Materials Innovation for Sustainable Agriculture Center of Excellence:
A platform for scientists, growers, regulatory and industry partners to meet and exchange the knowledge and skills
available to combat agricultural diseases for long-term crop protection
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Welcome note from the Chairs Dear Friends,
It is our great pleasure to welcome you to MISA 2019 – the fourth symposium for materials innovation for sustainable agriculture! The demand for global food security and nutrition resulting from steady population growth continues to rise. It is critical for current agriculture practices to adapt and propose new solutions to meet this demand. In particular, food crop productivity and quality must be improved in the near, short and long terms. For instance, there is a pressing need to identify new approaches to improve the resiliency of crops against new conditions imposed by global climate change. The emergence of new threats must be met with the development of innovative and “smart” technologies. In recent years, new initiatives involving a systems approach with cross-disciplinary approaches in close collaboration with stakeholders have introduced a shift to modern practices in agriculture research.
The UCF MISA center was founded in 2016 in an effort to foster collaborations towards materials innovations for sustainable agriculture – in particular (but not limited) to tackle citrus greening and canker in Florida. Since then, our team has designed and developed several industrially-viable pesticide and fertilizer formulations that have shown good efficacy in the field. Our aim is to impact overall crop productivity and nutrition, while decreasing the environmental risk factors. These new products would alleviate the excessive use of conventional crop protection chemicals and subsequent risks of pathogen resistance development. Moreover, versatile designs suggest new avenues for multi-functional crop-protectant formulations, which are also expected to gain popularity in coming years.
Our center is committed to:
Promoting interdisciplinary research culture for stimulating innovations in materials research Developing new technologies for protecting sustainability of agriculture industry challenged
by emerging threats of plant diseases and unpredictable weather patterns Establishing industry-academia collaborations with a strong emphasis on educational and
technological knowledge sharing Establishing interdisciplinary education and extension programs to support growers in
making informed decision Promoting sustainable outcomes by adopting innovative and synergistic approaches through
engagement of stakeholders, scientists, engineers, industry partners and regulatory agencies
Since our last meeting, our activities at the MISA center have expanded in several countries including India and Bangladesh. Our material designs now encompass pesticides and fertilizers. We have expanded our partnerships with industry, government and non-profit organizations to explore new initiatives for a positive impact on food production.
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In this year’s program, we highlight some of this work and foster discussions to accelerate research and discoveries at the forefront of the field. Our presentations, in the next two days, will cover the topics of Fertilizers and Soil Interactions, Emerging Tools for Sustainable Agriculture and will introduce a new application sector, Industrial Hemp. The goal of the symposium is to provide a platform that will facilitate interactions between academics, industry partners, growers and government agencies by engaging participants from diverse expertise. Our common goal is to solve pressing agricultural problems by taking advantage of new technologies. We truly appreciate your support for the MISA Center and hope to continue this collaboration for years to come! We wish you a fruitful conference!
Dr. Swadeshmukul Santra Dr. Laurene Tetard (MISA Director and Program Chair) (MISA Program Co-Chair)
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Organizing Committee
Swadeshmukul Santra, Ph.D. (Chair) Professor
NanoScience Technology Center (NSTC), Materials Science & Engineering, Department of
Chemistry, Burnet School of Biomedical Sciences
Email: [email protected]
Phone: 407-882-2848
Laurene Tetard, Ph.D. (Co-Chair) Associate Professor
NanoScience Technology Center (NSTC) and Physics Department
Email: [email protected]
Phone: 407-882-0128
Our team of volunteers Kamry Samuel
Briana Lee
Dr. Hajeewaka Mendis
Dr. Maria Campos
Stephen Smith
Danya Belnour
Ava Milani
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Acknowledgements We gratefully acknowledge
the United States Department of Agriculture National Institute of Food and Agriculture (USDA-NIFA)
for recognizing the Materials Innovation for Sustainable Agriculture (MISA) Center of
Excellence, and the University of Central Florida (UCF) for providing financial support
for our MISA activities and Symposium.
We acknowledge our funding from the Citrus Research & Development Foundation (CRDF)
and our Industry Sponsors for their support of research projects.
University of Central Florida NanoScience Technology Center,
Office of Research and Commercialization
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Acknowledgements We gratefully acknowledge our MISA donors for their continued support in view of training the next generation of workforce of Materials Innovation in
Sustainable Agriculture
and
Estes Citrus Inc. Premier Citrus LLC
YARA North America
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MISA 2019 Program
October 24th, 2019 Registration opens at 8:00 am
8:00 – 9:00 am Breakfast
9:00 – 10:00 am Opening remarks
9:00 – 9:05 am Welcome address by Dr. S. Santra, Professor NSTC and Chemistry
9:05 – 9:10 am Welcome address by Dr. L. Zhai, NSTC Director
9:10 – 9:15 am Welcome address by Dr. E. Klonoff, Vice President for Research and Dean of Graduate Studies
9:15 – 10:00 am Keynote
9:15 – 10:00 am Dr. Gregory Lowry, Carnegie Mellon University Walter J. Blenko, Sr. Professor of Civil & Environmental Engineering Deputy Director, Center for Environmental Implications of Nanotechnology Title: Towards Highly Efficient Uptake and Translocation of Foliar Applied Environmentally Responsive Nano-agrochemicals
10:00 am – 12:15 pm Scientific program Fertilizers and Soil Interactions Moderator: Dr. Swaminathan Rajaraman and Dr. Davie Kadyampakeni
10:00 – 10:25 am Dr. Bill Easterwood, YARA North America Title: Balanced Nutrition for Citrus Tree Health and Productivity
10:25 – 10:50 am Dr. Davie Kadyampakeni, University of Florida Title: Nitrogen, Calcium and Magnesium management for Improving Performance of HLB-affected Orange Trees (Citrus sinensis)
10:50 – 11:15 am Dr. Fernando Alferez, University of Florida Title: Modulating lipid and auxin signaling through nutrition may influence fruit retention and root health in citrus.
11:15 – 11:40 am Dr. Tripti Vashisth, University of Florida Title: Soil-applied micronutrients can improve productivity of Huanglongbing-affected sweet orange
11:40 am – 12:05 pm Dr. Rhuanito S. Ferrarezi, University of Florida Title: Can higher planting density and enhanced fertilization increase fruit yield of HLB-infected grapefruit?
12:05 – 12:15 pm Discussion
12:15 – 1:45 pm Lunch/Networking
1:45 – 4:00 pm Scientific program Emerging Tools for Sustainable Agriculture Moderator: Dr. Mathews Paret, Dr. Jayan Thomas
1:45 – 2:10 pm Dr. Evan Johnson, University of Florida Title: Zinkicide improves health and yield of HLB-affected trees: limited by dilution and residual efficacy?
