................................ 41 Tal Hanuny, Moshe Inbar, Leah Tsror, Eric Palevsky

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The beneficial fungus Neozygites floridana for the control of Tetranychus urticae ................... 49 Ingeborg Klingen, Karin Westrum, Silje Stenstad Nilsen, Nina Trandem, Gunnar Wærsted

Exploration and evaluation of natural enemies for the invasive spider mite Tetranychus evansi ....................................................................................................................... 51

Markus Knapp Predatory mites associated with the coconut mite Aceria guerreronis in Brazil ......................... 59

L.M. Lawson-Balagbo, M.G.C. Gondim Jr, G.J. de Moraes, R. Hanna, P. Schausberger

Biological control in vineyards by means of a laboratory phytoseiid strain: a small scale experiment in Tuscany (Italy) ...................................................................................................... 65

Marialivia Liguori, Giuseppino Sabbatini Peverieri, Sauro Simoni, Laura Ferré Biological control of the newly introduce persea mite with indigenous and exotic predators ............................................................................................................................ 73 Yonattan Maoz, Shira Gal, Yael Argov, Martin Berkeley, Miriam Zilberstein, Mickey Noy, Yehonatan Izhar, Jonathan Abrahams, Moshe Coll, Eric Palevsky Biological control of the bulb scale mite Steneotarsonemus laticeps (Acari: Tarsonemidae) with Neoseiulus barkeri (Acari: Phytoseiidae) in amaryllis ........................................................ 81

G.J. Messelink, R. van Holstein-Saj Additivity versus interactions in mite-plant and predator-mite interactions ................................ 87

Jay Rosenheim, Valerie Fournier The history of the predacious Phytoseiidae mites in Israel .......................................................... 89 Amos Rubin A tritrophic perspective to thre biological control of eriophyoid mites ....................................... 91 Maurice Sabelis, Izabela Lesna, Nayanie Aratchige The effects of varieties and agronomic practices on acarine populations in Italian vineyards ...................................................................................................................................... 95 Sauro Simoni, Marisa Castagnoli Seasonal quality assessment of mass-produced Phytoseiulus persimilis .................................... 101 Shimon Steinberg, Hadasa Cain Biological control of Polyphagotarsonemus latus (Acari: Tarsonemidae) by the predaceous mite Amblyseius swirskii (Acari: Phytoseiidae) ...................................................... 111

Carmit Tal, Moshe Coll, Phyllis G. Weintraub

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Field evaluation of cotton seed treatments and a granular soil insecticide in controlling spider mites and other early-season cotton pests in Texas .......................................................... 117 Noel Troxclair Spider mite control by four phytoseiid species with different degrees of polyphagy ................. 123

Yvonne M. van Houten, Hans Hoogerbrugge, Karel J.F. Bolckmans The influence of Amblyseius swirskii on biological control of two-spotted spider mites with the specialist predator Phytoseiulus persimilis (Acari: Phytoseiidae) ....................... 129

Yvonne M. van Houten, Hans Hoogerbrugge, Karel J.F. Bolckmans Antibiosis of kidney bean cultivars to the carmine spider mite, Tetranychus cinnabarinus (Boisduval) (Acari: Tetranychidae) ...................................................................... 133

Carlos Vásquez, Mariela Colmenárez, Neicy Valera, Lisbeth Diaz Spatiotemporal within-plant distribution of the spider mite Tetranychus urticae confronted with specialist and generalist predators ................................................................... 139 Andreas Walzer, Karl Moder, Peter Schausberger Symbionts in mites and their relevance for pest control ............................................................ 147 Einat Zchori-Fein, Monika Enigl, Peter Schusberger, Netta Mozes-Daube, Yuval Gottlieb, Tal Hanuny, Eric Palevsky Transgenic crop-mite interactions .............................................................................................. 155 Rostislav Zemek

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Integrated Control of Plant–Feeding Mites IOBC/wprs Bulletin Vol. 30 (5) 2007

pp. 1-7

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Host selection behavior of broad mite, Polyphagotarsonemus latus (Acari: Tarsonemidae) Jeyasankar Alagarmalai 1 , Mor Grinberg1,2, Saadia Reneh1 Lital Sharon1, Rafael Perl-Treves2 , Victoria Soroker1 1 Dept. of Entomology, Agricultural Research Organization, Bet Dagan 50250, Israel; 2The Mina and Everard Goodman Faculty of Life Science, Bar-Ilan University, Ramat-Gan 52900 Israel, [email protected]

Abstract: This study presents evidence for host selection ability of broad mites, Polyphagotarsonemus latus (Banks) (Acari: Tarsonemidae), by both free-moving and phoretic individuals. Host selection by free-moving mites was monitored in two or four choice set up of young leaves (3rd leaf from the apex). Host choice between several plants was tested: cucumber (Cucumis sativus L.) cv Kfir, tomato (Solanum lycopersicon) cv. M82, two isogenic lines, cv Moneymaker and Motelle, that differ in resistance to whiteflies, and the cucurbits Lagenaria siceraria cv Sus and Cucurbita pepo cv Orangetti, previously shown to differ in resistance to Tetranychus urticae. A tomato mutant derived from cv Castlemart, def-1 (defenseless-1), and its wild-type isogenic line were also compared. Response of phoretic mites to the plants was tested in non choice bioassay. Performance on the hosts was determined by counting the progeny after one week. Our data indicate that broad mites are able to actively choose between host plants and usually successfully discriminate between susceptible and non-susceptible hosts.

Key words: Broad mites, Cucumis sativus, Solanum lycopersicon, Lagenaria siceraria, Cucurbita pepo, plant defenses

Introduction Plant herbivore relationships are extremely complex and include damage to the plant on one side and a variety of defense mechanisms on the other. Host selection is one of the most crucial stages in the life cycle of an organism, especially minute herbivores such as mites which have limited locomotory abilities. The broad mite (BM) Polyphagotarsonemus latus (Banks) (Acari: Tarsonemidae) is a minute, 0.2 µm, polyphagous mite found on plants from almost 60 different families. It causes severe, economic damage to many greenhouse crops, especially cucurbitaceae, but common tomato varieties are usually resistant. Susceptible crops suffer mainly from growth inhibition and distortion in leaf and fruit (Gerson 1992).

Mite dispersal occurs through various means. Within the same infested plant, broad mites continuously move upwards; they may walk over short distances between plants, or be displaced by wind. In addition mated females of BM have a specific phoretic association with whiteflies (Heteroptera: Aleyrodidae) (Palevsky et al. 2001, Soroker et al. 2003). Most information on the relationship between BM and their host has been inferred from the prevalence or degree of damage on certain hosts, but very few studies have directly addressed host resistance mechanisms or BM choice behavior. On the other hand, plant defense against broad mites’

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phoretic host, Bemisia tabaci (Gennadius), was extensively studied. For example, the resistance of commercial and transgenic tomato varieties bearing the resistance gene Mi against B. tabaci has been characterized (Nombela et al. 2000, 2001, 2003).

The aim of this study was to investigate the host selection ability and host acceptance of phoretic and free-moving broad mites on different plants and to evaluate the possible involvement of the jasmonic acid (JA) signaling pathway in tomato defense against BM.

Material and methods Broad mite and whitefly culture Broad mite cultures were established on young cucumber plants as described in Palevsky et al. (2001). Female B. tabaci B-biotype were collected from a laboratory colony and kept frozen for 24 hours at -20°C for the phoretic bioassay. Plant cultivation and preparation Experimental plants, of Cucumis sativus L. cv Kfir, Lagenaria siceraria cv Sus and Cucurbita pepo cv Orangetti, and tomato (Lycopersicon esculentum Mill) varieties: M82, Moneymaker (Mi+), Motelle (Mi-), cv Castlemart and its JA pathway mutant def-1, were grown in 1L pots with standard fertilization in a growth room at ca. 24ºC, as described by Grinberg et al. (2005). Young leaves (3rd leaf from the apex) were collected from different host plants. Bioassays Host preference of free-moving mites was examined using a choice test. The bioassay was conducted on 9 cm Petri dishes. Two or four different types of leaves were compared at each test. A BM infested leaf was placed in the centre of the plate and used as a source of BM population. Four discs of un-infested leaves (1.8 cm) were placed at an equal distance of 1.2cm on the four sides: two leaves of each type for the two-choice bioassay (four experiments), or one leaf of each kind for the four-choice bioassay. After 24 hours, the leaves were washed with 70% ethanol and the number of mites on each leaf disc was monitored using a stereomicroscope (Olympus SZX12, at 40 x magnification).

In order to evaluate host plant preference by phoretic mites, we monitored the detachment of phoretic mites from their vector, B. tabaci, on leaf discs from different host plants (four tests). In order to acquire phoretic broad mites, frozen B. tabaci adults were placed on BM-infested cucumber leaves, and mites (on average 7 individuals per whitefly) were allowed to attach to the whiteflies; the number of phoretic BM was recorded under the stereomicroscope. Subsequently, a single whitefly loaded with broad mites was placed on each leaf disc (2.8cm diameter), situated on wet filter paper in a Petri dish. Petri dishes were kept in a growth room (25 °C, 14h L:10h D photoperiod). The number of mites that detached from whitefly after 4 hour on a leaf disc was monitored. This experimental setup was replicated 10-37 times for each plant type.

To determine host suitability for broad mites, leaf discs with a specific number of female broad mites were incubated under the above environmental conditions on humid filter paper. After seven days, the leaf discs were washed with 70% ethanol and the total number of progeny (male, female and larva) was counted under the stereomicroscope. Data analysis Host selection by free-moving mites was calculated as the percentage of mites (out of the total number of mites that selected new leaf discs). Mite's discrimination between the two or four alternative leaf-types was analyzed by Wilcoxon Signed-Rank Test. Arcsin transformation was

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performed on the proportion of detached mites. Rate of mites' detachment on different hosts was compared on arcsin transformed data using GLM of SAS/STAT, or t-test. Differences in detachment from phoretic host between the leaf type groups were analyzed using Scheffe post-hoc test. Broad mite progeny on different leaf types (number of individuals per female) was compared on square root transformed data using GLM of SAS/STAT or t-test.

Results and discussion Free-moving mites were able to discriminate between different host plants in two-choice and four choice bioassays (Figures 1 & 2). In two choice assays, broad mites significantly preferred cucumber leaves over those of tomato Moneymaker, Moneymaker over M82, and the def-1 mutant over Castlemart wild type, but did not discriminate between Moneymaker (Mi-) and Motelle (Mi+) (Exp 3). In four-choice bioassays, BM mostly favored Moneymaker, followed by C. pepo, whereas the less-preferred leaves were from tomato M82 and L. siceraria. Figure 1. Broad mite host selection in four different two-choice tests after 24 hours. * indicates significant differences between number of mites selecting each leaf type, p<0.05, Wilcoxon Signed-Rank Test.

When phoretic mites were placed on leaves from different plants, the proportion of detached mites varied (Table 1). We hypothesize that higher detachment rates from the phoretic host reflect BM preference. In general, phoretic mites' preference agreed with that of free-moving mites. The fastest detachment was observed on cucumber leaves, followed by tomato Moneymaker, whereas the slowest detachment of broad mite was on M82. Like the free-moving mites, phoretic ones detached significantly more on def-1 compared to the wild type, but showed no differences in detachment between tomato Moneymaker and Motelle. However, some

Figure 1

0

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C. sativ

us (K

fir)

L. escu

lentum

(MM)

L. escu

lentum

(MM)

L. escu

lentum

(M82

)

L. escu

lentum

(MM)

L. escu

lentum

(Mote

lle)

Castlemart (

wild-ty

pe)

Castlemart (

def-1)

Plants

Perc

enta

ge o

f se

lect

ing

BM

* * NS

*

Exp.1 Exp.2 Exp.3 Exp.4

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Figure 2. Broad mite host selection in a four-choice bioassay after 24 hours. discrepancy was found in BM response to C. pepo and L. siceraria between free-moving and the phoretic mites. While free-moving mites preferred C. pepo (Orangetti) over L. siceraria (Sus), phoretic mites showed a bit higher, albeit not significantly different, detachment on the second.

Table 1. Broad mites’ detachment and performance on leaf discs from a number of host plants. Results from four experiments are presented as mean ±SE of at least 10 replicates. Different letters within each test indicate groups that are statistically different (p<0.05).

Host plant Host performance

Test Family Species %

detachment 4h*

Total progeny/female

Average sex ratio

Female/MaleCucurbitaceae Cucumis sativus (Kfir) 93 a 4.8 ± 0.48 a 2:1

Lycopersicon esculentum (Moneymaker) 83 b 3.6 ± 0.40 a 3:1 1

Solanaceae Lycopersicon esculentum

(M82) 64 c 1.1 ± 0.15 b 5:1

Lycopersicon esculentum (Moneymaker) 85 a 1.4 ± 0.37 a 7:1

2


(Motelle) 78 a 1.0 ± 0.20 a 7:1

Lycopersicon esculentum (Castlemart wt -type) 45 b 1.8 ± 0.27 b 4:1

3


(Castlemart- def-1 mutant) 72 a 3.9 ± 0.27 a 5:1

Lagenaria siceraria (Sus) 90 a 0.7± 0.09 a 5:1 4

Cucurbitaceae Cucurbita pepo

(Orangetti) 77 a 1.5 ± 0.24 b 4:1

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L. esculentum (MM) C. pepo (Orangetti) L. esculentum (M82) L. siceraria (Sus)Plants

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BM

Figure 2

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The reproductive performance of BM was host dependent and generally in accordance to host preference. The progeny in all tests was female-biased, which represents the usual situation in this species: the common female to male ratio is 4:1 (Gerson 1992). Fluctuations in sex ratio between tests and host plant remain to be further studied.

Although host related biotypes may be found in broad mites, in this study a cucumber-adapted BM population had similar reproductive success on leaves from cucumber and tomato Moneymaker. It is expected that mites’ performance (survival and fecundity) will be affected by plant quality and defense ability. We have observed some differences in mites performance within cucurbit and tomato varieties, for example in test 1 (Table 1 ) the progeny on tomato Moneymaker was significantly higher that on M82. In test 2, (Table1) the progeny that developed on Moneymaker (Mi-) was lower than in test 1, but did not differ from that developed on Motelle (Mi+). The latter suggest that although the Mi gene is responsible for the resistance of tomato to both B-and Q-biotypes of Bemisia tabaci (Nombela et al. 2001, 2002, 2003), root-knot nematodes (Meloidogyne spp.) (Roberts & Thomason, 1986) and the potato aphid, Macrosiphum euphorbiae (Thomas) (Goggin et al. 2001, Rossi et al. 1998), this gene does not provide resistance to broad mites. These data are interesting in respect to deferent host suitability between broad mite and its phoretic vector B. tabaci. Furthermore, in agreement with broad mites’ behavior, significantly higher progeny developed on Castlemart def-1 mutant than on its wild type isogenic line. Similarly, Tetranychus urticae (Acari: Tetranychidae) had a higher reproductive success on def-1 (Ament et al. 2004) while larvae of Spodoptera exigua (Lepidoptera: Noctuidae) showed increased survival and growth on def-1 plants compared to wild type (Thaler et al. 2002). Def-1 plants are known to have a reduced ability to produce JA, and thus lower expression of defense genes such as proteinase inhibitors after herbivore damage (Howe et al. 1996, Li et al. 2002).

Comparison C. pepo (Orangetti) with L. siceraria (Sus), revealed that reproductive performance was better on the C. pepo and corresponded well with its preference by the free-moving mites. Lower performance of BM on L. siceraria, suggests resistance of this species to BM. Similarly, Edelstein et al. (2000) reported that L. siceraria was more resistant to Tetranychus cinnabarinus.

It therefore appears that BM is able to discriminate between resistant and susceptible plants, and prefers the susceptible ones. The mechanisms and cues operating in broad mites host selection are unknown. In general, host-plant selection by herbivorous insects depends on the distance from the plant, and selection is modulated by olfactory, mechanosensory and gustatory information (Schoonhoven et al. 1998). Since broad mite vision is probably limited to light/dark differentiation, visual cues are highly unlikely to play a role in host selection. For example, in case of discrimination between Castlemart def-1 mutant and its wild type, significant differences in the volatile profile (Ament et al. 2004) are likely to provide discriminating cues. On the other hand, inability of phoretic BM to discriminate “correctly” between C. pepo and L. siceraria in detachment bioassay could reflect the difference in detection modalities operating during host analysis in both bioassays. While free-moving mites were able to assess host suitability both via olfactory, mechanosensory and gustatory information, phoretic mite detachment probably occurs prior to any physical contact with the leaf. The fact that a progeny correlates with host selection of free-moving mites emphasizes the significance of broad spectrum of information emanating from the host. Specific cues used by mites for host selection remain to be explored.

In conclusion, broad mite exhibits active host selection and host preference which mostly correlates with its reproductive performance. Broad mites were able to discriminate between

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isogenic varieties that differed in JA-mediated defense. However, the Mi gene does not seem to be responsible for the resistance tomato plants to BM. Acknowledgement Special thanks are addressed to Prof. U. Gerson and Dr. E. Palevsky for providing helpful discussions and Dr. P. Weintraub for her assistance with BM culture and IOBC/WPRS study group for supporting the participation of J. Alagarmalai and M. Grinberg in the current meeting. We would also like to thank S. Kontsedalov for providing the Bemisia tabaci, M. Edelstein for providing the Lagenaria siceraria and Cucurbita pepo seeds, J. Milo for providing the Moneymaker and Motelle seeds, M. Pilowsky for providing the M82 seeds and G. Howe for providing the Castlemart wild type and def-1 mutant seeds. The fellowship of J. Alagarmalai was financed by MASHAV (Center for International Cooperation, Israel). This manuscript is contribution no. 502/07 of the Institute of Plant Protection, ARO, Israel. References Ament, K., Kant, M.R., Sabelis, W., Haring, M.A. & Schuurink, R.C. 2004: Jasmonic acid is a

key regulator of spider mite-induced volatile terpenoid and methyl salicylate emission in tomato. Plant Physiol. 135: 2025-2037.

Edelstein, M., Tadmor, Y., Abo-Moch, F., Karchi, Z. & Mansour, F. 2000: The potential of Legenaria rootstock to confer resistance to the carmine spider mite, Tetranychus cinnabarinus (Acari: Tetranychidae) in cucurbitaceae. Bull. Entomol. Res. 90: 113-117.

Gerson, U. 1992: Biology and control of the broad mite, Polyphagotarsonemus latus (banks) (Acai: Tarsonemidae). Exp. Appl. Acarol. l13: 163-178.

Goggin, F.L., Williamson, V.M. & Ullman, D.E. 2001: Variability in the response on Macrosiphum euphorbiae and Myzus persicae (Hemiptera: Aphidae) to the tomato resistance gene Mi. Environ. Entomol. 30: 101-106.

Grinberg, M., Treves, R.P., Palevsky, E., Shomer, I. & Soroker, V. 2005: Interaction between cucumber plants and the broad mite, Polyphagotarsonemus latus : from damage to defense gene expression. Entomol. Exp. Appl. 115: 134-144.

Howe, G.A., Lightner, J., Browse, J. & Ryan, C.A. 1996: An octadecanoid pathway mutant (JL5) of tomato is compromised in signaling for defense against insect attack. The Plant Cell 8: 2067-2077.

Li, C., Williams, M.W., Loh, Y., Lee G.I. & Howe, G.A. 2002: Resistance of cultivated tomato to cell content-feeding herbivores is regulated by the octadecanoid-signaling pathway. Plant Physiology 130: 494-503.

Nombela, G., Beitia, F. & Muniz, M. 2000: Variation in tomato host response to Bemisia tabaci (Hemiptera: Aleyrodidae) in relation to acyl sugar content and presence on the nematode and potato aphid resistance gene Mi. Bull. Entomol. Res. 90: 161-167.

Nombela, G., Beitia, F. & Muniz, M. 2001: A differential interaction study of Bemisia tabaci Q-biotype on commercial tomato varieties with or without the Mi resistance gene, and comparative host response with the B-biotype. Entomol. Exp. Appl. 98: 339-344.

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Nombela, G., Williamson, V. M. & Muniz, M. 2003: The root-knot nematode resistance gene Mi-1.2 of tomato is responsible for the resistance against the whitefly Bemisia tabaci. Mol. Plant Microbe Interact. 16: 645-649.

Palevsky, E., Soroker, V., Weintraub, P., Mansour, F., Abo-Moch, F. & Gerson, U. 2001: How species-specific is the phoretic relationship between the broad mite, Polyphagotarsonemus latus (banks) (Acai: Tarsonemidae), and its insect hosts? Exp. Appl. Acarol. 25: 217-224.

Roberts, P.A. & Thomason, I.J. 1986: Variability in reproduction of isolates of Meloidogyne incognita and M. javanica on resistant tomato genotypes. Plant Dis. 70: 547-551.

Rossi, M., Goggin, F.L., Milligan, S.B., Kaloshian, I., Ullman, D. E. & Williamson, V.M. 1998: The nematode resistant gene Mi of tomato confers resistance against the potato aphid. Proc. Natl. Acad. Sci. U.S.A. 95: 9750-9754.

Schoonhoven, L.M., Jermy, T., van Loon, J.J.A. 1998: Insect-plant biology. Chapman and Hall. Pp 409.

Soroker, V., Nelson, R.D., Bahar, O., Reneh, S., Yablonski, S., Palevsky, E. 2003: Whitefly wax as a cue for phoresy in the broad mite, (Polyphagotarsonemus latus (banks) (Acai: Tarsonemidae). Chemoecology 13: 163-168.

Thaler, J.S., Farag, M.A., Pare, P.W. & Dicke, M. 2002: Jasmonate deficient plants have reduced direct and indirect defenses against herbivores. Ecol. Let. 5: 764-774.

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pp. 9-15

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Augmentation and conservation of indigenous generalist acarine predators for the control of citrus rust mite, Phyllocoptruta oleivora, in Israel Yael Argov1, Sylvi Domeratzky, Uri Gerson2, Shimon Steinberg3 Eric Palevsky4

1Israel Cohen Institute for Biological Control, Plant Production and Marketing Board, Citrus Division, POB 80 Bet Dagan, 50250, Israel, [email protected];2Department of Entomology, Faculty of Agricultural, Food and Environmental Quality Sciences, Rehovot 76100, ISRAEL, [email protected]; 3Bio-Bee Biological System, Kibbutz Sdeh Eliyahu, Beit Shean Valley 10810, Israel, [email protected]; 4Agricultural Research Organization (ARO), Newe-Ya’ar Research Center, Ramat Yishay, 30095, Israel, [email protected]. Abstract: The key acarine citrus pest in Israel is the citrus rust mite (CRM), Phyllocoptruta oleivora, which is probably the most cosmopolitan citrus pest. In this study we focused on the conservation and augmentation of two indigenous phytoseiids, found to be potential predators of CRM, namely Amblyseius swirskii and Iphiseius degenerans. In order to identify a chemical suitable for Medfly control that is also more selective for these acarine predators we compared the field effects of spinosad to malathion, and found the former to be more selective to A. swirskii than the latter. Field augmentation trials with A. swirskii and I. degenerans yielded significantly higher levels of predators in some of the trials, but had no effect on CRM populations. In unsprayed groves where CRM is under control, these predators subsist on a diet composed of alternate food sources, such as other mites and insect prey, pollen, honeydew and various fungi. We thus believe that habitat management and conservation should become part and parcel of an indigenous predator augmentative program. Key words: Phyllocoptruta oleivora, Phytoseiidae, Iphiseius, Amblyseius, pollen, spinosad. Introduction The key acarine citrus pest in Israel is the citrus rust mite (CRM), Phyllocoptruta oleivora (Ashmead) (Prostigmata: Eriophyidae) (Palevsky et al., 2003a). It is probably the most cosmopolitan citrus pest (Childers, 1994). Damage caused by CRM is usually negligible in minimally or unsprayed groves located in the central coastal plain of Israel, an indication that the conservation of indigenous natural enemies could be instrumental for CRM control. In contrast, in adjacent commercially sprayed groves, CRM populations resurge every year. Previously sprayed plots that go untreated suffer severe fruit damage and are not economical to harvest (personal observations). The augmentation of indigenous predatory mites in such plots could enhance mite control and expedite the return to equilibrium between CRM and its natural enemies. The conservation of these natural enemies could then be accomplished by using only selective pesticides.

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Materials and methods Conservation of indigenous phytoseiid predators Recent legislation in the United States and Europe is calling for the reduction of use and even removal of organophosphates. Accordingly substitutes for malathion, an organophosphate traditionally used for Ceratitis capitata Wiedemann (Mediterranean fruit fly, Medfly) control, are being sought. One such material is spinosad (Success™, Dow Agrosciences), a compound that has been found to be non-toxic to Neoseiulus cucumeris (Oud.) and slightly toxic to Iphiseius degenerans (Berlese) (Phytoseiidae) in greenhouse flower crops (Driesche et al., 2006). In contrast, malathion is known to be highly toxic to Euseius mesembrinus (Phytoseiidae) on citrus (Childers et al, 2001). To determine whether spinosad is more compatible than malathion for phytoseiids on citrus in Israel, we conducted a field experiment, performed in a grapefruit orchard (CV ‘Sunrise’) in a blocked design (replicate size: 9 trees long and three rows wide, the three central trees were monitored). Every 10 days from April through May 2006 aerial sprays of spinosad were applied at ultra low volume to the entire plot. To compare the effects of malathion+spinosad vs. spinosad alone, malathion was sprayed from the ground with an airblast sprayer on its respective sites; in total 5 sprays were applied. The plots were monitored bi-monthly from the end of March (before the experiment was initiated) through the end of July, two months after the last treatment. CRM were counted with a 10x magnifying glass, in 2 fields of view (1 sq cm) per leaf/fruit, 10 leaves/fruit per site. Phytoseiid populations were sampled by beating the tree with an irrigation pipe and collecting the mites from the beating tray with an aspirator (Palevsky et al., 2003b). Mites were stored in alcohol, in the lab all specimens were mounted and identified (Swirski et al., 1998). Augmentation of indigenous phytoseiid predators Porath and Swirski (1965) found nine species of the family Phytoseiidae on citrus in Israel. Amblyseius swirskii Athias-Henriot and I. degenerans were abundant in the humid coastal plain, A. swirskii being the dominant species. In a recent study the authors (Palevsky et al., 2003b) found higher I. degenerans populations in the cooler months whereas A. swirskii was more substantial in the summer. We thus considered these two phytoseiids as worthy candidates for evaluation as augmentative biological control agents.

Amblyseius swirskii – Release experiments were performed on ‘Marsh’ grapefruit and on ‘Shamouti’ oranges in two plots [one organic located in the inner coastal plain (Yesodot) and one conventional in the coastal plain (Hadera)]. Experiments were conducted in a paired t-test design, replicated 9-10 times, each replicate being three trees long and three rows wide. Experiments were initiated in the beginning of April. Amblyseius swirskii were reared and provided by Bio-Bee Biological Systems (http://www.seliyahu.org.il/eBiobee.htm). Predators were released on the first sampling date, when CRM populations were nil, 2000 mites/tree in the organic orchard and 4000 mites/tree in the conventional orchard. In the organic orchard a second release, 4000 mite/tree, was carried out when CRM populations increased.

Iphiseius degenerans – Trials were conducted in two conventional plots, in March on ‘Marsh’ grapefruit in the North Western Negev (Shibolim) and in June on ‘Shamouti’ oranges in the inner coastal plain (Zafariya), replicated five times, each replicate consisting of one release/control tree and two adjacent trees, and two rows as buffers, using the same experimental design as described above. Iphiseius degenerans were reared on pollen (Argov et al., 2002) at the Israel Cohen Institute for Biological Control, Plant Production and Marketing Board, Israel.

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Three releases of I. degenerans, each of 2000 mite/tree, were carried out once every two months, the first upon the initiation of the trials.

Predator and CRM populations for both phytoseiid species were monitored using the same methodology as described (see conservation of indigenous phytoseiid predators). Data were analyzed using ANOVA procedures with JMP5.0.1a (SAS Institute, Inc.). Results and Discussion Conservation of indigenous phytoseiid predators The interaction between time and treatment (spinosad vs. malathion+spinosad), upon considering all dates for both predators, was not significant (A. swirskii – P = 0.769; F = 0.509; DF = 5,60; I. degenerans – P = 0.789; F = 0.481; DF = 5,60). Thus we considered the values from all dates in a single analysis. On plots treated only with spinosad, indigenous populations of A. swirskii were higher than those treated with malathion and spinosad (P = 0.044; F = 4.24; DF = 1,60). In contrast, the populations of I. degenerans did not differ between treatments (P = 0.275; F = 1.214; DF = 1,60). Having said that, we believe it to be premature to conclude that malathion does not affect I. degenerans. Additional field trials conducted in the winter, when population levels of I. degenerans are usually higher (Palevsky et al., 2003b), are needed to confirm this result. The effect of predator conservation on CRM control could not be evaluated in this trial because pest populations remained nil throughout. As spinosad is still considerably more costly than malathion, it has not yet been adopted for Medfly control. Should the use of spinosad allow for substantially higher indigenous predator populations, resulting in lower CRM seasonal levels and thus fewer chemical treatments to control this pest, growers might reconsider and prefer spinosad over malathion. Augmentation of indigenous phytoseiid predators Amblyseius swirskii - In the two trials the interactions between time and treatment (release vs. control) for A. swirskii populations were not significant (Yesodot- P = 0.432; F = 0.996; DF = 6,112; Hadera – P = 0.080; F = 2.155; DF = 4,90), thus the data from all dates (for each experiment) were combined. The Yesodot releases of A. swirskii had no significant effect on A. swirskii populations (P = 0.177; F = 1.848; DF = 1, 112) or CRM (P = 0.796; F = 0.067; DF = 1, 112). In contrast at the Hadera trial, the release treatment had a very significant effect on A. swirskii populations (P < 0.0001; F = 36.403; DF = 1,90), but it had no effect on the CRM outburst that occurred more than 100 days post-release (P = 0.607; F = 0.267; DF = 1,90) (Figure 1).

Iphiseius degenerans - The interactions between time and treatment (release vs. control) in respect to the population levels of I. degenerans, in both trials, were very significant (Shibolim- P < 0.0001; F = 54.272; DF = 1,64; Zafariya – P = <0.0001; F = 3.699; DF = 7,64). Furthermore the effect of the release was also very significant in both experiments (Shibolim- P < 0.0001; F = 13.782; DF = 7,64; Zafariya – P < 0.0001; F = 54.099; DF = 7,64) (Figures 2 and 3).

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Figure 1. Mean numbers of Amblyseius swirskii collected from a beating tray (6 branches per tray) and of Phyllocoptruta oleivora (citrus rust mite, CRM) per lens view, in predator (A. swirskii) release and control sites, ‘Marsh’ grapefruit and ‘Shamouti’ oranges, Hadera. Arrows indicate times of release. However, once again, there was no significant interaction between treatment and time (Shibolim- P = 0.972; F = 0.246; DF = 7,64; Zafariya – P = 0.117; F = 1.732; DF = 7,64) nor any effect of treatment on CRM population levels (Shibolim - P = 0.707; F = 0.143; DF = 1,64; Zafariya – P = 0.235; F = 1.437; DF = 1,64).

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Figure 2. Mean numbers of Iphiseius degenerans collected from a beating tray (6 branches per tray) and of Phyllocoptruta oleivora (citrus rust mite, CRM) per lens view, in predator (I. degenerans) release and control sites on ‘Marsh’ grapefruit, Shibolim. Arrows indicate times of release. Despite the significant increase of predator levels on the release trees (A. swirskii in Hadera and I. degenerans in both plots) we observed no reductions in the CRM levels. The timing of the releases is clearly problematic. On the one hand, when predators are released before pest populations are present, predator levels might not be maintained due to lack of prey and/or alternate foods. On the other, when predators are released later, when CRM populations are escalating, generalist predators such as A. swirskii and I. degenerans, might not be able to prevent outbreaks of this problematic pest. In unsprayed citrus groves, where CRM is under control, these predators subsist on a balanced diet composed of alternate food sources, such as other mite and insect prey, pollen, honeydew and various fungi (Duso et al., 2005). We thus believe that habitat management and conservation must be part and parcel of an indigenous predator augmentative program. Pollen availability can be enriched by planting hedge rows, properly managing the flowering of wild species of gramineae or by adding a cultivated grass as a cover crop (Smith and Papacek, 1991). These measures could enhance the efficacy of CRM control by indigenous phytoseiids.

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Figure 3. Mean numbers of Iphiseius degenerans collected from a beating tray (6 branches per tray) and of Phyllocoptruta oleivora (citrus rust mite, CRM) per lens view, in predator (I. degenerans) release and control sites on ‘Shamouti’ oranges, Zafariya. Arrows indicate times of release.

Acknowledgements We are indebted to the citrus growers and to Tslila Ben David, Arnon Alush, Amnon Hadar, Yaakov Yistrov and Recardo Baram who assisted in the field trials. We would like to acknowledge the technical staff of ICIBC, Martin Berkeley, Wolf Kuslizky And Eti Melamid. This manuscript is a contribution of the Institute of Plant Protection, Volcani Center, ARO, Israel.

References Argov, Y., Amitai, S., Beattie, G. A. C. & Gerson, U. 2002: Rearing, release and establishment

of imported predatory mites to control citrus rust mite in Israel. BioControl 47: 399-409.

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Childers, C.C. 1994: Biological control of phytophagous mites on Florida citrus utilizing predatory arthropods. p. 255-288. In: D. Rosen, F.R. Bennett and J.L. Capinera (eds.), Pest management in the subtopics, biological control - a Florida perspective. Intercept Ltd., Andover, Hampshire, UK.

Childers, C. C., Aguilar, H., Villanueva, R. & Abou Setta, M. M. 2001: Comparative residual toxicities of pesticides to the predator Euseius mesembrinus (Acari: Phytoseiidae) on citrus in Florida. Flor. Entomol. 84: 391-401.

Duso, C., Pozzebon, A., Capuzzo, C., Malagnini, V., Otto, S. & Borgo, M. 2005: Grape downy mildew spread and mite seasonal abundance in vineyards: effects on Tydeus caudatus and its predators. Bio. Cont. 32: 143-154.

Palevsky, E., Argov, Y., Drishpoun, Y., Childers, C.C. & Gerson U. 2003a: Mite problems on citrus and control strategies in Israel. Proceedings of the International Society of Citriculture IX Congress 2000. Vol. II: 760-763.

Palevsky, E., Argov, Y., Ben David, T. & Gerson U. 2003b: Identification and evaluation of potential predators of citrus rust mite. Sys. Appl. Acarol. 8: 39-48.

Porath, A. & Swirski, E. 1965: A survey of phytoseiid mites (Acarina: Phytoseiidae) on citrus, with a description of one new species. Israel J. Agric. Res. 15: 87-100.

Smith, D. & Papacek, D. F. 1991: Studies of the predatory mite Amblyseius victoriensis (Acarina: Phytoseiidae) in citrus orchards in south-east Queensland: control of Tegolophus australis and Phyllocoptruta oleivora (Acarina: Eriophyidae), effect of pesticides, alternative host plants and augmentative release. Exp. Appl. Acarol. 12: 195-217.

Swirski, E., Ragusa di Chiara, S. & Tsolakis, H. 1998: Keys to the phyotseiid mites (Parasitiformes, Phytoseiidae) of Israel. Phytophaga 8: 85-154.

van Driesche, R.G., Lyon S. & Nunn C. 2006: Compatability of spinosad with predacious mites (Acari: Phytoseiidae) used to control western flower thrips (Thysanoptera: Thripidae) in greenhouse crops. Florida Entomol. 89: 396-401.

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pp. 17

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The genetic variability of the spider mites of Israel Tselila Ben-David1, Sarah Melamed2, Uri Gerson2 and Shai Morin2 1PPIS Ministry of Agriculture and Rural Development, Bet Dagan 50250, Israel; [email protected]; 2Department of Entomology, Faculty of Agriculture, The Hebrew University of Jerusalem, Rehovot 76100, Israel Abstract: Forty-three sequences of the second internal transcribed spacer (ITS2) of nuclear ribosomal DNA were obtained from 16 Israeli species of spider mites (Acari: Tetranychidae). The length of the ITS2 region was species-specific and ranged from 368-542 bp. Two species, Eutetranychus orientalis and Panonychus ulmi, showed extensive polymorphism in their ITS2 base composition (eleven and seven sequences, respectively), while eight species had only one ITS2 sequence. The interspecific variation ranged from 4.4-54.8%, and the intraspecific variation from 0.2 to 2%. Using a 2% threshold for species diagnosis in our data set, 14 out of 16 (87%) species, recognized by morphological criteria, would have been accurately identified. The only exceptions involved the low divergence, 0.011-0.015 (1.1-1.5%), between the closely related Tetranychus urticae and Tetranychus turkestani. Still, these species had fixed alternative rDNA-ITS2 variants, with five diagnostic nucleotide substitutions separating them. The maximum parsimony phylogenetic trees supported the monophyly of the Bryobiinae and the Tetranychinae and that of their genera, with the exception of Oligonychus, where monophyly was rejected. Descriptions of the experimental work can be found in: Ben-David, T., Melamed, S., Gerson, U. & Morin, S. 2007: ITS-2 sequences as barcodes for

identifying and analyzing spider mites (Acari: Tetranychidae) Exp. Appl. Acarol. Accepted: 29 January 2007

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pp.19

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Interplay between omnivory and intraguild predation: thrips spatial dynamics and damage to strawberry Moshe Coll1, Sulochana Shakya1 and Phyllis Weintraub2 1Department of Entomology, The Hebrew University of Jerusalem, Rehovot, Israel; 2Department of Entomology, Agricultural Research Organization, Gilat, Israel Abstract: Omnivory, the feeding on both plant and prey material, and intraguild predation (IGP) are common in ecological systems. For the most part, these complex trophic interactions have been studied separately. Yet some predators are often involved in both omnivory and IGP. We therefore tested these interactions together, in a system that consisted of strawberry plants, western flower thrips (WFT) and two of its predators, the mite Neoseiulus cucumeris and the bug Orius laevigatus. All three are omnivorous consumers that feed on strawberry pollen; WFT damages strawberry fruit, the mite preys on first instar WFT, and the bug feeds on WFT and the mites. We asked: (i) what is the effect of pollen feeding on intensity of IGP? (ii) what is the combined effect of omnivory and IGP on WFT suppression? and (iii) what is the importance of the within-plant distribution of the predators in relation to prey feeding site, for pest-inflicted damage to strawberry fruit?

Results show that (i) predation on N. cucumeris by O. laevigatus was significantly lower in the presence of pollen than in its absence; (ii) significantly fewer WFT were killed by the predators in the presence of pollen than in its absence; (iii) in the presence of pollen, WFT and both predators primarily reside in flowers rather than fruit and leaves; (iv) in the absence of pollen, WFT were recorded primarily on fruits; (v) in the presence of pollen, N. cucumeris is found in the flowers only when O. laevigatus is absent; else, the mites are found on the fruits or leaves; and (vi) when both predators are present, significantly lower fruit damage was observed in the absence of pollen than in its presence.

