Source: NORTH CAROLINA STATE UNIV submitted to NRP
PLANT VIRAL NANOPARTICLES FOR THE DELIVERY OF AGRONOMIC COMPOUNDS
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
HATCH
Reporting Frequency
Annual
Accession No.
1004987
Grant No.
(N/A)
Cumulative Award Amt.
(N/A)
Proposal No.
(N/A)
Multistate No.
(N/A)
Project Start Date
Jan 20, 2015
Project End Date
Sep 30, 2019
Grant Year
(N/A)
Program Code
[(N/A)]- (N/A)
Recipient Organization
NORTH CAROLINA STATE UNIV
(N/A)
RALEIGH,NC 27695
Performing Department
Plant Pathology
Non Technical Summary
Viruses are found in nearly all living organisms. They are simple infectious agents consisting of nucleic acid (DNA or RNA) wrapped in a protein coat and sometimes covered in a membrane. They are detrimental to agriculture when found in crops and the goal of our research is to understand how they do what they do so that we can find ways to eliminate/control them. We are focused on viruses that are found not only in North Carolina fields but throughout the world.The research we are conducting involves working with various scientists having different interests to tackle ongoing problems in a concerted, unified effort. We characterize the viral sequences to determine what proteins they are making and how they are different under differing conditions/circumstances. By knowing how the virus reacts, we can anticipate its next move and be one step ahead of it to eliminate it before it spreads uncontrollably. We are also working on exploiting certain viruses for doing good by using them to control other types of infections, namely tiny worms living in the soil called nematodes which are devastating to many crops as well as turfgrass (think golf courses, parks and athletic fields). The viruses that we are engineering for good are pumped full of chemicals which are toxic to the worms. Without the virus, the chemicals don't travel very far in the soil and a lot of chemical is used but is wasted. The virus allows us to use much less of the chemical to seep deep down into the soil where the worms reside to kill them. Some viruses are also found naturally in the worms and we are trying to understand their relationship with the worms in the hopes of turning them against the worms.
Animal Health Component
10%
Research Effort Categories
Basic
85%
Applied
10%
Developmental
5%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
21240301101100%
Knowledge Area
212 - Pathogens and Nematodes Affecting Plants;

Subject Of Investigation
4030 - Viruses;

Field Of Science
1101 - Virology;
Goals / Objectives
The major goal of this project is to elucidate the infection processes and physicochemical properties of viruses infecting plants and nematodes in order to develop unique control strategies based on our findings. The objectives to be undertaken involve three specific plant viruses and five viruses that infect soybean cyst nematode.Objectives:1. Develop Red clover necrotic mosaic virus (RCNMV) as a nanocarrier of agronomically important compounds2. Determine the thrips transmissibility determinant of Tomato spotted wilt virus (TSWV) through next generation sequencing of isolates and qRT-PCR of TSWV from thrips populations3. Investigate the seed transmissibility of Tobacco mosaic virus (TMV) in NC fields4. Elucidate the relationship between viruses infecting soybean cyst nematode (SCN) and their effects on the pathogenicity of SCN on soybeans
Project Methods
Experiments will be carried out with RCNMV as a nanocarrier of abamectin which has activity against root knot nematodes. The abamectin is infused into the virions to produce the formulated nanoparticle. The nanoparticle will then be assayed for activity against both C. elegans and root knot nematode in vitro and when placed with tomato seedlings. Turf core samples naturally infested with sting nematodes will also be tested with the nanoparticles. Efficacy will be determined by the decrease in galling of tomato roots or the healthy appearance of turf cores after treatment with the nanoparticles.The TSWV titer within the thrips vectors will be assayed via qRT-PCR to determine if there is a correlation between titers and transmission efficiency. The TSWV RNA genomes of various isolates will be sequenced from plants and thrips vectors to determine specific nucleotide changes that are responsible for phenotypic changes. These changes will be introduced into a reverse genetic system for TSWV infection (to be produced via Agroinoculation of the required viral components: L protein, nucleocapsid, NSs, a viral inhibitor of RNA silencing and the S RNA genome). This reverse genetic system will also allow us to rapidly screen various compounds for their antiviral activitiy.Total RNA from tobacco seeds will be extracted and subjected to RT-PCR and qRT-PCR to assay for the presence/absence of TMV. Sensitivity of the methodology will be determined based on the number of seeds required to produce a positive signal (if any). This assay will be more sensitive than the serological based methods currently employed and will allow definitive identification of the contaminating virus (if present) down to the specific sequence level.Soybean cyst nematodes (J2 stage) will be induced to take up silencing RNAs targeted to viral sequences with octopamine to determine if endogenous viruses are required for the pathogenicity of SCN. Also, artificial defective-interfering RNAs (containing the green fluorescent protein, GFP, gene) based on the ScNV viral sequences will be tested for its ability to replicate/express GFP in SCN after uptake/electroporation.