2:10 – 2:35 pm Dr. Natalia Peres, University of Florida Title: Use of UV light for control of strawberry powdery mildew
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2:35 – 3:00 pm Daniela Diaz Hernandez, University of Central Florida Title: Novel Silica-functionalized Quaternary Ammonium Nanocoating for Water Disinfection and Harmful Algal Bloom (HABs) Mitigation
3:00 – 3:25 pm Dr. Melanie Kalischuk, University of Florida Title: Early detection of crop health with multispectral sensing
3:25 – 3:50 pm Dr. Jayan Thomas, University of Central Florida Title: Introduction to plant wearables
Dr. Mathews Paret, University of Florida Title: Field performance of formulated Mg and LSP nanoparticles against bacterial spot of tomato
3:50– 4:15 pm Coffee Break 4:15 – 5:15 pm Roundtable
Industry-Academia Discussion Pros and cons of systemic delivery of agrochemicals Moderator: Mike Barry (TradeMark Nitrogen)
Participants: Tim Eyrich (VP of Research and Commercialization, Southern Gardens Citrus); Eli Partyka (Global Head of Marketing, Gowan Company LLC); Miles Armstrong (TradeMark Nitrogen, Inc.); Chris Albright (Helena Agri-Enterprises, LLC); Mike Harowitz (Certis USA); Brett Highland (Certis USA)
5:30 – 6:30 pm Poster Session and dinner
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October 25th, 2019
Registration opens at 8:00 am
8:00 – 9:00 am Breakfast
9:00 am – 10:00 am Scientific program
Young Scientists Series Moderator: Dr. Maria Campos, University of Central Florida
9:00 – 9:20 am Dr. Avra Kundu, University of Central Florida Title: Precision Vascular Delivery of Agrochemicals with Micromilled Microneedles (µMMNs)
9:20 – 9:40 am Dr. Dayse Pulici, University of Florida Title: Huanglongbing on roots: early movement of 'Candidatus Liberibacter asiaticus' and disease progression
9:40 – 10:00 am Marcus V. Merfa, Auburn University Title: Enhancing in vitro growth of ‘Candidatus Liberibacter asiaticus’ by culture medium optimization
10:00 – 10:30 am Coffee Break
10:30 am– 12:30 pm Scientific program
Industrial Hemp Moderator: Dr. Swadeshmukul Santra, University of Central Florida
10:30 – 10:45 am Ray Mazzie, Executive Director, Hemp Industries Association of Florida Title: About the HIAF organization
10:45 – 11:30 am David Hasenauer, Green Point Research LLC. Title: Industrial hemp in Florida
MISA Symposium Adjourns
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Abstracts - Invited Talks
October 24th, 2019
Keynote: Towards Highly Efficient Uptake and Translocation of Foliar Applied
Environmentally Responsive Nano-agrochemicals Gregory V. Lowry
Walter J. Blenko, Sr. Professor of Civil & Environmental Engineering
Abstract Engineered nanomaterials have the potential to revolutionize agrochemical efficacy and vastly improve use efficiency. Nanomaterial properties, including charge, size, and coating hydrophobicity can be engineered to promote efficient uptake and translocation of engineered nanomaterials in plants. We assessed the importance of each of these variables on the uptake and translocation of engineered nanomaterials in plants after foliar and root exposures. Spatially resolved synchrotron X-ray imaging tools demonstrated that coating hydrophobicity controls both route and extent of nanoparticle uptake across the plant leaf cuticle for foliar applied nanomaterials. A relatively more hydrophobic coating afforded nearly 100% uptake through the leaf cuticle due to hydrophobic interactions. Spatially resolved metal transport in leaf cross sections indicate that coating identity also impacts the ease of transport through leaf mesophyll into the plant vasculature. NP size up to 50nm did not influence NP uptake through the cuticle, but size did influence their leaf-to-root transport. For NP<50nm, approximately 20% of the foliar applied Au NPs were exuded from roots into the rhizosphere soil. Experiments exposing plant roots to CeO2 NPs indicates that NP charge affects the distribution of the NPs in plant leaves, with dicots and monocots showing different behaviors. Finally, we have developed a temperature-responsive star polymer that can release active ingredients in vivo under heat stress. Overall, the body of evidence indicates great potential for manipulating nanomaterial properties for beneficial applications in agriculture and for increasing agrochemical utilization efficiency and sustainability of food production. Bio: Greg Lowry is the Walter J. Blenko, Sr. Professor of Civil and Environmental Engineering at Carnegie Mellon University. He is the Deputy Director of the NSF/EPA Center for Environmental Implications of Nanotechnology (CEINT). His research aims to safely harness the unique properties of engineered nanomaterials for making crop agriculture and water treatment more resilient and sustainable. Recent work aims at understanding how a nanomaterial’s properties and environmental conditions influence their fate in soils, nanomaterial-plant interactions, nutrient uptake efficiency, and crop disease management. He has authored more than 160 peer-reviewed journal articles (H index=70) and one book. He served on the board of directors of the Association of Environmental Engineering and Science Professors, and on the US EPA Science Advisory Board (Environmental Engineering committee). He is a Board-Certified Environmental Engineer (BCEEM), Fellow of the American Association for the Advancement of Science, and was a member of the National Academy of Science Committee on Science Breakthroughs 2030: A Strategy for Food and Agricultural Research. Dr. Lowry holds a B.S. in Chemical Engineering from the University of California at Davis, an M.S. from the University of Wisconsin at Madison, and a Ph.D. in Civil & Environmental Engineering from Stanford University.
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10:00 am – 12:15 pm Scientific program Fertilizers and Soil Interactions Moderator: Dr. Swaminathan Rajaraman and Dr. Davie Kadyampakeni
Balanced Nutrition for Citrus Tree Health and Productivity
Bill Easterwood, Ph.D.
Director of Agronomic Services, Yara North America, Inc. [email protected]
Abstract As we endure the scourge of HLB infection in our citrus trees and await our talented scientists to produce a genetic response, we have found that the most productive and beneficial programs to date are psyllid control and balanced nutrition. We have learned that fertilization of HLB citrus is very complex and we must be more diligent and thorough as we determine the nutrition needs of our citrus trees as we minimize tree stress and maximize tree health and productivity. Our balanced nutritional programs should be coupled to 4R nutritional stewardship (right nutrient source, right nutrient rate, right nutrient timing and right nutrient placement) if we are to give our trees maximum management while minimizing nutrient loss to the environment. When we engage the 4R’s with balanced nutrition, we not only produce healthy productive trees but provide the nutrients that can initiate a plant protection response like our own immune system. We will address each of these issues based on the work in the literature performed by great scientists before us and the research Yara has conducted both nationally and internationally.
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Nitrogen, Calcium and Magnesium management for Improving Performance of HLB-affected Orange Trees (Citrus sinensis)
1*Davie Kadyampakeni, 1,2Eduardo Esteves, 2Gabriel Maltais-Landry1, Kelly Morgan3, Rhuanito Ferrarezi4
1University of Florida, Soil and Water Sciences Department, Citrus Research and Education Center, 700 Experiment Station Rd, Lake Alfred, FL33850; 2University of Florida, Soil and
Water Sciences Department, 2181 McCarty Hall, Gainesville, FL32611; 3University of Florida, Southwest Research and Education Center, 2685 SR 29 N, Immokalee, FL 34142; 4University
of Florida, Indian River Research and Education Center, 2199 S Rock Rd, Fort Pierce, FL34945.
*Corresponding author: [email protected] Abstract Citrus production has been declining in Florida since 1998, mainly because of Huanglongbing (HLB) disease. However, nutrients play an important role in defense mechanisms and new approaches attempting to manage the disease with a balanced nutrition are emerging. Nitrogen, calcium and magnesium have the potential for extending the productive life of HLB-affected citrus trees. As the interaction between these nutrients in HLB-affected citrus trees grown in Florida is still unclear, the main objective of this study was to determine the response of HLB-affected trees to N, Ca and Mg applications. The study was conducted in a Candler sand (hyperthermic, uncoated, Lamellic Quartzipsamment) with Valencia trees (Citrus sinensis) on Swingle citrumelo (C. paradisi Macf. X P. trifoliata) rootstock at a planting density of 1,111 trees ha-1. Applications of N at 168, 224 (currently the recommended rate) and 280 kg N ha-1 were the main plots. Split plots consist of a control treatment (C) receiving no Ca nor Mg; Ca applied at 45 kg Ca ha-1; Mg applied at 45 kg Mg ha-1; and Ca+Mg applied at 22.5 kg Ca ha-1 + 22.5 kg Mg ha-1. Phosphorus, potassium and micronutrients were added to all the treatments at recommended rates. Soil NH4
+, NO3- and exchangeable macronutrients and micronutrients; leaf
tissue nutrients; tree size, root growth, yield and fruit quality variables were measured. Yield was significantly lower at the highest N rate (280 kg N ha-1). Canopy volume, leaf Mg concentration and fruit acidity were significantly higher with Mg treatment compared to control. There were no significant differences for root growth variables. Soil nitrate was higher in treatments fertilized with Ca and Ca+Mg. Monitoring tree and soil variables continues to determine the effects of different nutrient management strategies on citrus productivity.