Taken together, results show that omnivorous feeding and differential response to heterogeneously distributed resources buffer herbivory and IG predatory interactions. This may allow for complex trophic interactions, such as omnivory and IGP, to persist and be common in nature.

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Tetranychus evansi Baker & Pritchard control in European solanaceous greenhouses: facts and perspectives Maxime Ferrero1, Marie-Stéphane Tixier1, Karel Bolckmans2, Serge Kreiter1 1 Montpellier Sup Agro, Laboratoire d'Acarologie, Unité Mixte de Recherche Centre de Biologie et de Gestion des Populations, Bâtiment 16, 2 Place Viala, 34060, Montpellier cedex 01, France; 2 Koppert BV, Veilingweg 17, P.O. Box 155, 2650 AD Berkel en Rodenrijs, The Netherlands Abstract: Tetranychus evansi, a pest of several garden market crops, especially solanaceous ones, is being of great matter for growers for years in Africa and now in southern Europe. The authors discuss here which methods were considered to control T. evansi, and what may be the best path to follow to get this severe pest out of economically important crops. Chemical control is the only possibility to control by now this spider mite, but it has a low efficiency due to some resistances developed by the mite, and lead to environmental and health problems. In European solanaceous greenhouse crops, biological control is widely use and thus the route of finding a biological agent to control T. evansi has been privileged by researchers, without success until recently. Plant resistances, fungi, generalist predators and predatory mites have been tested for more than 25 years but did not lead to field applications. In 2005, a predatory mite, Phytoseiulus longipes has been found to be efficient against the pest populations. Phytoseiulus longipes is a very promising predator and is currently being studied, but may not be the only way to protect solanaceous crops from T. evansi injuries. Keywords: Tetranychus evansi, Phytoseiulus longipes, biological control, solanaceous crops, predators, fungi, plant resistances

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pp. 23-28

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Interaction of the mango bud mite, Aceria mangiferae, with Fusarium mangiferae, the causal agent of mango malformation disease Efrat Gamliel-Atinsky1,2, Stanley Freeman1, Abraham Sztejnberg2, Marcel Maymon1, Eduard Belausov3 and Eric Palevsky4 1Dept. of Plant Pathology, ARO, The Volcani Center, Bet Dagan 50250, Israel e-mail: [email protected]; 2Dept. of Plant Pathology and Microbiology, The Hebrew University of Jerusalem, Faculty of Agricultural, Food and Environmental Quality Science, Rehovot 76100; 3Microscopy Unit, ARO, The Volcani Center; and 4Dept. of Entomology, Newe-Ya'ar Research Center, ARO, Ramat Yishay 30095, Israel Abstract: It has been suggested in the literature that the mango bud mite, Aceria mangiferae, plays an important role in epidemiology of mango malformation caused by Fusarium mangiferae. Current work was designed to study the role of the mites in carrying fungal conidia, vectoring them into the primary penetration sites and possibly assisting fungal penetration and dissemination. Carrying: bud mites were exposed to a gfp-marked fungal isolate. After 24 hours the mites were removed and mounted for microscopic observation. The gfp fluorescing conidia were observed on the examined mites and did not seem to cling to any particular part of the mite’s body. Vectoring: agar plugs bearing either bud mites and/or gfp-marked pathogen were placed on a leaf near an apical bud on potted mango plants according to the following treatment design: 1. bearing bud mites and gfp-marked pathogen; 2. bearing bud mites; 3. bearing the gfp isolate; 4. untreated control. Bud mites were found only in apical buds of treatments 1 and 2 and gfp conidia were found in bud bracts only in treatment 1. Penetration: potted mango plants were inoculated with gfp-marked conidia in two treatments with or without the presence of bud mites. The frequency of infected apical buds was higher in the presence of bud mites. Dissemination: spore and mite traps were placed in a diseased orchard for one year in order to trace possible association between windborne bud mites and windborne conidia. No windborne bud mite bearing conidia was found on the traps, although high numbers of windborne conidia were trapped. These results suggest that the mango bud mite can carry the pathogen conidia on its body, vector it to the apical bud and improve fungal penetration. It also appears that the bud mites do not play a role in aerial dissemination of conidia.

Key words: Aceria mangiferae, Fusarium mangiferae, mango malformation, Eriophyidae, Mangifera indica L. Introduction Mango malformation disease is one of the most destructive diseases of mango and is prevalent in most of the mango production areas worldwide (Kumar et al. 1993, Ploetz et al. 2002). The disease causes malformation of vegetative growth and inflorescence, and causes serious yield loss since malformed panicles do not bear fruit (Kumar et al. 1993, Majumder & Sinha 1972).

Fusarium mangiferae is the causal agent of mango malformation disease (Chakrabarti & Ghosal 1989, Freeman et al. 1999, Noriega Cantu et al. 1999). Little is known about the epidemiology of the disease, location of penetration sites, modes of infection and colonization of

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the plant tissue. One work suggests a wind dispersal mechanism of fungal conidia (Noriega Cantu et al. 1999), but since trapped airborne conidia were not “exclusively attributed to this (the pathogen) species”, more evidence needs to be provided.

Aceria mangiferae was first recorded and described in Egypt (Hassan 1944, Sayed 1946). It is commonly found inside generative and vegetative closed mango buds, in both malformed and non-malformed trees (Sternlicht & Goldenberg 1976). Many reports suggest the involvement of A. mangiferae in mango malformation as both the mite and the fungus are found together within the bud, however it is still not clear that these two organisms actually interact in the epidemiology of this disease (Ploetz et al. 2001). Possible reasons for the lack of information can be related to problems with identification of the pathogen and the lack of molecular tools for tracking the fungus.

In this study, we studied four possible interactions between the mango bud mite and the pathogen: 1. carrying F. mangiferae conidia on its body; 2. vectoring the pathogen’s conidia into the apical buds; 3. assisting conidial penetration to plant tissue; 4. aerial dissemination of conidia. Materials and methods Mites bearing the pathogen Mites collected from infested buds in the orchard were exposed to a gfp (green fluorescent protein)-marked isolate, using two different methods. In the first method 20 mango bud bracts bearing approximately 100 mites were dipped in a gfp-marked F. mangiferae suspension of 106 conidia per ml for 5 seconds. After allowing the bud bracts to dry, mites were removed from the bracts with an ultra fine paint brush and mounted on double-sided sticky tape. In the second method 30 mites were placed on an agar plug (5 mm2) which was inoculated 48 hours beforehand with the gfp-marked isolate. After 24 hours the mites were removed and mounted for microscopic observation. Images were acquired using a confocal laser-scanning microscope system OLYMPUS IX81. Confocal images were obtained via a PLAPO 40X WLSM immersion objective lens at an excitation wavelength of 488nm (Argon laser), a BA515-525 emission filter for gfp and BA660IF emission filter for auto-fluorescence. Transmitted-light images were acquired using Nomarski differential interference contrast. Vectoring the pathogen into apical buds The experiment was performed on potted mango plants, in a controlled environment growth chamber with 25 ± 2°C and 14:10 L:D. The plants were fumigated twice with dichlorvos (30ml/100m2; Makhteshim Chemical Works Ltd, Beer Sheva, Israel), two weeks before the beginning of the experiment to ensure that they were void of mites or insects. The base of the stem was ringed with a sticky barrier to prevent infestation by ambulant arthropods. Each plant was placed in a disinfected plastic cage and was treated with one of the following four treatments: treatment 1. 100 mites were placed on two 5 mm2 agar plugs with the gfp-marked isolate. The agar plugs bearing bud mites and the gfp-marked pathogen were then placed on a leaf, approximately 5 cm away from an apical bud; treatment 2. 100 mites were placed on two 5 mm2 agar plugs without the fungus and then placed near an apical bud as described above; treatment 3. two 5 mm2 agar plugs with the gfp-marked isolate were placed near an apical bud; treatment 4. untreated control. Four apical buds were inoculated in each treatment and the experiment was repeated 5 times. Two days following inoculation the apical buds were inspected under a stereomicroscope and the bud mites were counted. Then, the gfp-marked conidia (if present) were washed from the bud bracts and plated on a potato dextrose agar (PDA) selective

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media amended with 50 µg/ml hygromycin. After 5 days, the gfp-marked colonies were enumerated on the plates. Assisting fungal penetration Quantitative evaluation – The experiment was performed on potted plants in a controlled environmental chamber (as described above). Each plant was placed in a disinfected plastic cage and was treated with one of the following two treatments: treatment 1. 40 apical buds were inoculated with a 106 gfp-marked conidial suspension; treatment 2. 40 apical buds were inoculated as in treatment 1 and also with bud mites that were collected from the orchard (50 mites per bud). Three weeks post-inoculation apical buds were separated into bracts, surface-sterilized 10 seconds in 70% ethanol and 3.5 minutes in 0.1% NaClO3, and placed on PDA selective media. Fungal growth was evaluated after 5 days and two parameters were measured: 1. Frequency of infected buds was evaluated calculating the percentage of infected buds in the treatment, using a Pearson statistical test to compare the two treatments (P < 0.05); 2. Severity of infection was measured by calculating the average number of infected bracts per bud and the means of the two treatments were compared using Tukey-Kramer analysis (P < 0.05). Qualitative evaluation – Apical buds that were inoculated with both the pathogen and the mites, were separated into bracts and inspected with a confocal microscope (as described above) and Jeol (Tokyo, Japan) scanning microscope 5410 LV. Aerial conidial dissemination Both fungal conidia and mango bud mites were trapped using the following trapping methods: 1. Spore dissemination in the Volcani mango orchard was monitored using a BurkardTM volumetric spore trap (Burkard Manufacturing Co Ltd, Rickmansworth, England) placed in the orchard, sucking air at a speed of 10 liters per minute on adhesive coated transparent plastic, continuously, for periods of seven days. The adhesive tape was then washed and mounted on the selective Nash medium for enumerating F. mangiferae colonies. 2. Petri dishes containing the selective Nash medium for F. mangiferae were opened and exposed to the orchard’s environment overnight, then brought back to incubate in the lab and examined for the presence of F. mangiferae colonies. 3. For monitoring wind blown mites in the orchard, a freely rotatable wind trap made of a PVC pipe (9-cm in diameter) was constructed and mounted on a pole attached to a wind vane (Duffner et al. 2001). The pipe’s floor was covered with 70 sticky slides that were replaced monthly, then inspected under a stereo-microscope for the presence of mites and placed on Nash selective media for pathogen detection. Results and discussion Mites bearing the pathogen Mites from bracts removed from the conidial suspension did not bear conidia. However, gfp-marked conidia were observed on mites that were placed on agar plugs bearing the marked strain (Figure 1). Conidia of the pathogen did not seem to cling to any particular part of the mite’s body. Therefore, our observations indicate that mites can bear conidia of the pathogen.

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Figure 1: Mango bud mite, Aceria mangiferae, bearing gfp-marked conidia of Fusarium mangiferae, the causal agent of mango malformation disease. Vectoring the pathogen into apical buds Bud mites were found in 25% and 35% of the inoculated buds of treatments 1 (inoculation with bud mite and with gfp-marked conidia) and 2 (inoculation with bud mites), respectively, showing clearly that the bud mites could orientate themselves from the adjacent leaves to the apical bud. No bud mite was found in treatments 3 (inoculation with gfp-marked conidia) and 4 (untreated control) which confirms that the plants used in these experiments were not infested prior to the initiation of the experiment. The gfp-marked conidia were found in bud bracts of 25% of the inoculated buds in treatment 1 and not in buds of other treatments. This demonstrates that the mango bud mite is able to carry F. mangiferae conidia on its body and transfer them into the apical bud. Assisting fungal penetration A significantly higher degree of apical buds were infected in the gfp + mites combined treatment (50% in the gfp-inoculated treatment compared with 82% infected buds in the combined treatment - figure 2A). Moreover, the infection severity was also higher in the combined treatment where more bracts per bud were infected (figure 2B). This suggests that the bud mite may play a role in pathogen penetration of the bud tissue, perhaps through the feeding wounds it creates on the bud bracts, thereby facilitating conidial penetration. Microscopic observations supported our finding by demonstrating the physical proximity and the actual contact between the two organisms. Bud mites were observed touching gfp-glowing hyphae and conidia using the confocal microscope. SEM (scanning electron microscope) observations also illustrated bud mites bearing hyphae and conidia on their body. Aerial conidial dissemination Conidia of F. mangiferae were trapped successfully using both trapping methods. An annual peak of dissemination was found in the spring/early summer months (figure 3). Similar results were obtained with the BurkardTM trap where higher numbers of conidia were caught early in the summer months (May/June) declining towards the end of the summer/beginning of autumn. A. mangiferae was trapped throughout the season, but no fungal growth was detected after placing the slide-bearing mites on selective media. Our findings imply that conidia can reach the bud independently of the bud mite and thus the latter does not seem to play a role in the windborne dissemination of the pathogen.

20 um

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Figure 2: Frequency and severity of infection in mango apical buds inoculated with gfp-marked conidia of Fusarium mangiferae (treatment 1-GFP) or with gfp-marked conidia and with bud mites, Aceria mangiferae (treatment 2: GFP + mites). Figure 2A: Frequency of infected buds with/without the presence of bud mites. Figure 2B- Frequency of infected bracts per bud with/without the presence of bud mites.

Figure 3: Air-born conidia of Fusarium mangiferae trapped on selective media plates exposed overnight to Volcani orchard’s conditions.

Results from this study suggest that A. mangiferae can carry F. mangiferae conidia on its

body. Furthermore, A. mangiferae can vector the pathogen’s conidia into apical buds, which serve as penetration sites for the pathogen (unpublished data). The mite can also improve fungal penetration perhaps via its feeding wounds, and finally, it also appears that the bud mites do not

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play a role in aerial dissemination of conidia. These results support involvement of A. mangiferae in mango malformation epidemiology (Ploetz et al., 2001). Acknowledgements This research was made possible through support provided by the Bureau for Economic Growth, Agriculture, and Trade, U.S. Agency for International Development, under the terms of the Middle East Regional Cooperation Program Award No. TA-MOU-02-M21-030. We would also like to thank Mr. Martin Berkeley for his assistance.

References Chakrabarti, D. K. & Ghosal, S. 1989: The disease cycle of mango malformation induced by

Fusarium moniliforme var. subglutinans and the curative effects of mangiferin-metal chelates. J. Phytopathol. 125: 238-246.

Duffner, K., Schruft, G. & Guggenheim, R. 2001: Passive dispersal of the grape rust mite Calepitrimerus vitis Nalepa 1905: (Acari, Eriophyoidea) in vineyards. Anzeiger-fur-Schadlingskunde 74 :1-6.

Freeman, S., Maimon, M. & Pinkas, Y. 1999: Use of GUS transformants of Fusarium subglutinans for determining aetiology of mango malformation disease. Phytopathology 89: 456-461.

Hassan, A. S. 1944: Notes on Eriophyes mangiferae. Bull. Soc. Fouad Ier Entomol. 27 :179-182. Kumar, J., Singh, U. S. & Beniwal, S.P.S. 1993: Mango malformation: one hundred years of

research. Ann. Rev. Phytopathol. 31: 217-232. Majumder, P.K. & Sinha, G.C. 1972: Studies on the effect of malformation on growth, sex ratio,

fruit set and yield of mango. Acta Hort. 24: 230-234. Noriega Cantu, D.H., Teliz, D.G., Mora Aguilera, J., Rodriguez Alcazar, E., Zavaleta Mejia, G.,

Otero Colinas, E. & Campbell, C.L. 1999: Epidemiology of mango malformation in Guerrero, Mexico, with traditional and integrated management. Plant Dis. 83: 223-228.

Ploetz, R., Zheng, Q., Vazquez, A. & Sattar, M.A.A. 2002: Current status and impact of mango malformation in Egypt. Int. J. Pest Manag. 48: 279-285.

Ploetz, R.C., Summerell, B.A., Leslie, J.F., Backhouse, D., Bryden, W.L. & Burgess, L.W. 2001: Malformation: a unique and important disease of mango, Mangifera indica L. In Fusarium: Paul E. Nelson Memorial Symposium, 233-247 St. Paul; USA: American Phytopathological Society (APS Press).

Sayed, M.T. 1946: Aceria mangiferae, nov. spec. Bull. Soc. Fouad Ier Entomol. 30: 7-10. Sternlicht, M., & Goldenberg, S. 1976: Mango eriophyid mites in relation to inflorescence.

Phytoparasitica 4: 45-50.


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A tribute to the late Professor Eliahu Swirski, our foremost agricultural acarologist Uri Gerson Department of Entomology, Faculty of Agriculture, Hebrew University of Jerusalem, Rehovot 76000, Israel Abstract: The late Professor Eliahu Swirski studied the aphids of the Middle East, the local pests of subtropical fruit (date palms, avocado and citrus), and pestiferous mites along with their predators. He followed the biology of the citrus rust mite in the orchard and in the laboratory and looked for its natural enemies. He collected and described many predatory mites of the family Phytoseiidae from Israel, Italy, Greece, Turkey and Kenya. The biology of several indigenous predators (including Amblyseius swirskii) and their susceptibility to pesticides were also studied. Key words: date palm, avocado, citrus, Phytoseiidae The late professor Eliahu Swirski was an agricultural entomologist par excellence, whose scientific contributions were in three main areas: aphids, pests of subtropical crops and the Phytoseiidae. He began in the 1950’s, with studies on the biology, control and systematics of aphids. This early research culminated in a book on the Aphidoidea of the Middle East (Bodenheimer & Swirski 1957). Swirski maintained his interest in these insects throughout his career, and in 1999 summed up the aphids of Israel (Swirski & Amitai 1999). Concurrently Swirski investigated and published on the biology and control of the pests of avocadoes, date palms and citrus. He was a tireless proponent of integrated pest control (IPM) and led the teams that introduced exotic natural enemies for the control of pests that affect these crops. His efforts were recognized when, 1994, he was awarded the highest Israeli scientific honor, namely the Israel Prize. In 2002 Swirski published (with his long-time colleagues Manes Wysoki and Yehonatan Izhar) the monumental “Subtropical Fruits Pests in Israel”, the culmination and summing up of more than 40 years of research.

His interest in citrus pests naturally led him to the citrus rust mite (Phyllocoptruta oleivora), then (early 1950’s) as now, a major pest of citrus in Israel. At that time (the mid-1950’s) little was known about this pest and basic research tools had to be developed. Special methods of culturing the pest (which is an obligatory parasite) in the laboratory were devised, whether on citrus leaves or on small, raw fruit (Swirski & Amitai 1958). The population fluctuations of the mite were followed in the field, and the effect of various pesticides for its control was studied (Swirski et al. 1969). Concurrently, Swirski’s belief that only IPM programs could alleviate the damage caused by this mite naturally led him to seek and assay its natural enemies. Thus began his 40-years long studies of the predatory mites of the family Phytoseiidae.

During 1959 he began to collect phytoseiid mites from all over Israel, an effort that would continue for many years. By 1961 he began to publish on the phytoseiid fauna of Israel (Swirski & Amitai 1961), describing many new species along the way. His collections were not, as are

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those of many economic acarologists, restricted to crop plants; rather he looked at and obtained the mites from all plants in an area. Thus he collected from the very different climatic regions of Israel, for instance from the Dead Sea region (Swirski & Amitai 1985) or from around the Sea of Galilee (Swirski & Amitai 1990). In 1960 he visited Japan and Hong-Kong in order to search for the natural enemies of the citrus rust mite, a visit that resulted in the description of a new genus and several new species (Swirski & Shechter 1961). In addition Swirski also contributed to an understanding of the phytoseiid fauna of Italy (Ragusa & Swirski 1976), Greece (Swirski & Ragusa 1976), Turkey, The Philippines and even Kenya (Swirski & Ragusa 1978), describing new species along the way. His last publication (Swirski et al. 1998) about the Phytoseiidae was an annotated and illustrated key to all species found in Israel.

Along with his systematic studies, Swirski investigated the life history of several promising indigenous phytoseiids, their diets and susceptibility to pesticides. Of special interest are the two contributions (Swirski et al. 1967, Ragusa & Swirski 1977) about the biology of Amblyseius swirskii. This species, which is the dominant phytoseiid in citrus groves along the coastal plain of Israel, developed well on spider mite, false spider mite and citrus rust mite diets, as well as on moth eggs and the pollen of some plants. The addition of honeydew from several scale insects (Coccoidea) enhanced the fecundity of the predator. Swirski also studied the effect of various pesticides on these predators (Swirski et al. 1969), in order to find and recommend chemicals that would have the least disruptive effect on the phytoseiids, thus promoting the implementation of IPM programs.

Among other topics, Swirski also studied the effect of Phytoseiulus persimilis on spider mites damaging strawberry and banana, and the natural enemies of the invading citrus pest, Panonychus citri. (Swirski et al. 1986).

Eliahu Swirski was highly respected in the international community of agricultural entomologists, and several species (in addition to the above-mentioned Amblyseius swirskii) were named in his honor. Furthermore, the new genus Swirskiseius Denmark and Evans was dedicated to him.

Swirski’s acarological publications fall into three groups, quite similar in size: those dealing with mite (pest and beneficial) biology, with the control (chemical, integrated and biological) of pest mites and those devoted to the systematics of the Phytoseiidae (Figure 1). This serves to emphasize Swirski’s approach, which may briefly be summarized in two sentences.

1. One cannot study the biology of mite pests or their natural enemies without understanding their systematics.

2. One cannot study the systematics of mite pests or their natural enemies without understanding their biology.

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Figure 1. Eliahu Swirski’s contributions to Acarology: number of articles that deal with the control, biology and systematics of mites. Acknowledgements I thank Amos Rubin for suggesting this talk, Manes Wysoki for much valuable data and Danny Blumberg for visual material. References Bodenheimer, F.S. & Swirski, E. 1957: The Aphidoidea of the Middle East. The Weizmann

Science Press of Israel. Ragusa, S. & Swirski, E. 1976: Notes on predaceous mites of Italy, with a description of two new

species and of an unknown male (Acarina: Phytoseiidae). Redia 59: 179-196. Ragusa, S. & Swirski, E. 1977: Feeding habits, post-embryonic and adult survival, mating,

virility and fecundity of the predaceous mite Amblyseius swirskii (Acarina: Phytoseiidae) on some coccids and mealybugs. Entomophaga 22: 383-392.

Swirski, E. & Amitai, S. 1958: Contribution to the biology of the citrus rust mite (Phyllocoptruta oleivora Ashm.). A. Development, adult longevity and adult life cycle. Ktavim 8: 189-207.

Swirski, E. & Amitai, S. 1961: Some phytoseiid mites (Acarina: Phytoseiidae) of Israel, with a description of two new species. Israel J. Agric. Res. 11: 193-202.

Swirski, E. & Amitai, S. 1985: Notes on phytoseiid mites (Mesostigmata: Phytoseiidae) from the Dead Sea region of Israel. Israel J. Entomol. 19: 181-192.

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Swirski, E. & Amitai, S. 1990: Notes on phytoseiid mites (Mesostigmata: Phytoseiidae) from the Sea of Galilee region of Israel, with a description of a new species of Amblyseius. Israel J. Entomol. 24: 115-124.

Swirski, E. & Amitai, S. 1999: Annotated list of aphids (Aphidoidea) in Israel. Israel J. Entomol 33: 1-120

Swirski, E., Amitai, S. & Dorzia, N. 1967: Laboratory studies on the feeding, development and reproduction of the predaceous mites Amblyseius rubini Swirski and Amitai and Amblyseius swirskii Athias (Acarina: Phytoseiidae) on various kinds of food substances. Israel J. Agric. Res. 7: 101-119.

Swirski, E., Dorzia, N., Amitai, S. & Greenberg, S. 1969: Trials on the control of the citrus rust mite (Phyllocoptruta oleivora Ashm.) with four pesticides, and on their toxicity to predaceous mites (Acarina: Phytoseiidae). Israel J. Entomol. 4: 145-155.

Swirski, E., Gokkes, M. & Amitai, S. 1986: Phenology and natural enemies of the citrus red mite, Panonychus citri (McGregor) in Israel. Israel J. Entomol. 20: 37-44.

Swirski, E. and Ragusa, S. 1976: Notes on predaceous mites of Greece, with a description of five new species (Mesostigmata: Phytoseiidae). Phytoparasitica 4: 101-122.

Swirski, E. and Ragusa, S. 1978; Three new species of phytoseiid mites from Kenya (Mesostigmata: Phytoseiidae). Zool. J. Linnean Soc. 63: 397-409.

Swirski, E., Ragusa di Chiara, S. and Tsolakis, H. 1998; Keys to the phytoseiid mites (Parasitiformes, Phytoseiidae) of Israel. Phytophaga 8: 85-154.

Swirski, E. and Shechter, R. 1961: Some phytoseiid mites (Acarina: Phytoseiidae) of Hong-Kong, with a description of a new genus and seven new speciers. Israel J. Agric. Res. 11: 97-117.

Swirski, E., Wysoki, M. and Izhar Y. 2002: Subtropical Fruits Pests in Israel. Fruit Board of Israel. 285 pp.


pp. 33-40

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Acaricides and the integrated control of plant feeding mites

Richard M. GreatRex Syngenta Bioline Ltd., Telstar Nurseries, Holland Road, Little Clacton, Essex, CO16 9QG UK Abstract: In discussing compatibility of acaricides or other pest control products within Integrated Pest Management, we need first to understand how to interpret the information which is available. Laboratory and field results often differ, and many things can contribute to this difference. The interaction between the product and the plant, leaf expansion and degradation, application technique and equipment, and application timing can all affect compatibility. Examples of how these various parameters influence compatibility are given for abamectin. Details are given of some of the most recently registered products, and also some products which are still in the developmental pipeline.

The use of products is not only driven by efficacy: politics and public opinion are all driving a change in their use. Government policy throughout Europe is to reduce pesticide use. Supermarkets react to public opinion and perceived risk by restricting use of products on the fruit and vegetables which they will sell: even registered products can be excluded. The rate of registration of new active ingredients is also low: there are relatively few new chemical classes and modes of action in development. In this situation, it is critically important that we maintain the viability of the few products which are available. Careful use and management of these products within IPM systems is one way to do this. Keywords: acaricides, compatibility The BCPC Pesticide Manual 14th Edition (Tomlin, 2006) lists 85 active ingredients as acaricides. Of these, a substantial number are older products. Some have been or will be lost in Europe because companies will not support the re-registration costs for products which no longer have patent protection. Others will not be available in all countries, nor on all crops on which we might wish to use them. Commercial roll-out of a new product will focus on those markets with the highest perceived potential value in order to provide the shortest period of return on the investment in product discovery and development. Smaller markets and less valuable crops will follow, and products may never be registered in smaller countries, or on minor crops, where the market size does not justify the investment in registration of a product. Cotton, maize and cereals will probably get the first registrations, fruit, vegetables and ornamental crops will be further down the list. Thus, what users see as a new product in one country may have been in use elsewhere for many years. In deciding what to discuss then, we need to understand that not all of the products mentioned will be widely available, and that there are many reasons for this. We can take a slightly different approach and talk more about some general principles, and the context in which products will be used. What is the growers perception of products, and why do they chose to use them? Are they concerned solely with control of a single pest, or with management of the pest and disease complex on their crop? Do they care about indigenous natural enemies or introduced biological control agents, and if so, what value do they assign to these? What characteristics of a

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product make it compatible with natural enemies, and how do we interpret the published and anecdotal evidence which abounds on the subject of compatibility?

The majority of growers are business people. They grow a crop in order to earn a living for themselves, their families and perhaps their communities. They use acaricides, insecticides and fungicides to secure that living: their primary concerns are with yield and quality. In recent years, concerns about the environmental impact and the health implications of pesticide use have grown, and adverse publicity about this is driving changes in public opinion. This in turn has an impact upon production practices. In this situation, growers will of course use biological products in exactly the same way as conventional crop protection products. Their customers will positively encourage this, and may demand it. The trap for the grower exists where no biological alternatives are available, or where those alternatives are found to be less reliable or to have lower efficacy than conventional products. By using such products, the grower takes a financial risk. If yield or quality are lower, then the produce may be rejected, or have lower value. Any move away from conventional towards alternative products for pest control therefore requires the growers to have confidence. They must believe that the gain from making the change is greater than the risk incurred by doing so, or that there is no penalty. The manufacturers of alternative products, and consultants who promote their use, must be sure that that confidence is justified. If products perform poorly, then confidence will be lost, and growers will be unwilling to take that financial risk again. In order to provide relevant advice, we need to consider which is the most appropriate method of control for any given pest, and which combination of controls provides the grower with the greatest security. That requires an acceptance, irrespective of ideology, that the best option for a given pest may be a conventional product. The challenge then lies in understanding how that product can be integrated into an overall programme of pest and disease control. In order to do this, an understanding of the product is essential. How does it interact with the plant and the environment following application? What is the influence of application method? One of the older acaricides and insecticides can provide some insights here. Abamectin (Vertimec®, Agrimec®, Avid® or Dynamec®) is a naturally derived product. Fermentation of Streptomyces avermitilis produces 8 natural Avermectins: A1a, A1b, A2a, A2b, B1a, B1b, B2a, B2b. Abamectin, the active ingredient in Vertimec®, is a purified product containing 80% Avermectin B1a and 20% Avermectin B1b. On application to leaves, a proportion of the product is absorbed through the cuticle into the leaf lamina, where it is stable. It is locally translaminar, with limited lateral spread, and has no significant systemic activity. Residue remaining on the leaf surface is subject to rapid photo-degradation. Bull et al. (1984) applied abamectin to the surface of mature cotton leaves in a field crop, and found that only 5% of the applied product remained un-degraded on the leaf surface 4 days after application. Following application, therefore, the product moves into the leaf, where it will continue to provide protection against leaf feeding insects and mites. Leaf surfaces, at least in high light levels, rapidly become safe to beneficials as the residue degrades. Rapid leaf expansion in some crops further dilutes leaf surface residues (and of course also the active ingredient present in treated leaves). The lack of systemic activity means that new foliage contains no active ingredient; it is therefore unprotected from pests, but safe to all beneficial arthropods. Of course, abamectin also has strong contact activity, and direct contact will kill many predatory mites and insects. Nevertheless, careful timing and application make it a useful tool within ICM programmes. Application to the upper leaf surface avoids direct contact with

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predatory mites, whilst the translaminar activity ensures that pest mites feeding on the lower leaf surface are killed. Spraying specific parts of a crop can control small ‘emigrant’ populations of pests whilst leaving predators in core pest populations unscathed. This can help to deal with the problem of predators remaining within the pest focus while pests migrate to form new colonies. Pest populations spread in a wave from the initial colony, and damage continues to occur despite the presence of predators. The predators are always following the pest, but never quite providing control. Controlling that outward spread of the pest allows the predators time to catch up – they finish the available food and migrate outwards to find only small numbers of pests, rather than major colonies which might otherwise have developed. Spaying a horizontal band at the top of the crop protects the high value flowering stems from damage, but permits predators to survive in the canopy of foliage which exists below this. This technique was advocated by Sanderson and Zhang (1995) in roses, and has been used with great success in rose crops in California and elsewhere. This is perhaps contrary to conventional advice, especially for contact acting products. Good spray coverage and maximum contact with the target pest would be preferred. Air-blast sprayers or ruffler bars would be used to move the foliage and get better penetration of the spray into the canopy. This basically ignores the potential of predatory mites and insects which may already be present, or at best requires that the product is physiologically selective towards those beneficials. In an ICM context, we might propose that poor spray coverage can be an advantage. It will leave pest reservoir populations, but also reduce the impact on predatory mites, and in the long term provide a better population balance and enhanced overall control. There are thus several methods which permit a product which is apparently damaging towards predators to be successfully integrated. Many of these illustrate why field data on compatibility often give a more benign view of a product than do laboratory studies: the laboratory study does not provide the opportunity for residue degradation and dilution, nor provide untreated refuges. Having said that, even field and semi-field data can be misinterpreted. A simple reduction of predators or parasites on a treated crop does not imply a direct effect. If the target of the treatment is the food source of the affected predator, then the reduction may be a simple consequence of resource limitation. The real measure in this case has to be the ratio between the remaining pests and their predators or parasites. This has been demonstrated for abamectin in numerous internal trials: predator to prey ratios during the season were lower in plots treated early post petal fall than in control plots, although predator numbers were lower. We can conclude that the ratio has been changed in favour of long term suppression by the predatory mites. Even where the product applied does not target the obvious host of a predatory mite, we might expect to see secondary effects on populations, or on overall control. Duso et al. (2003) found a positive correlation between populations of Amblyseius andersoni and Downy Mildew (Plasmopara viticola) on grape vines. Populations increased markedly as the disease spread, numbers were highest on infected leaves, and numbers were positively correlated with the extent of the symptoms on those leaves. There was clear evidence from electrophoretic studies that individuals of A. andersoni were feeding upon the fungus. We should expect, then, that application of a fungicide to control Downy Mildew will decrease populations of A. andersoni, not because it is directly toxic, but because it reduces the supply of one major food.

On a similar note, there have been reports of resurgence of Tetranychus populations following treatment with imidacloprid and other neonicotinoid insecticides. There are conflicting reports on the causes of this problem. Several authors have found a direct effect upon some

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species of predatory mites, but not on others. James & Vogele (2002) found no significant mortality to Typhlodromus dossei or T. doreenae from field application rates; Mizell & Sconyers (1992) found no significant impact on Neoseiulus collegiae, or Phytoseiulus macropilis; Poletti et al. (2007) found low toxicity to adults of P. macropilis and Neoseiulus californicus from residues of imidacloprid, thiamethoxam or acetamiprid, but they did detect a difference in the consumption of treated prey; James (2003) found high levels of toxicity to Galendromus occidentalis and Neoseiulus fallacis, and lower levels of toxicity to A. andersoni; Castagnoli et al. (2005) report a decreased fecundity of N. californicus, and at the same time an increase in the fecundity of T. urticae; James (1997) showed repellency of residues of imidacloprid to Amblyseius victoriensis, but at the same time an increase in fecundity of the predator by up to 54%. James & Price (2002) have found a significant increase in fecundity from T. urticae treated with imidacloprid either as a foliar application or as a drench, with 10-19% more eggs during the first 12 days, and a lifetime increase of up to 26%. Villanueva and Walgenbach (2006) found similar changes. In contrast, Ako et al. (2006) found no similar increase in any of four strains of T. urticae tested. It is difficult to draw any firm conclusion from this mass of conflicting data. It seems likely that the test methodology, the application method and the species of predator are all significant factors: it also seems likely that field reports of resurgence of spider mites could be interpreted as a directly negative effect upon the predator complex, but may be due instead to an enhanced reproductive rate in T. urticae. As was stated earlier, The Pesticide Manual 14th Edition lists 85 active ingredients with acaricidal activity. It is clearly not possible or desirable to discuss all of these here. Many of the products are well known and characterised within IPM systems. The discussion is therefore limited to some of the more recently launched products, or those which are still in development. Information on the products has been variously extracted from manufacturers data sheets, web sites and published papers. Acequinocyl was developed by Agro Kanesho and first registered in Japan and Korea in 1999. It is being developed under various trade names in the US (www.arysta-na.com) and Europe for control of all stages of Tetranychidae on top fruit (Hulin et al. 2005), citrus, ornamental and vegetable crops and tea. Some sources suggest activity against Eriophyidae and Tarsonemidae, but this is not claimed in the marketing literature. The chemistry and mode of action are novel. It acts primarily by contact and ingestion, with no translaminar or systemic effects. It apparently has long residual activity, but is also reported to be rapidly photodegraded. It is reported to be safe to Amblyseius andersoni and Typhlodromus pyri, but moderately harmful to Heteroptera, Hymenoptera and Coleoptera 48 hours after application (Hulin et al. 2005). Reports on toxicity to predatory mites vary. Saenz-de-Cabezon & Zalom (2006) found that it greatly reduced longevity in Galendromus occidentalis, whilst it is reported to be safe to Amblyseius cucumeris (Kim et al, 2005) and Phytoseiulus persimilis (Yoo & Kim 2000, Kim & Yoo 2002) and to Amblyseius womersleyi (Kim & Seo 2001, Amano et al. 2004). Bifenazate was first launched in Japan in 2001, and subsequently in Europe and the US during 2002/2003. It is active against mobile stages by contact and feeding, with some ovicidal activity, but with no translaminar or systemic effect. The manufacturer states that it is effective against Tetranychus spp., Panonychus spp., and Oligonychus spp, and Steneotarsonemus pallidus, but not against ‘broad mite’ nor against Eriophyidae. The product literature also states that the product is safe to Phytoseiid and Stigmaeid mites and to the generalist predator Chrysoperla carnea. In the US, only two applications are permitted per crop per season, and only

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a single application is recommended before rotating the next two applications with acaricides with different modes of action. Etoxazole was first registered in Japan in 1999, and in Europe and the US in 2002. It is currently commercialised against Tetranychid mites on fruit, vegetables and tea, with a side effect on aphids. It has contact activity against eggs, and some moulting inhibition of mobile stages. It is generally reported to be safe to predatory mites (e.g. Amano et al. 2004, Kim et al. 2005)

Chlorfenapyr was launched in Japan in 1995 against Tetranychid and Eriophyid mites. It also has insecticidal activity against thrips and Lepidoptera, including Diamond Back Moth. It is reported to be safe to Phytoseiulus persimilis in comparison with other acaricides (e.g. Kim & Yoo, 2002) but there are no indications of planned registration in Europe or the US. Cyenopyrafen (NC 512) is a new active ingredient reported in Compendium of Pesticide Common Names, and also listed on the IR4 Project website. First expected launch is in Japan, but no other information is available. Cyflumetofen is a new active ingredient launched in Japan in 2006 under the trade name Danisataba. It is listed on the IR4 Project web site as an acaricide of interest on fruit and vegetable crops. Target pests are Tetranychus spp. and Panonychus spp. It is reported to have low efficacy against Eriophyidae. No information is available on compatibility Fluacrypirim was launched in Japan in 2002 for control of Tetranychus urticae, T. kanzawi, Panonychus ulmi and P. citri on apple and pear crops. It is active against adult and juvenile mites by contact and feeding activity. It is reported to be harmless to Stethorus japonicus and Scolothrips takahashi (Mori & Gotoh, 2001). There is no further compatibility information available. Milbemectin. Like abamectin, this is a product of bacterial fermentation. First launched in Japan in 1990, it has subsequently been launched in Europe (2004) and the US (2005). It is active against mobile stages of Tetranychid mites by contact and feeding. It is in the same class, and has the same mode of action, as abamectin, but is reported to have less translaminar activity. Pyrimidifen was launched in Japan in 1996. It has activity against spider mites and Eriophyidae, and insecticidal activity against whitefly and aphids. Active by contact and feeding, with no translaminar or systemic effect. Mori & Gotoh (2001) found no significant negative effects upon Stethorus japonicus or Scolothrips takahashi. It is harmful to predatory mites. Spirodiclofen is the first in a new class of product from Bayer, the tetronic acids or ketoenoles, all of which are reported to act by inhibition of lipid synthesis (Maeyer et al. 2002b, Bretschneider et al 2003). It was first launched in Europe in 2003. It has activity against Tetranychidae, Eriophyidae, (Sauzay & Ledoux, 2005) and Brevipalpus, and also has insecticidal activity against Psyllidae and Scale Insects (Maeyer et al. 2002a, 2005). It is reported to be slightly harmful to Phytoseiidae, but populations in orchards rapidly recover following application and long term control is maintained at good levels (Sauzay & Ledoux, 2005). Asihara et al. (2006) report good selectivity towards Phytoseiulus persimilis and Amblyseius andersoni. Spiromesifen is the second of the ketoenoles products launched by Bayer. First reported in 2002 (Nauen et al. 2002) and launched in the UK in 2003 and in Europe 2005. The main label claims are for whitefly control, but the product is widely promoted and used for spider mite control. It is mainly active against juveniles, but gives a strong reduction in adult fecundity. It has contact and translaminar activity and good rain-fastness. Target crops are vegetables, ornamentals, fruit, corn and cotton. There is also insecticidal activity against Psyllidae. It is relatively slow acting against all of the target pests, but reported to be selective towards predatory insects (Kavitha, 2006)

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Spirotetramat is the third Ketoenole product announced by Bayer, with no current registrations. Commercial launch is expected in 2008 to 2009. (Bayer 2005). It is primarily insecticidal, with effects against whitefly and aphids and some activity against thrips species, but poor efficacy against Frankliniella spp. and Thrips tabaci. As an acaricide, current information indicates that it is active against Tetranychus. As with the other ketoenoles, it inhibits lipid synthesis and is reported to be relatively slow acting.