Progress 01/20/15 to 09/30/19

Outputs
Target Audience:Trained graduate student on experimental techniques/analysis of data. Changes/Problems:It is uncertain at this time whether I can find a collaborator willing to continue working on Objective 4. In the meantime, we've continued working on a project to look at using plant viruses for good. What opportunities for training and professional development has the project provided?1 PhD graduate student worked on Objectives 2. Graduate training for 3 students at Virginia Tech as a scientific mentor and as a member of their graduate committees on work in Objective 3. How have the results been disseminated to communities of interest? Nothing Reported What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? 1. Develop Red clover necrotic mosaic virus (RCNMV) as a nanocarrier of agronomically important compounds Determined that RCNMV was not economically viable as a nanocarrier for agricultural compounds due to high cost of production All work terminated 2. Determine the thrips transmissibility determinant of Tomato spotted wilt virus (TSWV) through next generation sequencing of isolates and qRT-PCR of TSWV from thrips populations qRT-PCR revealed a correlation with virus titer in individual thrips and strain/host combinations responsible for transmission efficiency prepared manuscript for submission detailing the qRT-PCR results 3. Investigate the seed transmissibility of Tobacco mosaic virus (TMV) in NC fields Project completed but collaboration with Virginia Tech researchers continues to examine seedborne transmission of TMV Published paper demonstrating that embryo infection rate correlates with emerging seedling infection rates with TMV 4. Elucidate the relationship between viruses infecting soybean cyst nematode (SCN) and their effects on the pathogenicity of SCN on soybeans Viruses appear to be commensal with no adverse effects on SCN pathogenicity Potential to develop an RNA virus-based expression system to study SCN biology

Publications

  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Ruark-Seward CL, Davis EL & Sit TL. 2019. Electropermeabilization-based fluorescence in situ hybridization of whole-mount plant parasitic nematode specimens. MethodsX 6, 27202728. https://doi.org/10.1016/j.mex.2019.11.009.
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Sherman MB, Guenther R, Reade R, Rochon D, Sit T & Smith TJ. 2020. Near atomic resolution cryo-electron microscopy structures of cucumber leaf spot virus and red clover necrotic mosaic virus: evolutionary divergence at the icosahedral three-fold axes. J Virol. 94 (2) e01439-19 DOI: 10.1128/JVI.01439-19.
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Ellis MD, Hoak J, Ellis BW, Brown JA, Sit TL, Wilkinson CA, Reed TD & Wellbaum GE. 2020. Quantitative real-time PCR analysis of individual flue-cured tobacco seeds and seedlings reveals seed transmission of Tobacco mosaic virus. Phytopathology 110, 194-205. https://doi.org/10.1094/PHYTO-06-19-0201-FI
  • Type: Journal Articles Status: Accepted Year Published: 2020 Citation: Ruark-Seward CL, Davis EL & Sit TL. 2020. Localization of viral and host RNA within soybean cyst nematode via fluorescence in situ hybridization. Exp Parasitol, accepted.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Pirzada T, Salam A, Vogel N, Mathew R, Guenther RH, Sit TL, Byrd MV, Pal L, Opperman CH & Khan SA. 2018. Sustainable crop protection management in sub-Saharan Africa through wrap & plant technology. Materials Research Society Fall Meeting & Exhibit, Boston, MA November 25-30.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Sit TL & Ma B. 2019. Evaluation of SP2700 antiviral activities in three model patho-systems. American Phytopathological Society Annual Meeting, Cleveland, OH August 3-7.
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Farias BV, Pirzada T, Matthew R, Sit TL, Opperman C & Khan SA. 2019. Electrospun polymer nanofibers as seed coatings for crop protection. ACS Sustain Chem Eng 7, 19848-19856. https://doi.org/10.1021/acssuschemeng.9b05200.