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Modulating lipid and auxin signaling through nutrition may influence fruit retention and root health in citrus
Fernando Alferez and Daniel Boakye.
Corresponding author: [email protected] South West Florida Research and Education Center, University of Florida IFAS, Immokalee, Fl
Abstract
The role of zinc in auxin biosynthesis has been well known for decades. Zinc is required for maintaining normal auxin levels in plants, and zinc deficiency results in lower auxin levels. A decrease in auxin may result in senescence and fruit abscission. At the same time, there is an interplay between auxins and phospholipases in regulating different developmental processes. Phospholipases are a group of enzymes involved in membrane degradation after different stresses, including nutritional stress, to produce lipid signals that trigger downstream biological responses. To investigate the role of zinc and how this element can modulate the interplay between auxins and phospholipases in citrus fruit abscission we treated ‘Hamlin’ sweet orange fruit in an in vitro system with a solution of zinc sulfate. Also, we monitored fruit drop in the field on adult trees of ‘Hamlin’ grafted on Swingle rootstock during maturation. In vitro, May-harvested fruit (early- green fruit) abscission rate progressed faster in zinc-treated fruit than in controls: by14 days after treatment, only 13% of Zn-treated fruit had abscissed as compared to 53% of non-treated controls. In contrast, in July-harvested fruit (mature-green fruit) abscission dynamics was the opposite: by day 14, 93% of Zn-treated fruit had abscissed as compared to only 40% of non-treated controls. Fruit drop data collected in the grove from trees showed the same pattern. Both patterns for May and July were identical in auxin-treated fruit. This parallelism reinforces the link between auxins and zinc and points out to a seasonal component worth to be investigated, because it might result in different nutritional requirements during fruit maturation and development. In earlier and parallel experiments, we found also that by altering auxin transport we can alter lipid signaling, abscission and fruit drop.
At the same time, in model plants such as Arabidopsis thaliana and Oryza sativa, it has been reported that increased expression of different isoforms of phospholipase D (PLD) promote root hair elongation and primary root growth in conditions of nutrient deprivation. This fact strongly suggests that lipid signaling plays a role in translating membrane sensing of nutrient status into increased plant growth and root density. IAA is also involved in lateral and adventitious root initiation and development via phospholipase activation, and zinc addition can stimulate adventitious root formation in the presence of auxins. We are currently determining the involvement of different PLD isoforms in root patterning in citrus in response to zinc.
These two increasing bodies of experimental evidence are beginning to unravel how nutrients may modulate hormonal function via lipid signaling in citrus. Since lipid signaling is also seasonally regulated, taken together this accumulating knowledge will allow to fine tune nutrition to maximize fruit yield by reducing fruit drop, and to increase root density.
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Soil-applied micronutrients can improve productivity of Huanglongbing-affected sweet orange
Tripti Vashisth* and Jude Grosser
University of Florida, Citrus Research and Education Center, Lake Alfred, FL 33850 * corresponding author
Email: [email protected]
Abstract In last 15 years Florida citrus industry has been struggling with Huanglongbing (HLB). Field observation and preliminary studies have shown that constant supply of soil applied nutrients (macro and micro) are beneficial for HLB-affected trees. In addition, it has been well documented that HLB-affected groves that have bicarbonate-rich irrigation water and soil tend to decline rapidly as compared to groves with neutral to acidic soil and irrigation water pH. However, till date, there are very few scientific reports suggesting effect of soil-applied nutrients and low pH on improvement of health and productivity of HLB-affected trees. Meanwhile, Florida citrus growers have been dynamically changing their fertilization practices to enhanced soil-applied fertilization program based on any available information on citrus nutrition. Therefore, in 2016 we initiated a large scale field trial, to evaluate the effect of soil-applied micronutrients (with sulfur bentonite coating, ability to lower the soil pH) in combination with controlled release fertilizer (CRF). In total there were 9 treatments compared to grower control at two separate locations, Central Florida and Southwest Florida, respectively. As expected we found site-specific results however, grower control was consistently the poorest performer (lowest yield) at both sites for three consecutive years. In second and third year, significantly improved yield were observed for added manganese, boron, and iron treatment in southwest Florida site. Overall, in 3 years at central Florida site, the 20% higher manganese and boron treatment yielded 1,130 boxes per acre (1 box = 45 kg) as compared to control treatment which produced 893 boxes per acre whereas the added iron treatment produced 1310 boxes per acre compared to control (908 boxes per acre) at southwest Florida site. Altogether, the use of CRF with soil-applied micronutrient consistently was ranked second with 1263 and 1076 boxes per acres for southwest and central Florida site, respectively. In addition, in a consumer preference juice quality test we found that, consumer preferred the juice from added manganese and boron treatment over control. In sum, our 3 year long trial suggests that the use of CRF with soil applied micronutrient can improve productivity of HLB-affected trees.
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Can higher planting density and enhanced fertilization increase fruit yield of HLB-infected grapefruit?
Rhuanito S. Ferrarezi*, Dinesh Phuyal, Taylor Meadows and Hernan Soto
University of Florida, Indian River Research and Education Center, Fort Pierce, FL 34945 Davie Kadyampakeni
University of Florida, Citrus Research and Education Center, Lake Alfred, FL 33850 Kelly Morgan
University of Florida, Southwest Research and Education Center, Immokalee, FL 34142 * corresponding author
Abstract Citrus greening or Huanglongbing (HLB) disease is a serious threat to citrus production in Florida. Since a cure is still unknown, disease management is necessary to maintain trees in production. Management strategies rely on different approaches: controlling the insect vector, applying antimicrobials and enhance plant nutrition to extend tree survival. We are conducting a field trial on 5-year-old ‘Ray Ruby’ grapefruit (Citrus paradisi) on Kuharske citrange (Citrus sinensis·× Poncirus trifoliata) rootstock planted in the Flatwoods soils in Fort Pierce, FL. The objective is to understand how tree spacing, soil and foliar fertilization affect fruit yield and fruit quality on HLB-infected grapefruit trees in the Indian River citrus district. We tested three planting densities [standard spacing (300 trees/hectare), high-density single row (440 trees/hectare), and high-density double row staggered in diamond setting (975 trees/hectare)], two soil application blends of controlled-release fertilizer (CRF) (16N-1.31P-16.6K and 12N-1.31P-7.47K plus higher micronutrient rate), and four foliar rates of micronutrient (a blend of Mn, Zn, and B at 0×, 1×, 3×, and 6× IFAS recommendation). All the trees tested positive for HLB based on real-time quantitative PCR test. The preliminary results indicate the highest yield was obtained with high-density double row staggered at 170 boxes of 38.5 kilograms per hectare (P<0.001). Canopy volume was 43% higher with standard spacing than high-density double row staggered (P<0.0001). Trunk diameter was 2.3% larger on standard tree spacing than high-density double row staggered with no effect of nutrient treatment (P<0.0001). Fruit with high-density double row staggered were more acidic (average titratable acidity 1.16 mg/100 mL) than other treatments irrespective of the nutrient application (P<0.0001). Soluble solids content was 5.75% higher on high-density double row staggered trees than standard spacing (P=0.0278) with no effect of nutrient treatment. There was no effect on yield by both CRF formulations (P=0.8428) and foliar treatment (P=0.7126) in the first year. The 12N-1.31P-7.47K increased the canopy volume by 8% (P<0.0001). Canopy volume remained unaffected by foliar treatment (P=0.3782). Leaf nutrient concentration increased with the increase in foliar rate for all micronutrients, particularly Mn (P<0.0001) that exceeds the optimum range with 6× application of IFAS rate. The study shows that high-density double row staggered results in higher yield with lower canopy volume. CRF blends with higher micronutrients can contribute to greater canopy growth without increasing fruit yield. Foliar treatment of micronutrients can maintain tree health by improving nutrients status of the plant. Funding. Funding for this research was provided by UF/IFAS Citrus Initiative.