References Ahn, K.S., Lee, S.Y., Lee, K.Y., Lee, Y.S. & Kim, G.H. 2004: Selective toxicity of pesticides to

the predatory mite, Phytoseiulus persimilis and control effects of the two-spotted spider mite, Tetranychus urticae by predatory mite and pesticide mixture on rose. Korean J. Appl. Entomol. 2004 43: 71-79.

Ako, M., Poehling, HM., Borgemeister, C. & Nauen, R. 2006: Effect of imidacloprid on the reproduction of acaricide-resistant and susceptible strains of Tetranychus urticae Koch (Acari: Tetranychidae). Pest Manag. Sci.. 625: 419-424.

Amano, H., Ishii, Y. & Kobori., Y. 2004: Pesticide susceptibility of two dominant phytoseiid mites, Neoseiulus californicus and N. womersleyi in conventional Japanese fruit orchards. J. Acarol. Soc Japan 13: 65-70.

Ashihara, W., Kondo, A, Shibao, M., Tanaka, H, Hiehata K., & Izumi, K. 2006: Comparative toxicity of spirodiclofen and lambdacihalotrin to Tetranychus urticae, Tarsonemus pallidus and predatory mite Amblyseius andersoni in a strawberry site under field conditions. Ecology & Control of Eriophyid Mites Injurious to Fruit Trees in Japan Agronomy Research, Special Issue 4: 317-322.

Bayer. 2005: Press Release Monday, September 5. Bretschneider, T.; Benet-Buchholz, J.; Fischer, R.; Nauen, R 2003: Spirodiclofen and

Spiromesifen - novel acaricidal and insecticidal tetronic acid derivatives with a new mode of action. CHIMIA Intern’l J. Chem., 57: 697-701.

Bull, D.L., Ivie, W., MacConnell, J.G., Gruber, V.F., Ku, C.C., Arison, B.H., Stevenson, J.M. & VandenHeuvel, W.A. 1984: Fate of Avermectin B1a in soil and plants. J. Agric. Food Chem. 32: 94-101.

Castagnoli, M., Liguori, M., Simoni, S. & Duso, C. 2005: Toxicity of some insecticides to Tetranychus urticae, Neoseiulus californicus and Tydeus californicus. BioControl 50: 611-622.

Duso, C. Pozzebon, A. Capuzzo, C. Bisol, P. M. Otto, S. 2003: Grape downy mildew spread and mite seasonal abundance in vineyards: evidence for the predatory mites Amblyseius andersoni and Typhlodromus pyri. Biol. Cont. 27: 229-241.

Hulin, L, Lebrun-Destombres, M, Rouas, G and Kinoshita, S. 2005: Acequinocyl, a new compound for control of mites on top fruit. AFPP Second International Conference on Mites in Crops, Montpellier, France, 24-25 October

James, D.G. 1997: imidacloprid increases egg production in Amblyseius victoriensis. Exp. Appl. Acarol. 21: 75-82.

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James, D.G. 2003: Toxicity of imidacloprid to Galendromus occidentalis, Neoseiulus fallacis and Amblyseius andersoni from hops in Washington State, USA. Exp. Appl. Acarol. 31: 275-281.

James, D.G. & Price, T.S. 2002: Fecundity of two spotted spider mite (Acari: Tetranychidae) is increased by direct and systemic exposure to Imidacloprid. J. Econ. Entomol. 95: 729-732.

James, D.G. & Vogele, B. 2002: The effect of imidacloprid on the survival of some beneficial arthropods. Plant Prot. Quart.16: 58-62.

Kavitha, J. Kuttalam, S. Chandrasekaran, S. & Ramaraju, K. 2006: Effect of spiromesifen 240 SC on beneficial insects. Ann Plant Prot. Sci. 14: 343-345.

Kim, S.S. & Seo, S.G. 2001: Relative toxicity of some acaricides to the predatory mite, Amblyseius womersleyi and the two-spotted spider mite, Tetranychus urticae (Acari: Phytoseiidae, Tetranychidae).Appl. Entomol. Zool. 36: 509-514.

Kim, S.S., Seo, S.G., Park, J.D., Kim, S.G., & Kim, D. 2005: Effects of selected pesticides on the predatory mite, Amblyseius cucumeris (Acari: Phytoseiidae). J. Entomol. Sci. 40: 107-114.

Kim, S.S. & Yoo, S.S. 2002: Comparative toxicity of some acaricides to the predatory mite, Phytoseiulus persimilis and the two spotted spider mite, Tetranychus urticae. BioControl 47: 563-573.

Maeyer, L., de Peeters, D., Wijsmuller, J.M., Cantoni, A., Brueck, E. & Heibges, S. 2002a: Spirodiclofen: a broad-spectrum acaricide with insecticidal properties: efficacy on Psylla pyri and scales Lepidosaphes ulmi and Quadraspidiotus perniciosus. The BCPC Conference: Pests and diseases, Volumes 1: 65-74.

Maeyer, L., de Schmidt, H.W. & Peeters, D. 2002b: Envidor(R) - a new acaricide for IPM in pome fruit orchards. Pflanzenschutz-Nachrichten Bayer 55: 211-236.

Maeyer, L. de Schnorbach, H.J., James, D. & Elbert, A. 2005: Envidor ®, a new acaricide with an excellent selectivity against key beneficials in IPM orchards. AFPP Second International Conference on Mites in Crops, Montpellier, France, 24-25 October 2005

Mizell, R.F. & Sconyers, M.C. 1992: Toxicity of imidacloprid to selected arthropod predators in the laboratory. Florida Entomol. 75: 277-280.

Mori, K. & Gotoh, T. 2001: Effects of pesticides on the spider mite predators, Scolothrips takahashii (Thysanoptera: Thripidae) and Stethorus japonicus (Coleoptera: Coccinellidae). Intern’l J. Acarol. 27: 299-302.

Nauen, R., Bretschneider, T., Bruck, E., Elbert, A., Reckmann, U., Wachendorff, U. & Teiman, R. 2002: BSN 2060: a novel compound for whitefly and spider mite control. The BCPC Conference: Pests and diseases, 1: 39-44

Poletti, M., Maia, A.H.N. & Omoto, C. 2007: Toxicity of neonicotinoid insecticides to Neoseiulus californicus and Phytoseiulus macropilis and their impact on functional response to Tetranychus urticae. Biol. Cont. 40: 30-36.

Sáenz-de-Cabezón Irigaray, F.J. & Zalom, F.G. 2006: Side effects of five new acaricides on the predator Galendromus occidentalis (Acari, Phytoseiidae). Exp. Appl. Acarol. 38: 299-305.

Sanderson, J.P. & Zhang, Z.Q. 1995: Dispersion, sampling, and potential for integrated control of two-spotted spider mite on greenhouse roses. J. Econ. Entomol. 88: 343-351.

Sauzay, S. & Ledoux, F. 2005: Le spirodiclofen, nouvelle substance active pour controller les acariens en arboriculture. AFPP Second International Conference on Mites in Crops, Montpellier, France, 24-25 October 2005

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Tomlin, C.D.S. 2006: The Pesticide Manual 14th Edition. British Crop Production Council Villanueva, R.T. & Walgenbach, J. 2006: Duration of life stages and oviposition of Tetranychus

urticae after exposure to new-chemistry insecticides for several generations. ESA Conference http://esa.confex.com/esa/2006/techprogram/paper_26027.htm

Yoo, S.S. & Kim, S.S. 2000: Comparative toxicity of some pesticides to the predatory mite, Phytoseiulus persimilis (Acarina: Phytoseiidae) and the twospotted spider mite, Tetranychus urticae (Acarina: Tetranychidae). Korean J. Entomol. 30: 235-241.


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The interaction between Rhizoglyphus robini and plant pathogens on onion Tal Hanuny1, Moshe Inbar1, Leah Tsror2 and Eric Palevsky3 1Department of Evolutionary and Environmental Biology, University of Haifa, Mount Carmel, Haifa 31905, Israel [email protected]; 2Agricultural Research Organization (ARO), Gilat Research Center, D.N. Negev, 85280, Israel [email protected]; 3ARO, Newe-Ya’ar Research Center, Ramat Yishay, 30095, Israel [email protected] Abstract: We determined the pest status of the bulb mite, Rhizoglyphus robini, on young onion plants with and without the presence of different fungal species. First, we analyzed the degree of attraction of the mite to various fungi found on onion. We then assessed the effects of selected fungi and the mite on the germination and subsequent survival of young onion seedlings. Finally, we examined how the interactions between a weakly pathogenic fungus and the mite would affect the onion seedlings. The mites were always more attracted to colonized PDA plugs versus non-colonized PDA (except in the case of bi-nucleate Rhizoctonia AG-A). Onion seedling survival was significantly reduced by the fungi; the most pathogenic being the Fusarium moniliformae white strain and the least being F. moniliformae purple strain. Onion sprouts colonized by this fungus were significantly more attractive to the mites than healthy sprouts. At two days post-mite infestation, the effects on onion sprout length of the mites and the F. moniliformae purple strain and their interaction were significant. Mites had no effect on sprout length in the absence of this fungus, in contrast, in its presence, mites significantly reduced sprout length, more than the fungus alone. The importance of host-plant, fungi and mite interactions are discussed. Key words: Rhizoglyphus robini, Fusarium, highly- and weakly-pathogenic fungi, onion. Introduction The bulb mite Rhizoglyphus robini Claparède (Astigmata: Acaridae), a cosmopolitan soil-borne mite known as a pest of Liliaceae and associated with fungi (Diaz et al., 2000), is commonly found in agricultural soils in Israel, from the Negev in the south to the Hula Valley in the north. The biology, ecology and control of this pest were investigated in Israel on onion and garlic (Gerson et al., 1985). Recently there has been a substantial increase in complaints from growers and extension personnel of economic damage on onion (Allium cepa). Examination of the subsoil plant material revealed highly- and weakly-pathogenic fungi and varying levels of the bulb mite. Although the mite is considered to be a pest in its own right (Gerson et al., 1985), there is no action threshold, and control measures are either prophylactic or applied on the basis of presence/absence sampling. In our previous study (Ben-David et al., 2005), conducted in the winter in an unheated screen-house, we found that varying levels of R. robini had no effect on vegetative or yield parameters. We suggested temperature and plant pathogens were involved in the epidemiology of R. robini. Here we have focused on the interaction between pathogenic fungi of varying degrees and R. robini on onion (sown from seed) under temperature-controlled conditions. Our objective was to determine how fungi affect the pest status of the mite, specifically: 1. determine the degree of attraction of the mite to various fungi found on onion, 2.

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assess the effects of some fungi and the mite on the germination and subsequent survival of onion seedlings, 3. evaluate the interactions between a weakly pathogenic fungus and the mite on the host plant.

Materials and methods Attraction of mites to fungi The attraction of R. robini to six species/strains of fungi isolated from onion (Fusarium oxysporum (purple and white), F. moniliformae (purple and white), 2-nuclei Rhizoctonia AG-A and Pythium oligandrum) and one fungal species (Verticillium dahliae) that is not found on Liliaceae isolated from potato were tested at 25±1C°. Note - In this manuscript we refer to the different strains of F. oxysporum and F. moniliformae according to the color of their mycelia; their taxonomic status is now being determined and will be soon published. Rhizoglyphus robini were reared in darkness in the lab on peanuts in Petri dishes at 25 ± 1°C (Gerson et al., 1991). Four potato dextrose agar (PDA) plugs were placed at equal distances (adjacent to the rim) in a 5-cm Petri dish on wetted filter paper, two colonized with the fungus and two non-colonized as controls, six replicates per fungus. Ten females of R. robini were placed at the center of the plate and their location monitored once an hour for four hours. The effect of time and fungus was first checked for each pathogen (comparing the number of mites on colonized versus non-colonized agar plugs, within each dish). Attraction to various fungi species/strains were then compared using the mean number of mites found on colonized plugs (ANOVA). Pathogenicity of fungi found on onion To determine the degree of pathogenicity of fungi to onion seedlings we evaluated, following Koch’s postulates, four species/strains of fungi, namely, a purple strain of F. oxysporum, two strains of F. moniliformae (purple and white) and P. oligandrum, in eight replicates. Twelve seeds of onion CV ‘Ada 781’ were shallowly planted in a soil mix commercially prepared for house plant nurseries (“Shaham”, Givat Ada, Israel) in 360-cc pots. For the Fusarium species/strains, four cc of conidial suspension were added to each pot (4 x 105 conidia/pot), whereas for P. oligandrum, PDA plugs of mycelium from 3-days old colonies (1.8 x 103 per pot) were used, covered by a thin layer of soil mix. The experiment was conducted in a temperature-controlled greenhouse set at 25 ± 2°C. Forty days later, the number of seedlings per pot and plant height were recorded. Surface-sterilized plant material was then placed on PDA to assess fungal colonization levels. Effect of mites on onion The effects of four levels of Rhizoglyphus robini were assessed in 250 cc pots, replicated 6 times. Each pot was sown with 16 seeds (same CV and soil mix as listed above). To prepare the mites for quantitative inoculations, Petri dishes with mites and peanuts were washed with tap water through a coarse sieve. Mites/ml (all motile stages) were estimated by sampling 10 one-ml samples and counting them in a standard 3-cm Petri, equipped with a grid, under a dissection microscope (25x). Accordingly, the volume of the mite dilute was calculated to reach inoculation levels of 10, 100 and 1000 mites per pot. The experiment was performed in a incubator, L:D 11:13, day and night temperature, 24 ± 2°C and 17 ± 2°C, respectively, 68 ± 9 % RH. Germination was monitored weekly for four weeks. Mites were recovered from each pot using Berlese funnels.

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The effect of fungal-mite interactions on germinating onion seeds To study the interaction between mites and fungus we chose a moderate level of mites (20 adult females per sprout) and a relatively non-pathogenic fungus that was attractive to the mites namely, F. moniliformae purple. In regard to mite location, we hypothesized that more mites would be found on the colonized sprouts. With respect to sprout length we hypothesized that the interaction between mite and fungus would be significant, at least at the beginning of the experiment. The effects on the elongation of onion seeds (same CV as above) of mites, the weakly pathogenic fungus F. moniliformae purple strain (determined in the first experiment) and their combination were evaluated at 25 ± 1°C. Seeds were allowed to germinate on filter paper wetted with distilled water for one week (four 9-cm Petri dishes containing 16 seeds/dish). Half of the sprouts were then transferred to two Petri dishes containing PDA inoculated eight days earlier with F. moniliformae purple strain. After an additional three days, the experiment was initiated. The experimental design consisted of four treatments: seedlings were infested with mites (20/dish), fungus, fungus & mites and control (seedlings only), that were replicated ten times (each Petri dish being one replicate, totaling 40 dishes). Once a day all dishes were photographed with a digital camera, using the macro mode (Nikon 5600 Coolpix). The location of the mites (on the onion plant or not) and the presence/absence of fungal mycelia were noted. Daily plant growth was measured from the digital images using an Olympus Soft Imaging System (http://www.olympus-sis.com/). The experiment was monitored for six days. All data were analyzed using linear fit and Two Way ANOVA procedures with JMP5.0.1a (SAS Institute, Inc.). Results and discussion Attraction of mites to fungi When comparing between colonized and non-colonized PDA plugs (for each fungus separately), mites were always more attracted to colonized PDA plugs except in the case of bi-nucleate Rhizoctonia AG-A (Table 1).

Time and the interactions time X fungus were never significant. Similarly, in the analysis where all fungi were compared, the interaction between time and fungi was not significant (P = 0.99; F = 0.262; DF = 18,140). In contrast, both time and fungi were significant (Time – P = 0.004; F = 4.60; DF = 3,140; Fungi – P < 0.0001; F = 9.51; DF = 6,140). Mite attraction was similar to the fungi F. oxysporum white, V. dahliae, F. oxysporum purple, F. moniliformae purple. Attraction to F. moniliformae white was similar to the three latter species/strains but differed significantly from the first (F. oxysporum white) (Figure 1). Fusarium moniliformae white and P. oligandrum were similar in their attractivity to the mite, both differing from the least attractive fungus, 2-nuclei Rhizoctonia AG-A. A rather curious result was that V. dahliae, a fungus not associated with onion, was more attractive than Rhizoctonia AG-A, known to be pathogenic to onion. Pathogenicity of fungi Onion seedling survival was significantly affected by fungi (P < 0.0001; F = 57.016; DF = 4,35), the most pathogenic being the F. moniliformae white strain and the least being F. moniliformae purple strain, causing 95% and 18% reductions, respectively (Figure 2a).

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Table 1. F ratio and probability values for the effect of time and fungus (Fusarium oxysporum (purple and white), F. moniliformae (purple and white), 2-nuclei Rhizoctonia AG-A, Pythium oligandrum) and Verticillium dahliae, each fungus evaluated separately) and for the interaction between time and fungus on Rhizoglypus robini attraction to colonized PDA plugs (mites on colonized PDA plug vs. control PDA plug). (Degrees of freedom for the error, time, fungus (colonized or not) and interaction were 39, 3, 1 and 3, respectively).

F. oxysporum

(white) Verticillium

dahliae F. oxysporum

(purple)

F ratio Prob >F F ratio

Prob >F F ratio

Prob >F

Time 0.50 .6780 0.98 0.4127 0.46 0.7112 Fungus 121.04 <.0001 148.58 <.0001 176.54 <.0001 Interaction 0.70 0.5505 1.73 0.1766 1.94 0.1390

F. moniliformae

(purple)

F. moniliformae

(white) Pythium

oligandrum

2-nuclei Rhizoctonia

AG-A

F ratio Prob >F

F ratio

Prob >F

F ratio

Prob >F

F ratio

Prob >F

Time 0.08 0.9861 0.80 0.5004 0.34 0.7937 1.38 0.2622 Fungus 214.33 <.0001 74.25 <.0001 34.04 <.0001 1.93 0.1725 Interaction 0.76 0.5229 0.07 0.9738 1.89 0.1473 0.33 0.8066

Figure 1. Varying degree of attraction of Rhizoglyphus robini to seven species/strains of fungi (Fusarium oxysporum (purple and white), F. moniliformae (purple and white), 2-nuclei Rhizoctonia AG-A, Pythium oligandrum and Verticillium dahliae). Mean number of mites on colonized PDA plugs plus SE. Different letters indicate a significant difference between fungi (P≤0.05).

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Effect of mites on onion The rate of seedling germination was significantly affected by time (P < 0.0001; F = 15.310; DF = 3,79) and mites (P < 0.0001; F = 57.505; DF = 3,79) but the interaction between them was not significant (P = 0.998; F = 0.234; DF = 9,79), therefore all data were pooled (Figure 2b). Rate of germination in pots containing 10 mites was similar to the control and did not differ from the pots containing 100 mites, the latter being significantly different from the control on the one hand and pots containing 1000 mites on the other.

Figure 2. a) Onion seedling survival (%), 40 days post inoculation with Fusarium moniliformae (purple strain), Pythium oligandrum, F. oxysporum (purple strain) and F. moniliformae (white strain). b) Onion germination (%) four weeks post infestation with varying levels of Rhizoglyphus robini. Different letters indicate a significant difference between fungi (a) and mite levels (b) (P ≤ 0.05). Fungal-mite interactions on germinating onion seeds To study the interaction between mites and fungi we chose a moderate level of mites (20 adult females per sprout) and a relatively non-pathogenic fungus that was attractive to the mites namely, F. moniliformae purple. In regard to mite location, we hypothesized that more mites would be found on the colonized sprouts. With respect to sprout length we hypothesized that the interaction between mite and fungus would be significant, at least at the beginning of the experiment.

Mite attraction to onion sprouts was significantly affected both by time (P < 0.0001; F = 12.33; DF = 4,77) and the presence of fungus (P < 0.0001; F = 95.88; DF = 1,77). The

46

interactions between the factors was not significant (P = 0.0865; F = 2.12; DF = 4,77) (Figure 3a), indicating that mite attraction to the fungus did not decline over the 5 day experiment.

At two days post-mite infestation, the effect on onion sprout length of the mites (P = 0.0467; F = 4.34; DF = 1,27) and the F. moniliformae purple strain (P < 0.0001; F = 62.46; DF = 1,27) and their interaction (P < 0.0020; F = 11.70; DF = 1,27) were significant. Mites had no effect on sprout length in the absence of this fungus, in contrast, in its presence, mites significantly reduced sprout length, more than the fungus alone (Figure 3b). At four days post-mite infestation, the effects of mite (P = 0.03915; F = 4.70; DF = 1,27) and fungi (P < 0.0001; F = 95.34; DF = 1,27) were still significant but their interaction was not (P = 0.3978 F = 0.74; DF = 1,27). Reduction of sprout length by fungus was similar with or without mites. Mites without fungus did not affect sprout length.

Figure 3. a) Mites on onion sprouts with and without fungi (± SE). b) Sprout length two days post infestation with varying levels of Rhizoglyphus robini. Different letters indicate a significant difference between fungi-mite combinations (P ≤ 0.05).

The association and interaction between various fungi and R. robini have been reported for various crops such as lily (Ascerno et al., 1981) and rakkyo (Allium chinense). Rakkyo (Allium chinense) infested with F. oxysporum f. sp. allii was more attractive to R. robini than healthy or wounded plants (Shinkaji et al., 1988). Healthy plants dipped in alcohols isolated from culture

47

filtrates of Fusarium fungi were just as attractive to the mite as infested plants (Okabe & Amano, 1990) and the extent of bulb penetration and population development of the mite was substantially higher on Fusarium-infested bulbs (Okabe & Amano, 1991). These studies focused on highly pathogenic fungi. In the present study we have evaluated the attraction to both highly- and weakly-pathogenic fungi and found mite attraction to be similar. Mites preferred onion sprouts colonized by the weakly pathogenic fungus over onion sprouts that were not, suggesting that on the one hand, fungi are necessary for mite establishment and subsequent damage on onion sprouts but on the other, the fungus does not necessarily have to be highly pathogenic.

Abdel-Sater and Eraky (2001) suggested that the presence of pathogenic micro-organisms facilitate the penetration of the bulb mites and provides additional food supplements for its development. Moderate mite feeding can enhance mycelial growth, furthermore astigmatid mites dispersing from one patch to the next distribute fungal spores (Hubert et al., 2004). To improve the control strategy for R. robini, more research should be conducted to determine how disease control can affect the susceptibility of young onion plants to bulb mite attack. Acknowledgements The authors would like to thank Dr. Ronny Cohen, Carmela Horev, Sarah Laviush Mordechai, Aisheh Sadia and Shira Gal for their technical assistance. This paper is a contribution from the Agricultural Research Organization, Institute of Plant Protection, Bet Dagan, Israel. References

Abdel-Sater, M.A. & Eraky, S.A. 2001: Bulb mycoflora and their relation with three stored

product mites. Mycopathologia 153: 33-39. Ascerno, M.E., Pfleger, F.L. & Wilkins, H.F. 1981: Effect of root rot and Rhizoglyphus robini on

greenhouse-forced Easter lily development. Environ. Entomol. 10: 947-949. Ben–David T., Tsror, L,. & Palevsky, E. 2005: Developing an action threshold for the bulb mite,

Rhizoglyphus robini on lily, onion and garlic. IOBC/wprs Bulletin 28(10): 11-14. Diaz, A., Okabe, K., Eckenrode, C.J., Villani, M.G. & Oconnor, B.M. 2000: Biology, ecology,

and management of the bulb mites of the genus Rhizoglyphus (Acari: Acaridae). Exp. Appl. Acarol. 24: 85-113.

Gerson, U., Cohen, E. & Capua, S. 1991: Bulb mite, Rhizoglyphus robini (Astigmata: Acaridae) as an experimental animal. Exp. Appl. Acarol. 12: 103-110.

Gerson, U., Yathom, S., Capua, S. & Thorens, D. 1985: Rhizoglyphus robini Claparede (Acari: Astigmata :Acaridae) as a soil mite. Acarologia 26: 371-380.

Hubert, J., Jarosik, V., Mourek, J., Kubatova, A. & Zdarkova, E. 2004: Astigmatid mite growth and fungi preference (Acari: Acaridida): comparisons in laboratory experiments. Pedobiologia 48: 205-214.

Okabe, K. & Amano, H. 1990: Attractancy of Alcohols isolated from culture filtrates of Fusarium fungi for the robine bulb mite, Rhizoglyphus robini CLAPAREDE (Acari: Acaridae), in sand. Appl. Entomol. Zool. 25: 397-404.

Okabe, K. & Amano, H. 1991: Penetration and population growth of the robine bulb mite, Rhizoglyphus robini CLAPAREDE (Acari: acaridae), on healthy and Fusarium-infected rakkyo bulbs. Appl. Entomol. Zool. 26: 129-136.

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Shinkaji, N., Okabe ,K. & Amano, H. 1988: Reaction of rhizogyphine mites (Acarina:Acaridae) infesting rakkyo (Allium chinense G. Don) plants infested with Fusarium fungi as well as to the culture medium and filtrate of the fungi. Jpn. J. Appl. Entomol. Zool. 32: 37-42.


pp. 49

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The beneficial fungus Neozygites floridana for the control of Tetranychus urticae Ingeborg Klingen, Karin Westrum, Silje Stenstad Nilsen, Nina Trandem, Gunnar Wærsted Bioforsk, Norwegian Institute for Agricultural and Environmental Research, Plant Health and Plant Protection Division, Høgskoleveien 7, 1432 Ås, Norway. E-mail: [email protected] Abstract: Neozygites floridana is a fungus in the order Entomophthorales that infects and kills the two-spotted spider mite, Tetranychus urticae. The fungus is therefore of interest for the biological control of T. urticae. To obtain information that might help in the use of this fungus under practical conditions in strawberries and cucumbers we have tried to answer the following questions in a series of studies*): 1) When, and at what infection levels does N. floridana occur in T. urticae populations in field grown strawberries? 2) How and where does N. floridana survive harsh climatic conditions (i.e winter) in Norway? 3) How and where does N. floridana infected T. urticae move and sporulate on a plant? 4) How do commonly used pesticides in strawberries affect N. floridana and T. urticae? 5) How can N. floridana be inoculated in augmentative microbial control of T. urticae? Results show that N. floridana infected and killed T. urticae in 12 out of 12 Norwegian strawberry fields studied. Infection levels up to 90% were observed, and the highest levels were observed late in the season. The infection levels throughout a season varied considerably. N. floridana was observed to over winter as both hyphal bodies in hibernating T. urticae females from October to at least February at temperatures as low as -20o C. Cadavers with resting spores were found from October to the end of January. Cadavers then probably disintegrated, and resting spores were left on leaves, soil, etc. In a bioassay where a Norwegian N. floridana isolate was tested for numbers and distance of spores thrown at three different temperatures (13o, 18o, 23o C), preliminary results show that high numbers of spores (ca 1300-1900 per cadaver) were thrown at all three temperatures. Further, spores were thrown about the same distance (up to about 6 mm) at all three temperatures. The effects of pesticides used in strawberries on the N. floridana infection level were studied to evaluate factors that might be important for conservation biological control. The pesticides tested were three fungicides; Euparen (tolylfluanid), Teldor (fenhexamid), Switch (cyprodinil + fludioxonil) and one acaricide/ insecticide: Mesurol (methiocarb). The experiment indicated that all three fungicides affect N. floridana negatively but that Euparen might be the least harmful. Mesurol did not affect N. floridana. Our attempts to inoculate N. floridana artificially in a strawberry field has not yet been successful, but we now work on promising methods for inoculation of N. floridana in T. urticae populations in greenhouse cucumbers. More detailed results from the studies referred to in this abstract will soon be published elsewhere. Key words: Tetranychus urticae, Neozygites floridana, microbial control, natural enemy, pesticides, side effect, resting spores, overwintering, harsh climatic conditions, strawberry, cucumber, greenhouse *) Studies have been financed by the following Norwegian Research Council projects: “Use of beneficial fungi to control weeds, insect pests and plant pathogenic fungi”. Project leader: Richard Meadow, Bioforsk. “Reduced use of pesticides in open field strawberries”. Project leader: Nina Trandem, Bioforsk. “Optimizing greenhouse climate to improve growth, yield, quality and control of mildew and biological control of pests in cut roses and cucumber”. Project leader: Hans-Ragnar Gislerød, Norwegian University of life sciences and Nina Svae Johansen, Bioforsk (Sub project: Biological control).

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pp. 51-57

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Exploration and evaluation of natural enemies for the invasive spider mite Tetranychus evansi Markus Knapp Icipe – African Insect Science for Food and Health, P.O. Box 30772, 00100 Nairobi, Kenya, [email protected] Abstract: The spider mite Tetranychus evansi is an invasive pest of tomatoes in East and Southern Africa probably originating from South America. It is causing severe yield losses in small-holder tomato systems. No effective natural enemies are present in the region. Initial surveys for natural enemies in northeastern Brazil, from where the mite had been reported as a pest, did no reveal any potential biocontrol agent. Areas in South America, that are climatically similar to areas in Kenya and Zimbabwe where T. evansi is a problem, were identified using Desktop-GARP (Genetic Algorithm for Rule-set Production). Several natural enemies were found during extensive surveys in these areas. Evaluation of several natural enemies collected revealed a Brazilian strain of the phytoseiid mite Phytoseiulus longipes to be the only promising candidate for classical biological control in Africa so far. This predatory mite was imported into Kenya and a permit for experimental released was granted in December 2006. Key words: Tetranychus evansi, classical biological control, climate matching Introduction The spider mite Tetranychus evansi Baker & Pritchard is an important pest of tomatoes in East and Southern Africa (ESA) (Meyer 1996, Knapp et al. 2003). It is exotic to Africa and probably originates in South America (Gutierrez & Etienne 1986). More recently the mite has also been reported from Senegal in West Africa (Duverney et al. 2005). It is also known to occur in North Africa and southern Europe (Migeon & Dorkeld 2006). No effective natural enemies of T. evansi have been found in ESA and studies in other parts of the world have shown that this mite is not a suitable prey for several species of Phytoseiidae (Moraes & McMurtry 1985, 1986, Escudero & Ferragut 2005). Farmers in ESA exclusively rely on synthetic acaricides to control T. evansi. However, pesticide applications by small-holder farmers are often ineffective due to various reasons including choice of wrong products, under-dosage and substandard application techniques (Sibanda et al. 2000, Saunyama & Knapp 2003).

A project was initiated by the International Centre of Insect Physiology and Ecology (icipe) in 1998 to develop Integrated Pest Management (IPM) and biological control strategies for spider mites in small-holder tomato production in ESA. At this time it was thought that Tetranychus urticae Koch was the major spider mite species in tomato production. When it became clear that the exotic mite T. evansi is much more important, a collaborative programme with ESALQ-USP in Brazil was initiated to search for potential classical biological control agents in South America.

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Material and methods Search in areas where T. evansi had been reported as a pest Tetranychus evansi was reported as a serious pest in large scale irrigated tomato production around Petrolina, Pernambuco State in northeastern Brazil in the early 1980s (Moreas et al. 1986).

Tomato fields (0.1 ha) were planted at monthly intervals at EMBRAPA’s Mandacaru Research Station in Juazeiro near Petrolina between November 2000 and July 2001. Samples of 30 top leaflets, 30 middle leaflets and 30 bottom leaflets were taken every 15 days. The leaflets were taken to the laboratory and mites counted and identified under a dissecting microscope.

Between August 2000 and July 2001, mites were also collected from Datura stramonium, Physalis angulata and Solanum americanum growing naturally in irrigation schemes around Petrolina. At each sampling occasion, samples of 20-30 leaves per plant species were taken to the laboratory and mites collected under a dissecting microscope were identified and counted according to species.

A survey was conducted in October 2002 in the states of Pernambuco, Paraiba, Rio Grande do Norte and Ceará along the route between Recife (Pernambuco) and Crato (Ceará) with stops at about every 50 km. Native and cultivated Solanaceae were examined for mites either directly in the field or leaves were taken to the laboratory for examination under a dissecting microscope. Climate matching and surveys in areas climatically similar to Kenya and Zimbabwe Areas in South America, that are climatically similar to areas in Kenya and Zimbabwe where T. evansi is a problem, were identified using Desktop-GARP (Genetic Algorithm for Rule-set Production) (Stockwell & Noble 1992, Peterson 2001). The procedures used are described in detail by Fiaboe et al. (2006). Surveys for natural enemies were carried out along main roads in these areas with stops every 50-70 km. At each stop leaves were collected from all wild and cultivated Solanaceae that could be found. Additionally, wastelands at the periphery of towns were also sampled in some areas because it became evident that Solanum americanum and spontaneously growing tomatoes could frequently be found in such places and that these plants very often harboured mites. The leaves were taken to the laboratory in cool boxes where mites were collected from the leaves and counted under a dissecting microscope. All spider mites and predatory mites were mounted and identified; insects that are potential predators of spider mites were also collected and identified. Laboratory investigations on the biology of natural enemies The suitability of T. evansi as prey was determined for five predatory mite species that had been found associated with T. evansi near Recife in northeastern Brazil (Asca sp., Pronematus ubiquitus (McGregor), Paraphytoseius orientalis (Narayanan, Kaur & Ghai), Phytoseius guianensis De Leon and Phytoseius cismontanus De Leon) by evaluating their oviposition rates when fed T. evansi under laboratory conditions (26°C, 65-70% rH and 12:12h photoperiod). The T. evansi colony used originated from S. americanum growing at the university campus and was maintained on potted S. americanum in a greenhouse. For comparison, the same parameters were investigated with tomato russet mite, Aculops lycopersici Massee, and pollen of Ricinus communis as food, offered separately or together.

The life history of the coccinellid beetle Stethorus tridens Gordon was investigated at 27°C, 70-80% rH and 12:12h photoperiod. Fifty newly laid eggs were carefully transferred to a rearing unit containing a leaf disk of S. americanum infested with all stages of T. evansi. Larvae were supplied daily with excess prey by brushing mites from infested leaves into the arena. Once the

53

adult emerged, couples were formed and put in larger rearing units containing a leaf of S. americanum infested with all stages of T. evansi.

Immature development and survival, and oviposition of a strain of the predatory mite Phytoseiulus longipes Evans collected on T. evansi-infested tomatoes in Uruguaiana, Rio Grande do Sul with T. evansi and T. urticae as prey were studied in the laboratory at 25°C, 80-90% rH and 12:12h photoperiod. Newly laid eggs were isolated in experimental units consisting of a piece of a Canavalia ensiformis leaflet kept on a foam mat placed in a plastic tray. The foam mat was kept wet by daily additions of distilled water to the trays. Food was provided by placing a tomato leaflet infested with either T. evansi or T. urticae on the arenas. The arenas were observed every 6 hours until the predator started oviposition and every 24 hours thereafter. A male taken from the rearing colony was provided for each adult female. Results and discussion Search in areas where T. evansi had been reported as a pest Low numbers of mites of the families Phytoseiidae, Tenuipalpidae, Cheyletidae, Tarsonemidae and Trombiculidae were collected in the tomato fields at Juazeiro but no T. evansi were found. Mites of the family Tydeidae were found at high levels. The phytoseiid species collected were Euseius concordis (Chant), Neoseiulus idaeus Denmark & Muma and Typhlodromus (Anthoseius) sp.

On wild Solanaceae T. evansi was found on a single occasion on D. stramonium. A single specimen of each of the following phytoseiid species was collected in this study: Euseius sibelius (De Leon), P. guianensis and Typhlodromus (Anthoseius) sp. The former two species were found in association with T. evansi.

Tetranychus evansi was only found in five locations during the survey between Recife and Crato. The only phytoseiid species found in a T. evansi colony was Phytoseiulus macropolis (Banks) (Furtado et al. 2004).

The reasons for the scarcity of T. evansi in areas where it had been reported a severe pest before remain unclear. Large-scale tomato production for processing was abandoned in Petrolina/Juazeiro in the late 1990s due to very high and difficult to control populations of whiteflies (Bemisia tabaci (Gennadius)) and the South American pinworm (Tuta absoluta (Povolny)). However, T. evansi was not considered a key pest at this time anymore. An entomopathogenic fungus infecting T. evansi in Petrolina might also have contributed to the population decline (Humber et al. 1981). Climate matching and surveys in areas climatically similar to Kenya and Zimbabwe The areas in South America climatically similar to places where T. evansi occurs in Africa and the survey routes are shown in Figure 1. T. evansi was rarely found in all areas surveyed in Brazil but it was more common in Argentina. When the mite was present it frequently occurred at very high population levels.

In southern Brazil it was found in five locations on five different host plants. The predatory mites Neoseiulus californicus (McGregor), Phytoseiulus fragariae Denmark & Schicha, Phytoseiulus longipes Evans, Galendromus annectens (De Leon), Euseius ho (De Leon), Euseius inouei (Ehara & Moraes), and Neoseiulus idaeus Denmark & Muma were found at least once each associated with T. evansi. (Furtado et al. 2006). N. californicus, P. fragariae and P. longipes were taken to the laboratory for further studies.

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T. evansi was not found during the surveys in the Southeast and only on three occasions in the Northeast. No predatory mites were found directly associated with T. evansi in these surveys but the acarophagous ladybird beetle S. tridens was associated with the mite in all three locations where it was found (Fiaboe et al. 2007). In Argentina T. evansi was found in 21 out of 53 samples and eight species of Phytoseiidae were associated with it.

