Abstract booklet potato wart disease workshop june 2019 · Absrac Booklet - 3 - Background and...

Abstract Booklet

Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL

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Abstract Booklet

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Background and program

In June 2019, the Dutch National Plant Protection Organization (NPPO-NL), Wageningen University & Re-search (WUR), and Agriculture and Agri-Food Canada (AAFC) organize a workshop on Synchytrium endobi-oticum, the causal agent of potato wart disease. The workshop brings together biologists, researchers and policy makers working on all aspects of Synchytrium endobioticum and the interaction with its host potato.

Aims of the workshop are: 1. expanding professional networks; 2. sharing knowledge via presentations and posters, and; 3. identifying shared research needs.

Workshop attendees are encouraged to present their work, and presentations are grouped in three sessi-ons: Session 1. “Disease occurrence and management” Chair: Dr Nico Horn Director General, European and Mediterranean Plant Protection Organization, France

Session 2: Understanding the pathogen Dr Peter Bonants Senior scientist/group leader, Biointeractions and plant health, Wageningen University & Research, the Netherlands

Session 3: Host resistance and breeding Prof. Dr Richard Visser Professor of Plant Breeding, Wageningen University & Research, the Netherlands

A keynote lecture will be delivered by Prof. Dr Francine Govers, Professor of phytopathology from Wagenin-gen University & Research.


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Abstract Booklet

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Oral presentations


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Oral presentations Session 1. Disease occurrence and management

Pathotype identification of Synchytrium endobioticum (Schilb.) Perc. in Bulgaria and testing of potato varieties for resistanceLidia Dimitrova (1), Ani Becheva (1), Mariyana Laginova (1)

1. Bulgarian Food Safety Agency, Central Laboratory for Plant Quarantine, Sofia, Bulgaria

Potato wart disease (PWD) caused by Synchytrium endobiticum was identified for the first time in Bulgaria in the cool and humid summer of 2004 (region Samokov). In 2012 new outbreaks were reported in the sou-thern part of the country (region Smolyan). The pathogen was detected in tuber samples in small gardens with a long history of potato cropping. The area is mountainous, mainly with light sandy to sandy loam soils, low content of organic matter and pH below 5,5. The continuous growing of potatoes without crop rotation and the use of non certified planting material in courtyards were the main reasons for the spread of the pathogen.

A series of laboratory tests for pathotype identification of isolates from both regions were carried out. A set of differential cultivars (EPPO, 2004, 2017) was tested using the Glynne-Lemmerzahl method (IHAR-PIB protocol, Poland). The reaction types of the sprouts were scored after two or three weeks according to the classification scheme established by Langerfeld and Stachewiccz (1994). Based on the results the pathoty-pes 8(F1), 18(T1) and 38(Nevsehir) were identified.

A list of cultivars resistant to the local pathotypes is updated annually, according to Council Directive 69/464/EEC of 8 December 1969 on control of PWD, transposed into Bulgarian legislation through Ordinan-ce № 20 of 2 July 2001. Around 170 varieties were tested for resistance in order to be used in buffer zones surrounding infested fields. Most of the tested cultivars were susceptible. The resistant varieties have no commercial importance.

Abstract Booklet

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The role of the National Reference Centre, Wageningen in identification, collection maintenance and disease management of potato wart diseaseGerard C.M. van Leeuwen (1), Nico J.B. Mentink (1), Johan P. Meffert (1), Karin H.M. Rosendahl (1) & Patri-cia C.J. van Rijswick (1)

1. Dutch National Plant Protection Organization, National Reference Centre, section Mycology, Wageningen, the Netherlands

The study of potato wart disease in all its aspects has always been a major issue in the Netherlands Plant Protection Organisation. Since its first occurrence in 1915 in the north of the country one has tried to mi-nimise the detrimental effects of this disease. Planting resistant cultivars was ánd is still one of the corner-stones to build management of the disease on. In time, the dominant pathotype 1(D1) has been replaced by new pathotypes, e.g. 2(G1) and 18(T1) in the Netherlands. The focus in the past 10-15 years has laid on pathotypes 1(D1), 2(G1), 6(O1) and 18(T1); it is foremost with this series of pathotypes where efforts were done to harmonise methodology and choice of differential cultivars. In conjunction with Germany and Poland an interlaboratory comparison was done on pathotyping in 2009-2011. Later, the Euphresco-project SENDO was initiated by the National Reference Centre (NRC), including partners from nine different Euro-pean countries (2012-2015). Ultimately, this has lead to a new version of the EPPO Diagnostic Protocol on Synchytrium endobioticum, describing a standardized, validated set of differential cultivars for pathotype identification.

Descheduling of infested fields has got much attention in the Netherlands recently. Many of these fields are scheduled now for 20 years, and descheduling according to EPPO Standard PM 3/59 has started. Ana-lysis of soil samples by direct detection (after washing/sieving) ánd bio-assays are executed. Bio-assays are considered as less sensitive, direct examination (detecting resting spores under microscope) is hampered by doubts about the identity of particles seen. Recently, the NRC has started to compare two real-time PCR tests for use in molecular detection of resting spores in soil suspension resulting from the washing/sieving steps.


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To be announcedHale Gunacti (1)

1. Biological Control Research Institue, Turkey

To be announced

Abstract Booklet

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Potato wart disease in the Netherlands: a historic perspective from a molecular viewMarga P. E. van Gent-Pelzer (1), Bart T.L.H. van de Vossenberg (1,2), Patricia C.J. van Rijswick (2), and Theo A. J. van der Lee (1)

1. Wageningen UR, Biointeractions and Plant Health & Plant Breeding, Wageningen, The Netherlands2. Dutch National Plant Protection Organization, Geertjesweg 15, 6706EA Wageningen, The Netherlands

To be announced


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Oral presentations Session 2. Understanding the pathogen

To be announcedSyed M. Hoque (1)

