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Funded Project

Funding Program: Enhancement Grants - Special Projects

Project Title: Weeds as reservoirs of resistance breaking TSWV isolates in tomato and pepper systems of Georgia

Project Director (PD): Rajagopalbabu Srinivasan [1]

Lead State: GA Lead Organization: University of Georgia Research Foundation

Cooperating State(s): North Carolina, Washington

Undesignated Funding: $25,000

Start Date: Mar-01-2009 End Date: Feb-28-2010

No-Cost Extension Date: Aug-31-2010

Pests Involved: thrips,Tomato spotted wilt virus

Site/Commodity: Tomato and pepper

Area of Emphasis: vector-virus interactions, epidemiology

Summary: Tomato and pepper production in the southeastern United States has been severely impacted by Tomato spotted wilt virus (TSWV). For example, in 2000 TSWV caused an estimated $ 8.8 million loss in Georgia tomatoes alone. The wide host range of TSWV coupled with the presence efficient vector species viz., western flower thrips has made management of TSWV extremely difficult. The most efficient approach for managing TSWV in tomato and pepper in southeastern US is the use of resistant cultivars. A number of resistant cultivars are currently available and planted every year. These cultivars were obtained through breeding and all or most of them have the same gene (tomato Sw-5; pepper Tsw) conferring resistance to TSWV. Due to the wide host range of TSWV and its close biological association with its vectors (the virus multiplies in its thrips vectors) the virus is known to evolve rapidly, and resistance breaking strains have emerged frequently and affected the durability of resistant cultivars. Besides crop hosts, weed hosts in particular are reservoirs of TSWV. Research in North Carolina indicated that TSWV-resistance breaking isolates exist in weed reservoirs. No information on weeds as reservoirs of resistance breaking isolates exists in Georgia. Nevertheless, prevalence of reservoir weeds in Georgias agro-ecosystems and TSWV symptom development in some resistant cultivars indicates that resistance breaking strains might exist in Georgia. Through our transdisciplinary proposal, we intend to survey the existence of resistance breaking isolates in TSWV weed hosts in nine different counties (tomato and pepper production belt) in southwestern Georgia. Weeds will be evaluated as thrips reservoirs. TSWV isolates from various weed hosts will be assayed on tobacco, Nicotiana tabacum (L.) plants and their virulence will be analyzed. Subsequently, two highly virulent isolates will be evaluated against TSWV-resistant tomato and pepper cultivars through thrips inoculation. Resistance breaking isolates from North Carolina will also be evaluated against the same resistant tomato and pepper cultivars through thrips inoculation and their responses will be compared. Generated information will be useful to carry out additional in-depth studies aimed at understanding the unique and intricate interactions between thrips and resistance breaking isolates. In the following year, a grant proposal will be submitted to Southern region IPM (S-RIPM), addressing various aspects such as viz., 1) characterization of resistance breaking isolates from Georgia using molecular tools, 2) comparison of Georgia isolates with North Carolina isolates through phylogenetic analyses, 3) development of markers to screen for resistance breaking isolates, and 4) evaluation of weeds as reservoirs of resistance breaking isolates in the epidemiology of TSWV. Objectives: 1.Survey weed species as reservoirs of thrips and TSWV from nine different counties in the state of Georgia 2.Test the virulence of TSWV isolates from weeds on tobacco, Nicotiana tabacum (L.) plants and monitor symptom expression through mechanical inoculation 3.Evaluate highly virulent isolates on resistant tomato and pepper cultivars by thrips inoculation 4.Compare Georgia TSWV isolates to North Carolina TSWV isolates on the same tomato and pepper cultivars by thrips inoculation

Final Report:

Outputs This research proposal was initiated as a complementary project to a RAMP grant already awarded to CoPIs in this proposal (Drs. Riley and Kennedy). The RAMP project is involved in developing a reduced risk strategy to manage TSWV in the Southeastern US. The issue of resistance breaking strains in Georgia was not directly addressed in the RAMP project; this projects main goal was to fill that gap. Information pertaining to this SR-IPM seed grant was discussed in the RAMP workshop held at Southeast Fruit and Vegetable Conference at Savannah, January 2010. Information was also discussed during field days held as part of the RAMP project. Further information will also be posted on the TSWV website, www.tswv.org, this website is currently undergoing alterations to accommodate recent RAMP findings; information from this SR-IPM project will also be incorporated.