Figure 1. Areas in South America climatically similar to areas in Kenya and Zimbabwe where T. evansi is present (grey areas) and survey routes for the search for natural enemies.

Recife-Crato

Northeast

Southeast

South

Central West

Argentina

Laboratory investigations on the biology of natural enemies For all predatory mites tested in Recife, oviposition was very low and lower on T. evansi than on other prey except for Asca sp., which did not oviposit on T. evansi and on pollen (Table 1). Therefore, all these mites are not effective predators of T. evansi, however P. orientalis might have a potential as a control agent for A. lycopersici, which is also an important pest of tomatoes in many parts of the world.

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Table 1. Oviposition rates of different predatory mites fed T. evansi, A. lycopersici and/or pollen.

Oviposition (eggs/♀/day) ± SE

Predators T. evansi A. lycopersici

+ pollen A. lycopersici Pollen Asca sp. 0.0 ± 0.00bC 0.1 ± 0.04aC 0.1 ± 0.03aC 0.0 ± 0.00bC P. ubiquitus 0.0 ± 0.00cC 0.7 ± 0.15aB 0.8 ± 0.09aB 0.3 ± 0.04bB P. guianensis 0.1 ± 0.03bAB 0.5 ± 0.08aB 0.7 ± 0.07aB 0.6 ± 0.06aA P. cismontanus 0.2 ± 0.04bA 0.7 ± 0.08aB 0.7 ± 0.05aB 0.6 ± 0.06aA P. orientalis 0.1 ± 0.04cB 2.0 ± 0.12aA 1.8 ± 0.07aA 0.7 ± 0.09bA

Means followed by the same upper case letter in the same column or lower case letter in the same row are not significantly different (SNK P<0.05).

Egg to adult development of S. tridens took 16.2 days at 27 °C; adult females lived on average 72 days and laid 123 eggs. Despite high fecundity and prey consumption (68 T. evansi nymphs per day for ovipositing females) S. tridens did not have a significant effect on T. evansi populations in the field (Fiaboe et al. submitted).

The duration of immature development of P. longipes was similar on T. evansi (4.7 days) and T. urticae (4.8 days) as prey. Oviposition was significantly higher on T. evansi (3.9 eggs/female/day) than on T. urticae (3.0 eggs/female/day). When given the choice, P. longipes preferred to feed and to oviposit on tomato leaflets infested with T. evansi (Furtado et al. submitted).

Phytoseiulus longipes is the only promising candidate for classical biocontrol in Africa so far. It was imported into Kenya and is currently kept at icipe’s quarantine facilities awaiting a release permit. This study clearly shows that climate matching can play an important role in determining areas were the search for classical biological control agents should be concentrated. The scarcity of the pest in its suspected area of origin made explorations difficult. Understanding the reasons and mechanisms behind this scarcity could provide important information for the design of the classical biological control project. Phytoseiulus species are specialised predator of spider mites (McMurtry & Croft 1997) and are therefore dependent on spider mites to become established in the farming system. Small scale vegetable farming systems in ESA are rather diverse and chances that other spider mites will be available as alternative prey are good. Tomato is attacked by a large number of other pests and diseases that are currently controlled with synthetic pesticides. To be successful, biocontrol must be integrated into a comprehensive IPM system and biocontrol-compatible control strategies for these pests and diseases have to be developed.

Relying on a single biocontrol agent for the diverse tomato growing areas in ESA is rather risky and additional explorations should be conducted in other areas of South America identified by climate matching. The potential of an isolate of the mite pathogenic fungus Neozygites floridana Weiser & Muma that is attacking T. evansi in Brazil as a biocontrol agent is currently tested.

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Acknowledgements This project was funded by a grant of the German Federal Ministry for Economic Co-operation and Development to icipe. References Duverney, C., Kade, N. & Ngueye-Ndiaye, A. 2005: Essais preliminaries pour limiter les degats

de Tetranychidae sur les cultures maraicheres dans le Sine-Saloum (Senegal). In: Comptes Rendus de 2ième Colloque International sur les Acariens des Cultures de l’AFPP. Agro-Montpellier, France, 24-25 octobre 2005 (CD-ROM).

Escudero, L.A. & Ferragut, F. 2005: Life-history of predatory mites Neoseiulus californicus and Phytoseiulus persimilis (Acari: Phytoseiidae) on four spider mite species as prey, with special reference to Tetranychus evansi (Acari: Tetranychidae). Biol. Control 32: 378-384.

Fiaboe, K.K.M., Fonseca, R.L., Moraes, G.J., Ogol, C.K.P.O. & Knapp, M. 2006: Identification of priority areas in South America for exploration of natural enemies for classical biological control of Tetranychus evansi (Acari: Tetranychidae) in Africa. Biol. Control 38: 373-379.

Fiaboe, K.K.M., Gondim Jr., M.G.C., Moraes, G.J., Ogol, C.K.P.O. & Knapp, M. 2007: Surveys for natural enemies of the tomato red spider mite Tetranychus evansi (Acari: Tetranychidae) in northeastern and southeastern Brazil. Zootaxa 1395: 33-58.

Furtado, I.P., Kreiter, S., Moraes, G.J., Tixier, M.-S., Flechtmann, C.H.W. & Knapp, M. 2004: Plant mites (Acari) from northeastern Brazil, with descriptions of two new species of the family Phytoseiidae (Mesostigmata). Acarologia 45: 131-143.

Furtado, I.P., Moreas, G.J., Kreiter, S. & Knapp, M. 2006: Search for effective natural enemies of Tetranychus evansi Baker & Pritchard in south and southeast Brazil. Exp. Appl. Acarol. 40: 157-174.

Furtado, I.P., Moraes, G.J., Kreiter, S., Tixier, M.-S. & Knapp, M. 2006: Potential of a Brazilian population of the predatory mite Phytoseiulus longipes as a biological control agent of Tetranychus evansi (Acari: Phytoseiidae, Tetranychidae). Submitted to Biol. Control.

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Humber, R.A., Moraes, G.J. & dos Santos, J.M. 1981: Natural infection of Tetranychus evansi (Acarina: Tetranychidae) by Triplosporium sp. (Zygomycetes: Entomophthorales) in northeastern Brazil. Entomophaga 26: 421-425.

Knapp, M., Wagener, B. & Navajas M. 2003: Molecular discrimination between the spider mite Tetranychus evansi Baker and Pritchard, an important pest of tomatoes in southern Africa, and the closely related species T. urticae Koch (Acarina: Tetranychidae). Afr. Entomol. 11: 300-304.

McMurtry, J.A. & Croft, B.A. 1997: Life-styles of phytoseiid mites and their roles in biological control. Annu. Rev. Entomol. 42: 291–321.

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Moraes, G.J. & McMurtry, J.A. 1985: Comparison of Tetranychus evansi and T. urticae (Acari: Tetranychidae) as prey for eight species of Phytoseiid mites. Entomophaga 30: 393-397.

Moraes, G.J. & McMurtry, J.A. 1986: Suitability of the spider mite Tetranychus evansi as prey for Phytoseiulus persimilis. Ent. Exp. Appl. 40: 109-115.

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Saunyama, I.G.M. & Knapp, M. 2003: The effects of pruning and trellising of tomatoes (Lycopersicon esculentum Mill.) on red spider mite (Tetranychus evansi Baker and Pritchard) incidence and crop yield in Zimbabwe. Afr. Crop Sci. J. 11: 269-277.

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Predatory mites associated with the coconut mite Aceria guerreronis in Brazil L.M. Lawson-Balagbo1,4, M.G.C. Gondim Jr2, G.J. de Moraes3, R. Hanna1, P. Schausberger4

1International Institute of Tropical Agriculture, Biological Control Centre for Africa, Cotonou, Benin; 2Universidade Federal Rural de Pernambuco, Departamento de Agronomia, Recife-PE, Brazil; 3ESALQ-Universidade de São Paulo, Dept. Ent., Fitop.e Zool. Agrícola, Piracicaba, SP, Brazil; 4Institute of Plant Protection, Department of Applied Plant Sciences and Plant Biotechnology, University of Natural Resources and Applied Life Sciences, Vienna, Austria, Email: [email protected] Abstract: Coconut is an important crop in tropical and subtropical regions. Among the mites that infest coconut trees, Aceria guerreronis Keifer is economically the most important. We conducted surveys throughout the coconut growing areas of Brazil. Samples were taken from hanging nuts of coconut palms in 163 sites with the aim of determining the predatory mites associated with A. guerreronis. About 78% of all predatory mites belonged to the family Phytoseiidae, mainly represented by Neoseiulus paspalivorus De Leon, Neoseiulus baraki Athias-Henriot Amblyseius largoensis Muma and Neoseiulus recifensis Gondim Jr. & Moraes; 15% were Ascidae, mainly Proctolaelaps bickleyi Bram, Proctolaelaps sp nov and Lasioseius aff. queenslandicus Womersley. Bdella distincta Baker & Balogh (Bdellidae) was also common under the bracts of the nuts as well as few species of the family Cheyletidae. Six species are reported for the first time in Brazil. Neoseiulus paspalivorus, N. baraki and P. bickleyi were the most frequent and most abundant predators associated with A. guerreronis. These three predators seem promising candidates for future biocontrol efforts. Keywords: Coconut, Aceria guerreronis, predatory mites, biological control Introduction Coconut palm, Cocos nucifera L. (Arecaceae) is highly important for the livelihoods of farming communities in coastal areas of tropical and subtropical regions in the world (Foale 2003). Among the mites that infest coconut palms Aceria guerreronis Keifer (Acari: Eriophyidae) is economically the most important. It has been reported from many coconut growing regions in the Americas and West Africa and most recently from the Indian subcontinent, often causing enormous damage (Mariau 1977, Moore et al. 1989, Fernando et al. 2002). Despite the variable measures used to control this pest, none has been proven sufficiently effective, making this mite the most intractable coconut pest. Considerable attention is being given to the pathogenic fungus Hirsutella thompsonii Fisher to be used as biopesticide against this pest; however, there is not enough information about the influence of climatic factors on the effectiveness of that pathogen in coconut plantations (Cabrera 2002).

Aceria guerreronis is most probably of South American origin (Navia et al. 2005). Hence, classical biological control may offer a sustainable solution to the problem caused by this pest in

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regions outside of South America such as Africa and Asia (Moraes & Zaccarias 2002). Brazil is the largest country in South America and falls within the likely native home of A. guerreronis. However, little is known about the predatory mites associated with A. guerreronis in that country (Gondim et al. 2000, Navia & Flechtman 2002). The present work is part of a multi-institutional project with the broad objective of developing a biological control program against A. guerreronis in Africa and elsewhere.

The objective of this study is to determine the predatory mites associated with A. guerreronis in the coconut growing areas of Brazil with the aim of identifying potential candidates for biological control of the pest. Material and methods Sampling methods Three collecting routes were initially established through the coconut growing areas in northern and northeastern Brazil, each starting from the city of Recife. Surveys were conducted along each of those routes between August 2004 and September 2005. For logistical reasons, an average distance of 50 km between sampling sites was initially set. In areas showing high concentration of coconut fields, the distance between sampling sites was reduced to approximately 20 km. After the first survey, some sites were revisited and new fields situated between two previously visited sites were visited at the following sampling occasions. In each site thirty, two to five month old, nuts were collected from two to four randomly selected palms. For logistical reasons, nuts were cut so that only the part carrying the bracts was collected. Mite evaluation Due to time limitation, only 10% of the sampled nuts were directly examined in the field using hand lenses at 15x magnification. The remaining nuts were placed in plastic bags, stored in cool boxes and transported to the Laboratory of Acarology at UFRPE (Universidade Federal Rural de Pernambuco, Recife) for further processing. During processing, nuts were cut in several sections and the bracts were gradually removed. All mites other than A. guerreronis were collected individually with a paint brush and placed in 70% ethanol. All specimens of A. guerreronis from each sample (30 nuts) were brushed into vials containing 10 ml of 70% ethanol.

The number of A. guerreronis was estimated using a methodology similar to that described by Siriwardena et al. (2005). A 1:10 dilution of the eriophyids’ suspension was prepared, shaken and an aliquot (1 ml) drawn into a counting chamber. The chamber was similar to the one used by Seaman et al. (1996) and consisted of a thick glass slide with a U-shaped trough forming a counting area of 3×2 cm divided into 24 squares. The edges of the trough are raised to support a thinner cover slide. The aliquot was introduced into the chamber with a pipette and was allowed to settle for two minutes before counting. Mites were counted in six of the 24 squares. The number obtained was multiplied by four and the coefficient of dilution (10) to estimate the total number of mites per 30 nuts.

All predatory mites and a subsample (50 individuals) of A. guerreronis were mounted in Hoyer's medium for subsequent identification or confirmation of identification to species level when possible.

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Results and discussion Sampling was conducted in a total of 163 sites in 9 states of northeastern Brazil (Alagoas, Bahia, Ceará, Maranhão, Paraíba, Pernambuco, Piauí, Rio Grande do Norte, Sergipe) and one state in northern Brazil (Pará) (Figure 1). A total of 4890 nuts were examined in this study.

Aceria guerreronis was by far the most abundant mite (6,643,177) and was present in 87% of the sites visited. The predatory mites associated with A. guerreronis in the surveyed areas are listed in Table 1. Twenty-five species of predatory mites belonging to five families were collected in this study. By far the largest numbers of predatory mites found were Phytoseiidae, followed by Ascidae. The most abundant phytoseiid species were Neoseiulus paspalivorus DeLeon and Neoseiulus baraki Athias-Henriot. The phytoseiids Neoseiulus recifensis Gondim Jr. & Moraes, Amblyseius largoensis Muma and the bdellid Bdella distincta Baker & Balogh were also regularly found but with lower abundance. Proctolaelaps bickleyi Bram (Ascidae) was the most abundant ascid mite. Five species of Ascidae were collected for the first time in Brazil including a new species of Proctolaelaps sp. nov.

The above-mentioned phytoseiid mites except N. recifensis were also found in association with A. guerreronis in other parts of the Americas (Howard et al. 1990), in Sri Lanka (Moraes et al. 2004) and in Benin (Negloh & Hanna, personal communication). The present study shows that A. guerreronis is widely distributed on coconut palms in Brazil, where it is clearly the most numerous mite on nuts. Several species of predatory mites are associated with A. guerreronis under the bracts of coconuts and seem promising natural enemies of this pest. Our surveys revealed N. paspalivorus along with N. baraki, P. bickleyi and Proctolaelaps sp. nov. as well as N. recifensis as predators that deserve further investigations as potential biocontrol agents of A. guerreronis.

Fig. 1: Sites sampled from August 2004 to September 2005

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Table 1. List of predatory mites associated with A. guerreronis under the bracts of coconuts in one state of northern and nine of northeastern Brazil.

FAMILY Total STATE Ascidae Asca foxi DeLeon 3 Maranhão, Pará, Pernambuco Blattisocius keegani Fox¹ 1 Maranhão

Lasioseius aff. queenslandicus Womersley 34 Alagoas, Bahia, Ceará, Maranhão, Pará, Paraiba, Pernambuco, Piaui, Rio G. Norte, Sergipe

Melichares aff. agilis Hering¹ 1 Pernambuco

Proctolaelaps bickleyi Bram 178 Alagoas, Bahia, Ceará, Maranhão, Pará, Paraiba, Pernambuco, Piaui, Rio G. Norte, Sergipe.

Proctolaelaps coffeae Karg & Rodriguez¹ 1 Pernambuco Proctolaelaps intermedius Athias-Henriot¹ 1 Pernambuco

Proctolaelaps sp nov¹ 42 Alagoas, Bahia, Ceará, Maranhão, Pará, Paraiba, Pernambuco, Rio G. Norte, Sergipe.

Proctolaelaps sp 4 Alagoas, Paraiba, Pernambuco, Rio G. Norte Phytoseiidae

Amblyseius largoensis Muma 96 Alagoas, Bahia, Ceará, Maranhão, Pará, Paraiba, Pernambuco, Piaui, Rio G. Norte, Sergipe.

Amblyseius tamatavensis Blommers 3 Bahia, Ceará, Pará, Rio G. Norte, Sergipe

Euseius alatus DeLeon 26 Alagoas, Bahia, Ceará, Maranhão, Pará, Paraiba, Pernambuco, Rio G. Norte, Sergipe.

Neoseiulus baraki Athias-Henriot 350 Alagoas, Bahia, Ceará, Maranhão, Pará, Paraiba, Pernambuco, Rio G. Norte

Neoseiulus paspalivorus DeLeon 821 Alagoas, Bahia, Ceará, Maranhão, Pará, Paraiba, Pernambuco, Piaui, Rio G. Norte, Sergipe.

Neoseiulus recifensis Gondim Jr. & Moraes 41 Alagoas, Ceará, Paraiba, Pernambuco, Rio G. Norte

Prorpioseiopis canaensis Muma 1 Alagoas, Bahia Typhlodromina subtropica Muma & Denmark 1 Bahia, Pernambuco Typhlodromips cananeiensis Gondim Jr. & Moraes 4 Bahia Typhlodromus aff. vulgaris Ehara¹ 4 Alagoas, Pernambuco, Rio G. Norte Cheyletidae Cheletomimus sp 5 Piaui

Hemicheyletia sp 22 Alagoas, Bahia, Ceará, Maranhão, Pernambuco, Piaui, Rio G. Norte

Mexecheles sp 1 Ceará Bdellidae

Bdella distincta Baker & Balogh 78 Alagoas, Bahia, Ceará, Maranhão, Pará, Paraiba, Pernambuco, Piaui, Rio G. Norte, Sergipe.

Spinibdella sp 3 Alagoas, Pernambuco, Rio G. Norte, Sergipe Cunaxidae Armascirus sp 2 Ceará, Maranhão, Pernambuco

1New record in Brazil.

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Acknowledgements This work was supported by the International Institute of Tropical Agriculture (IITA) through a grant from the Austrian Ministry of Finance, and by in-kind contribution from IITA, FEALQ, Fundação de Estudos agrarios Luiz de Queiroz, Piracicaba-Sao Paulo, Brazil and BOKU. This work is part of the PhD thesis of the senior author.

References Cabrera, R.I.C. 2002: Biological control of the coconut mite Aceria guerreronis (Acari:

Eriophyidae) with the fungus H. thompsonii and its possible integration with other control methods. Pp 89-100 in Fernando, L.C.P., Moraes, G.J. & Wickramananda, I.R (eds.) Proceedings of the International Workshop on Coconut Mite (Aceria guerreronis). Coconut Research Institute, Sri Lanka. January 6-8 2000.

Fernando, L.C.P., Wickramananda, I.R. & Aratchige, N.S. 2002: Status of coconut mite, Aceria guerreronis in Sri Lanka. Pp. 1-8 in Fernando, L.C.P., Moraes, G.J. & Wickramananda, I.R (eds.) Proceedings of the International Workshop on Coconut Mite (Aceria guerreronis). Coconut Research Institute, Sri Lanka. January 6-8 2000.

Foale, M. 2003: The coconut odyssey: the bounteous possibilities of the tree of life. Australian Centre for International Agricultural Research 101,132 pp.

Gondim Jr, M.G.C., Flechtmann, C.H.W. & Moraes, G.J. 2000: Mites (Arthropoda: Acari) associates of palms (Arecaceae) in Brazil V. Descriptions of four new species in the Eriophyoidea. Syst. Appl. Acarol. 5: 99-110.

Howard, F.W., Abreu-Rodriguez, E. & Denmark, H.A. 1990: Geographical and seasonal distribution of the coconut mite, Aceria guerreronis (Acari: Eriophyidae), in Puerto Rico and Florida, USA. J. Agric. U. Puerto Rico 74: 237-251.

Mariau, D. 1977: Aceria (Eriophyes) guerreronis: un important ravageur des cocoteraies africaines et américaines. Oléagineux 32: 101-108.

Moore, D., Alexander, L. & Hall, R.A. 1989: The coconut mite, Eriophyes guerreronis Keifer in St. Lucia: yield losses and attempts to control it with acaricides, polybutene and Hirsutella fungus. Trop. Pest Manag. 35: 83-89.

Moraes, G.J. Lopes, P.C. & Fernando, L.C.P. 2004: Phytoseiid mites (Acari: Phytoseiidae) of coconut growing areas in Sri Lanka, with descriptions of three new species. J. Acarol. Soc. Jpn 13: 141-160.

Moraes, G.J. & Zacarias, M.S. 2002: Use of predatory mites for the control of eriophyid mites. Pp 78-88 in Fernando, L.C.P., Moraes, G.J. & Wickramananda, I.R (eds.) Proceedings of the International Workshop on Coconut Mite (Aceria guerreronis). Coconut Research Institute, Sri Lanka. January 6-8 2000. Coconut Research Institute, Sri Lanka.

Navia, D. & Flechtmann, C.H.W. 2002: Mites (Arthropoda: Acari) associates of palms (Arecaceae) in Brazil: VI. New genera and new species of Eriophyidae and Phytoptidae (Prostigmata: Eriophyoidea). Int. J. Acarol. 28: 121-146.

Navia, D., de Moraes, G.J., Roderick, G. and Navavajas, M. 2005: The invasive coconut mite Aceria guerreronis (Acari: Eriophyidae): origin and invasion sources inferred from mitochondrial (16S) and nuclear (ITS) sequences. Bull. Entomol. Res. 95: 505-516.

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Seaman, E.K., Goluboff, E. Barchama, N. & Fisch, H. 1996: Accuracy of semen counting chambers as determined by the use of latex beads. Am. Soc. Reprod. Med. 66: 662-665.

Siriwardema, P.H.A.P., Fernando, L.C.P. & Peiris, T.S.G. 2005: A new method to estimate a population size of coconut mite, Aceria guerreronis, on a coconut. Exp. Appl. Acarol. 37: 123-129.


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Biological control in vineyards by means of a laboratory phytoseiid strain: a small scale experiment in Tuscany (Italy) Marialivia Liguori1, Giuseppino Sabbatini Peverieri1, Sauro Simoni1, Laura Ferré2 1Istituto Sperimentale per la Zoologia Agraria, via Lanciola 12/A, 50125 Firenze, Italy,2 Azienda agricola Podere Forte, Loc. Petrucci 13, 53023 Castiglione d’Orcia (Si), Italy Abstract: Eotetranychus carpini (Oud.) outbreaks frequently occurred in a small vineyard of cv Petit Verdot in Val d’Orcia (Siena, Italy); during 2004, samples showed that phytoseiids were not present. To re-establish their presence, the release of Typhlodromus exhilaratus Ragusa from laboratory rearing was made by performing a small scale experiment. In 2005, from mid-May to mid-August, 2,250 mites/plant were released. On the release plants the phytophagous population was significantly lower, especially till the end of July; in this period, damage level was positively correlated to E. carpini population density. Key words: vine, Eotetranychus carpini, Typhlodromus exhilaratus, phytoseiid release Introduction Recently, in some viticulture areas of Tuscany, the presence of Eotetranychus carpini (Oud.), the yellow grape-vine mite, has increased substantially; its outbreaks were especially frequent in Brunello and Val d’Orcia vine areas (personal unpublished data, information from extension services).

The control of spider mites in biological vineyards is mainly performed by conservation and recurring inoculations of native phytoseiid species usually by pruned branches or leaves originating from other colonized vineyards (Duso & Girolami 1985, Duso 1991, Duso & Vettorazzo 1999, Prischmann et al. 2006, Duso 2006). Other techniques based on the release of mass reared mites are mainly achieved in glasshouses or tunnel cultures rather than in open field (Blumel & Walzer 2002, Kondo 2004).

Since autumn 2004, the laboratory of Acarology of the Istituto Sperimentale per la Zoologia Agraria of Florence has been involved in monitoring E. carpini outbreaks in a small vineyard of cv Petit Verdot in Val d’Orcia (Siena, Italy). Sequential samplings in the vineyard showed no presence of native phytoseiid species or other mites. In spring of 2005, a small scale experiment was performed to introduce the phytoseiid Typhlodromus exhilaratus Ragusa from laboratory mass rearing. This phytoseiid was chosen for its widespread presence in vineyards of Central Italy (Castagnoli & Liguor1986a, Castagnoli et al. 1999) and its association with E. carpini (Castagnoli et al. 1991). Furthermore, in the laboratory this predator showed good reproductive potential and ability to feed on all stages of this prey and some tolerance to low humidities (Castagnoli & Liguori 1986b, Castagnoli et al. 1989, Liguori & Guidi 1995).

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Material and methods Site description The trial was conducted from spring to late summer 2005 in a vineyard near Castiglione d’Orcia (Siena, Tuscany), in an area belonging to the Orcia Rosso DOC vine. In this vineyard (about 0.5ha), 36 rows of Petit Verdot, a French cultivar with pubescent leaves, are contiguous to 16 rows of Cabernet. A part of the vineyard borders on a hedge of Quercus pubescens L. while, along two sides, a road divides it from other vineyards (cv. Sangiovese); the other sides of the vineyard are contiguous to olive trees. The 7 year-old vineyard was pruned according to the spur cord system. Green pruning is performed during the vegetative period. Copper oxicloride (Cu 35%) and wettable sulphur are usually applied to control downy and powdery mildew; if necessary, bromopropylate is used to control tetranychid outbreaks. Weather data were recorded throughout the year. Phytoseiid rearing The laboratory strain of T. exhilaratus originated from a vineyard of the Chianti area (Tuscany) and has been reared since spring 2003 in climatic cabinets (25°C, 75% RH, 16 hours light) on plastic arenas (5x10cm) surrounded by wet cotton wool to prevent mite escape and to supply a water source. Quercus ilex L. pollen was used as food and supplied three times a week. Mite release Weekly, from mid-May to mid-August 2005, each of the vine chosen for the trials received the same number of T. exhilaratus specimens (120 females and 30 males). The number of phytoseiids released was slightly higher than those used by Duso & Vettorazzo (1999) mainly to compensate for differences in release timing and eventual impact/stress suffered by the phytoseiids during the transfer from the laboratory to the field. Each group of phytoseiids was arranged on a home-made structure of plastic film (Ø 5cm), with cotton wool fixed in centre and two threads fixed on each side. This was put over a plastic arena of the same size, surrounded by wet cotton wool to prevent mite escaping and provided with a small amount of Q. ilex pollen as food. The day before the release, adult specimens of the predator were taken from laboratory mass rearing units with a fine brush, put on the release units and kept in fridge (+ 5°C) during the night. Then, these units were stored in refrigerated large boxes to transport mites from laboratory to the vineyard. At the moment of release, cotton wool was eliminated and plastic films tied to the lower branches of the vine trees. Experiment set up In the vineyard, since E. carpini outbreaks only occurred on Petit Verdot, 15 vine trees of this cultivar were chosen on the same row: 12 contiguous were the release plants; 3, at a distance of approximately 25 meters, were the control. A row on both sides separated the plants participating in the experiment from the remaining part of the vineyard which, in the year 2005, was conventionally managed. From mid-May to mid-August, samples of 5 leaves/plant were weekly collected prior to the release of the phytoseiids. The number of leaves in the vine canopy was always recorded.

In the laboratory, the leaves were examined under a stereomicroscope to count motile stages of E. carpini and T. exhilaratus. Starting from June spider mite foliar damage was evaluated and scored according to the following percentages of damaged area: 0%, damage 0; 1–20%, damage 1; 21–40%, damage 2; 41–60%, damage 3; 61–80%, damage 4; 81-100%, damage 5. The intensity of damage was expressed by means of the following formula: 100*(1n+2n+3n+4n+5n)/(5*N) (Kondo 2004).

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At the vintage time, the grapes of the plants participating in the experiment were separately gathered and chemically analysed in the cave before wine-making, particularly in order to measure the reducing sugar, total acidity, alcohol content and grape berry weight. Data analysis The abundance of tetranychids was evaluated across the sampling period to determine the effect of time and phytoseiid release by means of the General Linear Model, Repeated Measures option (SPSS 1999). The Pearson coefficient was calculated to measure the correlation power between the tetranychid number and the level of damage observed. The intensity and distribution of the different damage levels were analysed by the general log linear model and difference analysed by means χ2 test. All these analysis procedures were performed by SPSS (1999). Results and discussion During the vegetative period, from the beginning of May to the end of October, the wind had an average speed of 4.96 km/h and was mainly oriented from south/south-west; the average temperature was 20.2°C and the average relative humidity was 64.1% (Figure 1).The rainfall was practically absent (0.4 mm). These temperature and humidity conditions were similar to the laboratory values of temperature and humidity allowing good increase of the population of the yellow grape vine mite (Bonato et al 1990). The average number of leaves per vine tree ranged approximately from 80 to 150 and the number of leaves/plant sampled every time constituted about 5% of the total number.

Figure 1. Average daily means of temperature and relative humidity in the experimental site.

From the beginning of May to mid-August, cumulatively, 27,000 T. exhilaratus specimens were released on the experimental vine plants, at the release rate of 2,250 predators/plant. At the first release, the E. carpini density was 4 motile stages/leaf and the average number of leaves 82/plant;

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therefore, the prey/predator rate determined was approximately 2:1. During the experiment, E. carpini and T. exhilaratus were always recorded while presence of other mites was not observed.

Notwithstanding the difference in values, the two E. carpini dynamics (on release and control plants) showed analogous trends (Figure 2). The analysis across the experimental time revealed that the effects of sample time and treatment were highly significant (F1,209 = 178.3, P < 0.0001; F14,209 = 18.6, P < 0.0001) and affected the number of tetranychids. Furthermore, from the end of May, the number of tetranychids increased and reached significantly higher levels on the untreated vine trees than on the treated plants.

Figure 2. Density of E. carpini on phytoseiid release and control vine plants during the sampling period.

On the control vine trees, the highest E. carpini density was 124.60 mites/leaf at the end of July; two weeks later, on the release plants, the highest number of tetranychids (41 mites/leaf) was recorded. On the control plants, throughout the sampling period, the density of phytophagous mites remained always above the economic threshold of 6-10 specimens/leaf (Girolami 1981, Girolami et al. 1989); on the release vines, this was observed for the first time at the end of June. The presence of T. exhilaratus reached its maximum (3.03 mites/leaf) one month after the first release. From the beginning of August, the phytoseiid population decreased and was always lower than 1 mite/leaf in spite of the repeated introductions. This could be due to periods of extreme dry conditions, plant water stress and progressive increase in damage causing unsuitable acarodomatia on Petit Verdot leaves. Furthermore, genotypic characteristics of the introduced phytoseiid species could play an important role in their ability to establish (Tixier et al. 2002).

Difference in phytophagous populations was reflected also in damage levels (Figure 3). A very high correlation was found between the number of E. carpini motile stages and the damage levels registered which were higher in the control than in the release plants (Pearson coefficient = 0.783, N = 225, P<0.0001). In the release plants, during the whole vegetative period, the

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percentages of leaves without damage ranged from 57 to 76% till mid-July and were always slightly higher than those registered in the control plants even if only sporadic differences were detected (χ2 test, P = 0.05). In the following period, till October, all leaves were damaged but, significant differences were registered in the distribution of the damage levels. The most damaged leaves (score 4 and 5) were always detected on the control plants (χ2 test, P = 0.01).

Figure 3. Foliar damage observed on phytoseiid release and control vine plants during the experimental period. The cave chemical analysis (Table 1) gave similar results between the grapes gathered from experimental plants and those from the vineyard traditionally managed.

Table 1. Analysis of some parameters of the Petit Verdot grapevine conventionally managed and of the Petit Verdot grapevine where T. exhilaratus was released (due do the lack of replicates, no statistical analysis could be performed).

analysis parameter measure unit conventional management

T. exhilaratus release

acidity potential g/L tartaric acid 10.01 10.2 PH pH unit 3.07 3.1

reducing sugar total g/L 234.9 230.9 alcohol content % vol. 13.4 13.2

grape-berry weight g 1.2 1.2

0

20

40

60

80

100

Dam

age

%

release control

June July August September October

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In this study the release of T. exhilaratus significantly lowered tetranychid populations and reduced leaf damage. This confirms the findings of Prischmann et al. (2006) demonstrating the efficacy of two generalist phytoseiids, Typhlodromus caudiglans (Schuster) and Galendromus flumenis (Chant), in slowing down spider mite population growth in grapes. The expectation from a generalist phytoseiid such as T. exhilaratus was mainly to re-establish a prey-predator equilibrium rather than to exert a curative effect. Considering this perspective, the crucial point may be the timing of the introductions. Earlier and multi-year introductions of this predator could enhance establishment and thus provide eventual tetranychid control. Acknowledgements We are grateful to Pasquale Forte for financing the study and allowing experiment in his farm; to Andrea Bruni, Cristian Cattaneo and Luciano Guidotti for their assistance during the experiment. References Blumel, S. & Walzer, A. 2002: Efficacy of different release strategies of Neoseiulus californicus

McGregor and Phytoseiulus persimilis Athias Henriot (Acari: Phytoseiidae) for the control of two-spotted spider mite (Tetranychus urticae Koch) on greenhouse cut roses. Syst. Appl. Acarol. 7: 35-48.

Bonato, O., Cotton, D., Kreiter, S. & Gutierrez, J. 1990: Influence of temperature on the life-hystory parameters of the yellow grape-vine mite Eotetranychus carpini (Oudemans) (Acari: Tetranychidae). Int. J. Acarol. 16: 241-245.

Castagnoli, M. & Liguori, M. 1986a: Ulteriori indagini sull’acarofauna della vite in Toscana. Redia 69: 257-265.

Castagnoli, M. & Liguori, M. 1986b: Tempi di sviluppo e ovideposizione di Typhlodromus exhilaratus Ragusa (Acarina: Phytoseiidae) allevato con vari tipi di cibo. Redia 69: 361-368.

Castagnoli, M., Liguori, M. & Nannelli, R. 1999: Influence of soil management on mite populations in a vineyard agroecosystem. In: Bruin, J., van der Geest, L.P.S., Sabelis, M.W. (eds) Ecology and Evolution of the Acari., Kluwer Academic Publishers, The Netherlands, pp. 617-623.

Castagnoli, M., Liguori, M., Amato, F. & Guidi, S. 1991: Dinamica spaziale e temporale di Eotetranychus carpini (Oud.) (Acarina: Tetranychidae) e dei fitoseidi suoi predatori sulla vite. Atti XVI Congr. Naz. Ital. Ent, Bari-Martina Franca (TA), 23-28 settembre 1991: 339-345.

Castagnoli, M., Amato, F. & Monagheddu, M. 1989: Osservazioni biologiche e parametri demografici di Eotetranychus carpini (Oud.) (Acarina: Tetranychidae) e del suo predatore Typhlodromus exhilaratus Ragusa (Acarina: Phytoseiidae)in condizioni di laboratorio. Redia 72: 545-557.

Duso, C. 1991: Attività predatrice e dispersione di Amblyseius aberrans (Oud.) e Typhlodromus pyri Scheuten (Acari: Phytoseiidae) in un vigneto attaccato da Eotetranychus carpini (Oud.) (Acari: Tetranychidae). Atti XVI Congr. Naz. Ital. Ent, Bari-Martina Franca (TA), 23-28 settembre 1991: 355-362.

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Duso, C. 2006: Il controllo biologico e integrato degli acari fitofagi associati alla vite. In: La difesa della vite dagli artropodi dannosi. Ragusa, S. & Tsolakis, H. (Eds.), Marsala, 10-11 ottobre 2005: 171-204.

Duso, C. & Girolami, V. 1985: Strategie di controllo biologico degli acari tetranichide su vite. Atti XIV Congr. Naz. Ital. Ent., Palermo, Erice, Bagheria, 1985: 719-728.

Duso, C. & Vettorazzo, E. 1999: Mite population dynamics on different grape varieties with or without phytoseiids released (Acari: Phytoseiidae). Exp. Appl. Acarol. 23: 741-763.

Girolami, V. 1981: Danni, soglie di intervento, controllo degli acari della vite. Atti “III Incontro su la difesa integrata della vite”, Latina, 3-4 dicembre 1981, Regione Lazio: 111-143.

Girolami, V., Duso, C., Refatti, E. & Osler, R. 1989: Lotta integrata in viticoltura. Malattie della vite. Iripa ED., Mestre: 1-100.

Kondo, A. 2004: Colonizing characteristcs of two Phytoseiid mites, Phytoseiulus persimilis Athias-Henriot and Noseiulus womersley (Scicha) (Acari Phytoseiidae) on greenhouse grapevine and effects of their release on the kanzawa spider mite, Tetranychus kanzawai Kishida (Acari: Tetranychidae). Appl. Entomol. Zool. 39: 643-649.

Liguori, M. & Guidi S. 1995: Influence of different constant humidities and temperature on eggs and larvae of a strain of Typhlodromus exhilaratus Ragusa (Acari Phytoseiidae). Redia 78: 321-329.

Prischman, D.A., James, D.G., Wright, L.C. & Snyder, W.E. 2006: Effects of generalist Phytoseiid mites and grapevine canopy structure on spider mite (Acari: Teranychidae) biocontrol. Environ. Entomol. 35: 56-67.

SPSS, Inc. 1999: SPSS for Windows, v. 9.0 Chicago, Ill. Tixier, M.-S., Kreiter, S., Croft, B.A. & Auger, P. 2002: Colonization of vineyards by

Kampimodromus aberrans (Oudemans) (Acari: Phytoseiidae): dispersal from surrounding plants as indicated by random amplified polymorphism DNA typing. Agric. Forest Entomol. 4: 255-264.

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Biological control of the newly introduced persea mite with indigenous and exotic predators Yonattan Maoz1, Shira Gal1, Yael Argov2, Martin Berkeley2, Miriam Zilberstein3, Mickey Noy3, Yehonatan Izhar4, Jonathan Abrahams5, Moshe Coll6 and Eric Palevsky1

1Agricultural Research Organization (ARO), Newe-Ya’ar Research Center, Ramat Yishay, 30095, Israel, [email protected]; 2Israel Cohen Institute for Biological Control, Plant Production and Marketing Board, Citrus Division, POB 80 Bet Dagan, 50250, Israel; 3Extension Service, Ministry of Agriculture and Rural Development, POB 30 Bet Dagan, 50250, Israel; 4Western Galilee R&D, MP Oshrat, 25212, Israel;. 5Soil Conservation and Drainage Unit, Upper Galilee Ministry of Agriculture, Sefad, Israel. 6Department of Entomology, The Hebrew University of Jerusalem, POB 12, Rehovot 76100, Israel,

Abstract: Oligonychus perseae was first discovered in Israel on avocado trees in the autumn of 2001; by 2004 it spread to most of the important avocado growing regions. While field monitoring for persea mite we observed Euseius scutalis (Phytoseiidae) feeding on O. perseae within torn nests and outside of the nests. Subsequently, laboratory studies were performed to evaluate the efficacy of this predator. To improve persea mite control, the exotic predatory mite Neoseiulus californicus was released in 2004 and 2005. To determine whether other generalist predators can feed upon and tear the nests of persea mite, insect and arachnid predators were collected from avocado trees using a beating tray technique placed individually on newly infested leaf discs and monitored for several days. Although E. scutalis reduced adult persea mite populations in the lab (on leaf discs) with or without torn nests, egg predation was improved by tearing the nests. Seasonal CMDs following N. californicus releases were reduced by 30%, but leaf damage was still considerable and similar to control trees. Furthermore Phytoseiid predators recovered from all release plots were mostly of the species E. scutalis ranging from 78-95%. In our no-choice bioassays on leaf discs we observed nest tearing and predation by green lace wing Chrysoperla carnea, dusty wing Conwentzia sp. and others. Developing methods for augmentation and conservation of E. scutalis and nest-tearing predators may prove valuable for enhancing persea mite control. Key words: Oligonychus perseae, Euseius scutalis, Neoseiulus californicus, avocado, pollen. Introduction The persea mite, Oligonychus perseae Tuttle, Baker and Abbatiello, is a pest of avocado in Central and North America. The mite colonizes the bottom of the leaf, spinning densely woven nests along the leaf veins. The Hass cultivar is very susceptible whereas Reed is resistant (Kerguelen & Hoddle, 2000).