1. Bangladesh Agricultural Research Institute, Gazipur, Bangladesh

To be announced

Abstract Booklet

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The linear mitochondrial genome of the quarantine pest Synchytrium endobioticum; insights in the evo-lutionary history of an obligate biotrophBart T. L. H. van de Vossenberg (1,2), Balázs Brankovics (1), Hai D. T. Nguyen (3), Marga P. E. van Gent-Pel-zer (1), Donna Smith (4), Kasia Dadej (3), Jarosław Przetakiewicz (5), Jan F. Kreuze (6), Margriet Boerma (7), Gerard C. M. van Leeuwen (2), C. André Lévesque (3) and Theo A. J. van der Lee (2)*

* Presenting author

1. Wageningen UR, Biointeractions and Plant Health & Plant Breeding, Wageningen, The Netherlands2. Dutch National Plant Protection Organization, Geertjesweg 15, 6706EA Wageningen, The Netherlands3. Agriculture and Agri-Food Canada, 960 Carling Avenue, Ottawa, Canada4. Canadian Food Inspection Agency, 93 Mount Edward Road, Charlottetown, Canada5. Plant Breeding and Acclimatization Institute, National Research Institute, Warsaw, Poland6. International Potato Centre, Avenida La Molina, 1895 Lima, Peru7. Hilbrands Laboratorium BV, Kampsweg 27, 9418 PD Wijster, Wijster, The Netherlands

Chytridiomycota (chytrids) inhabit terrestrial and aquatic environments, and represent a basal lineage in true fungi. Most of the described chytridiomycota are free living saprophytes, but several species are noto-rious pathogens for plants or amphibians. Synchytrium endobioticum is an obligate biotroph chytrid causing potato wart disease. Quarantine measures have been implemented worldwide to control the disease and to prevent its spread.

To determine taxonomical relationships, and to gain insights into the evolutionary history of this plant pathogen we assembled and annotated the mitochondrial genome of S. endobioticum and generated mitochondrial genomes for five additional chytrid species. The mitochondrial genome of S. endobioticum is a linear 72,865 bp molecule with terminal inverted repeats that encodes 14 mitochondrial genes typically found in fungi.

Polymorphisms in 30 S. endobioticum isolates shows clustering in four main mitochondrial lineages, and from our data we conclude that the pest was introduced at least three times in Europe. Strains of patho-type 2(G1) and 6(O1) were represented in two mitochondrial lineages, showing that these pathotypes emerged independently. Variations within a strain for polymorphic sites were observed and seem to be consistent in different mitochondrial lineages suggesting that S. endobioticum strains are communities of different genotypes with conserved composition.


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Identification of the AvrSen1 gene: towards molecular pathotypingBart T.L.H. van de Vossenberg (1,2), Charlotte Prodhomme (1), Gert van Arkel (1), Marga P.E. van Gent-Pel-zer (1), Marjan Bergervoet (1), Balázs Brankovics (1), Jarosław Przetakiewicz (3), Richard G.F Visser (1), Theo A.J. van der Lee (1), and Jack H. Vossen (1)

1. Wageningen University and Research, Droevendaalsesteeg 1, Plant Science Group, 6708 PB, Wageningen, the Netherlands2. Dutch National Plant Protection Organization, National Reference Centre, Geertjesweg 15, 6706EA, Wa-geningen, The Netherlands3. Plant Breeding and Acclimatization Institute, National Research Institute, Radzików, 05-870 Błonie, Warsaw, Poland

Synchytrium endobioticum, the pathogen causing potato wart disease, has a world-wide quarantine status and is included on the HHS and USDA Isolates of S. endobioticum are grouped as pathotypes based on their ability to evade host-resistance in a set of differential potato varieties. So far, thirty-nine pathotypes are acknowledged of which pathotypes 1(D1), 2(G1), 6(O1) and 18(T1) are found most frequently.

We hypothesize that differential resistance is caused by recognition of pathotype specific effectors or avi-rulence (Avr) genes. We set out to identify the effector (AvrSen1) recognized by the potato Sen1 resistance (R) gene product, resulting in pathotype 1(D1) resistance. AvrSen1 is expected to be present in pathotype 1(D1) isolates but absent in others.

A comparative genomics approach with fourteen isolates representing six different pathotypes was used to screen S. endobioticum specific secretome. This led to the discovery of an AvrSen1 candidate, because it was present in all pathotype 1(D1) isolates, but absent in others. Expression of this Avrsen1 candidate in potato plants showed a specific hypersensitive defence response in a Sen1 dependant manner. Identificati-on of the AvrSen1 gene represents a first step towards molecular pathotyping based on functional genetic markers.

Abstract Booklet

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Comparative genome analyses of Synchytrium endobioticum strains from Canadian and European ori-gin representing six different pathotypes reveal pathotype-specific genes and SNPs useful for molecular diagnosticsHai D.T. Nguyen (1), Kasia Dadej (1), Donna S. Smith (2), Wendy Findlay (1)

1. Agriculture and Agri-Food Canada, 960 Carling Avenue, Ottawa, Canada2. Canadian Food Inspection Agency, 93 Mount Edward Road, Charlottetown, Canada Isolates of Synchytrium endobioticum are grouped into pathotypes, which are defined by their phenotypic virulence or avirulence to a set of potato varieties. Labour intensive and time-consuming bioassays are used in the process of pathotyping isolates. So far, there are no quick molecular diagnostic methods able to discern between various pathotypes.

Using the available whole genome data of 14 S. endobioticum strains from Canadian and European origin, representing pathotypes 1(D1), 2(G1), 6(O1), 8(F1), 18(T1) and 38(Nevsehir), three major comparative ge-nomic analyses were performed. Genes in the core genome of the 14 isolates, as well as the genes unique to each pathotype, were determined. The phylogenetic relationships of genes in the core genome were assessed to determine whether certain genes would be good predictors of pathotypes based on grouping patterns. Genome wide SNP analysis was employed to find the union and intersection of pathotype specific sites.

Preliminary results show that certain genes and SNPs seem to be unique to a given pathotype although some may be strain-specific. This information will potentially be useful for designing a molecular test for faster pathotyping of isolates compared to the traditional bioassay methods.


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Oral presentations Session 3. Host resistance and breeding

Research on the formation of new virulent pathotypes of Synchytrium endobioticum (Schilb.) Perc., the causal agent of potato wart diseaseJarosław Przetakiewicz (1)

1. Plant Breeding and Acclimatization Institute - National Research Institute, Department of Plant Pathology, Radzików, Poland.