Outcomes The goal of this project was to assess whether weeds can serve as reservoirs of resistance breaking Tomato spotted wilt virus (TSWV) strains. TSWV-resistant cultivars are the most important management tactic currently available for TSWV management. Resistance in these cultivars is often governed by a major gene (Sw-5). Breakdown of major gene resistance can severely influence spotted wilt epidemics. Prior to this project no information was available on resistance breaking strains in Georgia and their influence on TSWV epidemiology. Surveys identified several winter and summer weeds as reservoirs of TSWV and thrips. Confirmation was performed through ELISA; but, subsequent mechanical inoculation of ELISA-positive weed tissue did not produce any TSWV-infection on an indicator host (Tobacco). Re-examination of the ELISA positive samples through Reverse transcriptase-polymerase chain reaction (RT-PCR) revealed the absence of TSWV in ELISA positive plants. RT-PCR with spiked RNA from TSWV-infected crop hosts produced amplicons of desired size indicating that there were no PCR inhibitory materials present in weed hosts. This information highlights the importance of detection tools and their influence on production of false positives. In order for a weed host to influence the epidemiology of a pathogen it has to be a host for the pathogen (TSWV) and also serve as a host for the vector (thrips). Weeds as reservoirs of TSWV: Four winter weed species, namely chick weed (Stellaria media (L.) Cyrillo), Carolina geranium (Geranium carolinianum, L.), spiny sowthistle (Sonchus asper (L.) Hill), and narrow leaf cudweed (Gamochaeta falcata Lam.) were selected and inoculated with TSWV through Frankliniella fusca from TSWV-infected peanut foliage. All the four weeds were infected with TSWV. Percentage of infection in weed species ranged from 10-61% reiterating that winter weed species can be infected with TSWV and can potentially serve as TSWV inoculum sources for the spring crops. Weeds as thrips reservoirs: In order for these weeds to serve as a TSWV inoculum source, the weeds should also serve as hosts for thrips that can transmit TSWV. TSWV is acquired by thrips larvae soon after egg hatching and in order for them to successfully transmit TSWV they have to complete an entire generation on a potential host plant. Our tests indicated that that F. fusca was able to complete an entire generation (adult  adult) on all winter weed species tested. Weeds as inoculum sources and their potential to affect TSWV incidence: The next logical factor that could influence TSWV epidemiology is the ability of competent vectors to back transmit the virus to crop hosts. Most of the experiments for this objective were conducted on peanut seedlings. Our results indicate that thrips can successfully transmit TSWV from weeds to crop host with very high inoculation efficiency (60-75%). TSWV N-gene diversity and potential for development of resistance breaking strains: TSWV isolates from resistant and susceptible cultivars of crop hosts (tomato, tobacco, pepper, and peanuts) were collected from at least nine different counties in South Georgia. The capsid protein gene (N-gene) of all isolated sequences was completely sequenced. Sequence analysis indicated that the TSWV isolates from various crops were identical to each other (>99%) both at the nucleotide level as well as at the amino acid level. These sequences were very similar to TSWV N-gene sequences already submitted from Georgia ten years ago. These findings suggest that the virus has not undergone any major changes in the last decade (at least in the gene tested). This also reiterates that the incidence of resistance-breaking strains,if any, is still very low and does not pose any immediate threat to resistant cultivars.

Impacts TSWV-resistant tomato and pepper cultivars are the most important management option available to vegetable growers in the Southeast. However, it is not unusual for a RNA virus, such as TSWV to adapt to such cultivars and overcome inherent plant resistance. This phenomenon has occurred in several places in the world, including North Carolina. Current research examined the diversity of TSWV among Georgia isolates and quantitatively evaluated the role of weeds in TSWV epidemiology. The results re-emphasize the need to follow management tactics diligently to avoid TSWV-induced losses. The main factors include the planting of resistant cultivars, at least when available and in high-incidence areas and reduction of weed populations in the field and nearby areas. Although, weed removal sometimes can lead to higher TSWV incidence, this aspect needs to be revisited with particular reference to timing of removal, and the coverage area. Planting resistant cultivars and proper weed management tactics could potentially lead to huge monetary gains in high-incidence areas. The absence of or lack of dominance of resistance breaking strains serves as an encouragement for growers to plant resistant cultivars whenever available and exploit them successfully, at least for the next few years.

Potential Impacts Weeds, given the favorable conditions can severely affect TSWV epidemiology. Though this has been known for a long time, this particular research attempted to quantify such an effect. Back transmission to crop hosts can produce up to 75% TSWV infection on crop hosts. This information could be used as a modeling parameter to indicate impending TSWV-infection for the ensuing spring crop. A risk index developed for peanuts has proven to be very effective in TSWV reduction. However, no such system is currently available for TSWV management in vegetables. Weed species and their relative TSWV and thrips carrying capabilities can be factored into temperature dependent TSWV risk prediction model (degree day model-Kennedy et al. NCSU). Such an effort would help to develop a risk index that can be used to forewarn or forecast an impending TSWV epidemic; this scenario can thus provide growers more options. Evaluation of diversity of TSWV isolates in the region did not provide any indication on presence of resistance breaking strains. This information further substantiates that the resistant cultivars are still viable and can be successfully used to combat TSWV.

Leveraged Funds Funds from SR-IPM grants made it possible to leverage research funds from other sources. Funds were obtained from the National Peanut Board in 2009 ($40,000), and in 2010 ($58,000) to address the issue of TSWV diversity and its potential impact on TSWV resistance. As a part of this effort, research grants were sanctioned by the Georgia Peanut Commission in 2009 ($5,000) and 2010 ($7,000). Funds were also obtained from the Tobacco commission ($4,000) to address some of the critical issues stated above. A full research proposal was also submitted to SR-IPM grant program (December 2009), but was not funded.

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