Oligonychus perseae was first discovered in Israel in the autumn of 2001 in several avocado plots located in the Western Galilee. Since then, it has spread to the growing areas (from North to South) of the Upper Galilee, Jezriel Valley, Efraim Hills, Carmel Coastal Plain, Hefer Valley and Rehovot-Lachish, causing extensive foliar damage and leaf drop in most of these regions.

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To identify the indigenous predatory mite fauna on avocado, a survey was conducted in 2002 and 2003. Subsequently, laboratory studies were performed to evaluate the efficacy of the dominant indigenous phytoseiid predator, Euseius scutalis (Athias-Henriot). To improve persea mite control, the exotic predatory mite Neoseiulus californicus (McGregor) was released in 2004 and 2005. Inundative releases of this predator have significantly reduced persea mite levels in California (Hoddle, et al. 2000).

Materials and methods Survey of indigenous phytoseiid predators In 2002-2003, pest and predatory mite populations were monitored fortnightly from 5 plots in the Upper and Western Galilee between April-November and once a month during the cooler months of December-March. At least 5 sites were sampled per plot, 1 tree per site, 10 leaves per tree, were randomly collected from tree perimeters. Pest mites were counted in situ using a quick field counting method (Machlitt 1998). For predatory mite count and identification, the 10 leaves were then washed in 400 cc of 70% ethanol, the plant matter discarded and the ethanol stored in plastic containers. In the laboratory, the ethanol wash was poured into a Petri dish and examined under a stereo microscope. Predatory mites were then removed with a micropipette, cleared in lactic acid and mounted in Hoyer’s medium. Phytoseiids were identified according to Swirski et al. (1998). Predation efficacy of Euseius scutalis on leaf discs While field monitoring for persea mite we observed E. scutalis feeding on persea mites within torn nests and outside of the nests. Preliminary laboratory leaf disc trials indicated that E. scutalis cannot enter intact nests of the persea mite. To determine the predation potential of E. scutalis when persea nests are intact vs. torn, we conducted efficacy trials on leaf discs. One day prior to the beginning of the experiment, 6 female persea mites were transferred to each leaf disc. By the next day, all mites were found within their densely woven nests with their freshly laid eggs. Four treatment combinations were compared: 1) intact nests with 1 female predator/disc; 2) intact nests without a predator; 3) torn nest with 1 female predator/disc; and 4) torn nest without a predator. The two-way factorial experiment (2 predation levels x 2 nest conditions) was replicated 20 times, The experiment was conducted for 5 days (the duration of the egg stage). Pest and predator eggs and adults were counted daily, then the nests in treatments 3 and 4 were gently torn using a fine needle, making sure not to damage any of the mites. Experiments were conducted at 24 ± 1°C, 43 ± 2% RH, 16:8 L:D. Data were sqrt(x+0.5) transformed before being subjected to ANOVA (JMP5.0.1a; SAS Institute, Inc.). Field evaluations of the exotic predatory mite Neoseiulus californicus One shipment of a few thousands mites of N. californicus were received from Koppert (Berkel en Rodenrijs, Netherlands) at the Israel Cohen Institute for Biological Control (ICIBC), November, 2002. This strain was originally collected in California, reared by Biotactics (Romoland, California) and then sent to and subsequently cultured by Koppert, Netherlands (KNL). At ICIBC, N. californicus were reared in small containers on Tetranychus cinnabarinus Koch eggs. Trials, conducted in plots in different geographic regions (5 plots in total), were initiated in spring to early summer when the first persea mites were detected. Two releases of 2000 mites/tree, with a fortnight interval between releases were performed. Pest and predator population levels were monitored fortnightly at each plot on 5 release and 5 control trees (blocked design, control tree located at least 2 trees away from release tree) as described under "survey of indigenous phytoseiid predators" above. Following the decline of pest mites, total

75

cumulative mite days (CMDs) of the pest were calculated (Palevsky et al. 1996). Using this parameter we evaluated the effects of predator release, location and their interaction. The block effect of each location was considered by nesting the block within location. Nest tearing and predation by insect and arachnid predators To determine whether predators can feed upon and tear the nests of persea mite, insect and arachnid predators were collected from avocado trees using a beating tray technique, placed individually on newly infested leaf discs (as described, under "laboratory predation efficacy trials of E. scutalis" above) and monitored for several days.

To determine the impact of these nest-tearing predators on field populations, the proportions of abandoned intact nests, ripped nests with and without live mites, were assessed in an organic avocado orchard (Kibbutz Gaaton) in 2006. Based on our laboratory observations, we interpreted: 1) the abandoned intact nests as nests that were not attacked by predators, 2) ripped nests with no live mite as nests that were attacked (motiles and eggs either eaten or fled) and 3) ripped nests that still contained live mites as nests that a predator frequented but did not consume all prey, creating an opportunity for intra-guild facilitation (i.e., providing opportunity for other predators to prey on the now exposed mites). In this orchard, as in the orchards that N. californicus were released (see above), no chemical treatment was applied for persea mite control.

Results and discussion Survey of indigenous phytoseiid predators In all five avocado plots, predatory mites were readily detected (Table 1). Euseius scutalis was by far the most predominant species accounting for more than 96% of all collected predatory mites (n = 1586). Table 1. Predatory mites found (number and percent of each species) in a survey of indigenous phytoseiid predators conducted in 2002-2003 in the Western and Upper Galilee, Israel.

Location Predatory mite species Amblyseius swirskii Typhlodromus athiasae Euseius scutalis Rosh Haniqra 0 1 67 Yechiam 0 46 154 Gaaton 0 3 634 Yodfat 2 5 45 Hagoshrim 0 2 627 Total 2 57 1527 Relative % 0.1% 3.6% 96.3%

Predation efficacy of Euseius scutalis on leaf discs For female adult persea predation, the interactive effects of predators (E. scutalis/control) and nests (intact/torn) were not significant (P = 0.384; F = 0.7; DF = 1,76). Euseius scutalis significantly lowered population levels of female adult persea mite (P < 0.0001; F = 732.6; DF = 1,76) regardless of whether the nests were torn (P < 0.0001; F = 448.5; DF = 1,38) or not (P <

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0.0001; F = 316.0; DF = 1,38) (Figure 1A, lower case letters). In contrast, the significant effect of tearing the nests on persea mite levels (P = 0.006; F = 7.9; DF = 1, 76) was probably due to handling as this effect was significant on the control discs (P = 0.013; F = 6.7; D F= 1,38) but not when the predators were present (P = 0.175; F = 1.9; DF = 1,38) (Fig. 1A, upper case letters).

The interaction between predator and nest treatments was significant for persea egg predation (P < 0.0001; F = 17.6; DF = 1,76). Euseius scutalis significantly lowered persea egg levels (P < 0.0001; F = 732.6; DF = 1,76) when nests were intact (P < 0.0001; F = 42.2; DF = 1,38) or torn (P < 0.0001; F = 246.6; DF = 1,38) but the effect of the latter was far more substantial (Figure 1B, lower case letters). Tearing the nests also affected predation efficacy (P < 0.006; F = 7.9; DF = 1,76); this is attributed to accessibility of the prey to the predator as the effect was significant on discs bearing the predator (P < 0.0001; F = 61.8; DF = 1,38) but not on the control discs (P = 0.151; F = 2.2; DF = 1,38) (Figure 1B, upper case letters).

Figure 1. Mean number of Oligonychus persea adults (A) and eggs (B) (plus standard errors) remaining after a 5 day period on leaf discs with and without one Euseius scutalis female adult. Nests were gently torn once a day, using a fine needle, taking care not to damage any of the mites. Within each panel (A and B), columns followed by a different lower case letter indicate a significant difference between predator and control treatments. Different uppercase letters are indicative of a significant difference between torn and intact nests within the same predator treatment (P < 0.05, Tukey’s test).

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Field evaluations of the exotic predatory mite Neoseiulus californicus The effects of N. californicus releases (P = 0.0022; F = 12.32; DF = 1,20) and location (P < 0.0001; F = 23.52; DF = 1,20) of release plots were both significant while their interaction was not (P < 0.0807; F = 1.33; DF = 1,20). Despite a significant reduction of 30% in seasonal CMDs following N. californicus releases, leaf damage was still considerable and similar to control trees. Phytoseiid predators recovered from all release plots were mostly of the species E. scutalis ranging from 78-95% (Figure 2). Nest tearing and predation by insect and arachnid predators In our no-choice bioassays on leaf discs we observed nest tearing and predation by green lace wing Chrysoperla carnea, dusty wing Conwentzia sp., a spider Chiracanthium mildei, a predatory thrips and heteropteran. The two latter species as well as the dusty wing are yet to be identified.

Figure 2. Proportion of phytoseiid predator species (indigenous E. scutalis and exotic N. californicus) recovered from five avocado release plots of N. californicus.

Nest tearing predators (see ripped empty nests, Figure 3) were active from the first detection of persea populations. From May through August 2006, equal proportions of nests were attacked (ripped empty nests, Figure 3) or not (intact empty, Figure 3) by predators. From September onwards, most of the nests were preyed upon, reaching a peak of 84% by the end of 2006. The level of ripped nest containing live mites was minimal throughout the season.

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Figure 3. Mean number of abandoned intact nests, and ripped nests with and without live mites per leaf (plus/minus standard errors) in an organic avocado orchard (Kibbutz Gaaton, 2006). The indigenous predatory mite E. scutalis is the dominant phytoseiid in avocado orchards in Israel. This was observed first in our survey in Western and Upper Galilee in 2002-2003 and then again when we released and monitored the recovery of the exotic N. californicus in all avocado growing regions in the country in 2004-2005. Although we did succeed in recovering N. californicus at all release sites, its effect on mitigating persea mite seemed minimal and its population levels were negligible compared to those of E. scutalis. Possibly, inter-specific competition between these two predators is preventing the establishment of N. californicus. While E. scutalis reduced adult persea mite populations in the lab (on leaf discs) with or without torn nests, egg predation was improved by tearing the nests. Intra-guild facilitation between nest-tearing predators and E. scutalis does not seem to be very important because the level of torn nests housing live prey was consistently low. Prey fleeing the torn nests, however, would serve as suitable food for leaf-grazing E. scutalis. Developing methods for augmentation and conservation of E. scutalis and nest-tearing predators may prove valuable for enhancing persea mite control. Euseius scutalis, as other species of Euseius, feed on avocado and other pollens that are abundant on avocado leaves from spring to early summer. Extending the period of pollen availability by the establishment of cover crops that would release wind-borne pollen, such as Rhodes grass, could be a viable way of keeping E. scutalis populations high, thereby preventing persea mite outbreaks (Smith & Papacek, 1991).

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Acknowledgements We are indebted to the avocado growers that participated and assisted in the field trials. We would like to acknowledge the Chief Scientist of Agriculture and the Plant Production and Marketing Board of Israel for their financial support. This manuscript is a contribution of the Institute of Plant Protection, Volcani Center, ARO, Israel.

References Hoddle, M.S., Robinson, L. & Virzi, J. 2000: Biological control of Oligonychus perseae (Acari:

Tetranychidae) on avocado: III. Evaluating the efficacy of varying release rates and release frequency of Neoseiulus californicus (Acari: Phytoseiidae). Int. J. Acarol. 26: 203-14.

Kerguelen, V. & Hoddle, M.S. 2000: Comparison of the susceptibility of several cultivars of avocado to the persea mite, Oligonychus perseae (Acari: Tetranychidae). Sci. Hortic. 84: 101-14.

Machlitt, D. 1998: Persea mite on avocados: quick field counting method. Subtropical fruit 6: 1-4.

Palevsky, E., Oppenheim, D., Reuveny, H. & Gerson, U. 1996: Impact of European red mite on Golden Delicious and Oregon Spur apples in Israel. Exp. Appl. Acarol. 20: 343-354.

Smith, D. & Papacek, D.F. 1991: Studies of the predatory mite Amblyseius victoriensis (Acarina: Phytoseiidae) in citrus orchards in south-east Queensland: control of Tegolophus australis and Phyllocoptruta oleivora (Acarina: Eriophyidae), effect of pesticides, alternative host plants and augmentative release. Exp. Appl. Acarol. 12: 195-217.

Swirski, E., Ragusa di Chiara, S. & Tsolakis, H. 1998: Keys to the phyotseiid mites (Parasitiformes, Phytoseiidae) of Israel. Phytophaga 8: 85-154.

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Biological control of the bulb scale mite Steneotarsonemus laticeps (Acari: Tarsonemidae) with Neoseiulus barkeri (Acari: Phytoseiidae) in amaryllis G.J. Messelink, R. van Holstein-Saj Wageningen UR Greenhouse Horticulture, P.O. Box 20, 2265 ZG Bleiswijk, The Netherlands; E-mail: [email protected] Abstract: The bulb scale mite Steneotarsonemus laticeps is a serious pest in the culture of amaryllis and some other flower bulbs. Earlier results indicated Neoseiulus barkeri to be a promising candidate for biological control of this pest. In this study we assessed the ability of N. barkeri and Amblyseius andersoni to control or restrict the bulb scale mite in amaryllis in a greenhouse experiment. The pest was introduced by interplanting infested bulbs, and the predators originated from laboratory rearing. Populations of bulb scale mites were monitored over a period of 15 weeks. Both phytoseiid mites reduced pest injury significantly, with N. barkeri being the more effective predator. Unlike A. andersoni, N. barkeri established a permanent population, although in low numbers. Key words: Steneotarsonemus laticeps, Neoseiulus barkeri, biological control, amaryllis Introduction The bulb scale mite, Steneotarsonemus laticeps (Halbert) (Acari: Tarsonemidae), is a serious pest in greenhouse cultures of amaryllis (Hippeastrum). This species is distributed throughout Europe, South Africa and on the West Coast of the USA (Lin & Zhang 2002). It also attacks other flower bulbs like Narcissus and Eucharis. These tiny mites (adult females about 200 µm long) cause dramatic growth inhibition and flower deformation. About 95 percent of the amaryllis nurseries in The Netherlands are contaminated with this pest. Since effective biological control agents are lacking, control relies heavily on chemicals. Phytoseiid predatory mites are the most obvious biological control agents for tarsonemids. Neoseiulus barkeri (Hughes) and Neoseiulus cucumeris (Oudemans) are known to be effective predators of Polyphagotarsonemus latus (Banks) in sweet pepper (Fan & Petitt 1994, Mizobe & Tamura 2004). Amblyseius andersoni (Chant), Neoseiulus californicus (Chant) and Neoseiulus fallacis (Garman) have potential to control Phytonemus pallidus (Banks) in strawberry (Croft et al. 1998). Efforts by growers to develop IPM strategies in amaryllis with the predatory mite N. cucumeris were not successful. In spite of repeated introductions of high rates, this predator never established. Consequently a search for new predatory mites suitable for amaryllis was started in 2005 (Messelink & van Holstein-Saj 2006). A survey of naturally occurring predatory mite species was conducted among Dutch amaryllis growers resulting in a list of 15 species. The most abundant was N. barkeri which was found both in soil and on bulbs and leaves, often associated with colonies of the bulb scale mite. A laboratory evaluation of a ranch of predatory mites showed N. barkeri to be the most effective predator of S. laticeps (Messelink & van Holstein-Saj 2006). Predation efficacy in this experiment was correlated with body size of the various species.

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The small body size and short hairs of N. barkeri are believed to enable this species to follow S. laticeps in hidden places deep inside the bulbs. The objective of this study was to assess the ability of predatory mites to control or limit the expansion of the bulb scale mite in amaryllis. Growers would appreciate to have a method for restricting “hot spots” of bulb scale mite during the vegetative stage of amaryllis, when chemicals are not effective. The phytoseiids A. andersoni and N. barkeri were evaluated for this purpose in a greenhouse experiment. Material and methods Cultures The predatory mites N. barkeri and A. andersoni were reared in a climate room on Acarus farris (Oudemans) and wheat bran. The bulb scale mite S. laticeps was reared on amaryllis bulbs cultivar “Mont Blanc” at 20ºC and 70% RH. Greenhouse experiment The effects of the predators N. barkeri and A. andersoni on bulb scale mite was examined in a greenhouse experiment in two separate compartments of 18 m2, each containing two tables (1 x 3 m) on which amaryllis bulbs were cultivated in plastic crates. The temperature (T) as well as the relative humidity (RH) were comparable in each compartment with only a minimal variation (mean T = 22ºC, mean RH = 63%). On each table three plastic crates (44 x 65 cm) were placed containing a 15 cm layer of clean potting soil. Hot water treated amaryllis bulbs of the white flowering cultivar “Mont Blanc” were placed in the potting soil in week number 2 of 2006. These bulbs were placed in 3 rows of 5 bulbs in each crate. In the central position was a tarsonemid contaminated bulb from the rearing colony, added in week number 5. Above each table a yellow sticky plate was hanged and weekly replaced. The plates were assessed under a binocular microscope and searched for bulb scale mites in association with flying insects.

The experiment was set-up as a randomized block design with four replicates. Each block contained one table with three treatments: the two predatory mite species and a control treatment. Every single crate was isolated by a continuously present water layer with the intention to avoid contamination between the treatments. Mites were released in week number 5 and 10 in rates of 1000 mites/m2 by sprinkling them on the soil between the bulbs. An extra release of 1000 mites/m2 was added for the A. andersoni treatment in week number 16.

Bulb scale mite damage (red coloring) was examined weekly by counting the number of damaged leaves or flower stems per bulb, without touching any plant, starting in week number 8. Flowers were removed during week number 12 and 13. A final assessment was done in week number 23 when every single bulb was uprooted and cut in half. Bulbs were examined thoroughly. The fraction of damaged leaves and the presence of bulb scale mites inside the bulbs (indicated by red coloring of plant tissue) was assessed. The second and fourth bulb from the middle row of each crate was more closely examined for the presence of predatory mites and other soil micro-arthropods. Bulbs and roots with attached soil were separately packed in plastic bags and transported to the laboratory. Micro-arthropods were extracted from these samples by heat and collected in ethanol, using Tullgren funnels, and counted under a binocular microscope (40x). Predatory mites were mounted on glass slides for microscopic determination.

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Statistics The fractions of damaged leaves at the final assessment were logit transformed. The numbers of micro-arthropods were transformed on a log scale. Analyses of variance (ANOVA) and Fisher least significant differences (LSD) tests were applied using GenStat Release 8.11. Results and discussion During the observation period the number of bulbs with visible damage strongly increased in the control plot, up to 70 % in week 22 (Figure 1). This increase was much lower in the mite treatments, reaching a maximum of 10 % in the plots with N. barkeri and about 30 % with A. andersoni (Figure 1).

0

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5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22

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s of

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b sc

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untreated controlN. barkeri A. andersoni

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1e release of predatory mites

Figure 1. Mean percentages of visibly damaged by bulb scale mites on amaryllis bulbs.

The number of damaged bulbs was much higher at the final assessment when all bulbs were sliced and examined carefully. In the control plots almost every single bulb was found colonized by the bulb scale mite. The number of damaged bulbs and leaves was significantly lower in the predator treatments, with N. barkeri significantly better than A. andersoni (Table 1). In the N. barkeri treatment only 4 % of the leaves were damaged compared to 57 % in the control. Still 45 % of the bulbs were colonized by the bulb scale mite. Neither N. barkeri nor A. andersoni appear to be able to completely eliminate a bulb scale mite infection in amaryllis. Neoseiulus barkeri colonized the amaryllis crop throughout the observation period, though in low numbers, A. andersoni could not be detected anymore from week 22, in spite of the additional introduction (Table 2). Some N. barkeri had contaminated the A. andersoni plots, in spite of the water barriers. Two other predatory mite species were detected, namely: Leioseius bicolor (Berlese) (♀ 310-400 µm long) and Lasioseius ometisimilis (Westerboer) (♀ 520 µm long). The effect of these predators on bulb scale mite is unknown. The lower numbers of L. bicolor in the predator plots

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suggest some kind of competition with the phytoseiids. Neoseiulus barkeri significantly reduced the number of oribatid and astigmatid mites, suggesting these mites might be an alternative food source, but the number of collembolans was not affected (Table 3). The results of this study show the potential of N. barkeri to establish in an amaryllis crop and restrict hot spots with bulb scale mites. Bulb scale mites could not be detected on sticky plates, suggesting that colonization of new bulbs occurs mainly via the soil and is thus a relatively slow process. Ways should be found to improve the establishment of this predator for further improving the quality of bulb scale mite control. Table 1. Mean percentage of leaves and bulbs injured by the bulb scale mite at the final assessment in week number 23. Means within a column are significantly different (p<0.05) if not followed by the same letter. mean percentage of injury treatment leaves bulbs control 57 a 98 a N. barkeri 4 c 45 c A. andersoni 19 b 70 b

Table 2. Mean number of predatory mites (±se) per amaryllis bulb at the final assessment in week number 23. treatment Neoseiulus

barkeri Amblyseius andersoni

Leioseius bicolor Lasioseius ometisimilis

control 0.4 (0.3) 0 (0) 11.5 (5.6) 0.5 (5.4) N. barkeri 0.9 (0.2) 0 (0) 0.9 (0.3) 0.4 (0.7) A. andersoni 1.6 (0.5) 0 (0) 3.0 (1.0) 0.6 (1.8)

Table 3. Mean number of soil micro-arthropods per amaryllis bulb at the final assessment in week number 23. Means within a column are significantly different (p<0.05) if not followed by the same letter. treatment Oribatic mites Astigmatic mites Collembolans control 45.5 a 29.6 a 40.0 a N. barkeri 7.1 b 10.6 b 25.0 a A. andersoni 17.3 a 10.8 b 23.5 a

Acknowledgements This study was supported by the Dutch Product Board for Horticulture. Pierre Ramakers is acknowledged for his comments.

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References Croft, B.A., Pratt, P.D., Koskela, G. & Kaufman, D. 1998: Predation, reproduction, and impact

of phytoseiid mites (Acari: Phytoseiidae) on cyclamen mite (Acari: Tarsonemidae) on strawberry. J. Econ. Entomol. 91: 1307-1314.

Fan, Y. & Petitt. F.L. 1994: Biological control of broad mite, Polyphagotarsonemus latus (Banks), by Neoseiulus barkeri Hughes on pepper. Biol. Cont. 4: 390-395. Lin, J.-Z. & Zangh, Z.-Q. 2002: Tarsonemidae of the World: Key to genera, Geographical distribution, Systematic Cataloque & Annotated Bibliography. Systematic & Applied Acarology Society, London. 440 pp. Messelink, G.J. & van Holstein-Saj R. 2006: Potential for biological control of the bulb scale mite (Acari: Tarsonemidae) by predatory mites in amaryllis. Proc. Neth. Entomol. Soc. Meet. 17: 113-118. Mizobe, M. & Tamura, I. 2004: Biological control of the broad mite Polyphagotarsonemus latus

(Banks) by Amblyseius cucumeris (Oudemans) on greenhouse sweet pepper. Kyushu Pla. Prot. Res..50: 62-65.

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Integrated Control of Plan –Feeding Mites IOBC/wprs Bulletin Vol. 30 (5) 2007

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Additivity versus interactions in mite-plant and predator-mite interactions Jay Rosenheim, Valerie Fournier Department of Entomology and Center for Population Biology, University of California, Davis, 95616, USA; [email protected] Abstract: A general question in ecology is whether the dynamics observed in communities of multiple interacting species can be predicted from an understanding of pairwise interactions among the component species. That is: can we predict the behavior of the ‘whole’ by examining the function of the ‘parts’. Here we address this question by examining two case studies. In the first case study, we examined how a plant, papaya (Carica payayae) responds to attack by three organisms, a rust mite Calacarus flagelliseta, a foliar pathogen, powdery mildew Oidium caricae, and a root pathogen, Phytophthora palmivora. Because a plant’s ability to resist infection or compensate for herbivory may be non-linear, we expected that interactions might be important. When all three antagonists were present simultaneously, the plant appeared to be overwhelmed and plant death was observed. In the absence of the root pathogen, however, rust mites and powdery mildew had largely additive effects on plant performance. In the second case study, we examined how an herbivore, the spider mite Tetranychus cinnabarinus, responded to attack by three organisms, a coccinellid beetle Stethorus siphonulus, a predatory mite Phytoseiulus macropilis, and a web-building spider Nesticodes rufipes. Because the web-spider eats both Tetranychus and the other predators, theory suggests that interactions may be important. Experiments demonstrated that the web-spider caused spider mite population growth rates to increase, apparently by suppressing populations of Stethorus. Populations of Phytoseiulus, however, appeared to be insensitive to the presence of the web-spider, and produced robust spider mite control. Thus, both of our case studies suggest that important multi-species interactions can occur, and that the emergent behavior of speciose communities may not be readily inferred from a study of its component parts. Descriptions of the experimental work can be found in these papers: Rosenheim, J.A., Limburg, D.D., Colfer, R.G., Fournier, V., Hsu, C. L., Leonardo, T.E. &

Nelson, E.H. 2004: Herbivore population suppression by an intermediate predator, Phytoseiulus macropilis, is insensitive to the presence of an intraguild predator: an advantage of small body size? Oecologia 140: 577-585.

Rosenheim, J.A., Glik, T.E., Goeriz, R.E. & Rämert, B. 2004: Linking a predator’s foraging behavior with its effects on herbivore population suppression. Ecology 85: 3362-3372.

Fournier, V., Rosenheim, J.A., Brodeur, J., Diez, J.M., & Johnson, M.W. 2006: Multiple plant exploiters on a shared host: testing for nonadditive effects on plant performance. Ecol. Appli. 16: 2382-2398.

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The history of the predacious Phytoseiidae mites in Israel Amos Rubin 14, Rahavat Ilan, Givat Shmuel 54056 Israel e-mail: [email protected] In the mid-1950’s the successful biological control of the Florida red scale, Chrysomphalus aonidum, by the imported parasitic wasp Aphytis holoxanthus left only the citrus rust mite Phyllocptruta oleivora as a main pest in citrus orchards. The need for research on this pest and its natural enemies, lead to research on Phytoseiidae that started in 1959 by the late Professor Eliahu Swirski (1921-2002). Until 1966 six Phtoseiidae were imported from Hong Kong, Chile, USA, Italy and India. Nine Israeli species were exported in these years to USA, Switzerland and USSR. In the 1990's five species were introduced from Australia, and two from USA, Italy and Spain. Mass produced of Euseius victoriensis was established in the north of Israel on citrus.

The biology of Israeli phytoseiids: Amblyseius swirskii, A. rubini=scutalis, Iphiseius degenerans, Typhlodromus athiasae and the imported Phytoseiulus persimilis, T. occidentalis and Phytoseius finitimus were studied intensively. Some other factors such as over-wintering and chromosome numbers were also researched. Few of these species were mass produced and released (for example, 200,000 specimens of P. persimilis were released in 200 locations, with good recovery). Thirty-four Phytoseiidae were found on wild vegetation. A rare phenomenon of female viviparous was observed in one phytoseiid species: Paragigagnathus tamaricus.

Some Israeli Entomologists who have been involved in phytoseiid mite research are (in alphabetical order): Shlomo Amitai, Yael Argov, Natan Dorzia, Uri Gerson, Tova Grinberg, Yonatan Maoz, Eric Palevski, Chaim Reuveni, Amos Rubin, Eliahu Swirski, Carmit Tal, Phyllis Weintraub and Manes Wysoki. In 1998 a key of 51 phytoseiid mites of Israel was published by E. Swirski, S. Ragusa and H. Tsolakis.

The research of Israeli Phytoseiidae was important to understand their role as a control factor in rust, bud and red mite pests, and also as a control measure of whiteflies and thrips in several vegetable and orchard crops. They are mass produced and released in glasshouses, to control pests instead of pesticides. As a result healthy food products are sold in the markets.

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A tritrophic perspective to the biological control of eriophyoid mites Maurice W. Sabelis, Izabela Lesna & Nayanie S. Aratchige Section Population Biology, Institute for Biodiversity and Ecosystem Dynamics, University of Amsterdam, Kruislaan 320, 1098 SM Amsterdam, The Netherlands Abstract: Vagrant eriophyoid mites are so small that they can move into narrow plant structures. Here, they would normally be safeguarded from attack by predators, such as phytoseiid mites, because these are much larger than the eriophyoid mites. Recent experimental evidence has been obtained that plants do not passively undergo the attack by these plant parasites, but respond by altering the relevant plant structure so as to promote access of phytoseiid predators. Not all species of phytoseiid mites can take advantage of this plant response, but some species can. This suggests particular morphological and behavioural traits that enable these phytsoseiid species to get access to the sites where eriophyoid mites hide. These new insights may provide a new tritrophic perspective to the biological control of eriophyoid mites. Introduction Eriophyoid mites are about the smallest arthropods on Earth (Lindquist et al. 1996). Their worm-like body has a cross-section diameter of ca 50 µm, which is at least five times smaller than that of phytoseiid mites, one of their most significant predators (Sabelis 1996). This minute size of the eriophyoid mite is the key to their ecological success. It enables them to reach places, small enough to be free of predators (Sabelis 1996, Sabelis & Bruin 1996). Moreover, it allowed them to develop a plant-parasitic life-style quite different from other herbivorous arthropods (Lindquist et al. 1996). Many eriophyoids live in the plant galls they induce, but the ones of interest in this overview have a vagrant life-style, in that they frequently change feeding sites that vary in the degree of protection against predators. Under agricultural conditions this type of eriophyoid mites may easily reach pest status when predatory mites are lacking (Lindquist et al. 1996). Examples are Aceria tulipae on tulip bulbs, Aceria guerreronis on coconut palms and Aculops lycopersici on tomato plants. Chemical control is also bound to be less effective due to the (partially) hidden life style of the eriophyoid mites. Hence, biological control with predatory mites is an option. However, several attempts to apply this method of control have failed and there are very few known cases of successful biological control (Lindquist et al. 1996). We will argue that this is not so much due to an inadequate functional and numerical response of phytoseiid predators to the density of eriophyoid mites. It is also not due to the absence of phytoseiid species that have a preference for eriophyoid mites as prey. But, as we expect, the perspectives for phytoseiid predators as biological control agents of eriophyoid mites will improve once we understand how host plants respond to attack by eriophyoid mites, and thereby promote the effectiveness of phytoseiid predators, and once we understand which and how phytoseiid predators take advantage of the herbivore-induced changes in the host plant.

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Tulip bulbs and Aceria tulipae Consider tulip bulbs under attack by Aceria tulipae, an eriophyid mite so tiny that it can move

in between the scales of the bulbs (Conijn et al. 1996). Herbivory induces these bulbs to modify their internal structure. The resulting changes in distance between bulb scales are microscopic, yet sufficient to allow the phytoseiid predator Neoseiulus cucumeris to enter the interior space of the bulb. The consequence of this plant ‘behaviour’ is dramatic: with predators the inside of the bulb is cleared from herbivores, but – without – the bulb is eaten from within and dries out. The crucial changes in bulb morphology enabling predator access are controlled by ethylene, a plant hormone released upon herbivore attack (Lesna et al. 2005). This plant hormone simultaneously induces the release of plant volatiles that attract predatory mites. Changes in attractivity and accessibility of bulbs were demonstrated by a combination of chemical analysis (Aratchige 2007), olfactometry (Aratchige et al. 2004) and experiments on predator-prey dynamics in which the effect of ethylene was either promoted or blocked (Lesna et al. in prep.). Coconuts and Aceria guerreronis

Consider the coconut mite, Aceria guerreronis (Aratchige 2007) that lives under the perianth (collectively, the tepals that are often referred to as bracts) of the fruit of the coconut. The perianth of the nut protects the meristematic zone of the female flower and the developing nut thereafter. The edge of the perianth is so closely oppressed to the fruit that it leaves even less space for any mite to enter. However, the coconut mite is able to creep through to enter under the perianth. However, some predators can move under the perianth of the coconuts and attack the coconut mite. In Sri Lanka, the phytoseiid mite, Neoseiulus baraki, is the most common predatory mite that is found in association with the coconut mite. This predatory mite is ca 3 times larger than the coconut mite. Nevertheless, taking this predator’s flat body and elongated idiosoma with short distal setae into account, it is - relative to many other phytoseiid mites – better able to reach the narrow space under the perianth of infested nuts. On uninfested nuts, however, they are hardly ever observed under the perianth. Prompted by earlier work on the accessibility of tulip bulbs to another eriophyoid mite and its predators, we hypothesized that the nuts change their morphology in response to damage by eriophyoid mites and as a result allow predatory mites to enter under the perianth of infested nuts. This was tested in an experiment where we measured the distance between the perianth and the coconut fruit surface in 3 cultivars (Ordinary Tall, Dwarf Green and Dwarf x Tall hybrid) that are cultivated extensively in Sri Lanka (Aratchige 2007). In the uninfested nuts this distance was large enough for the coconut mite to creep under the perianth, yet too small for its predator N. baraki. However, when the nuts were infested by coconut mites, the perianth-fruit distance increased to such an extent that also the predatory mites could move under the perianth. Discussion The lessons from analysing the tulip bulb response to eriophyoid mites do not only seem to carry over to coconuts, but they may also apply to host plants protected by dense covers of glandular hairs, such as tomato. Glandular hairs are a very effective defense against many herbivorous mites (Chatzivassileiades et al. 1997, 1998, 1999, 2001), but they also hinder predatory mites (Van Haren et al. 1987). Eriophyoid mites, such as Aculops lycopersici, are so small that they can move in between the glandular hairs without being affected. This eriophyoid mite is currently an important pest in tomato crops and the use of phytoseiid predators for their biological control has failed so far. We are currently investigating whether tomato plants alter the glandular hairs in

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response to attack by Aculops lycopersici and whether these changes positively affect the access to these herbivores for some species of phytoseiid predators.

References Aratchige, N.S. 2007: Predators and the accessibility of herbivore refuges in plants. PhD Thesis,

University of Amsterdam, ISBN 978-90-76894-70-6. Aratchige, N.S., Lesna, I. & Sabelis M.W. 2004: Below-ground plant parts emit herbivore-

induced volatiles: Olfactory responses of a predatory mite to tulip bulbs infested by rust mites. Exp. Appl. Acarol. 33: 21-30.

Chatzivasileiadis, E.A. & Sabelis M.W. 1997: Toxicity of methyl ketones from tomato trichomes to Tetranychus urticae Koch. Exp. Appl. Acarol. 21: 473-484.

Chatzivasileiadis, E.A. & Sabelis, M.W. 1998: Variability in susceptibility among cucumber and tomato strains of Tetranychus urticae Koch to 2-tridecanone from tomato trichomes: Effects of host plant shift. Exp. Appl. Acarol. 22: 455-466.

Chatzivasileiadis, E.A., Boon J.J. & Sabelis, M.W. 1999: Accumulation and turnover of 2-tridecanone in Tetranychus urticae Koch and its consequences for resistance of wild and cultivated tomatoes. Exp. Appl. Acarol. 23: 1011-1021.

Chatzivasileiadis, E.A., Egas, M. & Sabelis, M.W. 2001: Resistance to 2-tridecanone in Tetranychus urticae Koch: effects of induced resistance, cross-resistance and heritability. Exp. Appl. Acarol. 25: 717-730.

Conijn C.G.M., van Aartrijk J. & Lesna I. 1996: Flower bulbs. - In: E.E. Lindquist, M.W. Sabelis, J. Bruin (eds.), “Eriophyoid mites - Their biology, natural enemies and control.” Elsevier Science Publishers, Amsterdam, 651-658.

Lesna, I., Conijn C.G.M. & Sabelis, M.W. 2005: From Biological Control to Biological Insight: Rust-mite induced change in bulb morphology, a new mode of indirect plant defence? In: G. Weigmann, G. Alberti, A. Wohltmann & S. Ragusa (eds), Acarine Biodiversity in the Natural and Human Sphere. Phytophaga (Palermo) 14: 285-291.

Lindquist, E.E., Sabelis, M.W. & Bruin J. (eds.) 1996: Eriophyoid Mites - Their Biology, Natural Enemies and Control. World Crop Pest Series Vol. 6, Elsevier Science Publishers, Amsterdam, The Netherlands, 790 + xxxii pp.

Sabelis, M.W. 1996: Phytoseiidae. In: E.E. Lindquist, M.W. Sabelis & J. Bruin (eds.). Eriophyoid Mites - Their Biology, Natural Enemies and Control. World Crop Pest Series Vol. 6, Elsevier Science Publishers, Amsterdam, The Netherlands, pp. 427-456.

Sabelis, M.W. & Bruin, J. 1996: Evolutionary ecology: life history patterns, food plant choice and dispersal. In: E.E. Lindquist, M.W. Sabelis & J. Bruin (eds.). Eriophyoid Mites - Their Biology, Natural Enemies and Control. World Crop Pest Series Vol. 6, Elsevier Science Publishers, Amsterdam, The Netherlands, pp. 329-366.

Van Haren, R.J.F., Steenhuis, M.M., Sabelis, M.W. & de Ponti O.M.B. 1987: Tomato stem trichomes and dispersal success of Phytoseiulus persimilis relative to its prey Tetranychus urticae. Exp. Appl. Acarol. 3: 115-121.

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The effects of varieties and agronomic practices on acarine populations in Italian vineyards Sauro Simoni, Marisa Castagnoli C.R.A., Istituto Sperimentale per la Zoologia Agraria, lab. Acarology, via di Lanciola 12/a, 50125 Florence Italy, email: [email protected] Abstract: The effects of the varieties, Nero d’Avola, Verdicchio, Fiano, Refosco and management practices (conventional, organic, no pesticides) on acarine species colonization and abundance were assessed in vineyards in different growing regions from Northern to Southern Italy. Verdicchio was the most inhabited variety by phytoseiids, Refosco by tetranychids and tydeids while Fiano was the less colonized by eriophyids. Phytoseiid mites showed significantly higher densities per leaf in untreated vineyards, while no difference was detected between biological and conventional vineyards. On the contrary, tydeids and tetranychids had the highest density in conventional vineyards. Key words: mites, organic vineyard, conventional vineyard, cultivars Introduction The predominant trends in Italian viticulture are to increase the quality and the typicality of the final product. Fine local varieties and the shift from conventional to organic grape production are the two important elements in this process.

