Source: WASHINGTON STATE UNIVERSITY submitted to NRP
EFFECTS OF TRI-TROPHIC INTERACTIONS ON TRANSMISSION OF A LEGUME PLANT PATHOGEN
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
Reporting Frequency
Annual
Accession No.
1015667
Grant No.
2018-67012-28083
Cumulative Award Amt.
$165,000.00
Proposal No.
2017-07165
Multistate No.
(N/A)
Project Start Date
Mar 15, 2018
Project End Date
Mar 14, 2020
Grant Year
2018
Program Code
[A7201]- AFRI Post Doctoral Fellowships
Recipient Organization
WASHINGTON STATE UNIVERSITY
240 FRENCH ADMINISTRATION BLDG
PULLMAN,WA 99164-0001
Performing Department
Entomology
Non Technical Summary
Plant pathogens present a major challenge to agricultural production because disease outbreaks can be so costly to control and dramatically reduce crop yield. A large number of damaging plant pathogens are carried by insects and spread when these insects feed on crop plants. In the Palouse region of Washington State, a leading producer of legumes (dry pea, lentil, chickpeas), the "Pea Enation Mosaic Virus" or PEMV, is spread by Pea Aphids and can cause extensive damage to crops in outbreak years. Currently preventative sprays of pesticides targeting aphids are the only effective way to prevent outbreaks, but this is costly and negatively impacts environmental health. The research we outlined in this proposal will look at how food web interactions, such as predation by natural enemies and aphid use of weedy plants as alternative hosts, impacts the spread of PEMV within farms. This results will provide guidance on actions growers can take, such as biological control targeting ladybird beetles or weed control of non-crop plants that harbor PEMV.To reach these applied goals, we much employ a series of experiments with three fundamentally different approaches. First, we will need to do field surveys to see what host-plant species (both crop and non-crop plants) can harbor PEMV that may become sources of future outbreaks. Second, we need to do highly-controlled greenhouse experiments to quantify the impacts natural enemies and alternative hosts have on the Pea Aphid abundance and movement. Third, we will run a series of whole-farm experiments to test if biological control and weed management are effective ways to reduce the spread of PEMV within fields. In all, this series of experiments will provide invaluable information to growers and ecologists tackling the economically and environmentally important issues of insect and pathogen management in farms.
Animal Health Component
50%
Research Effort Categories
Basic
50%
Applied
50%
Developmental
0%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
21114121070100%
Knowledge Area
211 - Insects, Mites, and Other Arthropods Affecting Plants;

Subject Of Investigation
1412 - Peas (dry);