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1:45 – 4:00 pm Scientific program Emerging Tools for Sustainable Agriculture Moderator: Dr. Mathews Paret, Dr. Jayan Thomas
Zinkicide improves health and yield of HLB-affected trees: limited by
dilution and residual efficacy?
E. G. Johnson, M. M. Murata, S. Santra, M. M. Dewdney
Abstract Huanglongbing (HLB), caused by the phloem-limited bacterium ‘Candidatus Liberibacter asiaticus’ (CLas), is the yield limiting factor in many citrus producing regions. Because of the difficulty to reach phloem tissue with chemical treatments, a 2-3 nm zinc oxide-based nanoparticle, Zinkicide, was designed for systemic activity with traditional foliar spray or soil drench application. A canker bioassay demonstrated systemic efficacy. Greenhouse trials have confirmed significant reduction in CLas populations by an average 2.54 log in 20 weeks when applied at a biweekly schedule to maintain an effective dose in the phloem. However, tree size and application frequency affected phytotoxicity thresholds that also altered efficacy. Multiple field trials on grapefruit and sweet orange were carried out for multiple years on trees of varied age and size. Surprisingly, when the nanoparticle synthesis was reformulated to tolerate impurities in agricultural grade starting materials, nanoparticle size became smaller and field efficacy improved leading to yield increases up to 73%. Young grapefruit trees have responded better than mature sweet orange trees. Dilution effects, preventing effective doses existing systemically, are the likely cause of this apparent tree size dependent efficacy. Hurricane Irma provided a unique opportunity to observe that Zinkicide reduced leaf loosening, leading to much less leaf drop damage from hurricane force winds in young non-bearing trees. Zinkicide nanoparticles show promise as a management tool for young plantings to reach profitability. Changes in application rate and frequency currently under evaluation may improve efficacy on larger plantings.
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Use of UV light for control of strawberry powdery mildew
N. Peres University of Florida
Abstract Strawberry powdery mildew, caused by Podosphaera aphanis, can be destructive in both covered and open field production. Fungicides are the principal means used to suppress this disease. Ultraviolet light (UV) has been used to effectively suppress diverse powdery mildews in greenhouse and other indoor facilities. In our studies, we designed a tractor-drawn apparatus to apply UV over the surface of strawberry plants in open fields. Over three growing seasons, we exposed plants of mildew-susceptible strawberry plants to UV light (254 nm, FWHM < 5 nm) approximately 1 hr after sunset either once or twice weekly. In all three seasons, UV has suppressed powdery mildew equally to or better than standard fungicide treatments. No deleterious effects of UV exposures have been noted on plant growth or yield.
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Novel Silica-functionalized Quaternary Ammonium Nanocoating for Water Disinfection and Harmful Algal Bloom (HABs) Mitigation
Daniela Diaz1, Jared Church1, Mikaeel Young2,3, Keug Tae Kim4, Jungsu Park5, Yun Bin
Hwang4, Swadeshmukul Santra2,3,6,7, and Woo Hyoung Lee1,* 1Department of Civil, Environmental, and Construction Engineering, University of Central
Florida, 12800 Pegasus Drive, Orlando, FL 32816, USA 2NanoScience Technology Center, University of Central Florida, 12424 Research Parkway,
Orlando, FL 32826, USA 3Burnett School of Biomedical Sciences, University of Central Florida, 6850 Lake Nona Blvd,
Orlando, FL 32827, USA 4Department of Environmental & Energy Engineering, Suwon University, 17 Wauan-gil,
Bongdam-eup, Hwaseong-si, Gyeonggi-do, 445-743, Republic of Korea 5K-water Institute, Korea Water Resources Corporation, 200 Sintanjin-Ro, Daedeok-Gu,
Daejeon, 34350, Republic of Korea 6Department of Material Science and Engineering, 4000 Central Florida Blvd, University of
Central Florida, Orlando, FL 32816, USA 7Department of Chemistry, 4111 Libra Drive, University of Central Florida, Orlando, FL 32816,
USA *Corresponding author: Woo Hyoung Lee, email: [email protected]
Abstract
Intensification of pollution loading and the lack of appropriate treatment of wastewater have been related to detrimental impacts in the quality of receiving surface waters. Nutrient overloading from sources such as fertilizer runoff and industrial effluents have potentially stimulated the incidence of harmful algal blooms (HABs) over many water bodies around the world. HABs are either considered toxic or the cause of major ecological impacts such as oxygen depletion and alteration of food web dynamics. Although many physical, chemical and biological treatments have demonstrated to be effective for the control of microbial pathogens, the undesired formation of harmful disinfection byproducts (DBPs) has revealed an urgent need to reevaluate and innovate conventional methods. Quaternary ammonium compounds (QACs) are cationic surfactants known to be effective at the ppm level against a broad range of microorganisms. As a potential way to utilize the biocidal properties of QACs without directly releasing them into the environment, the synthesis of modified surfaces impregnated with QACs can provide a potential alternative for water disinfection.
In this study, a fiberglass mesh was utilized as a substrate, containing silica, which covalent attachment of QACs forms an active antimicrobial surface. The silica-nanocoated mesh was fabricated via sol-gel technique using a composite of silica-modified QACs (Fixed-Quat), and evaluated for the control of E. coli and Microcystis aeruginosa in water. When using E. coli as a model bacterial pathogen, Fixed-Quat coated fiberglass mesh showed effective microbial inactivation performance with a rate of 1.3 × 10-3 log reduction cm-2 min-1. The modified surface also showed successful mitigation of M. aeruginosa with an 81% reduction of chlorophyll-a contents in a short term (less than 9 hours). Additionally, the developed antimicrobial mesh was evaluated with wild-type microalgal species collected from a water body experiencing HABs, obtaining a 97% removal efficiency. Overall, the silica-functionalized Fixed-Quat nanocoating showed promising antimicrobial properties for water disinfection and cyanobacteria growth control, while eliminating the formation of regulated DBPs and minimizing the release of nanomaterials (NMs) into the environment.
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Early detection of crop health using multispectral sensing
Kalischuk, M., Paret, M. University of Florida
Abstract Multispectral imaging is increasingly used in specialty crops, but its benefits in assessment of plant health relative to disease, fertilizer-flow, and irrigation-flow rates is unknown. Data obtained from an unmanned aerial vehicle carrying a multispectral sensor were classified using normalized difference index values (NDVIs) and supported early detection of limited nitrogen flow rates in watermelon and cucumbers. Disease severities of Gummy stem blight (Stagonosporopsis cucurbitacearum) and Phytophthora fruit rot (Phytophthora capsica) of watermelon and Downy mildew (Pseudoperonospora cubensis) of cucumber, were evaluated in plantations by conventional and UAV-assisted scouting. Disease incidence and severity ratings were significantly different between conventional and UAV-assisted scouting (P<0.01, Bhapkar/exact test). The UAV-assisted scouted locations had significantly lower green, red, and red edge NDVI values and higher stress index values than the conventional scouted areas (P<0.05, ANOVA/Tukey), and this corresponded to areas with higher disease severities. Multispectral imagery improved watermelon and cucumber field scouting owing to increased ability to identify disease foci and areas of concern more rapidly than conventional scouting practices with early disease detection possible using UAV-assisted scouting.