A multidisciplinary, national scale project (PROVIT), initiated in 2003, has focused on the effects of widespread Italian cultivars and vineyard management on quality of wine. Within this project the task of the laboratory of Acarology of the Istituto Sperimentale per la Zoologia Agraria of Florence was to evaluate cultivar and management influence on the diversity and abundance of the vineyard acarofauna.

The acarofauna of vine in Italy and its important role on the vineyard ecosystem has been intensively studied in some growing regions of Italy (Castagnoli 1988, Castagnoli et al. 1999, Duso et al. 2003, 2004, Nicotina & Capone 2003, Ragusa & Ciulla 1991). To increase our understanding, additional studies are needed. These should encompass all growing regions, use standardized methodologies, evaluate the relevant local varieties now in demand and assess the effect of new pest management practices presently being adopted.

In this paper we assessed the effects of three management practices (conventional, organic and no pesticides) and four standardized important autochthonous varieties of vine on mite populations in different growing regions from the Northern to Southern Italy devoted to high quality viticulture.

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Material and methods Mites and varieties – Provit A In 13 different geographical areas (Figure 1), we sampled four experimental 15-year-old vineyards where IPM was adopted for pest control. In each vineyard only one variety was grown, i.e. Nero d’Avola, Verdicchio, Fiano or Refosco. All varieties are autochthonous and high quality wine producing in the considered geographical areas. Two times per year (at first grape and ripening), 16 leaves of vine per 3 replicates were randomly sampled in the middle of the canopy. All mites found on the plant material were grouped according to the dominant acarine families found in vineyards, namely, Phytoseiidae, Tydeidae, Tetranychidae, Eriophyidae and others. The study was conducted for three years. Mites and pest management - Provit B The experiment was conducted with procedures analogous to the Provit A set up (16 leaves x 3 replicates, experimental time three years). The samples were three per years (at first grape growing, at the beginning of ripening and at the end of ripening). We compared three different management systems (conventional or IPM, organic and untreated) in 4 site locations (Figure 1) at the north-west, north-east, centre and south of Italy. The varieties chosen, among the most common in Italy, were different in each locality and were in the order Barbera, Merlot, Sangiovese, Malvasia.

Provit A - Nero d’Avola

- Verdicchio

- Fiano

- Refosco

Provit B - Ba, Barbera

- Me, Merlot

- Sa, Sangiovese

- Ma, Malvasia

Figure 1. Site locations and vine varieties involved in Provit Data analysis General Linear Model Analyses were applied to data obtained both from Provit A and B trials to evaluate effects of factors considered. First, in Provit A, it was evaluated how the cumulative number of mites was affected by sample time, leaf/replicate, site location, vine variety. In Provit

63 5Me

Ba

Sa

Ma

2 4

7 8

9

11 12

13

10

1

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B, it was analysed how the same response was affected by sample time, leaf/replicate, site location, treatment. Then, Univariate Analysis of Variance was performed to evaluate the significant factors emerged from the general analysis on abundance of single groups of mites: phytoseiids, tydeids, tetranychids, eriophyids, other mites in Provit A; phytoseiids, tydeids, tetranychids in Provit B. The difference in the separate analyses of groups was due to the sporadic presence of eriophyids and other mites in Provit B: eriophyids were 0.14% of the cumulative number of mites, other mites 0.09%. All data were transformed by ln(y+1), and all statistical procedures were performed with SPSS (1999). Results and discussion Figure 2 reports the main mite groups collected. On the whole, more than 100,000 mites were sampled on 10,384 vine leaves in Provit A project development, more than 15,800 on 4,320 leaves in Provit B. Although in Provit A the cumulative number of eriophyids recorded (only the rust mite was considered) appeared the highest, it was strongly affected by their consistent abundance in only one location of Central Italy where 99% of these mites were collected. The phytoseiid predators followed by tydeids, mites with unspecialized diets, and the tetranychid herbivores were the most representative groups in both the experiments. The ‘other mites’ represented a heterogeneous group, including mites with very different feeding habits and behaviour; on the whole, these species were always rare and with very low populations. Figure 2. Cumulative number of mites collected in the two trials Provit A Tests of between-subjects dffects showed a significant incidence of the sample time during vegetative period, the location and the vine cultivar (for these 3 factors, P < 0.0001) on the abundance of mites, while neither replicate nor leaf were significant (P = 0.337 and 0.068,

1

10

100

1000

10000

100000

phytoseiids tydeids tetranychids eriophyids other mites

Provit AProvit B

ln (N

. mite

s)

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respectively). The same test separately performed for each mite group, showed high significance of the same factors (P < 0.0001). In addition replicate showed significant incidence in all groups (P = 0.01 in phytoseiids, tydeids and other mites, P < 0.0001 in tetranychids and eriophyids). Furthermore, for phytoseiids, probably due to their more clumped distribution, leaf (P = 0.01) was a noteworthy factor affecting the density even if the interaction leaf*replicate was not significant (P = 1.000). This interaction, at the same, was insignificant in the other mite groups.

By leaving out the eriophyids and considering the other groups and the different localities, on the whole, phytoseiids were recorded in all samples and showed the highest density as well as the largest range of values with the greatest stratifications of significance. Considering the distribution of the different site locations, in Northern Italy (see points 1-6 in Figure 1) the average highest presence was registered with 3.40 phytoseiids/leaf and the wider range, 0.75-7.14; in Central Italy (points 7-9, Figure 1) 2.65 phytoseiids/leaf was the mean, range 1.94-3.44; in Southern Italy (points 10-13, Figure 1), 1.73 phytoseiids/leaf was the mean, range 0.28-4.21. The cumulative means per geographical area seemed to suggest a decrease of phytoseiids density from north to south, probably coincidently with the more and more dry ambient condition. However, it was not possible to tie the Anova results to the different latitude ranges of site locations: other more complex factors interfered with climatic condition of the site location.

Verdicchio was the variety most colonized by phytoseiids, Refosco by tetranychids and tydeids; Fiano was the less colonized by eriophyids. The heterogeneous group of other mites, recorded always with very low density, was found at the highest density on Verdicchio (Table 1).

Table 1. Provit A: varieties and mite groups. (mites/leaf; Anova, Tukey test, P=0.05)

Variety Phytoseiids (mean±SE)

Tydeids (mean±SE)

Tetranychids(mean±SE)

Eriophyids (mean±SE)

other mites (mean±SE)

Nero d'Avola (N=2464) 2.55±0.01a 0.24±0.02a 0.09±0.01a 8.57±2.68a 0.02±0.004a

Verdicchio (N=2816) 3.26±0.10c 0.36±0.02b 0.11±0.02a 9.09±2.52a 0.04±0.005b

Fiano (N=2608) 2.96±0.09b 0.82±0.05c 0.15±0.02a 0.0004±0.000

4b 0.01±0.003a

Refosco (N=2496) 2.44±0.08a 0.55±0.04d 0.23±0.04b 10.72±3.26a 0.01±0.003a

Provit B The cumulative number of mites recorded was significantly affected by sample, locality/variety and treatment (main effects for all factors, P<0.0001). By considering separately the most important groups, also replicate was significant, but not leaf and its interaction with replicate. Similarly to what happened in Provit A, this could be explained by the patchy distribution of the single group in comparison with the totality of specimens found. The highest number of phytoseiids and tydeids was

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Table 2. Provit B: sample time, site location, treatment and mite groups (mites/leaf; Anova, Tukey test, P=0.05)

Sample time N Phytoseiids

(mean±SE) Yydeids

(mean±SE) Tetranychids (mean±SE)

I 1488 1.20±0.07a 1.23±0.13a 0.92±0.08a II 1536 1.23±0.06a 0.78±0.06a 0.75±0.09b III 1296 2.63±0.14b 2.20±0.19b 0.21±0.03c

Location/variety

North-western Italy, Ba 288 3.79±0.22a 4.18±0.39a 0.00±0.00a North-eastern Italy, Me 1200 0.96±0.05b 1.63±0.08b 1.88±0.15b

Centre Italy, Sa 1728 1.07±0.66b 1.51±0.16c 0.30±0.04c South Italy, Ma 1104 2.70±0.17c 0.08±0.01d 0.007±0.003a

Treatment

organic 1456 1.29±0.07a 1.01±0.09a 0.84±0.06a conventional 1216 1.50±0.09a 1.71±0.22b 1.18±0.13a

untreated 1648 2.04±0.11b 1.40±0.08ab 0.08±0.01b sampled at the end of ripening, while that of tetranychids at the beginning of grape increase (Table 2).

Concerning locality/variety, the density of phytoseiids and tydeids are highest on Barbera, while tetranychids on Merlot (Table 2). Only phytoseiid mites showed a density per leaf significantly higher in untreated vineyards, while no difference was detected between biological and conventional vineyards. On the contrary, tydeids and tetranychids surprisingly had the highest density in conventional vineyards; furthermore tetranychid population was significantly lower where phytoseiids density was higher.

In IPM vineyards the phytophagous mite populations usually remained below the damage threshold. Both in Provit A and B, tetranychids rarely overcame one specimens/leaf and eriophyid 10 specimens/leaf. Their natural enemies, the phytoseiids, were consistently present as were the tydeids, both playing important roles in the maintenance of mite population equilibrium on grapevine (Duso et al. 2005). The well known role of variety to determine mite distribution (Castagnoli & Liguori 1985, Duso & Vettorazzo 1999) is confirmed. Although the variety significantly affected the abundance of all mite groups, none of the varieties in Provit A seemed to dramatically affect the density of mites: on the average, neither phytophagous mites were particularly favoured nor beneficial mites particularly lowered and always the ratio was such as to guarantee favourable control by the predators. In Provit B different varieties from Provit A were considered, but in this case their effects were not separable from locality effects. The organic management of vineyards did not increase any problem with mites even if a small (but not

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significant) decrease was observed both in the phytoseiid and tetranychid populations in comparison with conventional management.

The next planned study of mite species composition of Provit A and B presumably will allow us to further gain considerable information on the effect of varieties and vineyard managements on grapevine mites.

References Castagnoli, M. 1988: Recent advances in knowledge of the mite fauna in the biocenosis of the

grapevine in Italy. In: Cavalloro R (ed) Influence of Environmental Factors on the Control of Grape Pest, Diseases and Weeds. Balkema, pp. 169-180.

Castagnoli, M & Liguori, M. 1985: Prime osservazioni sul comportamento di Kampimodromus aberrans (Oud.), Typhlodromus exhilaratus Ragusa, Phytoseius plumifer (Can. & Fanz.) (Acarina: Phytoseiidae) sulla vite in Toscana. Redia 79: 361-368.

Castagnoli, M., Liguori, M. & Nannelli, R. 1999: Influence of soil management in mite population in a vineyard ecosystem. In: Bruin J., van der Geest L.P.S., Sabelis M.W. (eds) Ecology and Evaluation of the Acari. Kluwer, pp.617-623.

Duso, C. & Vettorazzo, E. 1999: Mite population dynamic on different grape varieties with or without Phytoseiidae released (Acari: Phytoseiidae). Exp. Appl. Acarol. 23:741-763.

Duso, C., Fontana, P. & Malagnini, V. 2004: Diversity and abundances of phytoseiids mites (Acari: Phytoseiidae) in vineyard and surrounding vegetation in north-eastern Italy. Acarologia 44: 31-47.

Duso, C., Pozzebon, A., Capuzzo, C., Malagnini, V., Otto, S. & Borgo, M. 2005: Grape downy mildew spread and mite seasonal abundance in vineyard: effects on Tydeus caudatus and its predator. Biol. Cont. 32: 143-154.

Duso, C., Malagnini, V., Drago, A., Pozzebon, A., Galbero, G., Castagnoli, M. & de Lillo, E. 2003: The colonization of phytoseiid mites (Acari Phytoseiidae) in a vineyard and the surrounding hedgerows. IOBC-WPRS Bulletin 26(4): 37-42.

Nicotina, M. & Capone, C.G. 2003: Species and population density of grapes and surrounding vegetation in a vineyard of phytoseiid mites (Parasitiformes, Phytoseiidae) of grapes and surrounding vegetation in a vineyard of the Campania region (Southern Italy). Phytophaga XIII: 3-12.

Ragusa, S. & Ciulla, A.M. 1991: Phytoseiid mites associated with vine in Sicilian vineyards. In: Schuster R., Murphy P.W (eds) The Acari. Reproduction, Development and Life-History Strategies. Chapman & Hall, pp. 417-423.

SPPS 1999. SPSS for Windows, v. 9.0. SPSS Inc., Chicago, Ill.

Integrated Control of Plan –Feeding Mites IOBC/wprs Bulletin Vol. 30 (5) 2007

pp. 101-110

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Seasonal quality assessment of mass-produced Phytoseiulus persimilis Shimon Steinberg, Hadassa Cain Bio-Bee Sde Eliyahu Ltd. Kibbutz Sde Eliyahu, Bet Shean Valley 10810, Israel

Abstract: A life-time fecundity test was conducted with the commercially-produced predatory mite Phytoseiulus persimilis in order to assess its intrinsic quality in light of a new concept that categorizes egg laying females to young and old individuals according to the duration of their ovipositional period. Phytoseiulus persimilis females from two standard production batches were sampled for the test. Within each batch the fecundity of freshly harvested females (control treatment) was compared with that of stored predators, the latter simulating the logistic chain of the commercial product exported to Europe. Total egg production and duration of ovipositional period were closely related both in the control and storage treatments within the two batches. The distribution of reproductive age categories showed in the two batches ca. 50% non-reproducing and old 5-day ovipositing females in the freshly harvested material and ca. 40% of the same categories from the stored mites. Around 20-30% of the freshly harvested predators belonged to the young >20-day ovipositing females, whereas the stored individuals in this category consisted ca. 40% of the tested population. The average number of eggs per reproductive category increased gradually from old to young reproductive ages, both in the freshly harvested and stored mites, reaching a maximum of ca. 90 eggs at 25 oviposition days in the two batches. The reproductive rate of a female predator in each reproductive category met the IOBC standard of 2 eggs/female/day in all reproductive categories at both batches and in the stored and non-stored predators. The calculated potential yield of eggs from 2,000-female predators in a commercial product was 66,000 and 51,000 eggs for freshly harvested P. persimilis in batches A and B, respectively and ca. 82,000 eggs for the commercial product of stored material. The results of this study show that reproductively young females survived cold storage better than the old ones. Furthermore, they indicate good intrinsic quality and high colony vigor of the mass produced P. persimilis. Key words: Phytoseiulus persimilis, fecundity, product control, storage capacity Introduction During spring 2006, problems regarding field performance of the predatory mite Phytoseiulus persimilis (commercial product named SPIDEX) were reported from different places in north-western Europe. As part of the efforts to pin-point the possible reason(s) for those problems, a thorough study was launched at Bio-Bee to test the intrinsic quality of P. persimilis. Based on the reproductive biology of P. persimilis, Luczynski et al. (2006) presented a new interpretation of life-time fecundity test results, namely that egg laying females can be categorized to young and old individuals with respect to the duration of their ovipositional period. This categorization has a great impact on assessment of the quality of a given production batch of P. persimilis, especially regarding egg output of the commercial product and the effect on storage capacity of the predatory mite. The objectives of the study reported herewith are: i) to assess the intrinsic quality of P. persimilis by applying the standard IOBC fecundity test; ii) to analyze the results according to

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and compared to the analysis presented by Luczynski et al. (2006); iii) to interpret the results in light of the problems that were raised in the field. Materials and Methods Source of P. persimilis Predatory mites were sampled from two standard production batches produced under greenhouse conditions in a traditional tri-trophic system comprising of bean plant, red spidermite and the predatory mite: i) batch A harvested on 28/4/2006; ii) batch B harvested on 7/5/2006. Treatments From every production batch individual P. persimilis females were sampled according to the following treatments: i) control – freshly harvested predators; ii) storage – P. persimilis kept in storage as bulk material for 2-3 days at 8 ºC, than formulated in bottles of the commercial product than kept for another 3 days at 8 ºC, simulating the logistic chain from Bio-Bee to end-users in Europe (Table 1). At the end of this period, female predators were sampled from the commercial bottle for the fecundity test. Test procedure The fecundity test was carried out in a controlled climate chamber according to the standard procedure described in the IOBC guidelines for product control of commercially produced natural enemies (van Lenteren et al. 2003). Test conditions were: temperature 25 ºC, RH 60 ± 10% and long day (16 hours light). The test arenas (individual brown bean leaf discs) were checked daily, eggs of P. persimilis were counted and the condition of the female predator (live or dead) was assessed. The trial continued for either 30 days or till the "natural" death of the female predator, whoever occurred first. Individual females (=replicates) that escaped or died due to handling were not taken into account. Table 1: A detailed time-table of the treatments from which P. persimilis adult females were sampled for the fecundity test. "n" refers to the number of individuals tested per treatment.

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in bottles

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Results and discussion In order to determine the reproductive age of P. persimilis females it was essential to study the relationship between total egg production and duration of the ovipositional period. These two

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parameters were very closely related in both the control and storage treatments within the two batches that were tested (Figures 1 and 2).

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y = -0.0098x3 + 0.3026x2 + 2.0597x - 3.0136R2 = 0.9691

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The predators were grouped into 5 categories: 0 was used for old mites that did not produce eggs at all whereas 5, 10, 15, 20, 25 and 30 categorize the reproductive age in increments of 5 days, five being old female predators and 30 very young females.

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In batch A it appears as if the population was composed of two separate generations of females, from both the non-stored and stored mites. One generation consisted of old females, from those that did not lay eggs at all (ca. 21% in the control treatment) to those that laid eggs for maximum 10 days. The majority of females from this generation were old individuals that laid

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eggs for maximum 5 days with the proportion of 37% and 41% for non-stored and stored mites, respectively (Figure 3A). The second generation included young females that laid eggs along 20-30 days, the largest proportion belonging to those that laid eggs along maximum 20 days, 25% and 32% for non-stored and stored mites, respectively (Figure 3A). In batch B the two-generation pattern was not evident. Old non-producing females were below 5% of the total population of females tested for both non-stored and stored predators. Still, the majority of female predators belonged to relatively old individuals that laid eggs up to a maximum of 5 days, 53% and 42% for non-stored and stored P. persimilis, respectively (Figure 3B). In this batch 37% of the stored females were of young reproductive age, i.e. laid eggs along 20 and 25 days (Figure 3B). Although the pattern of the highest proportion of females being of old reproductive age (5-day oviposition) was similar between the current study and the study of Luczynski (Anna Luczynski, personal communication), the proportion of females in young reproductive age (20, 25 oviposition days) was by far higher in the current study compared to Luczynski's figures (20-41% vs. <5%, respectively). The average number of eggs per reproductive category increased gradually from old to young reproductive ages, reaching a maximum of 95 and 88 eggs at 25 oviposition days in batches A and B, respectively (Figure 4). The number of eggs per category did not differ significantly between non-stored and stored female predators (Figures 4A and 4B). The reproductive rate per female predator (= number of eggs per day) in each reproductive category met the IOBC standard of 2 eggs/female/day in all reproductive categories at both batches and in the stored and non-stored predators (Figure 5). The only exception was the oldest females of the freshly harvested material in batch B. Their average eggs/day value was 1.6 (Figure 5B). The younger females (from 10 oviposition days onwards), at both batches in both treatments, exceeded the standard by far, yielding 3 eggs/day and above (Figures 5A and 5B). These figures are a positive indication to the culture's vigor and health. To determine the average number of eggs produced by 2,000 P. persimilis females (the content of a single commercial bottle), the proportion of each age category was multiplied by the average number of eggs produced in each category. Figure 6 demonstrates that the females from the non-stored freshly harvested material produced ca. 66,000 and 51,000 eggs per bottle in batches A and B, respectively whereas the stored females of both batches, following the logistic chain, yielded around 82,000 eggs per bottle. The high numbers obtained in this study are due to the relatively high proportion of young-fecund females. This is especially reflected in the stored material that contains high proportion of young and fecund female which survived the logistic chain. A potential source for variation in results between different P. persimilis products/producers is the different methodologies that are used to choose the female predators for the fecundity test. Luczynski (personal communication) selects the predatory mites randomly following their dispersal upward in a collection unit. In the present study the mites were selected by eye of a skilled technician either straight from the storage container of the freshly harvested bulk material

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According to Luczynski et al. (2006) a mass production procedure that will enhance a reproductive stable age distribution of P. persimilis females skewed towards young-fecund mites is the key to quality product. Luczynski (personal communication) claims that a laboratory mass

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production where P. persimilis culture is maintained continuously and only surplus of predators are harvested, will yield a fairly stable reproductive age distribution with a relatively high proportion of young-fecund females. On the other hand, the traditional mass production where the entire culture is harvested after the predators have increased by 2-3 generations, will lack stability in reproductive age distribution and the mite population will be skewed towards old females. The results reported in the current study show that a relatively high proportion of young-fecund females can be present also in a product of "traditional/standard" mass culture. Yet, a similar study should be conducted with other species of predatory mites that are mass reared in a continuous culture system in order to verify the above-mentioned hypothesis. The results of our study corroborate those of Luczynski et al. (2006) that reproductively young females survived cold storage better than the old ones and their rate of reproduction following storage was not significantly different from predators of the same age that have not been stored.

Finally, our results firmly indicate that the intrinsic quality of P. persimilis during spring 2006 was reasonable in all objective terms and apparently the problems reported from the field have to do with local field conditions and not with the intrinsic quality of the commercial product per se. Acknowledgements We thank Anna Luczynski of Quadra Consulting Ltd. and Elmer van Baal of Koppert Biological Systems for their useful comments and remarks. References Luczynski, A., Nyrop, J.P & Shi, A. 2006: The quality of mass produced Phytoseiulus persimilis

prior to and after storage. Technical report submitted to BC Greenhouse Growers Association. Project Number: W009.

van Lenteren, J. C., Hale, A., Klapwijk, J.N., van Schelt, J. & Steinberg, S. 2003: Guidelines for quality control of commercially produced natural enemies. In: Quality Control and Production of Biological Control Agents: Theory and Testing Procedures, ed. van Lenteren, J.C. CABI Publishing CAB International Wallingford UK.


pp. 111-115

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Biological control of Polyphagotarsonemus latus by the predaceous mite Amblyseius swirskii Carmit Tal1, 2, Moshe Coll2, Phyllis G. Weintraub1 1Agricultural Research Organization (ARO), Gilat Research Center, DN Negev, 85280, Israel; e-mail: [email protected]; Hebrew University, Department of Entomology, Rehovot, Israel Abstract: The broad mite, Polyphagotarsonemus latus, is a major pest on many crops in tropical and subtropical regions and in greenhouses world-wide. The phytoseiid predatory mite, Amblyseius swirskii, is able to feed on a wide range of plants and arthropods. However, its ability to control populations of P. latus is yet unknown. The object of this research was to determine the ability of A. swirskii to feed on broad mites in the laboratory on leaf discs and in the field on covered sweet pepper. Female A. swirskii were starved for 24 hours before being allowed to feed individually on different densities of broad mites. After 24 hours, broad mite mortality was assessed. Each broad mite density was replicated 15 times. We found that A. swirskii demonstrates a type II functional response to varying densities of P. latus. Based on these positive results we explored A. swirskii as a biological control agent against broad mites by determining their ability to control them on covered sweet pepper. Predators were released at two rates, 50 and 100/m2, and were compared to non-treated and acaricide-treated controls. The higher release rate was comparable to acaricide treatment.

Keywords: biological control, Polyphagotarsonemus latus, Amblyseius swirskii Introduction The broad mite, Polyphagotarsonemus latus (Banks) (Acarina: Tarsonemidae), although very small, is a major pest on many greenhouse crops in tropical and subtropical regions causing damage to plants and fruit by piercing and feeding on soft tissues. It is an important pest of vegetables, and peppers have a particularly low tolerance for the mite (de Coss-Romero and Peña, 1998). Damage caused by broad mites is expressed by leaves curving downward, plant growth is stunted and blossoms aborted. Fruit may become distorted and get a silvery cast upon it. There are several chemical treatments known to control its populations. Due to growing consumer demands for healthy and green produce, non-chemical solutions, foremost among which is biological control, are being sought. Neoseiulus cucumeris (Oudemans) (Acarina: Phytoseiidae) was shown to be a highly effective predator of the broad mite (Weintraub et al, 2003), but it is no longer available in Israel.

Field observations in Israel with the predatory mite, Amblyseius swirskii Athias-Henriot, an oligophagous, type III predator, have shown high efficiency against pest mites and other insects (Ragusa & Swirski, 1975). Based on laboratory research made in Europe the ability of the predatory mite had been proven as a biological control agent against tobacco whitefly Bemisia tabaci (Nomikou et al, 2005; Hoogerbrugge et al, 2005) and western flower thrips Frankliniella

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occidentalis (Messelink et al, 2005; van Houten et al, 2005). In this research the potential of the predatory mite A. swirskii as a biological control agent against P. latus was tested in laboratory and field trials.

Material and Methods

Broad mites were collected from basil (Ocinum basilicum) and cultured on bean plants (Phaseolus vulgaris) in controlled conditions (25 ± 3º C, 14:10 L:D). The predatory mites, A. swirskii, were obtained from Bio-Bee Biological Systems, Kibbutz Sedeh Eliyahu, Israel. Amblyseius swirskii were reared on plastic arenas with narrow-leaf cattail (Typha domingensis) and castor plant (Ricinus communis) pollen as the food source (Weintraub et al, 2006).

The laboratory trials were conducted in the Gilat Research Center, Israel. One percent polyacrylamid gel was prepared and poured into 10 x 15 cm plastic trays, and was used as a substrate to support and maintain leaf turgor throughout the trial. Predatory mites were isolated on clean bean leaf disks and starved for 24 hours before each trial. The number of adult female broad mites on each 5 cm2 leaf was counted before a single predator was released. After a period of 24 hours (rearing room conditions and 60 % RH), broad mite mortality was determined. Broad mites were grouped by 10s (i.e., 10 ± 2, 20 ± 2, etc) and each group was replicated 15 times.

All field trials took place at the Yair Research and Development Farm in the Arava Valley (between the Dead Sea and Red Sea), Israel. Plants were fertilized and watered by a drip irrigation system, according to standard organic agricultural practices for the area. Sixteen walk-in tunnels (7 x 15 m) were planted with 150 Celica variety of sweet pepper seedlings (red fruit) in three double row beds on 27 August 2006. At the beginning of the trials, there was a low-level broad mite infestation in all tunnels. Approximately 50 predators/m2 were released on the top leaves of each plant in 4 tunnels on 8 September and 100/m2 in another 4 tunnels. One set of 4 tunnels was treated with Evisect (thiocyclam hydrogen oxalate, Syngenta) on 26 September, to control mites, in one set of 4 tunnels was left untreated. Samples of 20 leaves each from the upper and middle portion of plants, and 20 flowers from each tunnel were taken weekly. Upper leaves were sampled from 12 September, middle leaves from 19 September and flowers from 27 September until the end of the trial. Sampled leaves and flowers were placed in jars containing 80% EtOH. In the laboratory, the leaves were removed and the contents of each container were examined under a dissecting microscope at 25x for presence of all mites.

Percent mortality data were normalized using arcsine transformation followed by ANOVA analysis with JMPIN 5.0.1 (SAS Institute, Inc, 2002). Mite field data were analyzed by completely randomized multiple ANOVA. Means were separated by Tukey HSD at α = 0.050.

Results and discussion The laboratory results on the mortality broad mites after a of 24 hour period isolated with a single, female A. swirskii are summarized in Figure 1. During the trial a number of female predators escaped from the arena and were not retrieved, and thus were eliminated from the trial. Figure 1 shows that as the number of P. latus per leaf increased, the predatory mite increased its consumption rate until about 30 broad mites/day. The trial yielded a polynomial fit with R2 ranging to 0.9277. Data fitted to a binomial curve describes a Hollings Type II functional

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Figure 1. Mortality of broad mites after 24 hours on leaf discs with 1 A. swirskii. Solid line, fitted polynomial trend of A. swirskii consumption.

The results of releasing A. swirskii (at two rates: 50 and 100 mites/m2) on broad mite

population in covered sweet pepper are shown in Figure 2. Broad mites were found primarily on young, upper leaves (Fig. 2A), but also occurred in the flowers and on middle leaves. At the first sampling time, shortly after release of the A. swirskii, there was no difference in the broad mite populations. By the second sampling date there were significant differences between populations of broad mites in insecticide and 100 mites/m2 versus non-treated control and 50 mites/m2 (P = 0.0002, F = 15.767, df = 3, 12). At the third sampling date, A. swirskii released at the low rate were able to control broad mites, while populations in non-treated control continued to grow. Populations of broad mites eventually collapsed in control tunnels; plants were very stunted and not producing new leaves.

Although broad mites were found on middle level leaves (Fig. 2B), their populations were substantially smaller. These leaves were collected starting one week after the upper leaves, and showed no difference in broad mite populations in insecticide-treated or A. swirskii-released tunnels. Populations of broad mites in flowers (Fig. 2C) were similar to those found in middle leaves. Flowers were collected from the time that there were a minimum available for sampling. Broad mites in flowers declined throughout the trial.

The use of insect predators is an integral component of biological control, and often requires importation from another country or continent. In these trials, we examined consumption rate of individual A. swirskii on leaf-discs infested with P. latus and different levels of release of A.

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swirskii for controlling the broad mite on sweet peppers. We demonstrated a positive correlation between raising population densities of P. latus and consumption rate of the predatory mite A. swirskii in laboratory leaf-disc trials. Based on these positive results we moved to field trials to determine the predator's ability in covered sweet peppers.

The Arava Valley has moderate winters and hot dry summers; agricultural pests often thrive, but not all natural enemies can perform well. Amblyseius swirskii is naturally found in the coastal region of Israel, a much more humid environment than the Arava Valley. We achieved excellent results with 100 predatory mites/m2 – comparable with chemical control. With a release of 50mites/m2 a slightly longer time was required to bring the broad mite population under control and because the plants were in a rapid growth period there was some noticeable stunting. Preliminary observations in other, more humid, areas of Israel indicated that the fewer predators were required to establish and control whiteflies (Shimon Steinberg, personal communication). Indeed, when A. swirskii was released in northern Europe greenhouses, control of B. tabaci on pepper was achieved with 25 mites/m2 (Hoogerbrugge et al, 2005) and control of F. occidentalis in cucumber at a comparable rate of 10 mites/plant (Messelink et al, 2005). Ultimately, this highly effective predator may be most effectively utilized with different application rates depending on climatic conditions. References de Coss-Romero, M. & Peña, J.E. 1998: Relationship of broad mite (Acari: Tarsonemidae) to

host phenology and injury levels in Capsicum annuum. Florida Entomol. 81: 515-526. Hoogerbrugge, H., Calvo, J., van Houten, Y. & Bolckmans, K. 2005: Biological control of the

tobacco whitefly Bemisia tabaci with the predatory mite Amblyseius swirskii in sweet pepper crops. IOBC/wprs Bull. 28(1): 119-122.

Messelink, G., van Steenpaal, S. & van Wensveen, W. 2005: Typhlodromips swirskii (Athias-Henriot) (Acari: Phytoseiidae): a new predator for thrips control in greenhouse cucumber. IOBC/wprs Bull. 28(1): 183-186.

Nomikou, M., Meng, R., Schraag, R. Sabelis, M.W. & Janssen, A. 2005: How predatory mites find plants with whitefly prey. Exp. Appl. Acarol. 36: 263-275.

Ragusa, S, & Swirski, E. 1975: Feeding habits, development and oviposition of the predacious mite Amblyseius swirskii Athias-Henriot (Acarina: Phytoseiidae) on pollen of various weeds.

vanHouten, Y.M., Ostlie, M.L., Hoogerbrugge, H. & Bolckmans, K. 2005: Biological control of western flower thrips on sweet pepper using the predatory mites Amblyseius cucumeris, Iphiseius degenerans, A. andersoni and A. swirskii. IOBC/wprs Bull. 28(1): 283-286.

Weintraub, P.G., Kleitman, S., Mori R., Shapira, N. & Palevsky. E. 2003: Control of broad mites (Polyphagotarsonemus latus (Banks)) on organic greenhouse sweet peppers (Capsicum annuum L.) with the predatory mite, Neoseiulus cucumeris (Oudemans). Biol. Cont. 27: 300-309.

Weintraub, P.G., Kleitman, S., Shapira, N., Argov, Y. & Palevsky, E. 2006: Efficacy of Phytoseiulus persimilis versus Neoseiulus californicus for controlling spider mites on greenhouse sweet pepper. Bull. IOBC/wprs 29(4): 121-125.

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Field evaluation of cotton seed treatments and a granular soil insecticide in controlling spider mites and other early-season cotton pests in Texas Noel Troxclair Texas Cooperative Extension, Texas A&M Research and Extension Center, P.O. Box 1849, Uvalde, Texas 78802-1849 Abstract: This on-farm, large-plot study involved a comparison of untreated control plots versus thiamethoxam and imidacloprid cotton seed-treatment insecticides and in-furrow aldicarb in controlling in early-season cotton arthropods. Early-season cotton arthropods were monitored on three dates and significant differences among treatments were detected for thrips, aphids, red imported fire ants and spider mites, on one or more dates. Key words: cotton, seed treatments, spider mites, thrips Introduction Over the last decade or so, insecticidal seed treatments have gained wide acceptance around the world for the management of arthropod pests of a variety of crops including barley, canola, cotton, corn, grain sorghum, potatoes and wheat (Yue et al. 2003). Systemic insecticide seed treatments, in lieu of granular insecticides such as aldicarb, have been widely adopted within the last three years in the United States for the management of early-season cotton pests such as thrips and aphids. Beginning in 2004, through 2006, infestations of two-spotted spider mites, Tetranychus urticae Koch, on seedling cotton, Gossypium hirsutum L., in some Winter Garden (Texas) fields reached population levels sufficient to severely damage or kill plants smaller than four-leaf stage. In some cases, rescue applications of acaricides were required to prevent economic stand losses and in a few cases where acaricides were not applied, growers incurred stand losses serious enough to require replanting (personal observations). This phenomenon of spider mite outbreaks on seedling cotton was unprecedented in the author’s experience. Follow-up conversations with producers revealed that those fields had been planted with seed receiving either imidacloprid or thiamethoxam seed treatments. Also starting in 2004, growers and others in pest management disciplines in other, southeastern, U.S. states began to observe the same phenomenon. Over a four-year period, cotton requiring acaricide applications to control T. urticae, on seedling cotton, in the Upper Mississippi Delta in Mississippi went from none in 2003 to approximately 20,200, 68,000 and 68,000 ha in the years 2004 to 2006, respectively (Angus Catchot, personal communication). Similarly, pecan leaf scorch mite, Eotetranychus hicoriae (McGregor) outbreaks have been observed following foliar applications of imidacloprid in Texas pecan, Carya illinoinensis (Wangenh.) K. Koch, orchards (William Ree, Jr., personal communication). Sclar et al. 1998, observed significantly greater damage by T. urticae to marigold, Tagetes erecta L., plants in the

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field that were treated with soil-applied imidacloprid. They also found populations of honeylocust spider mites, Platytetranychus multidigituli, were 3-4 times higher on honeylocust, Gleditsia triacanthos L., plants receiving an imidacloprid soil drench than those not treated. Raupp et al. 2004, found that hemlocks, Tsuga spp., treated with soil-applied imidacloprid for control of hemlock woolly adelgid, Adelges tsugae Annand, had higher populations of both spruce spider mite, Oligonychus ununguis (Jacobi), and hemlock rust mites, Nalepella tsugifolia Keifer, and more severe needle damage in terminals. James and Price, 2002, reported increased fecundity in T. urticae when either treated directly with, or ingesting, imidacloprid. They also observed increased longevity in the mites that ingested imidacloprid but not in those treated directly. Conversely, Ako et al. (2004) found lower fecundity, preimaginal survivorship and female sex ratio of offspring in T. urticae bioassayed on French beans, Phaseolus vulgaris L., that were treated with four different neonicotinoids. Several studies have determined that neonicotinoids exhibit both lethal and non-lethal negative effects on beneficial arthropods such as coccinellid predators (Smith and Krischik 1999, Vincent et al. 2000) and predatory Heteroptera (Elzen 2001, Studebaker & Kring 2000, Mizell et al 1992, James & Vogele 2001, Boyd & Boethel 1998). Mizell et al. (1992) found little toxicity to three species of predatory mites at concentrations of imidacloprid near the recommended field rate but that similar rates were toxic to most of the insect predators tested (mirids, coccinellids, chrysopids, and lygaeids). After nearly 15 years of studies, there is still no single factor to which the increases in spider mite populations following applications of neonicotinoids can be attributed. Based on their studies, Ako et al. (2004) suggest that the observed increases in mite populations following applications of imidacloprid may be dependent on several factors and their interactions, one of which could be a reduction in interspecific competition among surviving herbivores that favors phytophagous mites. The trial reported herein was not undertaken specifically to investigate the effects that neonicotinoid seed treatments would have on spider mite populations but data was recorded when the opportunity arose. For the sake of brevity and due to space limitations for the bulletin, only the spider mite data will be presented in detail; only cursory data for other taxa will be presented. Materials and methods FiberMax® variety, FM832LL, cotton was planted from 29 to 31 March 2005, in three fields (designated north, middle and south fields) which served as three replications for the trial. FM832LL cotton is a medium- to full-season okra-leaf cultivar with Liberty LinkTM transgenics. Treatments included thiamethoxam (Cruiser® 5FS) and imidacloprid (Gaucho® Grande) insecticide seed treatments, aldicarb (Temik® 15G) granular insecticide and an untreated control. Thiamethoxam was applied at 0.30 to 0.34 mg ai. per seed, imidacloprid was applied at 0.375 mg ai. per seed and aldicarb was applied in-furrow at planting, at 3.95 kg/hectare. Cotton was planted on a 96.5-cm width row spacing with an eight-row John Deere 7100 MaxEmerge® planter at a seeding rate of approximately 123,552 seeds per hectare. Plots with insecticide treatments were 4.8 hectares in size and untreated controls ranged in size from 0.8 to 2.4 hectares, depending on which field or position in the center pivot the plot was located. Treatments were arranged in a randomized, complete block design with three replications per treatment.