Field Of Science
1070 - Ecology;
Goals / Objectives
This research will examine biotic factors that affect the movement of pea aphids, which transmit a devastating viral pathogen in legume crops. Pea aphids are the primary vector for many damaging crop pathogens, and this project's objective is to better understand how agricultural communities affect aphids and viruses they transmit, which will allow us to improve predictive models for PEMV (pea enation mosaic virus) outbreaks. In our project we have outlined three objectives: (i) Employ field surveys to determine which alternative pea aphid host plants can harbor PEMV and the role of natural enemies and competitors in determining pea aphid population densities on those host plants. (ii) Use a series of greenhouse experiments to see how PEMV can spread from alternative hosts to crop plants. We will also quantify the contribution of natural enemies and competitors in mediating the spread of this virus. (iii) Develop and run a whole-farm experiment will measure direct impacts on plant performance in mixed plant experiment that also introduces biological control agents (ladybeetles). Each of these experimental approaches will tell us which host-plant species are reservoirs for PEMV, how quickly this can spread to crop plants, and what ecological factors determine the speed of spread at multiple spatial scales. In all, this will dramatically improve the ability to determine the source of PEMV outbreaks which is necessary to minimize their economic impacts.
Project Methods
For objective 1, our field surveys will leverage collaborations developed by mentor Crowder with scientists at the University of Idaho, who have organized a network of spring pea fields in the Palouse region that are sampled weekly for pea aphids and virus. This network, will consists of 15 commercial pea fields in each year, will form the basis of our observational studies. In each of these 15 pea fields, we will sample insect communities four times in 2018 in May, June, July, and August. We will use a DVac suction sampler to collect insects from 100 plants in each field at each time point. After collection, insects will be stored in the freezer until later identification. All predators and herbivores in the samples will be identified to species and their abundance will be recorded. Each plant will also be visually examined for characteristic PEMV symptoms, and leaf tissue will be collected for later confirmation of disease symptoms with ELISA. We will also sample the vegetation surrounding each pea field. Aphids are believed to acquire PEMV from native legumes, such as vetch and clover. We will digitize satellite photos within a 100m radius of each pea field and determine the area occupied by non-crop host plants. These data will be ground-truthed by PD Clark in July 2018. These data will allow us to determine the abundance of non-crop hosts near each pea field. Additionally, we will visually determine disease symptoms in any vetch plants we observe and collect leaf tissue for later disease confirmation using ELISA. We will use piecewise structural equation modelling to evaluate how predator, herbivore, and plant communities affect pea aphid abundance and the incidence of PEMV. By examining communities at four-time points, we will also be able to determine how variation in communities over the course of the season affects aphids and PEMV.For objective 2, we will conduct a greenhouse experiment to assess how tri-trophic interactions affect aphid movement and PEMV in microcosms. Preliminary data suggest that increased aphid plant-to-plant movement should accelerate the rate of pathogen transmission. Our experiment will be a 2 × 2 × 2 factorial design (Fig. 4) with host-plant variation, non-vector herbivores, and predators. Each experimental replicate will include two plants, a source plant where aphids are released, and a recipient plant placed 30 cm away (typical of field conditions, Fig. 4b). The recipient plant will always be pea, but the source plant will be either pea (representing in-field movement and transmission) or vetch (representing movement and transmission between crop and non-crop hosts). Each source plant will receive 20 inoculative adult aphids at the beginning of the experiment. After 24h of establishment, the aphids will be counted and then the recipient plant will be added to cages. At this time, the predator (2 lady beetles or 0 lady beetles) and herbivore (2 pea leaf weevils or 0 pea leaf weevils) treatments will be applied. Thus, the full experiment has 8 unique treatments (2 types of recipient plants [pea or vetch] × 2 predator treatments [lady beetle present or absent] × 2 herbivore treatments [weevil present or absent]). Each treatment will be replicated 10 times (totaling 80 experimental units), and the experiment will be replicated in two separate blocks, for a total of 160 experimental units in total. In each experimental cage, we will monitor the movement of aphids to the recipient plants over 72h, with a measurement each 12h. Only adults will be used in analyses of movement. At the end of 72h, we will apply a systemic insecticide (imidacloprid) to kill all aphids from plants and stop virus transmission. Fourteen days later, when virus symptoms can be identified, we will harvest the plant tissue of each recipient plant and check for PEMV infection using ELISA. We will use generalized linear models (Bolker et al. 2009) to evaluate the impacts of host plant, competitors, and predators on aphid abundance, movement (i.e., the proportion of adult aphids on the recipient plant), and PEMV incidence. Aphid abundance will be evaluated with parametric models, while movement and PEMV incidence will be evaluated using logistic models (as these response variables are binary). One of the key predictions of this experiment is that the presence of predators or competitors will increase the transmission of PEMV by increasing aphid plant-to-plant movement. If this hypothesis is verified, it would suggest a potential trade-off of biological control within PEMV fields; these results might also suggest that growers should manage pea leaf weevils to aid in managing the spread of PEMV.For objective 3, starting in May 2019, we will set up a 2 × 2 × 2 factorial experiment, with the same treatments described in Obj. 2 (i.e., variation in source plants, competitor presence, and predator presence) (Fig. 5). In each experimental cage, we will create a 5 × 5 array of plants. The plant in the center of this grid (i.e., the source plant) will be either a pea plant or a vetch plant, and the remaining 24 plants (i.e., the recipient plants) will all be pea plants. At the beginning of the experiment, we will release 40 inoculative adult aphids on the source plant. After 24h, we will count the number of aphids and apply the herbivore (10 pea leaf weevils present or absent) and predator (10 lady beetles present or absent) treatments. These densities of competitors and herbivores (0.4 per plant) and aphids (40 per plant) are typical of field densities in the Palouse region. Aphids, weevils, and lady beetles will then be allowed to interact within the field cages for the duration of the experiment. Every 2wk, we will count the number of aphids on 10-randomly selected plants in each cage (2 in each row - counting aphids on all 25 plants is logistically infeasible). We will also visually record disease symptoms. Plants will be grown through to yield, at which point they will be harvested. Plant dry weight will be recorded and the incidence of PEMV will be determined using ELISA to confirm prior visual observations. Aphid abundance will be evaluated with parametric models, while movement and PEMV incidence will be evaluated using logistic models (as these response variables are binary). As with Obj. 2, the key predictions of this experiment are that the presence of predators or competitors will increase the transmission of PEMV by increasing aphid plant-to-plant movement. However, the presence of vetch as a source plant would slow PEMV incidence due to the challenge of host-switching for aphids.