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Introduction to plat wearables
Jayan Thomas University of Central Florida
Abstract Currently, human wearable devices attract significant interest due to its applications in communication, on-body sensing, and general health care. However, wearable devices capable of predicting plant health are not that common. This presentation will provide a general overview of plant sensors that can be used to improve plant productivity, pathogen-related stress detection and plant physiology monitoring on real time. Since all these electronic devices require energy to operate, fibre-type wearable energy devices which can be used along with these devices will be introduced. The presentation will conclude with a discussion about the future prospects of plant wearables.
Magnesium nanomaterial, a novel alternative to bacterial spot of tomato disease management
Liao, YY.1,2, Strayer-Scherer, A.1,2, Huang, Z.3, Santra, S.3, White, J.C.4, De La Torre-Roche, R.
4, Fan, Q. 1,2, Da Silva, W.S. 2, Vallad G. E5., Freeman, J.2, Jones, J. B.1 and Paret, M. L.1,2
1Department of Plant Pathology, University of Florida, Gainesville, FL, USA 2North Florida Research and Education Center, University of Florida, Quincy, FL, USA
3Biomolecular Science Center, University of Central Florida, 12424 Research Parkway, Suite 400, Orlando, FL 32826, USA
4Department of Analytical Chemistry, The Connecticut Agricultural Experiment Station, New Haven, CT, USA
5Gulf Coast Research and Education Center, University of Florida, Wimauma, FL, USA
Abstract Bacterial spot of tomato, incited by four Xanthomonas species, is a devastating bacterial disease worldwide. There has been no effective chemical control strategy in Florida following the appearance of copper (Cu)-tolerant strains, which rendered Cu bactericides ineffective. In addition, the EPA has suggested that Cu bactericide applications should be limited on crops due to ecological risks. Therefore, finding an effective alternative is critical. Our research has focused on novel antibacterial properties of magnesium oxide nanoparticles (nano-MgO) against Cu-tolerant X. perforans in vitro, greenhouse, and the field. In addition, nano-MgO applications did not significantly alter levels of Mg, Cu, Ca, K, Mn, P and S compared to fruit in the untreated control. In our current study, newly formulated Mg nano-materials (i.e., SgMc, Mg-Cu, Mg double coated) with enhanced solubility significantly increased antibacterial efficacy compared to nano-MgO in vitro. In two greenhouse studies, SgMc when applied at 100 µg/mL reduced disease severity compared to the grower’s standard, Cu-EDBC, and the water control (p = 0.05). Based on the preliminary findings, we hypothesize that Mg nanomaterials will be an effective alternative to the current grower standard for control of bacterial spot. Keywords – Agriculture, Copper-tolerant bacteria
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Abstracts - Posters
October 24th, 2019
Comparative Study of Antimicrobial efficacy of Nano-Zinc oxide and Bulk-Zinc oxide on Pseudomonas syringe
Danya Belnoura,b, Hajeewaka Mendisb, Swadeshmukul Santraa,b,c,d
aBurnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, FL 32827
bNanoScience Technology Center, Orlando, FL 32826 cDepartment of Chemistry, dDepartment of Materials Science and Engineering, University of Central
Florida, Orlando FL 32816
Abstract Huanglongbing (HLB) also called Citrus greening disease, is a disease caused by Candidatus liberibecter asiaticus which is transmitted to plant by Asian citrus psyllid. HLB is responsible for devastating economic losses to the citrus industry in the United States and worldwide. Zinkicide is a micronutrient based biocide that was designed and developed to treat citrus greening. Zinkicide is a Zinc Oxide based nanomaterial (Nano-ZnO) which exhibit systemic antimicrobial activities. Ultra-small size (<5nm) is a key quality of Zinkicide for the treatment of HLB infected plants. In this study, a comparison of an equal concentration of Zinc in Zinkicide, rotovaped Zinkicide (R-Zinkicide), and bulk Zinc Oxide (bulk-ZnO) was completed to investigate the antimicrobial activities including membrane damage capabilities of those materials on Pseudomonas syringe. P. syringe is a gram-negative plant pathogen that serve as a common model system for bacteria pathogenesis. The antimicrobial activities were investigated using three standard techniques. Minimal inhibitory concentration assay (MIC) was used to determine the lowest concentration of material that inhibits bacterial growth. Alamar Blue assay was then used to determine the growth inhibition percentage of each material. Minimal bactericidal concentration (MBC) was completed to determine the minimal concentration that causes killing of the pathogen. The membrane damage study (DNA based assay) was completed to determine reagents membrane damage capabilities by measuring the absorbance of intercellular content such as DNA absorbance at 260nm and proteins absorbance at 280nm. Zinkicide has higher antimicrobial activity compared to R-Zinkicide and bulk-ZnO. Zinkicide minimal inhibitory concentration/minimal bactericidal concentration is 8ppm/64ppm. On the other hand, bulk-ZnO has an MIC/MBC of 16ppm/256ppm, and R-Zinkicide has an MIC/MBC of 32ppm/256ppm. Zinkicide and R-Zinkicide have a higher 260 and 280nm absorbance compared to bulk-ZnO. This comparative analysis has demonstrated an increase in antimicrobial efficacy of Nano-ZnO compared to its bulk counterparts.
Keywords- Zinc Oxide nanoparticles, Zinkicide, Pseudomonas Syringe, Huanglongbing, citrus greening
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Evaluation of antimicrobial effects and plant uptake of novel antimicrobial formulation MS3T to control surface and systemic
pathogens in citrus. Hajeewaka Mendis1; Maria Campos1, Ali Ozcan1,2, Mikaeel Young4, Swadeshmukul Santra1,2,3,4
1NanoScience Technology Center, 2Department of Chemistry, 3Department of Materials Science
and Engineering and 4Burnett School of Biomedical Sciences, University of Central Florida, Orlando, FL, USA.
Abstract MS3T (multifunctional surface/sub-surface/systemic therapeutic) is a novel plant nutrient-based antimicrobial formulation developed for controlling citrus bacterial pathogens including ‘Candidatus Liberibacter asiaticus’ (CLas) and Xanthomonas citri (Xcc). MS3T components fixed-quat (DDAC) and Zn chelate (TSOL) are designed to control plant surface pathogens and systemic pathogens respectively. This study investigated the antimicrobial effects of MS3T and its components in vitro on CLas and Xcc surrogates X. alfalfae (Xa) and E. coli (Ec). Ec and Xa were grown in 96-well and 24-well plates in LB and NB with serial dilutions of MS3T, Zn(NO3)2
and DDAC to determine Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). Systemic movement of TSOL in citrus plants was evaluated by leaf-spraying of TSOL followed by acid digestion and atomic absorption spectroscopy (AAS). Phytotoxicity of MS3T was evaluated in tomato seedlings. The MIC of Zn(NO3)2 and DDAC for Xa is 16 ppm and 0.5 ppm whereas MBC is 32 ppm and 1 ppm respectively. The MIC of MS3T for Xa is 1.5 ppm and 6 ppm DDAC and Zn. The MIC of Zn(NO3)2 and DDAC for Ec is 128 ppm and 4 ppm whereas MBC is 512 ppm and 4 ppm respectively. The MIC of MS3T for Ec is 6 ppm and 24 ppm DDAC and Zn. No phytotoxicity was observed in tomato seedlings after MS3T leaf-spraying and AAS analysis revealed that TSOL is uptaken by citrus leaves and transported to roots through phloem within 3 days. Keywords – MS3T, TSOL, ‘Candidatus Liberibacter asiaticus’, Xanthomonas citri subsp. Citri, Xanthomonas alfalfae, Escherichia coli.
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Magnesium nanomaterial, a novel alternative to bacterial spot of tomato disease management
Liao, YY.1,2, Strayer-Scherer, A.1,2, Huang, Z.3, Santra, S.3, White, J.C.4, De La Torre-Roche, R.