Three different cultivars of potato with varying degrees of resistance to pathotype 1(D1) were used in experience. Irga – extremely resistant, Erika – slightly susceptible and Evora – extremely susceptible to pathotyp1 1(D1) of S. endobioticum. Ten tubers of each cultivars were inoculated with winter sporangia of pathotype 1(D1), using the ring tests. After 2 months of incubation, big and small warts were visible on tu-bers of Evora and Erica, respectively. No warts were visible on tubers of cv. Irga. The warts were multiplied to check a virulence profile on EPPO differential set using the Glynne-Lemmerzahl method. In both cases the tests showed that the virulence profile was identical to the pathotype 1(D1). Winter sporangia obtained from warts of both cultivars were used to re-inoculate the same varieties as the next generation. After each passage the virulence of each generation were checked. After 2 or 3 passages, there was possible to select virulent pathotype on cv. Erika.

The galls obtained from Evora always corresponded to pathotype 1(D1). The virulence profile of isolate selected on cv. Erika was similar to pathotype 6(O1). This increased virulence could be the result of a shift in the heterogeneous 1(D1) population of S. endobioticum due to the selection.

Abstract Booklet

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Stacking of resistance loci to give full resistance to higher pathotypes of S. endobioticumCharlotte Prodhomme (1), Dong Zhang (1), Herman van Eck (1), Gert van Arkel (1), Marjan Bergervoet (1), Bart T. L. H. van de Vossenberg (1,2), Marga P. E. van Gent-Pelzer (1), Theo A.J. van der Lee (1), Jack H. Vos-sen (1), and Richard G.F Visser (1),

1. Wageningen University and Research, Droevendaalsesteeg 1, Plant Science Group, 6708 PB, Wageningen, the Netherlands 2. Dutch National Plant Protection Organization, National Reference Centre, Geertjesweg 15, 6706EA, Wa-geningen, The Netherlands

Synchytrium endobioticum, the soil borne parasitic Chytrid responsible for the potato wart disease, is under strict quarantine measures in Europe. Chemical control is not effective against this pathogen which pro-duces spores that can remain viable in the soil for more than 40 years. To contain the multiplication and spread of S. endobioticum, breeding for resistant varieties is required.

Breeding programs have been successful in breeding varieties resistant to pathotype 1(D1), but today, few varieties in the breeding germplasm are resistant to the higher pathotypes present in Europe. So far, efforts have been made to identify major resistance genes responsible for pathotype specific resistance, such as Sen1, or for a broader panel of pathotypes, such as Sen2 and Sen3.

In this talk, we will emphasize the importance of not using only these major genes in breeding programs and in the field but instead stacking major genes with loci having weaker effects on resistance. In a te-traploid population segregating for pathotypes 2(G1), 6(O1) and 18(T1) resistance, we mapped the Sen3 gene which brings a strong resistance to pathotypes 2(G1) and 6(O1) but a weaker resistance to the patho-type 18(T1) isolate used for the phenotyping.

Using a Bulked Segregant Analysis approach, we could identify another locus segregating in the popula-tion which, in combination with Sen3, improves the resistance to pathotype 18(T1). More efforts should be made to identify such minor effect loci and to use them in breeding programs. Indeed, the stacking of strong and weak effects resistance loci is necessary to bring a complete resistance, to contain the disease and to hamper the emergence of new pathotypes.


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Gene Sen2 – perspective for using in tetraploid potato breedingJarosław Plich (1), Jarosław Przetakiewicz (2), Beata Tatarowska (1), Bogdan Flis (1), Ewa Zimnoch-Guzowska (1)

1. Plant Breeding and Acclimatization Institute - National Research Institute, Department of Potato Genetics and Parental Lines, Młochów Research Center, 05-831 Młochów, Poland. 2. Plant Breeding and Acclimatization Institute - National Research Institute, Department of Plant Pathology, Radzików, Poland.

Synchytrium endobioticum causes potato wart disease imposing severe losses in potato production and, as a quarantine pathogen in many countries, it results in lost trade markets and land for potato cultivation. Chemical control of the pathogen in field conditions is not possible and therefore cultivation of resistant cultivars is the best solution to restrict its spreading. The main limitation of this method is scarcity of an attractive, market-tailored potato cultivars resistant to most important virulent pathotypes of S. endobioti-cum.

Recently the gene Sen2, which confers potato resistance to common (1(D1)) and at least seven virulent pa-thotypes of S. endobioticum (2(G1), 6(O1), 8(F1), 18(T1), 2(Ch1), 3(M1) and 39(P1)) was mapped to potato chromosome XI. The gene was identified in diploid potato clone and successfully introduced into tetraploid potato genepool via interploid crosses. Our researches confirmed that gene Sen2 provides the same very high level of resistance against the same broad spectrum of potato wart pathotypes in tetraploid potato clones as it was observed in diploid potato clones. A new DNA markers, suitable for marker assisted selec-tion (MAS) in tetraploid potato clones, were also developed. These results clearly show that gene Sen2 is highly promising and can be used in breeding of a new set of potato cultivars resistant to virulent pathoty-pes of S. endobioticum.

This research was funded by National Science Centre in Poland (grant 2013/11/B/NZ9/01959) and The Nati-onal Centre for Research and Development (grant Sen2Breed).

Abstract Booklet

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Development of Diagnostic Tools for Resistance Screening and Pathotype DifferentiationDennis Reckwell (1), Friederike Chilla (1), Annette Bartkiewicz (1), Diro Terefe-Ayana (1), Marian Meyer (1), Kerstin Flath (2), Marcus Linde (1), Thomas Debener (1)

1. Leibniz Universität Hannover, Institute of Plant Genetics, Department of Molecular Plant Breeding, Han-nover, Germany2. JKI, Federal Research Centre for Cultivated Plants, Institute of Plant Protection of Field Crops and Gras-sland, Kleinmachnow, Germany

Resistance against pathotype 1(D1) is included in potato breeding for more than hundred years. The locus for resistance (Sen1) is located on chromosome XI, but the nature of the responsible genes is unknown. To characterise resistance to pathotype 18(T1) a segregating monoparental dihaploid potato population was established to locate the respective locus via resistance screening of recombinant genotypes. The resistan-ce genes were located close to the Sen1 locus on an interval of 777 kbp on the reference genome. Additi-onal genotypes were analysed which offer the potential to locate the genes in an interval of less than 500 kbp. Furthermore, the genome of a highly resistant genotype was sequenced to detect candidate resistance genes.