Progress 03/15/18 to 03/14/20

Outputs
Target Audience:For this project, our target audiences are entomologists working in agricultural extension, crop commissions, and pulse (legume seed crop) growers in the Palouse region of eastern Washington and northern Idaho. The research products have been presented at crop commission annual meetings, including the dry pea and lentil council, shepherds grain (Columbia basin grain growers association), and as part of the research review at the Washington grain commission. Our findings were also presented at the Washington State University (WSU) entomology department colloquium, making this information available to WSU faculty working in extension in Washington state. Some of the data generated by this project were donated to the "Aphid Tracker" extension support system run by University of Idaho, a web platform which openly reports insect outbreaks in the Palouse to aid growers in pest management. Furthermore, example data from this project was used in statistics workshops for graduate students and faculty at WSU through the Center for Environmental Research, Education, and Outreach (CEREO) to teach about using structural equation modeling and generalized linear mixed models in insect community ecology. Finally, this project also worked in concert with WSU College of Agricultural, Human, and Natural resources summer internship programs to provide research experiences for undergraduates at WSU, who then presented research findings at a campus-wide undergraduate research symposium. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Fellowship recipient Clark received training in multiple skills as part of the project goals, including structural equation modeling, time series statistics, extension support tools, undergraduate and graduate mentorship opportunities, attendance of regional and national entomology and ecology meetings, teaching experience in statistics courses, and guidance on technical writing for grants and peer-review manuscripts.Undergraduate research internships associated with Clark's work provided suppport and mentorship for three undergraduates at WSU in entomology and plant pathology. How have the results been disseminated to communities of interest?Events related to this project included invited talks at the US Dry Pea and Lentil Council in Moscow, ID, Shepard's Grain in Spokane, WA, and WA Grain Commission Research Review in Pullman, WA. At these events, Clark presented information on alternative host plant use by pest aphids and vectored pathogens that attack pulse crops. Services included in this project include mentoring two undergraduate research internships (summer 2019 and spring 2020) through WSU College of Agricultural, Human, and Natural Resources, including research experiences for undergraduates in extension. Experiments and statistical analyses completed during the duration of this project have contributed to multiple publications by the Crowder lab group. PI Clark published "Tri-trophic interactions mediate the spread of a vector-borne pathogen" in the journal Ecology in summer 2019. PI Clark also made contributions to projects on aphid-virus-legume research that has been published in the past year, including "Plant-mediated interactions between a vector and a non-vector herbivore promote the spread of a plant virus" in the journal Proceedings of the Royal Society B in summer 2019. The results of this research were to be presented in an invited talk at the Pacific Branch of the Entomological Society of America in spring 2020 (cancelled due to Covid19). What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? All the goals of this project have been completed as outlined above. For objective I, we completed 2 years of surveys in 2018 and 2019, identifying the food web components driving the prevalence of PEMV (pea enation mosaic virus) in plant populations. We discovered that the presence of an important alternative host, hairy vetch, was a critical pathogen reservoir at the landscape level. For objective ii, greenhouse experiments were completed in 2018, and further demonstrated that natural enemies can suppress aphid vectors, but the impacts of alternative hosts as a source of PEMV offsets this benefit. The results of this experiment are now published in the journal Ecology. For objective (iii), we completed fieldwork in 2018 and molecular assays on crop plant infection in 2019. We directly manipulated predator abundance in a whole-farm experiment at the Palouse Conservation farm in Pullman, WA, and manipulated plant diversity with an intercropping regiment of clover, vetch, and pea. Overall, we were able to rule out multiple potential weedy hosts for PEMV, highlighting the importance of vetches (genus Vicia) as a source of aphid-borne pathogens. We provided evidence that ladybird beetles are important components of conservation biological control that can reduce vector abundance and pathogen spread as long as weedy hosts are not in proximity to crop plants. These patterns were demonstrated at the greenhouse, farm, and landscape scale. This information is not integrated into a large decision support system for growers and further work will determine how monitoring of hairy vetch can be used to predict risk of pathogen outbreak up a month ahead of time.