4, Fan, Q. 1,2, Da Silva, W.S. 2, Vallad G. E5., Freeman, J.2, Jones, J. B.1 and Paret, M. L.1,2
1Department of Plant Pathology, University of Florida, Gainesville, FL, USA 2North Florida Research and Education Center, University of Florida, Quincy, FL, USA
3Biomolecular Science Center, University of Central Florida, 12424 Research Parkway, Suite 400, Orlando, FL 32826, USA
4Department of Analytical Chemistry, The Connecticut Agricultural Experiment Station, New Haven, CT, USA
5Gulf Coast Research and Education Center, University of Florida, Wimauma, FL, USA
Abstract Bacterial spot of tomato, incited by four Xanthomonas species, is a devastating bacterial disease worldwide. There has been no effective chemical control strategy in Florida following the appearance of copper (Cu)-tolerant strains, which rendered Cu bactericides ineffective. In addition, the EPA has suggested that Cu bactericide applications should be limited on crops due to ecological risks. Therefore, finding an effective alternative is critical. Our research has focused on novel antibacterial properties of magnesium oxide nanoparticles (nano-MgO) against Cu-tolerant X. perforans in vitro, greenhouse, and the field. In addition, nano-MgO applications did not significantly alter levels of Mg, Cu, Ca, K, Mn, P and S compared to fruit in the untreated control. In our current study, newly formulated Mg nano-materials (i.e., SgMc, Mg-Cu, Mg double coated) with enhanced solubility significantly increased antibacterial efficacy compared to nano-MgO in vitro. In two greenhouse studies, SgMc when applied at 100 µg/mL reduced disease severity compared to the grower’s standard, Cu-EDBC, and the water control (p = 0.05). Based on the preliminary findings, we hypothesize that Mg nanomaterials will be an effective alternative to the current grower standard for control of bacterial spot.
Keywords – Agriculture, Copper-tolerant bacteria
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Nano Zinc Oxide Coated Urea Fertilizer for Efficient Delivery of Zinc Micronutrient to Plants
Maria Campos1,*, Ali Ozcan1,2, Christian Dimkpa3, Swadeshmukul Santra1,2,4
1- University of Central Florida, NanoScience Technology Center, Orlando, FL 2- University of Central Florida, Department of Chemistry, Orlando, FL
3- International Fertilizer Development Center, Muscle Shoals, AL 4- University of Central Florida, Department of Materials Science and Engineering and
Burnett School of Biomedical Sciences, Orlando, FL
Abstract Food production to feed the estimated 9 billion world population by 2050 is a big food security challenge. In addition, improving the nutritional value of food crops is also critical. Zinc (Zn) is an important micronutrient for plants and animals. Zn-deficiency causes stunted growth, leaf size reduction and yield loss in crops. As half of world’s arable soils is Zn-deficient, zinc oxide (ZnO) is commonly blended with urea fertilizer and applied to soil. However, the effectiveness of Zn uptake by plants is controlled by Zn bioavailability and water-solubility. Dissolution rate and solubility of ZnO can be related to particle size and surface area. Therefore, this study aims to coat urea with ZnO nanoparticles (NPs) synthesized with a combination of two capping agents: urea (U), sodium salicylate (SAL) and n-acetyl cysteine (NAC); to improve ZnO solubility and bioavailability. NAC-SAL, NAC-U and U-SAL capped ZnO nanoparticles were synthesized using a sol-gel method at room temperature. Samples were characterized by Atomic Absorption Spectroscopy (AAS), Infrared Spectroscopy (IR) , Dynamic Light Scattering (DLS), Zeta potential, and High Resolution Transmission Electron Microscopy (HR-TEM). Urea granules were coated with ZnO NPs and bulk ZnO. Total and water-soluble Zn in the coated urea were determined by AAS. Zn content (w/w) in U-SAL, NAC-SAL and NAC-U capped NPs was 22, 18 and 19%, respectively. NAC-SAL ZnO and NAC-U ZnO have negative surface charge, while U-SAL ZnO has negative zeta potential. The average hydrodynamic diameter of all NPs was around 100nm (DLS). HR-TEM analysis showed less than 5nm ZnO particles for all samples. The Zn-O band (1386cm-1) and C=O peak (1700cm-1) were found in the IR spectra of all samples. The S-H characteristic band (2548cm-1) was present in both NAC-U and NAC-SAL capped ZnO spectra. The peaks for N-H2 (U) were found at 3450 and 1625cm-1. The primary amide group of U and secondary amide group of SAL showed peaks at 1585 and 1535cm-1, respectively. Carboxylic acid group of SAL and NAC were related to the peak around 3100cm-1. C-H vibration peak in the benzene ring was found in both NAC-SAL ZnO and U-SAL ZnO spectra (810cm-1). The percentage of soluble Zn released by the NPs after 24h dialysis (3.5 kDa cutoff) against DI water was at least double, compared to bulk ZnO. Therefore, dissolution rate of ZnO can be controlled by particle size using a combination of capping agents, and may improve Zn uptake by plants due to higher bioavailability.
Keywords - Zinc, urea, coating, nanoparticles
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Systemic Tracking of Small Molecules in-planta via Fluorescence Lifetime Imaging
Gregory Millera,b, Maria Camposa, Tyler Maxwella,b, Swadeshmukul Santraa,b,c,e, Andre
Gesquierea,b,c,d, aNanoScience Technology Center, University of Central Florida, Orlando, FL 32826,
USA. bDepartment of Chemistry, University of Central Florida, Orlando, FL 32826, USA cDepartment of Materials Science and Engineering, University of Central Florida,
Orlando, FL 32826, USA dThe College of Optics and Photonics (CREOL), University of Central Florida, Orlando,
FL 32826, USA e Burnett School of Biomedical Sciences University of Central Florida, Orlando,
FL 32826, USA E-mail: [email protected]; [email protected]
Abstract Systemic bacterial diseases, such as Huanglongbing (HLB), have been decimating the agricultural industry. The citrus industry in Florida, for example, has seen a 75% decrease from 2005, when the bacteria causing HLB, Candidatus Liberibacter asiaticus (CLas), was first detected in citrus orchards. Current methods of treating systemic bacterial diseases use antibiotics or new nanoparticle drugs, which are both believed to be systemic. This idea arises from the knowledge of the systemic disease affecting the plant, and subsequent mitigation of disease symptoms. Direct systemic tracking, however, has proven difficult, due to either lack of spatial resolution or the convolution of natural plant fluorescence. We have demonstrated that Fluorescence Lifetime Imaging (FLIM) has the spatial resolution required to detect localization of small molecules in-planta, and its novel temporal separation allows the target to be detected simultaneously, but distinguishably, from plant background. This method isn’t without its own limitations: the target to be detected must be highly fluorescent and possess an excited state lifetime significantly longer than the ~7 ns lifetime collected from plant background. We used tris(bipyridine)ruthenium(II) chloride hexahydrate ([Ru(bpy)3]Cl2·6H2O) as our surrogate for small molecule antibiotics to demonstrate the capabilities of this method. This compound was selected, because it is a highly fluorescent dye molecule with a long lifetime (~1 µs), due to metal-to-ligand charge transfer effects. Direct delivery to the plant vasculature was accomplished with a microneedle roller. After incubation with dye and cross sectioning, we could determine the localization and translocation kinetics of the dye in-planta. We found that within 3 hrs of delivery, the dye begins moving through the xylem towards the leaves. Within 24 hrs, the dye is present in every cross section above the treated section. Interestingly however, little to no translocation into the roots was observed, even after 1 week of incubation. Keywords- Fluorescence Lifetime Imaging (FLIM), Systemic Translocation, Systemic Imaging
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A Minimally-invasive 3D-printed Microneedle Array Applicator System (µNAAS) for the Delivery of Therapeutics to Citrus Leaf Tissue