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No pre-plant nor pre-emergence herbicide was used. The seedbed was fine with good soil moisture at the time of planting and ambient air and soil temperatures at 10.16-to 20.32-cm depths were suitable for planting. However, several days after planting temperatures dropped and remained sub-optimal for an extended period. The cotton was slow in achieving a full stand and subsequent seedling growth progressed slowly. At beginning of squaring, the crop, physiologically, was considered to be three to four weeks behind where it should have been. On 21 and 25 April and 2 May, early-season arthropod counts were made from 25 randomly-selected plants from each of two locations in each plot. Taxa for which data was recorded included western flower thrips (Frankliniella occidentalis Pergrande), cotton aphids (Aphis gossypii Glover), two-spotted and carmine spider mites (Tetranychus urticae Koch and T. cinnabarinus Boisduval), cabbage loopers (Trichplusia ni Hübner), beet armyworms (Spodoptera exigua Hübner), red imported fire ants (RIFA) (Solenopsis invicta Buren) and cotton fleahoppers (Pseudatomoscelis seriatus Reuter). Other data collected included thrips damage ratings on 1- to 3-leaf stage cotton and 4- to 5-leaf stage cotton (25 plants per plot for each rating) and cotton yields which were determined by hand-harvesting 0.0004 hectare from three different areas in each plot. On 27 April, all plots in the middle and south fields were treated with a banded application of 0.63 kg per hectare of dimethoate for control of spider mites in the plots with imidacloprid and thiamethoxam seed-treatments. On 28 April, all plots in the north field received an application of 0.63 kg per hectare of dimethoate and 0.56 kg per hectare of amitraz (Ovasyn®) for control of spider mites in the plots with imidacloprid and thiamethoxam seed-treatments. Also, on 2 to 4 May and subsequent to insect counts being made, all plots (all fields) were treated with 0.28 kg of Acephate (Orthene®) per hectare for thrips control. Data analyses included ANOVA and Duncan’s Multiple Range Test for mean separations for insect counts. Mite data were analyzed using Chi-square analysis since only their presence or absence on plants was recorded. Results and discussion Overall, arthropod numbers were relatively low yet significant differences were observed for thrips, aphids, RIFA and spider mites; there was no difference in numbers of loopers, beet armyworms, nor fleahoppers among the different treatments. Cotton fleahopper numbers were very low during the time arthropod counts were recorded and insecticide treatments seemed to have lost their effectiveness when fleahopper numbers reached economic threshold levels. In spite of significant differences in thrips numbers among insecticide treatments, no difference was found in thrips damage ratings. The lack of differences in thrips damage ratings may have been due to the high variability in damage when the ratings were made; a sample size larger than 25 plants per plot for thrips damage ratings may have shown differences. Significant differences were observed among treatments for aphids and RIFA (Table 1) on all dates and for thrips on all but the first date (Table 2). All insecticide treatments controlled aphids when compared to the untreated controls on all dates. RIFA were observed feeding on aphid honeydew and their numbers paralleled aphid numbers and with one exception, were significantly higher on untreated plants than they were on any insecticide-treated plants. There was no difference in numbers of RIFA among any insecticide treatments.

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Table 1. Mean numbers of cotton aphids and red imported fire ants per seedling cotton plant on individual dates and for all dates combined.

aphids/plant RIFA/plant Treatment 4/21 4/25 5/02 all dates 4/21 4/25 5/02 all datesControl 2.474a 2.347a 1.080a 2.085a 0.527a 0.380a 0.293a 0.400a Thiamethoxam 0.220c 0.440bc 0.213b 0.291c 0.033a 0.080b 0.013b 0.042b Imidacloprid 0.813b 1.000b 0.167b 0.640b 0.047b 0.184ab 0.000b 0.071b Aldicarb 0.120c 0.240c 0.140b 0.167c 0.007b 0.080b 0.040b 0.042b

Means within a column, followed by the same letter, are not significantly different (P ≤ 0.0001), Duncan’s multiple range test. Table 2. Mean number of thrips per seedling cotton plant on individual dates and for all dates combined.

Thrips/plant Treatment 4/21 4/25 5/02 all dates Control 0.04a 0.587ab 0.493c 0.373b Thiamethoxam 0.07a 0.720a 1.373a 0.722a Imidacloprid 0.05a 0.448bc 1.553a 0.696a Aldicarb 0.01a 0.287c 0.867b 0.389b

Means within a column, followed by the same letter, are not significantly different (P ≤ 0.0001), Duncan’s multiple range test. Significant differences were observed in spider mite numbers among treatments on all dates. The percentages of plants from the plots with thiamethoxam and imidacloprid seed treatments infested with spider mites were significantly higher than the percentages of plants from the aldicarb-treated on every date and were different from the untreated control plots on two dates (Table 3).

Table 3. Mean percentages of seedling cotton plants infested with two-spotted and carmine spider mites on individual dates and for all dates combined. Treatment 04/21 04/25 05/02 all dates Control 4.0b 23.3a 4.0bc 10.44b Thiamethoxam 20.7a 26.7a 20.0a 22.44a Imidacloprid 32.0a 36.0a 15.3a 27.29a Aldicarb 0.0b 4.7b 8.7b 4.44bc

χ2=81.95, P≤ 0.0001

χ2=42.48, P≤ 0.0001

χ2=21.34, P ≤ 0.005

χ2=109.28, P≤ 0.0006

Means within a column, followed by the same letter, are not significantly different at the given P value below the column. For the given degrees of freedom in the first three columns, a χ2 value equal to or greater than 4.575 would be significant at P ≤ 0.05; for the fourth column, a χ2 value equal to or greater than 20.14 would be significant at P ≤ 0.05.

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On 25 April, a higher percentage of plants from the untreated control plots had spider mites than did plants from aldicarb-treated plots. On every other date, there was no difference between the untreated and aldicarb-treated plots in percentage of plants infested with spider mites. Chi-square analyses with large χ2 values ranging from 21.34 to 109.28 indicate that the differences, among treatments, in the percentages of plants infested with spider mites, show an insecticide treatment bias, with higher numbers of spider mites on plants from plots with seed treatments. There was no difference in spider mite numbers between the two seed treatments. Although actual numbers of mites was not recorded, the average spider mite colony on cotyledonary stage plants was about 10 to 12 mites with an occasional colony reaching numbers of about 30 mites. Spider mites reached levels that were causing leaf drop and plant death in some plots. Although all plots had spider mites after miticides were applied, spider mite populations eventually decreased to levels that were not damaging to the cotton plants. No significant difference among insecticide treatments was observed in cotton yields but the highest yields, in plots with imidacloprid-treated seed, had 59.95 kg more lint cotton than the untreated cotton and 62.19 kg more than yields from the plots with thiamethoxam-treated seed (Table 4). Cotton turnout in all three fields averaged 35%, so lint yields were excellent, ranging from a low of 1855.41 kg/ha in the plots with thiamethoxam-treated seed to a high of 1917.4 kg/ha in plots with imidacloprid-treated seed. Table 4. Mean lint cotton yields per hectare.

Treatment Lint cotton – kg/ha Control 1857.65a Thiamethoxam 1855.41a Imidacloprid 1917.60a Aldicarb 1895.31a

Means within a column, followed by the same letter, are not significantly different (P ≤ 0.0001), Duncan’s multiple range test. Yield data do not reflect the effect that the spider mites may have had because rescue insecticides/miticides were applied soon after mite damage to the seedlings became evident. Spider mites were present almost as soon as plants emerged, because they were observed on numerous plants which were only in the cotyledonary stage. Very few beneficial arthropods were observed during the time that arthropod counts were being made; otherwise their numbers would have been recorded. It appears that there was some factor that caused higher numbers of spider mites on plants treated with the seed treatments (neonicotinoids) because there was a relative void of beneficial arthropods across all treatments at that early stage. If the phenomenon could be explained by the lack of predators and parasites, plants in the untreated control plots should have had equally high numbers of spider mites. Additional research is needed to try to determine what factor(s) is/are involved in this phenomenon. This report reflects data from one study, conducted in one year, at one location, so no definitive conclusion should be drawn from the results reported herein.

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Acknowledgments I want to thank Mark Landry for his time, labor, and patience in working with me in the establishment of this trial and for his continuing support for various projects through the years. A thank you also is extended to Terry Pitts, Gustafson, Inc. and Brad Minton, Syngenta Crop Protection for their funding support and for providing the insecticides for the seed treatments used in this trial. References Ako, M., Bourgemeister, C., Poehling, H.-M., Elbert, A. & Nauen, R. 2004: Effects of

neonicotinoid insecticides on the bionomics of twospotted spider mite (Acari: Tetranychidae). J. Econ. Entomol. 97: 1587-1594.

Boyd, M. & Boethel, D.J. 1998: Residual toxicity of selected insecticides to heteropteran predaceous species (Heteroptera: Lygaeidae, Nabidae, Pentatomidae) on soybean. Environ. Entomol. 27: 154-160.

Elzen, G.W. 2001: Lethal and sublethal effects of insecticide residues on Orius insidiosus (Hemiptera: Anthocoridae) and Geocoris punctipes (Hemiptera: Lygaeidae). J. Econ. Entomol. 94: 55-59.

James, D. G. & Vogele, B. 2001: The effect of imidacloprid on survival of some beneficial arthropods. Plant Protect. Q. 16: 58-62.

James, D. G. & Price, T. S. 2002: Fecundity in Twospotted spider mite (Acari: Tetranychidae) is increased by direct and systemic exposure to imidacloprid. J. Econ. Entomol. 95: 729-732.

Mizell, R. F, III & Sconyers, M. C. 1992: Toxicity of imidacloprid to selected arthropod predators in the laboratory. Scientific Note: Fla. Entomol. 75: 277-280.

Raupp, M.J., Webb, R.E., Szczepaniec, A., Booth, D. & Ahern, R. 2004: Incidence, abundance, and severity of mites on hemlocks following applications of imidacloprid. J. Arboricult. 30: 108-113.

Sclar, D.C., Gerace, D. & Cranshaw, W.S. 1998: Observations of population increases and injury by spider mites (Acari: Tetranychidae) on ornamental plants treated with Imidacloprid. J. Econ. Entomol. 91: 250-255.

Smith, S.F. & V.A. Krischik. 1999: Effects of systemic imidacloprid on Coleomegilla maculata (Coleoptera: Coccinellidae). Environ. Entomol. 28: 1189-1195. Studebaker, G.E. & King., T.J. 2000: Lethal and sublethal effects of early-season insecticides on

insidious flower bug (Orius insidiosus): an important predator in cotton. Proceedings of the 2000 Cotton Research Meeting: AAES Special Report 198: 221-225.

Vincent, C., Ferran, A., Guige, L., Gambier J. & Brun, J. 2000: Effects of imidacloprid on Harmonia axyridis (Coleoptera: Coccinellidae) larval biology and locomotory behavior. Eur. J. Entomol.97: 501-506.

Yue, B., Wilde, G.E. & Arthur, F. 2003: Evaluation of thiamethoxam and imidacloprid as seed treatments to control European corn borer and Indianmeal moth (Lepidoptera: Pyralidae) larvae. J. Econ. Entomol. 96: 503-509.


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Spider mite control by four phytoseiid species with different degrees of polyphagy Yvonne M. van Houten, Hans Hoogerbrugge & Karel J.F. Bolckmans Koppert Biological Systems, P.O. Box 155, 2650 AD Berkel en Rodenrijs, The Netherlands Abstract: Four commercially available phytoseiid mite species with different life-styles; Phytoseiulus persimilis, Neoseiulus californicus, Amblyseius swirskii and Amblyseius andersoni are all predators of two-spotted spider mite. These species were compared with respect to the following features: oviposition rate on T. urticae, incidence of reproductive diapause under short day conditions and their performance as biological control agent of T. urticae on sweet pepper. The results showed that P. persimilis exhibited the highest oviposition rate followed by N. californicus, A. swirskii and A. andersoni in descending order. A. andersoni was the only species to enter diapause under short day conditions. In a cage experiment A. swirskii and A. andersoni strongly slowed down the population growth of T. urticae on sweet pepper, whereas P. persimilis and N. californicus were able to control T. urticae successfully. Key words: Phytoseiulus persimilis, Neoseiulus californicus, Amblyseius swirskii, Amblyseius andersoni, Tetranychus urticae, sweet pepper Introduction The specialist predator of tetranychid mites, Phytoseiulus persimilis Athias-Henriot, is the most important augmentative biological control agent of Tetranychus urticae (Koch) in greenhouses. After controlling the pest however, P. persimilis often disappears from crops because it can only reproduce on tetranychid mites; consequently, new releases are sometimes necessary in case of new spider mite outbreaks. Phytoseiulus persimilis does not enter diapause and is released together with spider mites (CPR controlled pest release) in Dutch greenhouses from February onwards to build up a population in the crop preventively.

Another predator of spider mites is Neoseiulus californicus McGregor (Castagnoli & Simoni 2003). Although N. californicus prefers tetranychid mites, it is able to feed on other genera of mites as well as on thrips (van Baal et al. 2007) and plant pollen.

The generalist predator A. swirskii (Athias-Henriot) is an important biological control agent of whiteflies and thrips (Bolckmans et al. 2005) but also feeds on two-spotted spider mites (Momen & El-Saway 1993) and pollen. The generalist predator A. andersoni (Chant) feeds on T. urticae (Overmeer 1981), thrips (van Houten et al. 2006) and pollen (Overmeer 1981).

In the absence of spider mites N. californicus, A. swirskii and A. andersoni can establish in the crop on the other food sources. In this study we compare these predatory mite species with P. persimilis with respect to the following features: oviposition rate on T. urticae, incidence of reproductive diapause under short day conditions and their performance as biological control agent of T. urticae on sweet pepper.

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Material and methods The A. andersoni originated from Applied Plant Research, Wageningen and was collected in The Netherlands under short-day conditions in 2001. The A. swirskii was collected in Israel and has been mass produced by Koppert BV since 2004. The N. californicus originated from California and has been mass produced by Koppert BV since 2005.

Amblyseius andersoni was reared on pollen of iceplant Mesembrianthemum sp., A. swirskii was reared on pollen of Zea mays and N. californicus on two-spotted spider mites in a climate room for 2 months before testing. Phytoseiulus persimilis was directly obtained from the mass rearing. Diapause For the duration of the experiments, the mites were kept in a temperature, humidity and photoperiod controlled incubator. The temperature was maintained constant at 18° ± 0.5°C. The photoperiod was a short-day regime (light: dark = 10:14) and relative humidity 75%.

Eggs of N. californicus were placed on cucumber leaf discs infested with two-spotted spider mites. Newly emerged females mated with males on the same leaf disc. Thereafter, the females were removed and single females were placed on small cucumber leaf discs (4.5 cm2) infested with two-spotted spider mites. Subsequently, the females were examined three times a week for egg laying.

The experiments with A. andersoni and A. swirskii were conducted on plastic arenas according to the method described by van Houten et al. 1995. The mites were fed with fresh cattail pollen and with purple pollen of iceplant.

Females that had mated but did not produce any eggs over a 2 week period were regarded as 'being in diapause'. To confirm the latter: after 2 weeks without eggs, these mites were transferred to 16L:8D regime at 25°C; egg laying should resume after about 4 days. Oviposition rate Oviposition rates were measured on small cucumber leaf discs (4.5 cm2) infested with spider mites, in a climate room at 15°C and 25°C. The mites originated from cohorts of eggs that were reared on pollen (A. andersoni and A. swirskii) or spider mites (N. californicus and P. persimilis). One young gravid female was placed on each leaf disc. At the start of the experiment the mites had been laying eggs for 2 days. During 14 days, the predators were transferred to new leaf discs infested with spider mites. The old leaf discs were examined to record the number of oviposited eggs. Data from the first day were omitted from calculations of ovipostion rates. Cage experiment The experiment was done in 14 cages (3x1x2 m) with six sweet pepper plants each. In wk 43 all plants were infested with two-spotted spider mites. In wk 44, when plants started to flower, 10 female predators were introduced per plant: A. andersoni and P. persimilis were released in 3 cages, N. californicus and A. swirskii were released in 4 cages. To monitor spider mite and predator populations, samples of 10 leaves of the upper part of the plants were taken every week from each cage. Because of the extremely high spider mite population, all plants were treated with Nissorun (hexythiazox) in week 45 to lower the spider mite populations. Daylength during this experiment was extended to 16 hours light per day with supplemental artificial lighting.

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Results and discussion Amblyseius andersoni entered diapause under short-day conditions. Consequently, the establishment and spider mite control during winter will be less successful. N. californicus and A. swirskii showed a total absence of diapause (Table 1) and therefore they can establish in a crop all year round on different kinds of food sources.

Phytoseiid population

0

50

100

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43 44 45 46 47 48

week number

Mea

n nu

mbe

r per

10

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es

P. persimilis A. swirskii N. californicus A. andersoni

T P N

Tetranychus urticae population

0

400

800

1200

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2000

43 44 45 46 47 48

week number

Mea

n nu

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P. persimilis A. swirskii N. californicusA. andersoni Untreated control

T P N

Figure 1. Population fluctuations of Tetranychus urticae and 4 predatory mite species on leaves of the upper part of sweet pepper plants in 13 cages. T: release of T. urticae; P: release of the predatory mites; N: application of Nissorun.

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Table 1. Diapause incidence in N. californicus, A. andersoni and A. swirskii under short-day conditions (Light:Dark = 10:14 h) at 18°C. N= number of females tested.

Phytoseiid species N Diapause incidence

Neoseiulus californicus 56 0%

Amblyseius swirskii 64 0%

Amblyseius andersoni 44 100% Table 2. Rates of oviposition of four predatory mite species on a diet of T. urticae, on cucumber leaf discs at two different temperatures. oviposition rate: mean number of eggs laid per female per day. N= number of females; s.e. standard error Phytoseiid species

N

Temperature: 25°C oviposition rate (mean ± s.e)

N

Temperature: 15°C oviposition rate (mean ± s.e)

P. persimilis N. californicus A. swirskii A. andersoni

27 18 16 15

5.4 ± 0.1 a1

4.0 ± 0.0 b 1.8 ± 0.1 c 1.6 ± 0.2 c

3025

1.7 ± 0.0 a1 1.4 ± 0.0 b Not tested Not tested

1 Means within a column followed by different letters are significantly different (Mann-Witney test, P<0.05) The specialized predator P. persimilis shows the highest oviposition rate of 5.4 eggs/day at 25°C and 1.7 eggs/day at 15°C followed by the selective predator N. californicus with 4.1 eggs/day at 25°C and 1.4 eggs/day at 15°C. The generalist predators A. swirskii and A. andersoni showed the lowest oviposition rates at 25°C, viz., 1.8 and 1.6 eggs/day respectively (Table 2). In the cage experiment the populations of A. swirskii and A. andersoni increased slowly. Both predators were adversely affected by dense spider mite webbing and did not enter webbed colonies. In spite of this, both A. andersoni and A. swirskii were able to inhibit the increase of T. urticae. In wk 48, an average of 6 predatory mites/leaf was found in the A. andersoni cage and an average of 9 predatory mites/leaf was found in the A. swirskii cage (Figure 1). P. persimilis and N. californicus were able to control the spider mites. Both predator populations increased very rapidly to 30 mites/ leaf in wk 46 for P. persimilis and in wk 47 for N. californicus. One week after the peak of the predator populations, the spider mites were completely under control in these cages. Based on these results, N. californicus can be regarded as a promising candidate for spider mite control throughout the season: it showed good reproduction capacities, even at 15°C and was not hampered by the webbing, it does not enter into diapause and can establish in the crop on alternative food sources.

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References Baal, E. van, Houten, Y. van, Hoogerbrugge, H. & Bolckmans, K. 2007. Side effect on thrips of the

spider mite predator Neoseiulus californicus (McGregor). Proc. Neth. Entomol. Soc. Meet. 18: 37-42.

Bolckmans, K., Houten, Y.M. van & Hoogerbrugge, H. 2005: Biological control of whiteflies and western flower thrips in greenhouse sweet peppers with the phytoseiid predatory mite Amblyseius swirskii Athias Henriot (Acari Phytoseiidae). Second International Symposium on Biological Control of Arthropods: 555-565.

Castagnoli, M. & Simoni, S. 2003. Neoseiulus californicus (McGregor) (Acari Phytoseiidae): Survey of Biological and Behavioural Traits of a Versatile Predator. Redia 86: 153-164.

Houten, Y.M., van, Østlie, M.L., Hoogerbrugge, H. & Bolckmans, K. 2005: Biological control of western flower thrips on sweet pepper using the predatory mites Amblyseius cucumeris, Iphiseius degenerans, A. andersoni and A. swirskii. IOBC/wprs Bull. Vol. 28 (1): 283-286.

Momen, F.M. & El-Saway, S.A. 1993: Biology and feeding behaviour of the predatory mite, Amblyseius swirskii (Acari: Phytoseiidae). Acarologia. 34(3): 199-204.

Overmeer, W.P.J. 1981: Notes on breeding Phytoseiid mites from orchards (Acarina: Phytoseiidae) in the laboratory. Med. Fac. Landbouw. Rijksuniv. Gent. 46(2): 503-509.

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The influence of Amblyseius swirskii on biological control of two-spotted spider mites with the specialist predator Phytoseiulus persimilis (Acari: Phytoseiidae) Yvonne M. van Houten, Hans Hoogerbrugge & Karel J.F. Bolckmans Koppert Biological Systems, P.O. Box 155, 2650 AD Berkel en Rodenrijs, The Netherlands Abstract: The biological control of Tetranychus urticae with Phytoseiulus persimilis was examined on sweet pepper plants in presence of the generalist phytoseiid mite Amblyseius swirskii. At a moderate density of T. urticae, A. swirskii had a negative effect on the population increase of P. persimilis. At a high density of T. urticae, no effect of A. swirskii on the P. persimilis population was found. Both at high and low densities of T. urticae, P. persimilis was able to control two-spotted spider mites when A. swirskii was present on the plants. Key words: Tetranychus urticae, Phytoseiulus persimilis, Amblyseius swirskii, competition for food, intraguild predation, sweet pepper Introduction The predatory mite Phytoseiulus persimilis (Athias-Henriot) is an effective biological control agent of Tetranychus urticae (Koch) in greenhouses and is used by greenhouse growers worldwide. This specialist predator only feeds on spider mites and can not establish in the crop without the presence of tetranychid mites.

Another predatory mite, Amblyseius swirskii (Athias-Henriot), is an important biological control agent of whiteflies and western flower thrips in sweet pepper (Bolckmans et al. 2005) and also preys on two-spotted spider mites. The generalist predator A. swirskii can slow down T. urticae populations but is not able to control hot spots of T. urticae because they do not enter the heavily webbed colonies (van Houten et al. 2007). Amblyseius swirskii can establish in sweet pepper by preying on different food sources and a density of 5 predators/leaf is not uncommon. Since 2005, A. swirskii has been used in commercial greenhouse crops.

Amblyseius swirskii and P. persimilis may potentially interact through competition for prey and intraguild predation. In the absence of prey, A. swirskii and P. persimilis are able to prey on each other (unpublished results). We investigated the effect of the presence of A. swirskii on biological control of T. urticae with P. persimilis on sweet pepper plants. The experiments were done at moderate and high T. urticae levels on plants. Material and methods

The first experiment was carried out in three cages (3 x 1 x 2 m). Three sweet pepper plants with 15 leaves were placed in each cage. The plants were placed in small containers with water and soap as a barrier to prevent escape and invasion of mites and did not touch each other nor the

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screen of the cage. T. urticae and predatory mites were released in numbers as shown in Table 1. Each cage contained one replicate of each treatment (3 replicates per treatment). To monitor the spider mite and predator populations, 5 leaves per plant were monitored every week from wk 28 (day 13 after the first introduction) onwards. Table 1. Release rate of predatory mites and spider mites per treatment. Treatment Release rate of T. urticae Release rate of A. swirskii Release rate of P. persimilis

A 200 mobile stages on day 1 100 females on day 6 10 females on day 7 B 200 mobile stages on day 1 10 females on day 7 C 200 mobile stages on day 1

The second experiment was conducted in 2 cages. Six sweet pepper plants with 60 leaves were placed in the cages in the same way as the first experiment. In wk 38 (day 1) and wk 41 (day 18) each plant was infested with high numbers of spider mites. Releases of 5 P. persimilis per plant were compared with combined releases of 50 A. swirskii and 5 P. persimilis per plant (table 2). Each cage contained one or two replicates of a treatment (3 replicates per treatment). To monitor the spider mites and predator populations, 5 leaves per plant were monitored every week from wk 40 (day 12) onwards. Table 2. Release rate of predatory mites and spider mites per treatment. Treatment Release rate of T. urticae Release rate of A. swirskii Release rate of P. persimilis

A 350 mobile stages on day 1 200 mobile stages on day 18

50 females on day 4 5 females on day 5

B 350 mobile stages on day 1 200 mobile stages on day 18

5 females on day 5

Results and discussion In the first experiment the P. persimilis population increased more rapidly in the treatment without A. swirskii than in the treatment with combined releases of the two species (Figure 1). One week after releasing the predators, an average of 5 P. persimilis was found in the P. persimilis treatment and an average of 2.2 P. persimilis /leaf and an average of 4 A. swirskii /leaf were found in the combined P. persimilis and A. swirskii treatment. At the same time, an average of 31 T. urticae/leaf was found in the P. persimilis treatment and an average of only 15 T. urticae /leaf were found in the combined treatment. One week later the spider mites were under control in both treatments. Subsequently, the P. persimilis populations decreased rapidly in both treatments, whereas the A. swirskii population established on the plants with pollen and thrips as food.

In the second experiment, the T. urticae population densities were much higher than in the first experiment. On these plants with a surplus of T. urticae, the presence of A. swirskii had no effect on the increase of the P. persimilis population (figure 2). In both treatments, P. persimilis populations increased to an average of 10 predators/leaf in week 42 (day 28). The A. swirskii population reached an average of 4.5 mites/leaf. Amblyseius swirskii was often found on the leaves with low T. urticae density and hardly found in the webbing. From week 42 onwards the condition of the plants

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decreased rapidly due to the high T. urticae infestation and therefore the experiment was stopped one week later.

When the population density of T. urticae is low, the presence of A. swirskii on the plant has a negative effect on the population development of P. persimilis. This effect could be attributed to competion for food and to predation by A. swirskii on P. persimilis. However, in both experiments the concurrent release of both A. swirskii and P. persimilis had no negative effect on the control of T. urticae. At low densities of two-spotted spider mites a combined release of P. persimilis and A. swirskii even resulted in a lower population of spider mites. Moreover, A. swirskii contributes to the biological control of two-spotted spider mites (van Houten. et al. 2007). When the population density of spider mites is high, the presence of A. swirskii does not affect the population development of P. persimilis.

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Figure 1. Population fluctuations of Tetranychus urticae and the phytoseiid mites Phytoseiulus persimilis and Amblyseius swirskii at moderate density of T. urticae on leaves of the upper part of sweet pepper plants in cages. T: release of T. urticae; P: release of the predatory mites

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Figure 2. Population fluctuations of Tetranychus urticae and the phytoseiid mites, Phytoseiulus persimilis and Amblyseius swirskii at high density of T. urticae on leaves of the upper part of sweet pepper plants in cages. T: releases of T. urticae; P: release of the predatory mites References Bolckmans, K., Houten, Y.M. van & Hoogerbrugge, H. 2005: Biological control of whiteflies and

western flower thrips in greenhouse sweet peppers with the phytoseiid predatory mite Amblyseius swirskii Athias Henriot (Acari Phytoseiidae). Second International Symposium on Biological Control of Arthropods: 555-565.

Houten, Y.M. van, Hoogerbrugge, H. & Bolckmans, K.J.F. 2007: Spider mite control by four phytoseiid species with different degrees of polyphagy. In this proceedings


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Antibiosis of kidney bean cultivars to the carmine spider mite, Tetranychus cinnabarinus (Boisduval) (Acari: Tetranychidae) Carlos Vásquez1, Mariela Colmenárez1, Neicy Valera1, Lisbeth Diaz2 Universidad Centroccidental Lisandro Alvarado, Decanato de Agronomía. 1Departamento de Ciecnias Biológicas. 2Departamento de Ingeniería Agrícola. Núcleo Tarabana. Via Agua Viva, Municipio Palavecino, Estado Lara, Venezuela. E-mail: [email protected]. Abstract: The antibiosis of three Phaseolus vulgaris L. cultivars to the carmine spider mite, Tetranychus cinnabarinus (Boisduval), was evaluated under laboratory conditions in Venezuela. Oviposition and survival of the carmine spider mite were evaluated on 22 or 55 day-old-leaf disks. Results showed that lower oviposition was observed in 22-day-old leaf disks, while on 55-day-old leaf disks oviposition increased in 43.1, 58.9 and 95.9% in Tacarigua, Coche and ICA-Pijao cultivars, respectively. On the other hand, survival was significantly lowered in 55-day-old leaf-disks from ICA-Pijao, thus suggesting that this cultivar produces feeding deterrent. Our results showed that ICA-Pijao could be used in developing of resistance programs to obtain inbred elite lines in Venezuela. Key words: plant resistance, Phaseolus vulgaris, tetranychid mites. Introduction Traditionally, the kidney bean (Phaseolus vulgaris L.) constitutes one of the main food sources in Latin America, due to it has high protein content (Cárdenas et al. 2000). In spite of its importance, decreasing yield has been observed because non-efficient pest control techniques are being used. Several tetranychid mite species have been reported as associated to this crop including Tetranychus cinnabarinus (Boisduval), T. ludeni Zacher, T. marianeae McGregor, T. mexicanus (McGregor), T.s neocaledonicus André and T. urticae Koch (Ochoa et al. 1994). However, Zhang and Jacobson (2000) pointed out T. urticae and T. cinnabarinus as the most common tetranychid species affecting crops worldwide.

Most of the effort to control pest population densities under threshold levels have relied on chemical tactics. However, in last decades attention has been paid to other management strategies, including resistant cultivars. So far, kidney bean has shown moderate resistance to the melon trips, Thrips palmi Karni (Cardona et al. 2002) and it seems to be underlying on antixenosis and antibiosis mechanisms (Frei et al. 2003).

In general, these defense mechanisms are considered plant responses to stressing conditions either abiotic (drought, salinity, etc) or biotic (herbivore or pathogens attacks) which induce physical barriers to feeding or secondary metabolites affecting oviposition or survival (Tomczyk & Kropczynska 1985, Gardner & Agrawal 2002). Boom et al. (2003) demonstrated that performance of T. urticae is determined by presence of certain toxic substance in the host plant. Seemly, De Ponti (1977) found that varietal resistance of cucumber plants to T. urticae is associated to cucurbitacine presence which caused a lowering of pest reproductive parameters.

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In Venezuela, Morros and Aponte (1995a, b) found that T. ludeni produced growth decreasing on kidney bean plants when attacked during vegetative or flowering stages. However, information about resistance potential of kidney bean cultivars to spider mite species, including T. cinnabarinus, is lacking, thus herein antibiosis of three of the most commonly cultivated P. vulgaris cultivars was evaluated thus providing growers alternatives to manage populations of this pest mite other than agrochemical substances in small farms in Lara state, Venezuela.

Material and methods Mite rearing Specimens were collected on infested Desmodium sp. in Núcleo Tarabana, Universidad Centroccidental Lisandro Alvarado, Lara state, Venezuela. In the laboratory, leaflets were examined under magnification (Leica MS 5) to select tetranychid mites using a fine hairbrush and then transferred to 3-4-week-old kidney bean plants grown in pots to maintain a colony for further studies. Microscope slides with male and female mites were prepared to determine species based in taxonomical keys provided by Gutierrez (1985) and/or comparison of aedeagus morphology (Ochoa et al. 1994). Plant material Oviposition and survival of T. cinnabarinus were evaluated on Tacarigua, Coche and ICA-Pijao. Tacarigua and Coche have been traditionally cultivated in Venezuelan highlands, however so far, just Tacarigua had been investigated and it has been found to be susceptible to other arthropod pests as Empoasca kraemeri Ross & Moore (Lozada et al. 1995). On the other hand, ICA-Pijao has shown to have promising resistance features to be considered in breeding programs (Lozada 1997). Oviposition and survival rates of T. cinnabarinus Resistance of P. vulgaris cultivars to T. cinnabarinus was evaluated as mite’s oviposition and survival on 22-day-old and 55-day-old disks under laboratory conditions (28 ± 2 ºC, 60 ± 10% R.H. and 12:12 L:D). Mites were reared following Helle & Overmeer (1985). Five leaf disks (3 cm diameter) were punched from each cultivar and placed on a circular foamed plastic. Two 1-day-old females were transferred to each leaf disk from the original stock. Then, they were put in a 30-mm Petri dish.

Rearing units were observed daily under a magnification 40X to determine the number of eggs laid by female during a five-day period. After incubation time, rearing units were used to evaluate survival rate of mite’s offspring on each cultivar. Five replicates per cultivars were used. Experimental design Studies were carried out in a completely randomized design with treatments arranged in split-split plot, being cultivars the main plot, leaf age the subplot and female age the sub-subplot. Oviposition data were transformed by 5.0+y (Steel & Torrie 1980), means were separated by Tukey’s mean test using Statistix version 8.0.

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Results and discussion T. cinnabarinus oviposition In all cultivars mean oviposition was significantly lower on 22-day-old- leaf disks, ranging from 0.98 to 1.17 eggs/female/day on ICA-Pijao and Coche, respectively, while increasing about 58 and 95% was detected on oviposition of females reared on 55 day-old leaf (Table 1). Similar results where showed by Zhang et al. (2001) evaluating oviposition rate in two Schizotetranychus bambusae Reck groups: one being reared on young bamboo leaves, whilst the second one was reared on old leaves. These authors found higher mean daily oviposition when old leaves were used (2.9 eggs/female/day) when compared to 1.9 eggs/female/day laid on young leaves. Previous studies have demonstrate that host plants produce jasmoic acid (JA) as response to T. urticae feeding, it seems to play an important role in expression of plant resistance eliciting alternative metabolic ways to affect herbivore metabolism. Li et al. (2002) reported that mutant tomato plants treated with methyl-jasmonate recovered resistance ability, thus reducing T. urticae fecundity. Similarly, Choh et al. (2004) also found that applying 0.1 mM JA reduced oviposition of T. urticae in 25%. Table 1. Oviposition (mean ± S.D.) of T. cinnabarinus on 22 and 55 day-old leaf disks from different kidney bean cultivars.

Number of eggs/female

22 day-old

55 day-old

Cultivar N Tacarigua 25 1.16 ± 0.7342 a (34) 1.66 ± 0.8119 b (72) Coche 25 1.17 ± 0.5905 a (30) 1.86 ± 0.7126 b (86) ICA-Pijao 25 0.98 ± 0.3852 a (15) 1.92 ± 0.8907 b (99)

Values in a file followed by same letter did not show significant differences (Tukey’s test; P<0.05). Number in parenthesis represent total egg number during a five day period.

In addition, female age affected oviposition rate irrespective of cultivar. Daily oviposition showed significant differences in 1, 2, 3, 4 or 5-day-old females when compared oviposition on 22 or 55-day-old leaf disks (Table 2). Females reared on 22-day-old leaf reached higher oviposition rate during days 4 and 5. Conversely, oviposition pattern on 55-day-old leaf disks was higher during the whole evaluation period. Read et al. (2002) found that younger leaves might have higher concentration of total phenols, which could act as oviposition deterrent, thus lower herbivore oviposition is expected.

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Table 2. Effect of female age of T. cinnabarinus on daily oviposition rate on 22 and 55 day-old leaf disks.

Female age(1)

Day 1 Day 2 Day 3 Day 4 Day 5

22 day-old 0.77±0.1821c 0.81±0.2143c 0.87±0.2897bc 1.67±0.7831a 1.38±0.5883ab

55 day-old 2.27±0.5430a 1.58±0.7096ab 2.06±0.7924a 1.94±0.8980ab 1.21±0.6709b

Values in a file followed by same letter did not show significant differences (Tukey’s test; P>0.001) (1) Values transformed by 5.0+y

T. cinnabarinus survival In general, higher survival was observed on 22-day-old leaves ranging from 5.67 to 6.63 live individuals in Tacarigua and Coche leaves, while on 55-day-old leaves survival varied from 5.42 to 1.80 in Coche and ICA-Pijao, respectively. Significant reduction of survival was observed on mites reared on 55-day-old leaf disks from ICA-Pijao cultivar (Figures 1A, B), suggesting that this cultivar could be able to produce dissuasive to mite feeding thus affecting mite survival. Zhang et al. (2001) found that survival of S. bambusae on bamboo diminished according plant age. Those authors hypothesized that probably old bamboo leaves lack of certain chemical substances that S. bambusae needs to properly digest plant nutrient. Heilker & Meiners (2002) stated that host plants showed direct defense mechanisms which or negatively affect access to plant’s nutrients or, otherwise, can produce anti-feeding compounds or toxins, both of them affecting herbivore survival.

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Based on results, ICA-Pijao could be an alternative for growers from Lara state because of it showed deterrence for feeding of T. cinnabarinus as compared with traditionally used cultivars; Tacarigua and Coche. Thus, the former should be included in breeding programs. Further studies on resistance mechanisms related to mites should be conducted in order to provide better understanding on the components contributing to this phenomenon.

Acknowledgements We thank the Government of República Bolivariana de Venezuela for partial foundation to this project through the Fondo Nacional de Ciencia, Tecnología e Innovación (FONACIT); Carlos Lozada and Carlos Najul for providing kidney bean seeds. References Boom van den, C.E.M., Beek van, T.A. & Dicke M. 2003: Differences among plant species in

acceptance by the spider mite Tetranychus urticae Koch. J. Appl. Entomol. 127: 177-185. Cárdenas, H., Gomez C., Díaz J. & Camerena F. 2000: Evaluación de la calidad de la proteína de

cuatro variedades mejoradas de frijol. Rev. Cubana Aliment Nutr. 14: 22-27. Cardona, C., Frei, A., Bueno, J.M., Diaz, J., Gu, H. & Dorn, S. 2002: Resistance to Thrips palmi

(Thysanoptera: Thripidae) in beans. J. Econ. Entomol. 95: 1066-1073. Choh, Y, Ozawa, R. & Takabayashi J. 2004: Effects of exogenous jasmonic acid and benzo

(1,2,3) thiadiazole-7-carbothioic acid S-methyl ester (BTH), a functional analogue of salicylic acid, on the egg production of a herbivorous mite Tetranychus urticae (Acari: Tetranychidae). Appl. Entomol. Zool. 39: 311–314.

De Ponti, O. 1977: Resistance in Cucumis sativus L. to Tetranychus urticae Koch search for sources of resistance. Euphytica. 27: 167-176.

Frei, A., Gu, H., Bueno, J., Cardona, C. & Dorn, S. 2003: Antixenosis and antibiosis of common beans to Thrips palmi Karny (Thysanoptera: Thripidae). J. Econ. Entomol. 96: 1577-1584.

Gardner, S.N. & Agrawal, A.A. 2002: Induced plant defense and the evolution of counter-defences in herbivores. Evol. Ecol. Res. 4: 1131-1151.

Gutierrez, J. 1985: Systematics. In: Spider Mites: Their Biology, Natural Enemies and Control. Vol 1A, eds. Helle & Sabelis: 75-90.

Heilker, M. & Meiners, T. 2002: Induction of plant responses to oviposition and feeding by herbivorous arthropods: a comparison. Entomol. Exp. Appli. 104: 181-192.

Helle, W. & Overmeer, W.P.J. 1985: Rearing techniques. In: Spider Mites: Their biology, Natural and control.Vol. 1A, eds. Helle & Sabelis. 331-335.