Publications

  • Type: Journal Articles Status: Accepted Year Published: 2019 Citation: "Tri?trophic interactions mediate the spread of a vector?borne plant pathogen" Robert E. Clark Saumik Basu Benjamin W. Lee David W. Crowder, Ecology, Volume 100, Issue 11, September 2019


Progress 03/15/18 to 03/14/19

Outputs
Target Audience:This project has currently communicated results to twotargetaudiences: First, collection data from summer 2018 has been made available for decision support tools. We have contributed collection records if pest insects (pea aphids) and presence of plant pathogens (pea enation mosaic virus) has gone towards online tracking data. These openly available reportsserve the larger Palouse region's legume agricuture sector. Second, the results from two experimentswere presented at the 2018 Ecological Society of America annualmeeting in New Orleans, LA. A 30-minute talk presented findings in an ecologically releveant framework toprofessionalscientists. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?PI Clark attended a 1-week workshop on statistical modeling hosted by PR Stats LTD following professional development goals in proposal. Additionally, this grant has provided trainingfor four undergraduate studentsworking onfield experiments and laboratory work. This training includesskills such as field taxonomy (plants and insects), molecular detection of virus (rtPCR), and managment of greenhouse experiments. How have the results been disseminated to communities of interest?We have presented results from objective (ii) at a national conference (Ecological Society of America), and provided insect and pathogen data for online decision support tool resources. Further engagement of stakeholders in legume agricutural sector will occur in the next reporting year. What do you plan to do during the next reporting period to accomplish the goals?The goals of the next reporting period are focused on publication and outreach, with a minor fieldwork component to supplement current data. In terms of research, we currently plan to complete one more series of field surveys for objective (i) and analyze samples in the laboratory. Data analysis for objective (iii) needs to be completed. The results of objective (ii) will be prepared for publication. We are currently opening communication with stakeholders and plan to dissimenate these findings to organizations such as the Dry Pea & Lentil Council in the next reporting period.

Impacts
What was accomplished under these goals? The three experimental objectives, outlined above, include:(i) field surveys, (ii) greenhouse experiments, (iii) whole-farm experiments.All fieldwork and labwork has been completed forobjectives (ii) and (iii), and we are currently analyzing data and preparing two manuscripts. Objective (i) has completed a single field season, but requires additional field data in 2019.

Publications