More than 40 pathotypes of Synchytrium endobioticum are known. Pathotypes 1(D1), 2(G1), 6(O1) and 18(T1) are the most important ones in Germany. Genome and transcriptome sequences were used to de-velop diagnostic markers to distinguish between pathotypes, including SCAR markers and microsatellites. Addition of these markers to biotests is capable to speed up the process of pathogen identification.

Effector proteins have a crucial role to overcome the plant defence system. Transcriptome sequences of potato warts were analysed to identify effector candidate genes. Four genes supported the spread of To-bacco Rattle Virus in transformed tobacco leaves, indicating the potential to decrease plant defence.


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Oral presentations Keynote lecture

Dissecting the biology and pathology of the late blight pathogen Phytophthora infestansFrancine Govers (1)

1. Laboratory of Phytopathology, Wageningen University, Droevendaalsesteeg 1, 6708 PB Wageningen. The Netherlands. [email protected]

The plant pathogen Phytophthora infestans is the causal agent of late blight, a devastating disease in potato worldwide. Its first appearance in Europe in the mid-nineteenth century led to the Irish potato famine and to the emergence of plant pathology as a scientific discipline. P. infestans belongs to the oomycetes, a diverse group of organisms comprising pathogenic species infecting plants, animals or microbes, as well as saprophytic species. It has a hemibiotrophic life style and exploits a variable repertoire of effector proteins for manipulating plant defence and facilitating colonization.

Comparative genomics revealed features illuminating the success of Phytophthora species as plant patho-gens, such as a massive expansion of genes encoding secreted proteins and peculiar gene innovations resul-ting in proteins with oomycete-specific domain combinations. For successful infection Phytophthora secre-tes a variety of proteins including a large number of effectors that share the host-cell targeting motif RXLR. Inside host cells these RXLR effectors promote virulence by manipulating the cell machinery via interaction with host targets thereby suppressing host defence. However, in plants carrying matching resistance genes RXLR effectors trigger defence and thus act as avirulence factors.

In this presentation I will summarize our current knowledge on the biology and pathology of P. infestans. I will also highlight some aspects of RXLR effectors and the discovery of oomycete specific features that might be instrumental for designing a next generation crop protection agents.

Abstract Booklet

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Poster presentations


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Wart Potato Synchytrium endobioticum (Schilb.) Perc. impact on plant host.Avreliya G. Zelya (1), Mykhailo P. Solomiychuk (1), George I. Vorobets (2)

1. Ukrainian Scientific- Research Plant Quarantine Station Institute of Plant Protection National Acadeny of Agrarian Sciences, Ukraine2. Chernivtsi National University, Ukraine

Wart potato is one of the dangerous diseases of potato in Ukraine. The researchers of Ukrainian scienti-fic-research plant quarantine station during 2009-2019 determined changes in biosynthesis of crude pro-tein. The peculiar protein spectrum was received by the way of isoelectric focusing in polycrylamide gel in gradulent pH 3,5-10,0. 59 components were identified in protein spectrum in non-infected by wart potato variety Poliska rozheva. The following results of protein components were recorded on infected potato: 28- by common pathotype 1(D1), 30 by 11 (Mizhhirya ), 42 by 13 (Rachiv), 51 by 18 (Yasinya ), 53 by 22 (Bystry-tsya ). The extracted protein fractions are differed by quantity and their isoelectric points location.

The amino acid content of synthesiszing protein change was recorded during the infecting by wart disease agent. The amount of non-changed amino acide was consisted of 42,3 %, defeated by wart potato ,-35,5% in non-defeated potato variety. The increasing of following components was recorded on 21-st day: gluta-mic acid from 10,6 to 13,1 %, cysteine and cystine from 1.0 to 2,1%, tyrosine from 5,3 to 6,8%, The decre-ase of following components content was recorded: lysine from 8.3 to 7.9%, aspartic acid from 8,0 to 7,9%, leucine and isoleucine from 8.6 to 6.9%.

There were determined difference during the activity analysis of oxidation-reduction enzyme (peroxidase, catalase). They considered “avarium enzymes”. They are immediately react on any stress on plan: tempea-rature drop, disease casuative agents impact . It was determined the peroxidase activity 0,62 micromole/min on infected potato sprout and 0,98 micromole/min on 21 st day after infecting. The analysis of biochemical and biophysical indexes of potato plant’s infecting processes is continued.

Abstract Booklet

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Choice of potato varieties resistance to Synchytrium endobioticum.George V. Zelya (1), Tetyana M. Oliynyk (2), Volodymyr M. Gunchak (1)

1. Ukrainian Scientific Research Plant Quarantine Station Institute of Plant Protection National Academy of Agrarian Sciences, Chernivtsi, Ukraine2. Institute for potato research, National Academy of Agrarian Sciences, Ukraine

Ukrainian Scientific- Research Plant Quarantine Station Institute of Plant Protection National Academy of Agrarian Sciences provides researches by evaluation and selecting new potato resistant varieties to wart of Ukrainian and foreign breeding for more than 80 years. The resistance test have been conducted by the old methods for evaluation and selection. The testing is conducted as per standard EPPO PM 7/28 after the providing training by Dutch National Plant Protection Organization in 2013 (Wageningen, the Netherlands), by Spieckermann and Glynne-Limmerzahl methods in laboratory test.

Field trials on naturally infected soils were conducted in the area of Chernivtsi. Zakarpattia (11- Maydan, Mizhhirya district, 13- Rakhiv, 11- Yasinia) and Ivano- Frankivsk region (22- Bystrets, Verkhovyna district). 1677 samples of potato breeding material were received from 6 scientific- research institutes and breeding centres of Ukraine.