Progress 10/01/17 to 09/30/18

Outputs
Target Audience:Trained graduate students on experimental techniques. Changes/Problems:As mentioned previously, funding for Objective 1 research has ended and that project has been terminated. Objective 2 is mostly completed with only further analysis of the next generation sequencing data by a fellow faculty member. Objective 3 has been completed but I continue to collaborate on research related to the original objective. Objective 4 has been mostly completed save for a few wrapup experiments dealing with the in situ hybridization experiments. It is uncertain at this time whether I can find a collaborator willing to continue working on this project. In the meantime, we've started working on a newly funded grant to look at using plant viruses for good. What opportunities for training and professional development has the project provided?2 PhD graduate students worked on Objectives 2 and 4. Graduate training for 3 students at Virginia Tech as a scientific mentor and as a member of their graduate committees. How have the results been disseminated to communities of interest?Some results from Objective 3 were presented at the 48th Tobacco Workers' Conference, Myrtle Beach, SC January 16-18. Results from Objective 4 were presented at the NCSU Graduate Student Research Symposium, March 21, 2018 and at the NCSU Plant Pathology PhD Symposium, April 28, 2018. What do you plan to do during the next reporting period to accomplish the goals?Funding for Objective 1 was completed in 2/17. Project was terminated and personnel shifted to work on related project involving nematode control in Sub Saharan Africa. Objective 2 is complete save for further analysis of next generation sequencing data while graduate student finishes her dissertation research. Results from initial experiments have been prepared as a manuscript for submission. Objective 3 is complete as funding ended but research on seedborne transmission continues as a collaboration with Virginia Tech researcher Carol Wilkinson. I serve as a committee member for her 3 MS students and design/analyze experiments with them. Objective 4 is mostly complete with the graduation of the PhD student. A few small scale experiments on in situ localization studies of the various viruses within SCN will be completed and the results packaged in a manuscript for submission.

Impacts
What was accomplished under these goals? 1. Develop Red clover necrotic mosaic virus (RCNMV) as a nanocarrier of agronomically important compounds Determined that RCNMV was not economically viable as a nanocarrier for agricultural compounds due to high cost of production Pursued development of RCNMV as a nanocarrier for biomedical applications in association with NanoVector after promising early work Developed collaboration with Kentucky BioProcessing to produce scaleable quantities of RCNMV using their industrial scale processes Unfortunately, NanoVector folded due to lack of interest by pharmaceutical companies in utilizing nanotechnology (vs. immunolgics) 2. Determine the thrips transmissibility determinant of Tomato spotted wilt virus (TSWV) through next generation sequencing of isolates and qRT-PCR of TSWV from thrips populations Next generation sequencing revealed the presence of a 10 nucleotide sequence that was present in viral RNA isolated from plants but missing from viral RNA isolated from the thrips insect vector Overall tighter constraint on genetic variation observed in plant vs. insect hosts 3. Investigate the seed transmissibility of Tobacco mosaic virus (TMV) in NC fields Project completed but collaboration with Virginia Tech researchers continues to examine seedborne transmission of TMV Determined that levels of TMV infestation varied throughout any given seed pod (even between 2 halves of the same pod) TMV infestation results in the production of some smaller defective seeds (lack embryo) not found in healthy pods Majority of TMV is found within the seed coat (testa) of infested seeds TMV found to be present in the embryo of some infested seeds Germination studies of TMV infested seeds demonstrates the presence of TMV in roots and leaves of emerging seedlings demonstrating definitive seed transmission of TMV 4. Elucidate the relationship between viruses infecting soybean cyst nematode (SCN) and their effects on the pathogenicity of SCN on soybeans Viruses appear to be commensal Developed in situ staining technique of intact J2 nematodes with fluorescent DNA probes to visualize locations of viral accumulation