Laboni Santra1,2, Avra Kundu2, Swaminathan Rajaraman2, 3, 4, 5
1Oviedo High School, Oviedo, FL, 32765, USA 2NanoScience Technology Center (NSTC), University of Central Florida, Orlando, FL, 32826,
USA 3Department of Materials Science & Engineering, University of Central Florida, Orlando, FL,
32816, USA 4Department of Electrical & Computer Engineering, University of Central Florida, Orlando, FL,
32816, USA 5Burnett School of Biomedical Sciences, University of Central Florida, Orlando, FL, 32827, USA
Laboni Santra: [email protected] Avra Kundu: 321-948-1982; [email protected]
Swaminathan Rajaraman: 407-823-4339; [email protected] Abstract This work reports the design, fabrication, and testing of a novel minimally-invasive mechanical delivery system that can potentially transport therapeutics to Huanglongbing (HLB) affected trees. HLB has devastated Florida’s 9-billion-dollar citrus industry. Since HLB is caused by phloem-restricted bacteria, treatment must reach phloem tissue to be effective. Direct delivery to phloem is extremely challenging, demanding innovation. It is hypothesized that a microneedle-based applicator will be suitable for creating punctured channels on leaves through which treatment can reach phloem. A microneedle array was designed (using CAD), 3D printed and fixed onto a mechanical applicator to fabricate the μNAAS device. As a proof-of-concept, a treatment containing cadmium (not present in leaves naturally), was delivered to citrus leaves by this applicator. Treated leaves were subsequently washed thoroughly and characterized using scanning electron microscopy-energy dispersive spectroscopy (SEM-EDS) for cadmium uptake, which confirmed treatment delivery qualitatively and the creation of punctured channels. X-Ray fluorescence spectroscopy quantified concentrations of cadmium in plant tissue. A 45% increase was observed in microneedle treated plants compared to control (statistically significant). This study successfully demonstrated the potential for microneedle applicators to directly deliver therapeutics and other useful substances (such as genetic materials) to citrus phloem. Future steps include designing and fabricating an efficient biodegradable microneedle-embedded staple carrying therapeutic cargoes that will be applied onto trees with a staple gun, creating minimally-invasive, cost-effective rapid therapeutic application suitable for testing in greenhouse/field condition. Keywords: Microneedles, 3D-printing, stapler applicator system, citrus greening
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Synthesis of a Fluorescent Antibiotic Conjugate to Monitor Translocation in planta via Fluorescence Lifetime Imaging
aRyan Parente, abGregory Miller, eDanya Belnour, bDr. Titel Jurca,
abceDr. Swadeshmukul Santra, abcdDr. Andre Gesquiere
aNanoScience Technology Center, University of Central Florida, Orlando, FL 32826, USA. bDepartment of Chemistry, University of Central Florida, Orlando, FL 32826, USA cDepartment of Materials Science and Engineering, University of Central Florida,
Orlando, FL 32826, USA dThe College of Optics and Photonics (CREOL), University of Central Florida, Orlando,
FL 32826, USA e Burnett School of Biomedical Sciences University of Central Florida, Orlando,
FL 32826, USA
[email protected]; [email protected]; [email protected] [email protected]; [email protected]; [email protected]
Abstract Antibiotic treatment of systemic bacterial plant pathogens has been an established practice in many crops; however, in citrus it has only recently become available for growers to utilize against citrus greening. The preliminary efficacy of these treatments is indeterminate at best, due to the inability to track their presence in phloem. The need to monitor their movement in planta, especially their presence in vascular tissue, is a necessary step in clarifying their effectiveness. Our previous work has shown the value of Fluorescence Lifetime Imaging (FLIM) in distinguishing between fluorescent probes and plant tissue autofluorescence, which is normally a barrier in photochemical studies in plants. Our aim in this study was the synthesis and characterization of a fluorescent antibiotic conjugate that could be utilized for tracking in citrus tissue. In our prior work, we demonstrated the efficacy of tris(bipyridine)ruthenium(II) chloride hexahydrate ([Ru(bpy)3]Cl2·6H2O) as a fluorescent dye that could be tracked in planta by utilizing FLIM. This ruthenium dye has a notably long (~1 µs) lifetime that can be easily distinguished from the short (<10 ns) plant autofluorescence lifetime. Synthesis of a modified ruthenium bipyridine complex was accomplished and subsequent conjugation to streptomycin sulfate, a common commercially available antibiotic, was shown. Optical and photophysical characterization showed both exceptional fluorescent properties as well as extended (43±6 ns, 105±13 ns) lifetime beyond that of plant autofluorescence. Likewise, Fourier-Transform Infrared Spectroscopy (FTIR) indicated successful conjugation with a distinct peak at 1705 cm-1 indicative of an amide linkage not present in any of the synthetic reagents. Ultimately, synthesis of a fluorescent probe with desirable lifetime was accomplished and subsequent conjugation to an agrochemical of great interest was successful. Further plant trials are in progress, and our expectation is that we will be able to follow the translocation of this antibiotic conjugate utilizing FLIM. Keywords – Fluorescence Lifetime Imaging, Systemic Translocation, Antibiotic Conjugation, Citrus Greening
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Evaluating the antimicrobial mechanisms of ZinkicideTM against Liberibacter crescens as a model for ‘Candidatus Liberibacter spp.’.
Eber Naranjoa, Marcus V. Merfaa, Swadeshmukul Santra b,c,d,e, Mikaeel Youngb,e, Evan
Johnsonf , Leonardo De La Fuentea.
aDepartment of Entomology and Plant Pathology, Auburn University. bNanoScience Technology Center, University of Central Florida. cDepartment of Chemistry, dDepartment of Materials Science and Engineering & eBurnett School of Biomedical Sciences, University of Central Florida, fCitrus Research and Education Center, University of Florida.
Address correspondence to: Leonardo de La Fuente, [email protected]. 209 Rouse Life Sciences Building, Auburn, AL, 36849-5413
Abstract Liberibacter spp. are causal agents of incurable plant diseases worldwide. Their intracellular life style, insect-vectored and unculturable nature, precludes the screening of antimicrobial compounds in vitro. Here, the antimicrobial activity of ZinkicideTM (Znk), a zinc-based nano-formulation developed to control bacterial citrus pathogens, was evaluated in vitro using Liberibacter crescens (Lcr) as a biological model, since it is the only culturable species of the genus. ZinkicideTM outperformed bulk zinc oxide (ZnO) in minimum bactericidal concentration (7 log10 reduction for Znk at 150 ppm; < 3 log10 reduction for ZnO at 200 ppm) and “time to kill” assays (5 log10 orders reduction at 24 hours for ZnK; < 2 log10 reduction at 48 hours for ZnO). When applied at 150 ppm against 109 Lcr cells, ZinkicideTM antimicrobial mechanisms included cell membrane disruption, release of Zn ions and an increase in reactive oxygen species. ZinkicideTM also displayed a stronger biofilm inhibition activity than bulk ZnO in batch cultures (50% more reduction at 25 ppm). Neither ZinkicideTM nor ZnO disrupted Lcr preformed biofilms in batch systems, but ZinkicideTM killed 20% more sessile cells in such biofilms than ZnO. In microfluidic chambers resembling plant vascular systems, ZinkicideTM outperformed ZnO in Zn ions translocation and qualitative cell death, as assessed by the LIVE/DEAD® BacLight Bacterial viability kit. Proved enhanced bactericidal activities in batch systems and microfluidic chambers for ZinkicideTM here make this compound a promising product to control plant diseases caused by Liberibacter spp. Keywords - HLB, Liberibacter, Bactericidal, ZnO nanoparticle, viability, biofilm, microfluidics, reactive oxygen species, membrane disruption.