3700 samples of potato breeding material were extracted from 4677 samples during 2011-2018. They were resistant to common pathotype (D1) S. endobioticum in previouis testing and 130 samples were approved in state testing. Among all samples tested, 75 (49%) were identified as resistant to 11- Mizhhirya pathoty-pe,5 (33,9%) samples were resistant to 13 Rakhiv pathotype, 55 (35,9%) samples were resistant to 8-Ya-sinya, 74 (48,3%) were resistant to 22-Bystrets pathotype.

The tests revealed that the national breeding program targeted on resistance against S. endobioticum, com-mon pathotype (D1), 11-Mizhhirya, 22-Bystrets, was the most effective, but it was least effective against 13, 18 pathotypes of wart potato. The already registered potato varieties Bozhedar, Solokha, Khortytsa, Bazys, Santarka which were found to have a multiple resistance to all pathotypes were recommended for use in the breeding process as sources of resistance and also for the eradication programs in sources of wart pota-to in Ukraine.


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Potato varietal resistance cannot always reduce the impacts caused by the introduction of Synchytrium endobioticum (Schilb.) Perc. into a new area - the case of GreeceIrene Vloutoglou (1); Konstantinos B. Simoglou (2); Helias Eleftheriadis (2); Christos Kritikos (1); Dimitrios Tsirogiannis (1); Ioanna Sarigkoli (2); Nikolaos Nikolaidis (2); Christos Arampatzis (3) and Dimitra Gkilpathi (4)

1. Benaki Phytopathological Institute, Department of Phytopathology, Kifissia, Attica, Greece.2. Region of Eastern Macedonia & Thrace, Regional Unit of Drama, Department of Quality and Phytosani-tary Control, Drama, Greece.3. Hellenic Ministry of Rural Development and Food, Department of Phytosanitary Control, Athens, Greece.4. Hellenic Ministry of Rural Development and Food, Department of Biocide Products, Athens, Greece

Potato wart caused by the quarantine soil-borne fungus Synchytrium endobioticum (Schilb.) Perc. was first reported from Greece in one of the main ware potato-producing areas (Kato Nevrokopi, Northeastern Gree-ce), during the 2011 official surveys. The pathotype present in the contaminated fields was identified as pathotype 18(T1). Since then, strict phytosanitary measures, including the demarcation of a safety zone (c. 200 ha) around the contaminated fields to prevent further spread of the pathogen, have been implemen-ted in the area, in compliance with the EU and National Plant Health legislation. In addition, official testing under field and controlled environmental conditions was performed to identify potato varieties resistant to pathotype 18(T1) to be used in the safety zone. Fifty European potato varieties of various end-uses (table varieties, processing industry/starch varieties), nine of which were reported by other EU Member States and/or potato breeding companies as resistant to pathotype 18(T1), were screened for resistance in a natu-rally contaminated field over four consecutive years.

Varieties showing field resistance were further evaluated in pot tests using artificially contaminated sub-strate. Results showed that, only two starch varieties constantly exhibited field resistance to pathotype 18(T1) and furthermore provided adequate protection against secondary infections by the pathogen, in line with Council Directive 69/464/EEC. Nevertheless, in the absence of potato starch manufacturers in the country, the farmers are very reluctant to grow these two varieties in the safety zone. As a result, the impacts caused by the introduction of S. endobioticum in Greece remain very high.for use in the breeding process as sources of resistance and also for the eradication programs in sources of wart potato in Ukraine.

Abstract Booklet

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The occurence and testing method of Synchytrium endobiticum in the Czech RepublicIveta Svobodová (1); Vladimír Gaar (2)

1. Central Institute of Supervising and Testing in Agriculture, Division of Diagnostics, Department Diagnostic Laboratory Olomouc, Czech Republic2. Central Institute of Supervising and Testing in Agriculture, Division of Diagnostics, Department Diagnostic Laboratory Prague, Czech Republic

Synchytrium endobioticum, the agent of potato wart disease, is an A2 pest for the EPPO region. The oc-curence of potato wart disease Synchytrium endobioticum in the territory of the present Czech Republic has been known since 1915. Many pathotypes (races) of S. endobioticum are known to occur, in the Czech Republic 19 pathotypes of Synchytrium endobioticum have been identified. However, the last finding of Synchytrium endobioticum in the Czech Republic on the field was in 2010.


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Occurrence of potato wart in GeorgiaSopio I. Ghoghoberidze (1); Tsisana M. Tsetskhladze (1); Ketino T. Sikharulidze (1)

1. Institute of Phytopathology and Biodiversity, Batumi Shota Rustaveli State University, the Autonomous Republic of Adjara, 6010, Batumi, Georgia

Potato wart is an important disease in Georgia causing yield loss of potato crops along with other diseases. The disease was first found in the private plots of Khulo municipality in 2009. Nowadays more infested foci were detected in two separately located western (Khulo region) and northern (Zemo Svaneti region, Mes-tia district) areas. Among them, 17 infested foci located at an altitude of about 183 meters above sea level were found in Khulo region, and 2 infested foci located at an altitude of about 2100 meters above sea level were detected in Mestia district, where the abiotic factors are respectively different.

To identify the pathotypes, the wart material collected from the private plot of Didajara village, Khulo dis-trict (the western part of Georgia) was sent to the Netherlands, and the laboratory, pot and field tests were also conducted in the Institute of Phytopathology and Biodiversity according to the EPPO standards. The results obtained from both laboratories showed the presence of the pathotype 38 (Nevsehir) in the wes-tern part of Georgia. By the end of 2018, the wart tissue, collected in the northen part of Georgia (Mestia district) was again sent to the Netherlands, and as well as the pot and lab (using Glynne–Lemmerzahl me-thod) tests for the same wart tissue is being conducted in Georgia.

Abstract Booklet

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Preliminary Screening of Potato Cultivars for Resistance to Potato Wart in GeorgiaZoya V. Sikharulidze (1); Sopio I. Ghoghoberidze (1); Tsisana M. Tsetskhladze (1)

1. Institute of Phytopathology and Biodiversity, Batumi Shota Rustaveli State University, 6010, Batumi, Ge-orgia

The average yield of potato in Georgia is low due to many reasons. Quarantine disease potato wart, caused by fungal pathogen Synchytrium endobioticum, is an important constraint. The pathogen is widely distribu-ted in the world including Georgia where it was first found in 2009 in Khulo district. The most effective way to control the disease spread is the cultivation of varieties resistant to the pathotypes present in the infe-sted fields.