Publications

  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Ruark CL, Gardner M, Mitchum MG, Davis EL, Sit TL. Novel RNA viruses within plant parasitic cyst nematodes. PLoS One. 2018 Mar 6;13(3):e0193881. doi:10.1371/journal.pone.0193881.
  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Diepenbrock LM, Lundgren JG, Sit TL, Burrack HJ. Detecting Specific Resource Use by Drosophila suzukii (Diptera: Drosophilidae) Using Gut Content Analysis. J Econ Entomol. 2018 May 28;111(3):1496-1500. doi: 10.1093/jee/toy077.
  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Steckelberg AL, Akiyama BM, Costantino DA, Sit TL, Nix JC, Kieft JS. A folded viral noncoding RNA blocks host cell exoribonucleases through a conformationally dynamic RNA structure. Proc Natl Acad Sci U S A. 2018 Jun 19;115(25):6404-6409. doi: 10.1073/pnas.1802429115.
  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Guenther RH, Lommel SA, Opperman CH, Sit TL. Plant Virus-Based Nanoparticles for the Delivery of Agronomic Compounds as a Suspension Concentrate. Methods Mol Biol. 2018;1776:203-214. doi: 10.1007/978-1-4939-7808-3_13.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Pirzada T, Salam A, Mathew R, Guenther RH, Byrd M, Pal L, Opperman CH, Khan SA, Sit TL & Vogel N. 2017. Biodegradable seed wraps for sustained release of pesticides for crop protection in Sub-Saharan Africa. American Institute of Chemical Engineers (AIChE) Annual Meeting, Minneapolis, MN October 29 - November 3.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Ruark CL, Mitchum MG, Gardner M, Koenning SL, Opperman CH, Lommel SA, Davis EL & Sit TL. 2017. Viral infections of soybean cyst nematode. Regional Southeast Meeting of Nematologists, Raleigh, NC November 13.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2018 Citation: Ellis BW, Wilkinson CA, Brown JA, Ellis MD & Sit TL. 2018. Quantitative real-time PCR analysis of individual flue-cured tobacco seeds reveals novel insights into seedborne transmission of Tobacco mosaic virus. 48th Tobacco Workers Conference, Myrtle Beach, SC January 16-18.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2018 Citation: Ruark CL, Mitchum MG, Gardner M, Koenning SL, Opperman CH, Lommel SA, Davis EL & Sit TL. 2018. A Multi-Pronged Approach to Identify and Characterize Novel RNA Viruses within Soybean Cyst Nematode. NCSU Plant Pathology PhD Symposium, Raleigh, NC April 27
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2018 Citation: Ruark CL, Gardner M, Mitchum MG, Davis EL & Sit TL. 2018. Novel RNA viruses within plant parasitic cyst nematodes. NCSU Graduate Student Research Symposium, Raleigh, NC March 21


Progress 10/01/16 to 09/30/17

Outputs
Target Audience:Provided research training experience for undergraduate student during summer 2017 (May-August). Changes/Problems:As mentioned previously, funding for Objective 1 research has ended and that project has been terminated. Objective 2 continues but there were some graduate student mental health issues during the course of this reporting period that produced a prolonged impact on data analysis. Those issues have been resolved for the time being and she is making continued progress towards completion of her dissertation. Objective 3 has been completed but I continue to collaborate on research related to the original objective. I will seek funding from industry sources to continue the work, if possible. Objective 4 has been proceeding without much drama but the graduate student should be completing her dissertation within the next reporting period. It is uncertain at this time whether I can find a collaborator willing to continue working on this project. In the meantime, we've started working on a newly funded grant to look at using plant viruses for good. What opportunities for training and professional development has the project provided?PhD graduate students are working on Objectives 2 and 4. The undergraduate mentioned earlier worked partly on Objective 3. How have the results been disseminated to communities of interest?Results from Objective 2 were presented at the NCSU Plant Pathology PhD Symposium, April 28, 2017. Results from Objective 4 were presented to various national stakeholders at the 2016 Soybean Cyst Nematode Conference, Coral Gables, FL December 13-15, 2016. What do you plan to do during the next reporting period to accomplish the goals?Funding for Objective 1 was ended in 2/17. Project was terminated and personnel shifted to work on related project involving nematode control in Sub Saharan Africa. Objective 2 continues while graduate student finishes her dissertation research. Analysis of data ongoing while results from initial experiments are being prepared as a manuscript for submission. Objective 3 is complete as funding ended but research on seedborne transmission continues as a collaboration with Virginia Tech researcher Carol Wilkinson. I serve as a committee member for her 3 MS students and design/analyze experiments with them. Objective 4 continues with in situ localization studies of the various viruses within SCN.