Li, C., Williamns, M.M., Loh, Y. T., Lee, G.I & Howe, G. A. 2002: Resistance of cultivated tomato to cell content feeding herbivores is regulated by the octodecanoid-signaling pathway. Plant. Physiol. 130: 494-503.

Lozada, C. 1997: Evaluación de 14 cultivares de caraota (Phaseolus vulgaris L.) y estimación de la estabilidad del rendimiento en zonas altas del Estado Lara. Bioagro. 9: 12-19.

Lozada, C., Borges, O. & Montagne A. 1995: Resistencia de cinco cultivares de caraota a Empoasca kraemeri (Homoptera: Cicadellidae) y su respuesta a diferentes dosis de insecticidas. Bioagro. 7: 3-9.

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Moros, M.E. & Aponte, O. 1995a: Efecto de dos niveles de infestación de Tetranychus ludeni Zacher sobre las fases de desarrollo de la caraota. I. Nivel de campo. Agron. Trop. 54: 189-194.

Moros, M.E. & Aponte, O. 1995b: Efecto de dos niveles de infestación de Tetranychus ludeni Zacher sobre las fases de desarrollo de la caraota. II. Nivel de invernadero. Agron. Trop. 54: 195-202.

Ochoa, R., Aguilar, H. & Vargas, C. 1994: Phytophagous Mites of Central America: An Illustrated Guide.

Read, J., Grus, E., Sanson, G., Clissold, F. & Brunt, C. 2003: Does chemical defence decline more in developing leaves that become strong and tough at maturity. Austral. J. Bot. 51: 489-496.

Steel, R. & Torrie, J. 1980: Principles and Procedures of Statistics. A Biometrical Approach. 2nd

ed. Tomczyk, A. & Kropczyńska, D. 1985: Effects on the host plant. In: Spider Mites: Their biology,

Natural and control. Vol 1A, eds. Helle & Sabelis: 317-329 Zhang, Z. & Jacobson, R.J. 2000: Using adult female morphological characters for differentiating

Tetranychus urticae complex (Acari: Tetranychidae) from greenhouse tomato crops in UK. Syst. Appl. Acarol. 5: 69-76.

Zhang, Y., Zhang, Z., Lin J., Ji, J. & Hou, A. 2001: Oviposition and survival of Schizotetranychus bambusae females (Acari: Tetranychidae) feeding on young and old bamboo leaves. Syst. Appl. Acarol. Special Publications. 9: 1-9.


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Spatiotemporal within-plant distribution of the spider mite Tetranychus urticae confronted with specialist and generalist predators Andreas Walzer1, Karl Moder2, Peter Schausberger1 1Institute of Plant Protection, Department of Applied Plant Sciences and Plant Biotechnology, University of Natural Resources and Applied Life Sciences, Peter Jordanstrasse 82, Vienna, Austria; E-mail: [email protected]; [email protected]; 2Institute of Applied Statistics and Computing, Department of Landscape, Spatial and Infrastructure Sciences, University of Natural Resources and Applied Life Sciences, Peter Jordanstrasse 82, Vienna, Austria; E-mail: [email protected] Abstract: The single and combined effects of the predatory mites Phytoseiulus persimilis and Neoseiulus californicus on the spatiotemporal of Tetranychus urticae on bean plants were investigated in separate greenhouse compartments. Four treatments were conducted: (1) T. urticae, (2) T. urticae + P. persimilis, (3) T. urticae + N. californicus, (4) T. urticae + both phytoseiid mites. Population development and distribution of the spider mites were compared among treatments. The spider mites were suppressed to zero density in the predator combination treatment but not in the single predator treatments. The predators determined the spatiotemporal distribution of the spider mites through density-mediated effects (density reduction) and behavior-mediated effects (triggering anti-predation behavior) and these effects were linked to diet specialization. The specialist P. persimilis exerted stronger density-mediated effects on the spider mite distribution than the generalist N. californicus. Both predators triggered similar anti-predation behavior in the spider mites, which was manifest in earlier bottom-up migration when predators were present. In combination the predators were somewhat more dispersed than when alone reducing the predator-free space on the plant and leading to local extinction of T. urticae. Key words: Phytoseiulus persimilis, Neoseiulus californicus, Tetranychus urticae, spatiotemporal distribution, density-mediated effects, behavior-mediated effects. Introduction Spatial distribution of prey is mainly influenced by behaviors related to foraging and/or search for mates and/or avoidance of predators (Lima 2002). Thus, predators are important determinants of distribution of prey individuals within populations (Lima & Dill 1990). In greenhouse crops the spider mite Tetranychus urticae (Acari: Tetranychidae) is often exposed to functionally different predator types such as the specialist Phytoseiulus persimilis and the generalist Neoseiulus californicus (Acari: Phytoseiidae) ( Schausberger & Walzer 2001, Blümel & Walzer 2002). In general, the predators may influence prey distribution via reducing prey density and via triggering behavioral changes in prey. The strength of these effects may be linked to diet specialization of the predators. First, the specialist P. persimilis is expected to have stronger density-mediated effects than the generalist N. californicus. Second, P. persimilis and N. californicus pose different threats to T. urticae with P. persimilis being the more dangerous predator. Antipredation behaviors such as reduced activity, avoidance of patches occupied by the

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predators and/or escape from predators by moving to predator free patches are linked with costs paid in time and energy at the expense of other fitness related activities. Thus, selection should favor T. urticae individuals that are able to adjust their antipredation behavior to the mortality risk posed by a given predator and this should be reflected in prey distribution (Sih 1986). In our study we evaluated the hypothesis that the specialist P. persimilis and the generalist N. californicus have different effects on the aggregation and distribution pattern of T. urticae. The experiments were conducted on single bean plants, divided in three vertical strata, named base, middle and top stratum. The spatiotemporal distribution of the mites was monitored over 30 days. Material and methods Experimental set-up Each experimental unit consisted of a plant-pot with french bean plants. Each treatment was replicated six times, i.e. 6 separate pots. The four treatments (T. urticae, T. urticae + P. persimilis, T. urticae + N. californicus, T. urticae + P. persimilis + N. californicus) were kept in separate computerized greenhouse compartments (environmental conditions: min. temp. 18°C during night and 25°C during day; 60±10% RH; 16:8 (L:D) photoperiod). Plants were infested with spider mites 2 weeks before releasing the predators. 16 gravid females each of P. persimilis and/or N. californicus were released per pot in the single- and two species systems. For data collection the plants (height: 80cm) were vertically sectioned into three virtual strata: the base stratum from 20 to 40cm, the middle stratum from 40 to 60cm and the top stratum from 60 to 80cm height. Starting on the 4th day after predator release, 3 leaves per replicate (1 leaf per stratum) were sampled each day and the number of mites was counted for 30 consecutive days. Statistical analysis First, the temporal variation in the overall T. urticae densities was compared among treatments using repeated measures analysis of variance (ANOVA). Second, for each treatment Taylor’s aggregation index b was calculated for T. urticae (Taylor, 1961). Third, to analyze the effect of time on the spatial distribution (hereafter termed spatiotemporal distribution) chi square tests for homogeneity of the spatial distribution of the spider mites among strata were calculated for all treatments and each day separately. To recognize spatial changes in the spider mite distribution over time a regression analysis based on the probability-levels of day wise chi square tests were performed with a Bonferroni adjustment of the significance level to lower the chance of an increased type I error due to multiple comparisons. Results and discussion Population dynamics of T. urticae In the first week the spider mite densities did not differ among treatments. In the predator free treatment the T. urticae population peaked at 194 spider mites per leaf on day 5. In the TU+NC treatment N. californicus was not able to completely suppress the spider mites within 30 days. T. urticae peaked at 79 spider mites on day 29. In every treatment with P. persimilis the spider mite populations were reduced to similar densities until day 12. In the TU+PP treatment the T. urticae density increased again after the disappearance of P. persimilis on day 14, whereas in the TU+PP+NC treatment the spider mite densities were suppressed to zero on day 14.

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Aggregation levels (Taylor’s b) Irrespective of treatment the spider mite populations were aggregated at the leaf level, i.e. Taylor’s b exceeded 1. Tetranychus urticae had a significantly higher aggregation level in treatments without predators (b=1.96) and with N. californicus (2.11) than in treatments with P. persimilis alone (1.46) and the predator combination treatment (1.46). Thus, P. persimilis but not N. californicus decreased the aggregation level of T. urticae, which was likely a consequence of P. persimilis’ stronger density-reducing effects.

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Spatiotemporal distribution of T. urticae In the predator free treatment the spatiotemporal distribution dynamics of T. urticae was characterized by an initial colonization of the base and middle stratum until day 7. Afterwards T. urticae was more frequently found in the top stratum than in the middle and base stratum (Figure 1A). This shift in vertical distribution over time may have been caused by spider mites moving up on the plants because of increased population densities in the base and middle stratum and the associated leaf damage (Bernstein, 1984). Additionally, population density and movement of the spider mites may have been affected by leaf age, with older leaves being less favorable for population growth than younger ones (Nachman & Zemek 2002). Thus, T. urticae migrated to the top stratum to search for unoccupied and/or young and more nutritious leaves.

The spatiotemporal distribution of T. urticae within strata and pooled over time was similar between the treatments TU vs. TU+NC, TU vs. TU+PP and TU+NC vs. TU+PP. Single data analysis however revealed significant differences in the spider mite distributions among treatments during certain time periods. In the TU+NC and TU+PP treatments the T. urticae fraction in the top stratum increased more rapidly than in the predator free treatment between day 2 and 6 (Figures 1B, 1C). At that time both predators mainly occupied the base and middle stratum (Figures 2A, 2B) and the overall spider mite densities did not differ between treatments with and without predators. Hence, the high T. urticae fraction in the top stratum was likely the result of antipredation behavior through premature upward migration.

It seems that T. urticae responds similarly to functionally different predators at close range (on leaves and among leaves within plants) but not at long range (among plants from the distance). At close range (on leaves) T. urticae females avoided oviposition in patches where cues from P. persimilis females were present (Grostal & Dicke 1999). This response was not predator specific because T. urticae behaved similarly when perceiving the cues of the generalist predators Euseius finlandicus (Oudemans), Amblyseius andersoni (Chant) and Iphiseius degenerans (Berlese) (Acari: Phytoseiidae) and even cues deriving from a parasitic mite of bees that poses no threat to T. urticae (Grostal & Dicke 2000). Similarly, in our experiments T. urticae avoided the strata occupied by the specialist P. persimilis or the generalist N. californicus and prematurely migrated to the top stratum irrespective of predator species and type. In contrast, Pallini et al. (1999) showed in olfactometer tests that T. urticae females avoid visiting plants with P. persimilis but do not avoid plants with N. californicus. It thus seems that the costs of patch avoidance for T. urticae increase with the spatial scale.

Avoiding plants with predators may only pay off for T. urticae when the enemy is a highly dangerous predator such as P. persimilis. The potential benefits from avoiding plants with the less dangerous predator N. californicus may not outweigh the high costs of a prolonged search for an alternative host plant. Contrary, at close range (among leaves within a plant) the costs of predator avoidance seem lower and migration to predator-free leaves may be an appropriate response to both a specialist predator such as P. persimilis and a generalist predator such as N. californicus. After day 6 the spatiotemporal distribution of T. urticae was similar in the TU and TU+NC treatments (Figures 1A, 1B). Contrary, following a rapid increase until day 9 the spider mite fraction decreased in the top stratum in the TU+PP treatment. Phytoseiulus persimilis disappeared on day 13. Consequently, the spider mite density increased again and reached higher fractions in the base and middle stratum than in the top stratum, which was different from the distribution pattern in the TU and TU+NC treatments (Figures 1A, 1B, 1C).

After the initial similar antipredation response in both single predator treatments, the spatiotemporal distribution of T. urticae was strongly influenced by density reduction caused by

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the predators. Density reduction by P. persimilis but not N. californicus was disproportionate among strata. As a consequence P. persimilis but not N. californicus had a pronounced density-mediated effect on the distribution of T. urticae. Phytoseiulus persimilis migrated earlier to the top stratum than did N. californicus. Additionally, the rapidly decreasing density of T. urticae in the middle stratum due to predation by P. persimilis and the associated increasing P. persimilis density from 0.2 (day 1) to 1.1 individuals per leaf (day 5) may have limited the chances of T. urticae to migrate to and colonize the top stratum. This conclusion is supported by the aggregation levels of T. urticae, which were lower with P. persimilis than with N. californicus. The latter difference may have been caused by (1) differing densities and associated variabilities due to predation (Onzo et al. 2005), or (2) differing responses to predators (Magalhães et al. 2002), or (3) both. Our experiment suggests that cause 1 had much more impact than cause 2. As mentioned before both predators initially caused a similar avoidance response in T. urticae yet density reduction by P. persimilis was stronger and more disproportionate among strata than density reduction by N. californicus. Moreover, the aggregation level of T. urticae was not affected by N. californicus and the spatial distributions of T. urticae and N. californicus were identical (Figures 1A, 2B). Hence, predation by N. californicus reduced the overall population densities of the spider mites but did not have an effect on their spatial distribution among strata.

The spatiotemporal spider mite distribution differed significantly between the treatments TU vs. TU+PP+NC, TU+NC vs. TU+PP+NC and TU+PP vs. TU+PP+NC. Single data analysis revealed two significant characteristics of the spatiotemporal distribution in the combined predator system: (1) between day 6 and 8 higher T. urticae fractions were found in the top stratum than in the other treatments; (2) the T. urticae distribution fluctuated more among strata when confronted with both predators (Figure 1D).

The predators in combination occupied more leaves than each species alone, which resulted in a more rapid movement of T. urticae from the base and middle stratum to the top stratum at the beginning of the experiments than in all other treatments (Figures 2C, 2D). Such non-lethal effects of predators may have similar stabilizing effects on the persistence of predator-prey systems as the use of prey refuges (Van Baalen & Sabelis 1999). However, vertical migration of T. urticae led only to a temporal release from predation because of the numerical response of the predators and the associated diminishment of predator-free space also in the top stratum. Consequently, in the further course of the experiment T. urticae was driven to local extinction.

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Figure 2. Vertical spatiotemporal within-plant distribution of P. perimilis (A) or in combination with N. californicus (C) and N. californicus (B) or in combination with P. persimilis (D) over 30 days. References Bernstein, C., 1984: Prey and predator emigration responses in an acarine system Tetranchus

urticae-Phytoseiulus persimilis. Oecologia 61: 134-142. Blümel, S. & Walzer, A., 2002: Efficacy of different release strategies of Neoseiulus californicus

McGregor and Phytoseiulus persimilis Athias Henriot (Acari: Phytoseiidae) for the control of two-spotted spider mite (Tetranychus urticae Koch) on greenhouse cut roses. Syst. Appl. Acarol. 7: 35-48.

Grostal, P. & Dicke, M., 1999: Direct and indirect cues of predation risk influence behavior and reproduction of prey: a case for acarine interactions. Behav. Ecol. 10: 422-427.

Grostal, P. & Dicke, M., 2000: Recognising one’s enemies: a functional approach to risk assessment by prey. Beha. Ecol. Sociobiol. 47: 258-264.

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Lima, G.A. & Dill, L.M., 1990: Behavioral decisions made under the risk of predation: a review and prospectus. Can. J. Zool. 68: 619-640.

Lima, S.L., 2002: Putting predators back into behavioral predator-prey interactions. TREE 17: 70-75.

Magalhães, S., Janssen, A., Hanna, R. & Sabelis M.W., 2002: Flexible antipredation behavior in herbivorous mites through vertical migration in a plant. Oecologia 132: 143-149.

Nachman, G. & Zemek, R., 2002: Interactions in a tritrophic acarine predator-prey metapopulation system IV: effects of host plant condition on Tetranychus urticae (Acari: Tetranychidae). Exp. Appl. Acarol. 26: 43-70.

Onzo, A., Hanna, R., Zannou, I., Sabelis, M.W. & Yaninek, J.S., 2003: Dynamics of refuge use: diurnal, vertical migration by predatory and herbivorous mites within cassava plants. Oikos 101: 59-69.

Pallini, A., Janssen, A. & Sabelis, M.W., 1999: Spider mites avoid plants with predators. Exp. Appl. Acarol. 23: 803-815.

Schausberger, P. & Walzer, A., 2001: Combined versus single species release of predaceous mites: predator-predator interactions and pest suppression. Biol. Control 20: 269-278.

Sih, A., 1986: Antipredator responses and perception of danger by mosquito larvae. Ecology 67: 434-441.

Taylor L.R., 1961: Aggregation, variance and the mean. Nature 189: 732-735. Van Baalen, M. & Sabelis, M.W., 1999: Nonequilibrium population dynamics of „ideal and free“

prey and predators. Am. Nat. 154: 69-88. Wilson, L.T., Zalom, F.G. & Smilanick, J.M., 1984: Sampling mites in almonds: I. Within-tree

distribution and clumping pattern of mites with comments on predator-prey interactions. Hilgardia 52: 1-13.

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Symbionts in mites and their relevance for pest control Einat Zchori-Fein1, Monika Enigl2, Peter Schusberger2, Netta Mozes-Daube1, Yuval Gottlieb1, Tal Hanuny1 and Eric Palevsky1 1Department of Entomology, Newe Ya'ar Research Center, ARO, Israel; 2Institute of Plant Protection, Department of Applied Plant Sciences and Plant Biotechnology, University of Natural Resources & Applied Life Sciences, Peter Jordanstrasse 82, A-1190 Vienna, Austria Abstract: Maternally-inherited symbionts of arthropods are known to influence many aspects of their host's biology. With the advance of the field, enough data has been collected to be of applied value and a novel approach, termed symbiont-based protection (SyBaP) or "symbiotic control", is being considered for combating economically and medically important pests. In an effort to assess the possible use of SyBaP methods against mite pests, one predatory mite - Neoseiulus californicus - and one phytophagous mite - Rhizoglyphus robini - was chosen, and their symbiotic complex was determined. Molecular fingerprinting techniques revealed Spiroplasma in N. californicus, but no significant influence on fitness was found. Similarly, Defluvibacter and another unnamed α-Proteobacterium were found in R. robini, and both were observed concentrated in the same areas of R. robini eggs using confocal microscopy. This study will serve as the first step toward the application SyBaP methods in mites.

Key words: symbiotic control, bacterial symbionts, predatory mites, bulb mites Introduction The past two decades have seen a rise in appreciation of the importance of intracellular symbionts of arthropods, because the advent of sensitive molecular techniques have provided tools for understanding the great ecological and taxonomic diversity of microorganisms that are transmitted from female hosts to their offspring. Some of these vertically-transmitted bacteria benefit their host’s fitness directly by providing critical nutrients on nutritionally-limited or unbalanced diets (Baumann, 2005). These primary symbionts are generally housed in specialized cells, are obligatorily associated with their host and carry a highly reduced genome. Others, facultative secondary symbionts, enhance their own transmission in two ways: 1) by benefiting host fitness through mediation of ecological interactions (e.g. Tsuchida et al. 2004; Oliver et al. 2005) and 2) by manipulating the host reproduction (O'Neill et al. 1997).

Until recently and as compared to insects, the literature available on the interaction between Acari other than ticks and their symbionts has been sporadic and mainly focused on diseases caused by fungi, viruses, bacteria and microsporidia (van der Geest et al. 2000). From the view point of protecting crops from phytophagous mites, the influence of such symbionts on their acarine host may be important for: 1) Improving the efficiency of phytoseiid predators (in mass rearing, inundative releases or classical biological control programs), and 2) Developing innovative control methods for the pest itself. The use of symbiotic bacteria for combating economically and medically important pests is a developing field and has been termed symbiont-based protection (SyBaP) or "symbiotic control". In an effort to assess the suitability of SyBaP

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methods for combating mites with agricultural importance, one predatory mite (the phytoseiid Neoseiulus californicus (McGregor)) and one phytophagous mite (the bulb mite Rhizoglyphus robini Claparède, Acaridae) was chosen, and the symbionts diversity associated with each one of them was determined.

Neoseiulus californicus is a generalist feeding on spider mites, insects and pollen. The mite is mass-reared and commercially applied as a part of biological control efforts on vegetables, fruits and ornamentals. Rhizoglyphus robini is a cosmopolitan soil-borne mite commonly found in agricultural soils. It is considered a pest of agricultural crops belonging to Liliaceae such as lily, onion and garlic and is associated with soil borne fungi (Diaz et al. 2000). Material and methods Mite origin and rearing Neoseiulus californicus: As a part of a multinational project funded by the European Union, N. californicus samples from different geographical origins were collected by Dr. Kreiter (ENSAM, Montpellier), and alcohol-preserved samples were sent to us (Table 1). A live population of the N. californicus "Sicily" strain was obtained from Dr. Kreiter, reared in the laboratory and then used for the fitness effects experiments. The mites were kept on artificial arenas, with either spider mites or pollen as a food source, at 20-25°C, 60-80% RH and 16:8 L:D. Rhizoglyphus robini: Mites were collected on lily bulbs from commercial screenhouses in the coastal plain of Israel in 2000 and have since been reared on peanuts in the laboratory of Dr. Palevsky (ARO, Israel) at 20-25°C, 60-80% RH and 16:8 L:D. PCR amplification and DGGE analysis To determine the diversity and identity of bacteria associated with N. californicus and R. robini, adults were ground in a lysis buffer, the 16S rDNA gene fragment (~550bp) was amplified by PCR using the general primers 341F and 907R and the products were subjected to denaturating gradient gel electrophoresis (DGGE) analysis as described in Gottlieb et al. (2006). Bands representing bacteria were eluted, cloned and sequenced (ABI 3700 DNA analyzer, Macrogen Inc. Korea). The sequences obtained were compared to known sequences in the data bases using the BLAST algorithm in NCBI. Sequencing the full length 16S rDNA To construct a larger segment of the 16S rDNA gene of the bacteria found in R. robini, a primer set that is known to amplify that gene from most known Bacteria (27F and 1513R, Weisburg et al. 1991) was used in combination with specifically-designed primers based on the sequences obtained by the DGGE (Defcon-F 5'-CAGTTTACTGTTCGGTGG and Licecon-F 5'-GGTTGGTAGTTAAGTATCGG) in a PCR. The two-spotted spider mite Tetranychus urticae Koch (Tetranichidae) served as a negative control for confirmation of primers specificity. The 16S rDNA contigs were assembled using DNAMAN (Lynnon Biosoft Vaudreuil, Quebec, Canada), so that the nearly-full length gene sequences were generated for the two R. robini symbionts. Fitness effects of Spiroplasma on its N. californicus host To determine the influence of Spiroplasma infection on sex ratio and fecundity, single pairs of Spiroplasma-infected and uninfected female deutonymphs and males were transferred to detached leaf arenas. Males were removed when the first egg was laid and the females were allowed to oviposit for 7 days. Eggs were counted and removed daily. Offspring were raised on separate arenas and sex ratio (proportion of female offspring) was determined. Influence of time

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(days 1 to 7) and infection status of mothers on egg production was analyzed through repeated measures analysis of variance and on sex ratio through a T-Test; the proportion of female offspring was square-root transformed prior to analysis. Fluorescent in situ hybridization (FISH) of R. robini eggs For the FISH analysis the reverse complement sequence of the specific PCR primers were used as specific probes. The probe for Defluvibacter-like symbiont was labeled with Cy3 and the probe for the second α-Proteobacterium was labeled with Cy5. The FISH procedure was performed on one-day-old eggs collected directly into Carnoy's fixative, and generally followed Gottlieb et al. (2006). Stained eggs were whole mounted and viewed under an IX81Olympus FluoView™500 confocal microscope. Specificity of the detection was confirmed using the negative controls of no probe, and eggs of T. urticae. Results and discussion As compared to insects, not much is known about the interactions between mites and their bacterial symbionts. Wolbachia and Cardinium have been demonstrated to cause effects such as cytoplasmic incompatibility, feminization and increased fecundity in several species of phytophagous mites (e.g. Breeuwer, 1997; Chigira & Miura, 2005) and phytoseiids (Weeks & Stouthamer, 2004), but the role of other symbionts is still unclear. The results presented here are an attempt to assess the diversity of bacteria associated with different mite species. Neoaliturus californicus Using DGGE analysis, the microbial community of six different N. californicus strains was established (data not shown). Of the 16 bands sequenced from the different N. californicus samples, nine had high sequence similarities to known bacteria (Table 1), three resembled the 18S rRNA of mites, two resembled chloroplasts and two were unknown. Wolbachia was found in two N. californicus populations, but since it could also be detected in the tetranychid prey and a sample from the Sicily line held on pollen tested negative for Wolbachia (data not shown), it was assumed that the symbiont originated from the gut (as demonstrated in Enigl et al. 2005). One result that was of particular interest was that two lines (i.e. Sicily and Chile) showed the presence of a Spiroplasma. Since Spiroplasma is known to be an invertebrate symbiont, we focused on that bacterium thereafter.

The genus Spiroplasma consists of motile, helical, wall-less bacteria belonging to the class Mollicutes, which it shares with other host-associated members such as the vertebrate-pathogenic mycoplasmas and the insect-vectored plant-pathogenic phytoplasmas (Gasparich et al. 2004). Species within the genus are known to exhibit a diverse array of relationships with their hosts, with respect to factors such as transmission modes, replication sites and number of hosts required for successful completion of the life cycle (Bové, 1997). Within the Acari, Spiroplasma has so far been identified in ticks but the influence of the symbiont on its tick host has not been determined. As far as we are aware, the data reported here is the first published record of Spiroplasma in mites other than ticks.

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Table 1. Origin of the different mite populations screened, and the bacteria found using DGGE Species and origin Collection data Received from Bacteria found

N. californicus USA (California) - Y. Argov, Israel Wolbachia Spain (Bolbaite) strawberries, 2000 S. Kreiter, France Wolbachia Chile (La Cruz) beans, 2000 S. Kreiter, France Spiroplasma, Acetobacter,

Serratia symbiotica France (Mauguio) eggplants, 2004 S. Kreiter, France None Italy (Florence) strawberries, 2004 S. Kreiter, France Pantoea sp. Italy (Sicily) strawberries, 2004 S. Kreiter, France Spiroplasma, Acetobacter

R. robini lily bulbs, 2000 E. Palevsky, Israel Defluvibacter, Providencia, Bacillus, Clostridium, unnamed Bacteroidetes, unnamed α-Proteobacterium

Spiroplasma-infected mothers produced the same total number of eggs (11) (F = - 0.065, DF

= 1, P = 0.800) and similar offspring sex ratios (62-63%) (T = -0.276, DF = 35, P = 0.784) as did non-infected mothers (Table 2). In contrast, Spiroplasma had several negative effects on pea aphids, where the growth, fecundity and longevity were adversely affected by its presence (Fukatsu et al. 2001). The phenotype of Spiroplasma in N. californicus is still unknown but in another arthropod-symbiont system, the corn stunt spiroplasma Spiroplasma kunkelli and its vector, the corn leafhopper Dalbulus maidis, it was shown that Spiroplasma-infected individuals had survival rates higher than non-infected ones under conditions of starvation and cold temperatures (Ebbert & Nault, 2001). Table 2. Fecundity of Spiroplasma infected and uninfected mothers of Neoseiulus californicus, and the offspring sex ratio.

Infection status of mothers

N Total # of eggs on Day 7 (Mean±SD)

N % female offspring (Mean±SD)

no 33 11 ± 5 29 62 ± 16 yes 8 11 ± 5 8 63 ± 19

Rhizoglyphus robini Using DGGE analysis the microbial community of R. robini was characterized. Bands representing six different bacteria were evident (Figure 1) and all of their sequences corresponded to known bacteria (Table 1). The two bacteria that were fully-sequenced during this study belonged to the α-Proteobacteria. One was most closely related to Defluvibacter sp. (99% similarity over 1451bp, acc. #AJ009467) and the other to a symbiont of a chewing louse found on pocket gopher (96% similarity over 690bp, acc. #AF467383).

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When one-day-old eggs were hybridized with specific probes for each of the above bacteria, a specific signal was observed under a confocal microscope. Both bacteria are localized throughout the egg, showing foci localization (Figure 2). The controls verified the specificity of the probes (data not shown). The concentrations of bacteria in specific foci may indicate the existence of bacteriocytes-like structures and suggest an intimate association of the symbionts with their host. The results obtained show that R. robini hosts a diverse assembly of symbionts, some of which are vertically transmitted to the next generation through the egg. Although the phenotype of these symbionts is yet to be determined, it is reasonable to assume they have an important role in their host's biology.

Figure 1. DGGE analysis of PCR-amplified, 16S rRNA gene fragments of bacteria found in two individual Rhizoglyphus robini. M – 50 bp marker.

Overall, the results presented illustrate the diversity of symbiotic bacteria harboured by both predatory and phytophagous mites and stress the importance of acknowledging their existance, identifying them and determining their influence on their acarine hosts. Such studies lay the foundation for applying symbiont-based control methods. Acknowledgements This study was part of a multi-institutional project funded by the European Community, within the sixth framework, Horizontal Research Activities involving SMEs, Co-operative Research (CRAFT), Project number and acronym: 508090, EUROMITE.

1 2 H2O M

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Transgenic crop-mite interactions Rostislav Zemek Institute of Entomology, Biology Centre AS CR, Branisovska 31, CZ-370 05 Ceske Budejovice, Czech Republic, [email protected] Abstract: Transgenic crops that are resistant to insect pests are grown on an increasing percentage of the global agricultural area. Understanding of the interactions between these crops and plant-feeding mites and their natural enemies is very important from both integrated pest control and risk assessment research point of view. So far, only a few laboratory and greenhouse studies on the effects of transgenic plants with insecticidal activity on mites have been reported. They used plants expressing lepidopteran- or coleopteran-active Bacillus thuringiensis toxins (maize, eggplants, potatoes), Galanthus nivalis agglutinin (potatoes) or proteinase inhibitor (tomato). Tetranychus urticae Koch was selected as a model herbivorous mite while Neoseiulus cucumeris (Oudemans), Phytoseiulus persimilis A.-H. and Typhlodromus pyri Scheuten were selected as predatory mites in these studies. Although no acute toxic effects of transgenic plants on any mites were reported, the obtained data revealed both positive and negative effects of these plants on mites. Whether the observed effects are due to the insecticidal proteins, or due to other changes in primary or secondary metabolites in transgenic plants, is unclear and remains to be elucidated. Key words: GMO, transgenic plants, mites, non-target effects Introduction Genetic engineering is one of several strategies adopted to generate resistance to biotic stress in crop plants. In this technology genes from other species are isolated and transferred into the plant genome using either gall-forming bacterium Agrobacterium tumefaciens or ballistic methods. Genetic engineering has been successful mainly in creating insect resistant plants (Jouanin et al. 1998). Transgenic plants expressing insecticidal proteins target the digestive system of insect pests and are either based on plant-derived genes or genes of entomopathogenic bacterium Bacillus thuringiensis Berliner (Bt). The former approach uses genetic information that code for enzymes like lectines, e.g. Galanthus nivalis L. agglutinin, proteinase or amylase inhibitors or cholesterol oxidase. The latter approach uses genes that code for Bt insect-toxic proteins, mostly crystal (Cry) type (Schuler et al. 1998).

Transgenic insect resistant plants are commercially available since the mid 1990s and the percentage of the global agricultural area where they are grown is increasing considerably (James 2005). Despite the proven effectiveness of transgenic plants in pest control, adoption of this new technology requires careful risk assessment and research into possible unintended side effects, for example on non-target organisms (Andow & Hilbeck 2004, Hilbeck 2001). In this paper, I review literature, in which effects of transgenic plants on mites were investigated.

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Transgenic plants containing B. thuringiensis genes The most spread transgenic plants which are either fully or partly resistant against specific insect pests are plants with incorporated genes from different subspecies of B. thuringiensis that encode Cry proteins. Among these Bt-plants, the most popular is transgenic maize carrying the lepidopteran-active Cry1Ab gene from B. thuringiensis kurstaki for control of stem borers (Koziel et al. 1993). New maize varieties with the coleopteran-active Cry3Bb for maize rootworm control have been developed recently (Ostlie 2001). Combination of several Bt genes that will provide various modes of action to allow more effective control of a broader range of the major insect pests will be used in new transgenic varieties. In addition to the commercially available Bt crops, Bt transgenes have been inserted into transgenic varieties of an array of other crop plant species that are not yet approved, e.g. eggplant, Solanum melongena L., expressing the coleopteran-active Cry3Bb toxin (Arpaia et al. 1997).

Contrary to insects, only little attention has, so far, been paid to the effects of transgenic Bt-plants on mites despite the known accumulation of Bt toxin in spider mites (Head et al. 2001, Dutton et al. 2002, Raps et al. 2001). The interactions between Bt plants and plant-feeding mites and their natural enemies have been studied using Bt-maize expressing the lepidopteran-active Cry1Ab toxin, Bt-potatoes expressing the coleopteran-active Cry3A toxin and Bt-eggplants expressing the coleopteran-active Cry3Bb toxin (Table 1). The results of these studies indicate that either Bt toxins or other changes in transgenic plants can have some effect on mites, although not acute toxicity was reported. Table 1. Summary of laboratory studies which tested the effect of transgenic Bt plants on mites. 0 = no differences between a non-Bt control and transgenic Bt-plant; - and + = negative and positive effects, respectively, on studied parameters. Parentheses indicate that the effect was only marginally significant.

Plant species Bt toxin Mite species Effect Reference Zea mays Cry1Ab Tetranychus urticae 0 (Lozzia et al. 2000) (-) (Dutton et al. 2002) Neoseiulus cucumeris 0 (Dutton et al. 2002) Phytoseiulus persimilis 0 (Zemek et al. 2003) Typhodromus pyri 0 (Vavrova 2005) S. tuberosum Cry3A Tetranychus urticae (-) (Zemkova & Zemek 2003) S. melongena Cry3Bb Tetranychus urticae + (Zemkova et al. 2005) Phytoseiulus persimilis - (Zemkova et al. 2005)

Transgenic plants containing snowdrop genes The snowdrop G. nivalis agglutinin lectin (GNA) has been shown to be insecticidal to a range of important pests. Genes that encode GNA have been incorporated into potatoes (Down et al. 1996), rice (Foissac et al. 2000, Rao et al. 1998, Wu et al. 2002), tobacco (Hilder et al. 1995),

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wheat (Stoger et al. 1999) and tomato (Wu et al. 2000) to increase crop resistance to aphids, leafhoppers, lepidopteran or coleopteran pests.

Zemkova & Zemek (2006) tested the effect of transgenic GNA potatoes on host plant preference of T. urticae using a two-choice disc test. Significantly more spider mites and their eggs were observed on control leaves than on transgenic leaves. The preference for control (non-transgenic) was more obvious than in aphids against which GNA potato was primarily aimed (Zemkova & Zemek 2006). Recent study (Zemek et al., in prep.) revealed negative effect of GNA potatoes on performance of T. urticae both in laboratory and small-scale greenhouse experiments. Transgenic plants containing proteinase inhibitor-coding genes Genes coding for proteinase inhibitors are considered to have a potential for improving defenses of plants against insect as well as pathogens (Ryan 1990). Recently, transgenic tomato containing the Kunitz gene (KT13), a serin proteinase inhibitor, has been developed (Castagnoli et al. 2003). This tomato line was shown to be tolerant to Helicoverpa armigera but it enhanced population growth of T. urticae, at least in the laboratory trials (Castagnoli et al. 2003). Discussion Although no detrimental effect of transgenic Bt plants was observed on both spider mites and predatory mites, long-term exposition to these plants might affect population dynamics of mites. For instance, Zemkova et al. (2005) observed that eggplants expressing Bt toxin for resistance against the Colorado potato beetle are more preferred by spider mites but are less preferred by their predator P. persimilis. Such a simultaneous shift in the preference of both the spider mites and their predators could result in lower effectiveness or even failure of biological control. It is also known that spider mites accumulate the Bt toxin in large amounts (Head et al. 2001, Dutton et al. 2002, Raps et al. 2001), yet the trophic impact of this accumulation has been little studied. The toxin is likely to be passed to the third trophic level, where unexpected effects might occur (Groot & Dicke 2002). Recent work by Hardwood at al. (2005) underlies the importance of knowing the transfer by trophic links of GM products.

Transgenic potatoes expressing snowdrop lectin negatively affected spider mites. When the observed effects were caused by GNA lectin then this lectin could be considered a possible target for genetic engineering into crops for broader pest resistance, including resistance against plant-feeding mites. On contrary, transgenic plants containing proteinase inhibitor-coding genes seems to have either stimulatory or none effect on spider mites.

Genetic transformation of plants can change metabolic pathways, and result in different chemical compositions of transgenic plants. For instance, a higher content of lignin was found in Bt-maize (Saxena & Stotzky 2001) and a lower level of leaf glycoalkaloids in transgenic potatoes (Birch et al. 2002). If the reported effects of transgenic plants on mites are due to introduced toxins, e.g. due to their possible anti-feedant properties, or due to other changes in primary or secondary metabolites in transgenic plants, remains to be elucidated.

The present paper demonstrates that the understanding of the interactions between GM plants, phytophagous and predatory mites will be very important from plant protection as well as from environmental risk point of view. To answer the questions whether transformation of plants

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incorporating insecticidal genes has any effect on plant-feeding mites, and whether the usage of this new pest management technology is compatible with biological control agents needs further research including greenhouse and field experiments. Acknowledgements This work was supported by the Grant Agency of the Academy of Sciences of the Czech Republic (A6007303). References Andow, D.A. & Hilbeck, A. 2004: Science-based risk assessment for non-target effects of transgenic

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Birch, A.N.E., Geoghegan, I.E., Griffiths, D.W. & McNicol, J.W. 2002: The effect of genetic transformations for pest resistance on foliar solanidine-based glycoalkaloids of potato (Solanum tuberosum). Ann. Appl. Biol. 140: 143–149.

Castagnoli, M., Caccia, R., Liguori, M., Simoni, S., Marinari, S. & Soressi, G.P. 2003: Tomato transgenic lines and Tetranychus urticae: changes in plant suitability and susceptibility. Exp. Appl. Acarol. 31: 177–189.

Down, R.E., Gatehouse, A.M.R., Hamilton, W.D.O. & Gatehouse, J.A. 1996: Snowdrop lectin inhibits development and decreases fecundity of the glasshouse potato aphid (Aulacorthum solani) when administered in vitro and via transgenic plants both in laboratory and glasshouse trials. J. Insect Physiol. 42: 1035–1045.

Dutton, A., Klein, H., Romeis, J. & Bigler, F. 2002: Uptake of Bt-toxin by herbivores feeding on transgenic maize and consequences for the predator Chrysoperla carnea. Ecol. Entomol. 27: 441–447.

Foissac, X., Loc, N.T., Christou, P., Gatehouse, A.M.R. & Gatehouse, J.A. 2000: Resistance to green leafhopper (Nephotettix virescens) and brown planthopper (Nilaparvata lugens) in transgenic rice expressing snowdrop lectin (Galanthus nivalis agglutinin; GNA). J. Insect Physiol. 46: 573–583.

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