The introduced potato cultivars were assessed for the reaction to potato wart using pot and field tests. When tubers formed, wart symptoms were evaluated in the pot test by the Spieckermann scale and in the fields conducted in Khulo villages (Skvana, Uchkho) according to the scale specified in the EPPO Diagnostic Protocol.

The majority of cultivars: Marfona, Pekaro, Panamera, Saturna, Spunta, Annalena, Nandina, Glorietta, Bernadette, Estela, Skrab, Uladar, Briz, Lileya, Europrima, Sylvana, Finca, Laura, Figaro, Impala, Jelly, Alwara, Artemis, Milva and Sante showed susceptible reaction and cultivars Arnova, Sofia, Arizona, Fabula, Caruso, Omega, Arinda were resistant in the pot test to pathogen isolate originating from Khulo, Didadjara village. The main part of cultivars showed the susceptibility in both field trials. Cultivars Catania, Leandra, Cardinia and cultivars Sylvana, Agria, Marabel were resistant in the Uchkho and Skvana trials, respectively.

As a result of preliminary screening, some potato cultivars showed the different reactions to potato wart in pot and field tests; therefore, further testing of these cultivars is needed to obtain more reliable results.


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The linear mitochondrial genome of the quarantine pest Synchytrium endobioticum; insights in the evo-lutionary history of an obligate biotrophBart T. L. H. van de Vossenberg (1,2), Balázs Brankovics (1), Hai D. T. Nguyen (3), Marga P. E. van Gent-Pel-zer (1), Donna Smith (4), Kasia Dadej (3), Jarosław Przetakiewicz (5), Jan F. Kreuze (6), Margriet Boerma (7), Gerard C. M. van Leeuwen (2), C. André Lévesque (3) and Theo A. J. van der Lee (2)

1. Wageningen UR, Biointeractions and Plant Health & Plant Breeding, Wageningen, The Netherlands2. Dutch National Plant Protection Organization, Geertjesweg 15, 6706EA Wageningen, The Netherlands3. Agriculture and Agri-Food Canada, 960 Carling Avenue, Ottawa, Canada4. Canadian Food Inspection Agency, 93 Mount Edward Road, Charlottetown, Canada5. Plant Breeding and Acclimatization Institute, National Research Institute, Warsaw, Poland6. International Potato Centre, Avenida La Molina, 1895 Lima, Peru7. Hilbrands Laboratorium BV, Kampsweg 27, 9418 PD Wijster, Wijster, The Netherlands

Chytridiomycota (chytrids) inhabit terrestrial and aquatic environments, and represent a basal lineage in true fungi. Most of the described chytridiomycota are free living saprophytes, but several species are noto-rious pathogens for plants or amphibians. Synchytrium endobioticum is an obligate biotroph chytrid causing potato wart disease. Quarantine measures have been implemented worldwide to control the disease and to prevent its spread.

To determine taxonomical relationships, and to gain insights into the evolutionary history of this plant pathogen we assembled and annotated the mitochondrial genome of S. endobioticum and generated mitochondrial genomes for five additional chytrid species. The mitochondrial genome of S. endobioticum is a linear 72,865 bp molecule with terminal inverted repeats that encodes 14 mitochondrial genes typically found in fungi.

Polymorphisms in 30 S. endobioticum isolates shows clustering in four main mitochondrial lineages, and from our data we conclude that the pest was introduced at least three times in Europe. Strains of patho-type 2(G1) and 6(O1) were represented in two mitochondrial lineages, showing that these pathotypes emerged independently. Variations within a strain for polymorphic sites were observed and seem to be consistent in different mitochondrial lineages suggesting that S. endobioticum strains are communities of different genotypes with conserved composition.

Abstract Booklet

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Sensitive detection of Synchytrium endobioticum in soil samples using zonal centrifugeKurt Heungens (1)

1. ILVO, Plant Unit, Merelbeke, Belgium

Detection of Synchytrium endobioticum in soil samples is relevant in surveys of potentially infected fields. Several methods have been described, including extraction of the winter spores from soil using a zonal cen-trifuge, filtering, DNA extraction and qPCR as described by Wander et al. (2007) and van Gent-Peltzer et al. (2010). This method is very sensitive, as it combines high efficiency extraction from 100 g soil samples with sensitive qPCR assays. We validated this method and included optimization steps relating to incubation time in the separation liquid, qPCR assay, and use of cloned DNA target for the standard curve.

Using these optimized methods we conducted recovery experiments from soil samples spiked with de-creasing numbers of resting spores. Even down to 2 spores per 100 g of soil, recovery was successful in all replicates. Especially at lower concentrations, the recovery rate was above 70%.


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Abstract Booklet

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Parallel sessions


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The workshop brings together biologists, researchers and policy makers working on all aspects of Synchytri-um endobioticum and the interaction with its host potato. Apart from knowledge exchange through pre-sentations, a tour of the NPPO facilities is organized, and research needs and experiences are discussed in six subgroups of approximately 10 persons. During registration you were assigned to a discussion group. On Thursday 27 June we will be active with three sessions.

A. A tour to the facilities of the NVWA demonstrating equipment and procedures with emphasis on spore extraction from soil suspensions, molecular methods and the wart disease compost collecti on.B. Workshop part 1: Discussion sessions on questions are grouped under three topics (Disease oc currence and management, Pathogen biology, and Host resistance/Breeding), which will serve as starting point for the discussion on needs in potato wart research and management. Discussion groups are invited to comment on the questions below which serve to start the discussion. Subgroups can put emphasis on a specific topic, or may address other relevant topics. C. Workshop part 2: groups are asked to summarize the items discussed in a PowerPoint presentation. Groups will be equipped with laptop computers and a template PowerPoint presentation.

Each group selects a person to plenary present (max. 4 slides, 10 min) the outcomes of the subgroup.