Impacts
What was accomplished under these goals? 1. Develop RCNMV as a nanocarrier of agronomically important compounds Performed mobility studies on turf plugs to demonstrate that RCNMV is able to penetrate below the thatch layer However, RCNMV did not moderate damage to turf caused by sting nematodes in limited test at local golf course putting green test site Determined that RCNMV is not a good candidate for agronomic applications due to its high cost of production scale up Might be more amenable for biomedical research purposes as a carrier for medical compounds 2. Determine the thrips transmissibility determinant of TSWV through next generation sequencing of isolates and qRT-PCR of TWSV from thrips populations In the midst of analyzing the next generation sequencing data but have so far found possible correlations between point mutations in the TSWV NSs gene and efficiency of thrips transmissibility 3. Investigate the seed transmissibility of TMV in NC fields Project completed but collaboration with Virginia Tech researchers continues to examine seedborne transmission of TMV Determined that TMV source is maternal only, not pollen transmitted despite presence of TMV in pollen 4. Elucidate the relationship between viruses infecting soybean cyst nematode (SCN) and their effects on the pathogenicity of SCN on soybeans Viruses found in SCN do not appear to directly influence pathogenicity as PA3 strain (less pathogenic) has similar complement/levels of RNA viruses as more virulent strains of SCN 2 new SCN infecting viruses were discovered through transcriptome analysis

Publications

  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Ruark CL, Koenning SR, Davis EL, Opperman CH, Lommel SA, Mitchum MG & Sit TL. 2017. Soybean cyst nematode culture collections and field populations from North Carolina and Missouri reveal high incidences of infection by viruses. PLOS ONE 12(1), e0171514. doi:10.1371/journal.pone.0171514.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Guenther RH, Lockney D, Opperman CH, Kerns JP, Lommel SA & Sit TL. 2016. Plant virus capsids as facile medical and agricultural nanoparticles. Biochemistry Society Conference: Taming Plant Viruses - Fundamental Biology to Bionanotechnology, Pitlochry, UK November 8-10.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Ruark CL, Koenning SR, Davis EL, Opperman CH, Lommel SA, Mitchum MG & Sit TL. 2016. Incidence and titer of viral infections within soybean cyst nematode culture collections and field populations. 2016 Soybean Cyst Nematode Conference, Coral Gables, FL December 13-15.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Pirzada T, Salam A, Vogel N, Mathew R, Guenther RH, Sit TL, Byrd M, Pal L, Opperman CH & Khan SA. 2017. Biodegradable matrix for sustained release of pesticides for crop protection. American Chemical Society National Meeting & Exposition, San Francisco, CA April 2-6.


Progress 10/01/15 to 09/30/16

Outputs
Target Audience:Provided mentoring of Garrett Johnson, a senior undergraduate in Biochemistry who worked on a research project during the spring semester. Changes/Problems:As mentioned previously, the funding for Objective 1 will be terminated shortly and additional funding will be sought to continue the project. If unsuccessful, will transition personnel onto an alternative project. Objective 2 and 4 are progressing as expected based on graduate student capabilities despite #4 being an unfunded project. Biggest challenge for #2 is a technical one in terms of amount of RNA being extractable from individual thrips for library generation. What opportunities for training and professional development has the project provided?PhD graduate students are working on Objectives 2 and 4. The undergraduate mentioned earlier worked partly on Objective 1. How have the results been disseminated to communities of interest?Results of the research described in Objectives 1, 2 and 4 were presented at the Annual American Phytopathological Society Conference in Tampa, FL. Results of Objective 3 were disseminated to several extension agents working with tobacco in NC. What do you plan to do during the next reporting period to accomplish the goals?The funding for Objective 1 will be ending on 2/28/17 and that project will probably not be continued. Objective 3 being tobacco related will no longer be included as a part of this Hatch report. Objectives 2 and 4 will continue with #2 looking at next generation sequencing of viral RNA isolated from thrips and #4 working on virus searching from nematode transcriptomes (self generated as well as publicly available ones).