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Nanoparticles for Sustained Antibiotic Release and Prolonged Antibacterial Activity for Crop Protection
Tyler Maxwell1,2 , Parthiban Rajasekaran2, Morgan Schaff 2, Mikaeel Young2,3, and
Swadeshmukul Santra 1,2,3,4 1Department of Chemistry, 2NanoScience Technology Center, 3Burnett School of Biomedical
Sciences 4Department of Material Science and Engineering, University of Central Florida
[email protected] 863.224.2634 Abstract Antibiotics such as streptomycin and oxytetracycline are currently used to control bacterial disease of plants. Recently, these antibiotics have received approval for use in combating diseases in citrus plants such as citrus canker. However, due to the water solubility of these antibiotics they are not able to leave a persisting residue on citrus trees due to frequent rainfall. Nanoparticles such as quantum dots (Qdots) have the potential to improve the leaf adhesion of antibiotics due to their high surface areas. Herein we report the design and synthesis of a nontoxic, heavy metal free ZnS:Mn Qdot for the sustained release, tracking, and prolonged activity of antibiotics for crop protection. The Qdots were synthesized in a one-step one-pot synthesis with EPA approved reagents and coated with N-acetylcystine (carboxyl containing biomolecule). Streptomycin was attached to the Qdot both covalently using EDC coupling and electrostatically using the negative charge of the Qdot coating and positive charge drugs. The Qdots were then characterized through spectroscopy (UV-Vis, fluorescence, and infrared) and electron microscopy. The therapeutic efficacy of the nanoparticles was evaluated against model pathogenic bacteria E. coli, P. Syringae, and X. alfalfae. The particles showed comparable antibacterial efficacy to free drug at the same concentrations. Drug release kinetics of the particles show a slow sustained release of free antibiotic over a 120 hour period. Rainfastness testing showed improved retention of streptomycin on a citrus leaf surface in combination with the Qdots compared to free streptomycin and Firewall WP (commercial streptomycin formulation). Keywords: Nanoparticle, Quantum Dot, Antimicrobial, Biocompatible, Antibiotic
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Abstracts - Invited Talks
October 25th, 2019 9:00 am – 10:00 am Scientific program
Young Scientists Series Moderator: Dr. Maria Campos, University of Central Florida
Precision Vascular Delivery of Agrochemicals with Micromilled Microneedles (μMMNs)
Avra Kundu1, Maria Gabriela Nogueira Campos1, Swadeshmukul Santra1, 2, 4, 5 and
Swaminathan Rajaraman1, 2, 3, 5*
1NanoScience Technology Center (NSTC), University of Central Florida, Orlando, FL 32826, USA.
2Department of Material Science & Engineering, University of Central Florida, Orlando, FL 32826, USA.
3Department of Electrical & Computer Engineering, University of Central Florida, Orlando, FL 32826, USA.
4Department of Chemistry, University of Central Florida, Orlando, FL 32816, USA. 5Burnett School of Biomedical Sciences, University of Central Florida, Orlando, FL 32827, USA.
*[email protected] Abstract We demonstrate use of makerspace techniques involving subtractive microtechnologies to fabricate micromilled microneedles (μMMNs) of stainless steel (SS) for precise delivery of agrochemicals into vascular bundles of plant tissue. Precision delivery is of immense importance for systemic pathogen control in specific areas of plant tissue. Optimization of the micromilling allows for selective removal of SS at the microscale and the microfabrication of a 5×5 array of μMMNs having both base width and height of 500 μm to enable precise puncture into the stem of citrus saplings. Atomic Absorption Spectroscopy reveals up to 7.5× increase in the uptake of a therapeutic cargo while Scanning Electron Microscopy reveals that specific sites of the vascular bundle; either xylem or the phloem can be uniquely targeted with customized μMMNs. Such rapid and cost-effective customization with intricate designs along with scalability is enabled by makerspace microfabrication. Additionally, a 19×20 array of micromilled mesoneedles has been fabricated and affixed to a paint roller as an applicator system for real-world field testing outside the laboratory. Initial results indicate reliable behavior of the applicator system and the technique can be applied to the systemic delivery of agrochemicals while conserving the loss of the agrochemical with increased application efficiency.
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Huanglongbing on roots: early movement of 'Candidatus Liberibacter asiaticus' and disease progression.
Jeane Dayse V. S. PULICI1, Mayara M. MURATA1 and Evan G. JOHNSON1
1University of Florida, Citrus Research and Education Center, Lake Alfred, FL, USA; Abstract ‘Candidatus Liberibacter asiaticus’ (CLas), an alpha-proteobacteria associated with Huanglongbing (HLB) and transmitted by Diaphorina citri is the most devastating pathogen of citrus worldwide. In Florida, citrus production declined more than 70% for all commercial citrus varieties compared to pre-HLB production. Characteristic canopy symptoms of HLB are blotchy mottle, sectored yellowing, leaf drop, twig dieback, preharvest fruit drop and low fruit quality. On roots, 30-50% root loss is observed prior to canopy symptoms development. Understanding how CLas moves within the plant and local and systemic effects of CLas on the different tissues is fundamental to improve HLB management. Using split root rhizotrons and one-side graft inoculation below the trunk split in late summer was used to study the role of root infection on disease development. Root dieback increased on both halves of the root system in as little as 6 weeks, even though CLas infection remained isolated in the inoculated half of the root system until the spring flush (8 months after inoculation). More root growth occurred in the non-infected half of the root system of HLB-affected trees during late summer and fall root flush increasing live root length compared to healthy trees after each seasonal flush. Increased root dieback removed this gain by the next root flush. This suggests the previously reported root growth stimulation in HLB-affected trees results from non-infected roots within the infected root system. The first detection of CLas in canopies occurred in the spring flush (9 months after inoculation), with 6 canopies CLas-infected at the end of the experiment (11 months). The results showed that movement of CLas from infected root systems is linked to season, where it can move up and around the trunk after the spring flush. This split root inoculation provides the ability to study local and systemic effects of CLas infection on roots as observed with root growth stimulation that occurred in the healthy half of the root system and systemic root dieback as the first roots are infected. Systemic effects of CLas on the root system are not dependent on CLas movement, complicating tree health responses to therapeutic treatments.
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Enhancing in vitro growth of ‘Candidatus Liberibacter asiaticus’ by culture medium optimization
Marcus V. MERFA1, Leonardo DE LA FUENTE1
1Department of Entomology and Plant Pathology, Auburn University, Auburn, USA Abstract ‘Candidatus Liberibacter asiaticus’ (CLas), a heretofore unculturable bacterium, is associated with citrus huanglongbing (HLB), the current most devastating citrus disease worldwide. In the US, this bacterium has been causing extensive damages in citrus orchards, mainly in Florida. Because CLas has only been established in transient cultures, and due to the absence of methods to transfer it to either insect or host plants, fulfilment of Koch’s postulates and effective screening of antimicrobials are impaired, which hampers the study of HLB. Recently, the environments where CLas lives, including the hemolymph of the insect vector (Asian Citrus Psyllid – ACP) and phloem sap content of host plants, were characterized. This information could be used towards the goal of culturing this bacterium, since these studies have elucidated the chemical makeup of these environments. Thus, our objective was to use this knowledge to increase the viability of CLas in laboratory conditions. In this study, we aimed at optimizing grapefruit juice (GJ), the culture medium previously used by our group to establish transient cultures of CLas, to enhance its growth in vitro. In our assays, CLas inoculum was obtained from seeds of infected citrus plants or guts of infected ACPs, and was inoculated in 24-well plates containing GJ, used here as the base medium, and GJ modified by addition of different compounds. Plates were incubated at 28ºC for 21 days, and the amount of CLas was determined by RT-qPCR. To obtain replicates, sub-cultures of each condition were performed. Results showed that increasing the pH of GJ from 3.5 to 5.85 enhanced growth of CLas. Furthermore, a highly repeatable growth pattern was observed in which CLas grows better starting from a low number of cells, but grows only to a certain limit before the culture starts to die. These results add information to the effort of culturing CLas in vitro. We believe that reaching this goal will ultimately lead to a better understanding of CLas, so appropriate management strategies can be developed.