Activities for the different subgroups during the parallel workshop and tour sessions11:00 – 12:30 13:30 – 14:45 14:45 – 16:00

Group 1 Tour Workshop 1 Workshop 2Group 2 Tour Workshop 1 Workshop 2Group 3 Workshop 1 Tour Workshop 2Group 4 Workshop 1 Tour Workshop 2Group 5 Workshop 1 Workshop 2 TourGroup 6 Workshop 1 Workshop 2 Tour

Abstract Booklet

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Questions for the discussionDiscussion groups are invited to comment on the questions below which serve to start the discussion. Sub-groups can put emphasis on a specific topic, or may address other relevant topics.

1. Disease occurrence and managementDo you think problems with potato wart disease are decreasing/increasing or remain similar in your coun-try?Do you think all infections are noted?Do you have indications of emerging/new pathotypes that “break” resistance?Do you think potato varieties offering partial resistance could be used in management strategies?Do you have indications of specific factors that might influence epidemics?Do you have indications of long lasting infestation in particular regions. If so do you have indications of par-ticular conditions that support the presence of S. endobioticum in these regions?Do you have indications alternative hosts may play a role in the epidemics?

2. Pathogen biologyHave you implemented pathotyping methods, do you pathotype all isolates, and which differential set and bioassay is used?Do you think the currently used protocol for bioassays are sufficiently harmonized?Do you think that there is a need of a standardized set of pathogen inoculum to determine potato resistan-ce for different pathotypes? Do you think the current used differential set is sufficient to reliably determine the pathotype identity?Would molecular characterization of isolates be something that would be of use?If yes”, for which characteristics would this be possible/which would be of interest to you?

3. Host resistanceDo you think your breeding material is sufficiently characterized for wart resistance?What are the main bottlenecks for testing wart resistance?Do you know of any mischaracterizations in of genotypes in public databases? Is the use of molecular markers required to obtain/characterize resistant material? Is the stacking of R genes required?How important is wart resistance for new cultivars?

Other topic related to potato wart disease


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Abstract Booklet

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Participants


- 34 -

Country Name Position Institute/Organization E-mailadresAustria Sonja Axmann Senior Expert Austrian Agency for Health and Food

Safety (AGES)

[email protected]

Bangladesh Syed Munerul

Hoque

Quarantine Patho-

logist

National Plant Quarantine Authority,

Department of Agricultural Extension,

Bangladesh

[email protected]

Belgium Kurt Heungens Senior researcher

mycology

Research Institute for Agriculture, Fis-

heries and Food (ILVO)Bulgaria Ani Becheva Head of depart-

ment

Bulgarian Food Safety Agency [email protected]

Lidia Dimitrova Chief expert Bulgarian Food Safety Agency [email protected] Hai Nguyen Research Scientist Agriculture and Agri-Food Canada (AAFC) hai.nguyen.1984@gmail.

comCzech Re-

public

Iveta Svobodová Mycologist Central institute for supervising and tes-

ting in agriculture

iveta.svobodova@ukzuz.

czDenmark Mette Vestergård Researcher Aarhus University [email protected]

Ea Riis Sundmark Breeder Danespo [email protected] Mikko Lehtonen Project Researcher Finnish Food Authority mikko.lehtonen@ruoka-

virasto.fiFrance Nico Horn Director-General European and Mediterranean Plant Pro-

tection Organization (EPPO)

[email protected]

Veronique Wilson French Agency for Food, Environmental

and Occupational Health & Safety (ANSES)

veronique.wilson@anses.

frGeorgia Zoya Sikharulidze Head of Depart-

ment, senior

scientist

Institute of Phytopathology and biodiver-

sity, Batumi Shota Rustaveli State Univer-

sity

[email protected]

Germany Dorothee Kaem-

merer

Head of the work

group ‘quarantine

measures pota-

toes’

Bayerische Landesanstalt für Landwirt-

schaft

Dorothee.Kaemmerer@lfl.

bayern.de

Peter Büttner labaratory head

mycology

Bayerische Landesanstalt für Landwirt-

schaft

Peter.Buettner@LfL.

bayern.deKarsten Buhr Phytopathologist Böhm-Nordkartoffel Agrarproduktion

GmbH & Co. OHG

mgelszat@boehm-kartof-

fel.deRafal Zgadzaj Project Coordi-

nator

Böhm-Nordkartoffel Agrarproduktion

GmbH & Co. OHGBettina Beerbaum desk officer Federal Ministry of Food and Agriculture

(BMEL)

bettina.beerbaum@bmel.

bund.deFriederike Chilla Wissenschaftliche

Mitarbeiterin

Julius Kühn-Institut (JKI) friederike.chilla@juli-

us-kuehn.de

Abstract Booklet

- 35 -

Country Name Position Institute/Organization E-mailadresGermany

(continued)

Kerstin Flath Senior Scientist Julius Kühn-Institut (JKI) kerstin.flath@julius-kue-

hn.deStephan Koenig Scientist - Labora-

tory Diagnostics

Julius Kühn-Institut (JKI) stephan.koenig@juli-

us-kuehn.deKarl-Heinz Pastrik Special consultant Landwirtschaftskammer Niedersachsen karl-heinz.pastruk@

lwk-niedersachsen.deDennis Reckwell Postdoc Leibniz Universität Hannover reckwell@genetik.

uni-hannover.deMarcus Linde Senior Scientist Leibniz Universität Hannover [email protected]

ver.deThomas Debener Professor Leibniz University Hannover debener@genetik.

uni-hannover.deJoachim Weinert Sachgebietsleiter

Mykologie

Pflanzenschutzamt joachim.weinert@lwk-nie-

dersachsen.deGreece Irene Vloutoglou Head of the Labo-

ratory of Mycology

Benaki Phytopathological Institute [email protected]

Ireland Andy Bourke Laboratory Analyst Department of Agriculture, Food and the

Marine, Ireland

Andy.Bourke@agriculture.

gov.ieLatvia Kristine Paruma Expert mycologist State Plant Protection Service of Latvia kristine.paruma@vaad.

gov.lvPoland Jarosław Plich Assistant professor Plant Breeding and Acclimatization Insti-

tute - National Research Institute (IHAR)

[email protected]

Jaroslaw Przeta-

kiewicz

Senior Scientist Plant Breeding and Acclimatization Insti-

tute - National Research Institute (IHAR)

[email protected].

plRussia Yulia Tsvetkova agronomist All-Russian Plant Quarantine Center (

FGBU “VNIIKR”)