Impacts
What was accomplished under these goals? 1. Develop RCNMV as a nanocarrier of agronomically important compounds Developed new loading protocols to expand the range of compounds that can be loaded into virus particles Demonstrated that RCNMV binding to soil is dependent on pH and soil composition RCNMV loaded with abamectin offers some protection to soybean seedlings against soybean cyst nematode challenge 2. Determine the thrips transmissibility determinant of TSWV through next generation sequencing of isolates and qRT-PCR of TWSV from thrips populations Determined that viral titer in individual thrips is not the sole determinant of transmissibility 3. Investigate the seed transmissibility of TMV in NC fields Determined that outbreaks in NC were not seedborne due to differing strains found in outbreak locations Have shown in collaboration with Virginia Tech researchers that TMV can be transmitted via seed coat 4. Elucidate the relationship between viruses infecting soybean cyst nematode (SCN) and their effects on the pathogenicity of SCN on soybeans Have demonstrated that previously reported viruses are present in field and greenhouse populations of SCN Viruses are found in all stages of SCN development but not in any soybean tissues Same viruses are also found in clover cyst and beet cyst nematodes

Publications

  • Type: Journal Articles Status: Published Year Published: 2016 Citation: Cao J, Guenther RH, Sit TL, Lommel SA, Opperman CH & Willoughby JA. 2016. Development of abamectin loaded lignocellulosic matrices for the controlled release of nematicide for crop protection. Cellulose 23, 673-687.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Brown JA, Jacobson AL, Kennedy GG, Lommel SA & Sit TL. 2016. Variation in transmission of Tomato spotted wilt virus (TSWV) from Thrips tabaci is not solely influenced by virus titer within individual thrips. American Phytopathological Society Annual Meeting, Tampa, FL July 30 - August 3.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Guenther RH, Opperman CH, Zhu J, Lindbo DL, Kerns JP, Lommel SA & Sit TL. 2016. Enhanced nematicidal actitivity of abamectin via nanoparticle formulation. American Phytopathological Society Annual Meeting, Tampa, FL July 30 - August 3.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Ruark CL, Koenning SR, Mitchum MG, Davis EL, Opperman CH, Lommel SA & Sit TL. 2016. Soybean cyst nematode culture collections and field populations from North Carolina and Missouri reveal high incidences of infection by viruses. American Phytopathological Society Annual Meeting, Tampa, FL July 30 - August 3.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Thammarat P, Sit TL & Lommel SA. 2016. Devising a custom 13K unigene-based Nicotiana benthamiana microarray to study host-pathogen interactions at the transcriptome level. American Phytopathological Society Annual Meeting, Tampa, FL July 30 - August 3.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Pirzada T, Mathew R, Guenther RH, Sit TL, Byrd M, Pal L, Opperman CH, Khan SA. 2016. Biodegradable Matrix for Controlled Release of Abamectin for Crop Protection in Sub Saharan Africa. Bayer CropScience Research Symposium, Durham, NC August 31.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Farias BV, Corder RD, Pirzada T, Mathew R, Sit TL, Opperman CH, Khan SA. 2016. Nanofibrous polymeric seed coating for crop protection via electrospinning. Bayer CropScience Research Symposium, Durham, NC August 31.


Progress 01/20/15 to 09/30/15

Outputs
Target Audience:A poster was presented describing our research findings at the Gordon Research Conference - Nanoscale Science & Engineering for Agriculture & Food Systems held in Waltham, MA June 7-12. A presentation was also made of our findings via a poster huddle session followed by direct interaction with attendees at the annual meeting of the American Phytopathological Society held in Pasadena, CA Aug.1-5, 2016. Changes/Problems:Objective 3 is problematic only in the sense that the project is currently unfunded. Otherwise, research progress has been great. What opportunities for training and professional development has the project provided?The NIFA funded Objective 1 supported 2 undergraduate researchers that aided in plant propagation, virus infection/purification. Objective 2 was carried out by a PhD graduate student supported by a USDA grant while Objective 3, funded by the NC Tobacco Research Commission supported the research efforts of an undergraduate student who graduated during the research period. How have the results been disseminated to communities of interest?The findings from Objective 1 were disseminated at a Gordon Research Conference and the annual meeting of the American Phytopathological Society. What do you plan to do during the next reporting period to accomplish the goals?We plan to continue work on Objective 1 with additional collaborations for field tests and expanded host/pathogen combinations. For Objective 2, we plan on completing the analysis of the next generation sequencing results along with development of a reverse genetic system to directly determine the effect of any discovered sequence polymorphisms. The research for Objective 3 was funded for another year and we plan to continue working towards a washing protocol to disinfect seeds. At the same time, we are ramping up efforts to monitor TMV infection of the flower prior to seed formation to determine the exact site of viral replication within the primordial seeds. We are currently pursuing funding for Objective 4 through private sources as well as intending to submit a NIFA application next year.

Impacts
What was accomplished under these goals? Viruses infect every living organism on earth and may play hidden roles in the life cycle of these organisms. For plant infecting viruses, most infections are economically damaging to growers resulting in significant yield losses. Many such viruses are vectored by insects which themselves cause significant losses due to feeding. To prevent further losses due to viral infections, it is necessary to understand the processes involved in establishing/propagating viral infections within and among different plants and/or plant pathogens. However, viruses have evolved over millenia to occupy a certain environment such as the soil. By taking advantage of inherent properties of viruses, we can repurpose them for beneficial needs. 1. Develop Red clover necrotic mosaic virus (RCNMV) as a nanocarrier of agronomically important compounds. This research is funded by a USDA-NIFA Nanotechnology grant. We've submitted annual reports to the program director but will summarize our current findings here. RCNMV has been loaded with abamectin (Abm; a nematicide) to produce a plant viral nanoparticle (PVN). This PVN has been shown to move Abm within the soil beyond its normally abilities. We have demonstrated that PVN-Abm demonstrates bioactivity against root-knot nematode infection of tomato seedlings. We are currently testing PVN-Abm against sting nematodes in golf course putting greens and plan to test loading of fungicides. 2. Determine the thrips transmissibility determinant of Tomato spotted wilt virus (TSWV) through next generation sequencing of isolates and qRT-PCR of TSWV from thrips populations. This project is a collaboration with George Kennedy and Alana Jacobson. We have developed an extraction procedure followed by a qRT-PCR protocol for determining viral titer within individual thrips. There does not appear to be a direct correlation between viral titer in individual thrips and transmissibility. We are currently analyzing the data obtained from next generation sequencing of viral RNA from infected plant populations to look for any sequence polymorphisms. 3. Investigate the seed transmissibility of Tobacco mosaic virus (TMV) in NC fields. We have determined that the TMV strains collected from infected tobacco in Stokes and Yadkin counties are different. Hence, it is highly unlikely that they arose from the same source, namely contaminated tobacco seeds. We are currently working on development of a seed washing procedure to potentially inactivate any virus present on seeds. 4. Elucidate the relationship between viruses infecting soybean cyst nematode (SCN) and their effects on the pathogenicity of SNC on soybeans. This project is currently unfunded but steady progress has been made regarding the survey of SCN populations collected in NC fields as well as populations obtained from our collaborator (Melissa Goellner-Mitchum, Missouri State University). Viruses are present at various titers in most SCN populations. There does not appear to be a direct correlation between viral titers and the pathogenicity of a given SCN population. Furthermore, viruses were also found in clover cyst nematode, a near relative of SCN.

Publications

  • Type: Journal Articles Status: Published Year Published: 2015 Citation: Cao J, Guenther RH, Sit TL, Lommel SA, Opperman CH & Willoughby JA. 2015. Development of abamectin loaded plant virus nanoparticles for efficacious plant parasitic nematode control. ACS Appl Mater Interfaces 7, 9546-9553.
  • Type: Journal Articles Status: Under Review Year Published: 2015 Citation: Cao J, Guenther RH, Sit TL, Lommel SA, Opperman CH & Willoughby JA. 2015. Development of abamectin loaded lignocellulosic matrices for the controlled release of nematicide for crop protection. Submitted to Cellulose.