[email protected]

Sweden Sofia Windstam Plant Health

Officer

Swedish Board of Agriculture sofia.windstam@jord-

bruksverket.seJohanna Boberg Pest risk analyst Swedish University of Agricultural Scien-

ces

[email protected]

Switzerland Stéphanie Schürch scientific collabo-

rator

Agroscope stephanie.schuerch@

agroscope.admin.chthe Nether-

lands

Alina Cristina

Cucerdean

Plant Breeder Agrico Research [email protected]

Gerrit Jan Brom-

mer

Plant Breeder Agrico Research [email protected]

Hilde Room Phytopathology

Technologist

Averis Seeds B.V. [email protected]

Jasper Tammes Breeding Specialist Averis Seeds B.V. jasper.tammes@avebe.

com


- 36 -

Country Name Position Institute/Organization E-mailadresthe Nether-

lands

(continued)

Nico Rookmaker Breeder Averis Seeds B.V. nico.rookmaker@avebe.

comBart van de Vos-

senberg

Molecular Biolo-

gist

Dutch National Plant Protection Organiza-

tion (NPPO-NL, part of NVWA)

b.t.l.h.vandevossenberg@

nvwa.nlDirk Jan van der

Gaag



[email protected]

Gabriela M. Ferrari

Cálcena

Supervision plant

health



g.m.ferraricalcena@nvwa.

nlGerard van Leeu-

wen

senior phytopa-

thologist



g.c.m.vanleeuwen@nvwa.

nlJos van Meggelen Senior Advisor

International Coo-

peration



[email protected]

Mieke Reyniers Head National

Reference Centre



[email protected]

Ineke van Holst analyst molecular

biology

HLB B.V. [email protected]

margriet Boerma projectleider HLB B.V. [email protected] Boomsma Program leader

Plant Pathology &

Cell Biology

HZPC Research B.V. doretta.boomsma@hzpc.

nl

Paul Schaap Solynta [email protected] Prod-

homme

Ph.D. Student Wageningen University & Research (WUR) charlotte.prodhomme@

wur.nlDong Zhang Ph.D. Student Wageningen University & Research (WUR) [email protected] Vossen Senior Scientist Wageningen University & Research (WUR) [email protected] van

Gent-Pelzer

Research techni-

cian

Wageningen University & Research (WUR) [email protected]

Peter Bonants Teamleader Wageningen University & Research (WUR) [email protected] GF Visser Professor and

Head of Depart-

ment

Wageningen University & Research (WUR) [email protected]

Theo van der Lee Senior Scientist Wageningen University & Research (WUR) [email protected] Hale Gunacti Senior Scientist Biological Control Research Institue [email protected] Avreliya Zelya Chief of Labora-

tory

Institute of Plant Protection - National

Academy of Agrarian Sciences of Ukraine

[email protected]

George zelya Scientific resear-

cher

Institute of Plant Protection - National

Academy of Agrarian Sciences of Ukraine

georgetrex7777777@

gmail.com

Abstract Booklet

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Country Name Position Institute/Organization E-mailadresUnited

Kingdom

Alexandra Sch-

lenzig

Senior Plant Pa-

thologist

Science and Advice for Scottish Agricultu-

re (SASA)

alexandra.schlenzig@sasa.

gov.scotMatthew Everatt Plant Health Policy Department for Environment, Food and

Rural Affairs

Matthew.Everatt@defra.

gov.ukUnited

States of

America

John Bienapfl Molecular Biolo-

gist

USDA APHIS PPQ S&T [email protected]


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Abstract Booklet

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Biorisk management at the National Reference Centre Plant Health

Hans Derks (Biorisk officer) National Plant Protection Organization, National Reference Centre, 6700 HC, Wageningen, the Netherlands,Email: [email protected]

Introduction.

To enable safe handling of plant pests and diseases, the National Reference Centre (NRC) of the National Plant Protection Organization (NPPO) has quarantine facilities and procedures that minimize the risk of escape of harmful organisms. The facilities consist of both laboratories and greenhouses of different containment levels to allow working with organisms with different risk profiles. Together these physical containment and procedures form the base of Biorisk management, including both biosafety and biosecurity.

- Organisation of biosafety and biosecurity- Supervision by Biorisk officer- Training of staff and external workers- Authorisation for access to containment areas- Rules of conduct- Maintenance of essential biosafety equipment

Procedures

The safe working procedures, embedded in the Biorisk Management System are an integral part of the ISO 17025 accreditation of the NRC. The desired practices are described in procedures and related documents and include the following topics:

Biorisk Plant

Class

Risk for

escape

Spread by / Survival Infrastructure

Characteristics

BPL-I,

BPG-I

Low risk

no spread: - closed transport and storage of harmful organisms

- decontamination of waste water before disposal - closed windows (insect screens for greenhouses) - no permanent administration working place

- authorised access - protective clothing –> white lab coat - disinfection laboratory - disinfection tools - disinfection hands on leave - treatment biological waste

BPL-II,

BPG-II

Medium

risk

spread by: - contact - water - soil - survival forms like cysts

- biosafety cabinet (laboratories) - windows air-tight (e.g. sealed) - functional arranged laboratories/greenhouses

- see level I - protective clothing –> blue-labelled lab coat plus gloves - paper work separated from lab work

BPL-III,

BPG-III

High risk

spread by: - air - vegetative reproduction - spores

- negative pressure - Hepa filtration of exhausted air - double door with vestibules in between

- see level II - protective clothing –> red-labelled lab coat plus gloves and shoe covers - paper not released unless treated as biological waste

Physical containment

Table 1 Characteristics of Biorisk Plant Classes

Figure 1 Facilities minimising the risk of escape Figure 2 Facilities ensure containment of laboratory BPL-III level

Biorisk Plant ClassesBased on the biology of the harmful organisms the facilities allow working at appropriate safety levels, based on different Biorisk Plant Classes for laboratories (BPL) and greenhouses (BPG). Tables 1 show the characteristics of these different classes: I, II and III, from low to high, respectively.

- Release of equipment for external service- Transport and storage of biohazard materials and waste- Emergency procedures, including:

- informing the NPPO, risk analysis- decontamination and corrective measures

- Change of containment level

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