Epidemiology of the aster yellows phytoplasma: the influence of non-crop hosts on geographic distribution and movement of the pathogen

Final Report for GNC08-091

Project Type: Graduate Student
Funds awarded in 2008: $9,873.00
Projected End Date: 12/31/2010
Grant Recipient: UW-Madison
Region: North Central
State: Wisconsin
Graduate Student:
Faculty Advisor:
Russ Groves
UW-Madison
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Project Information

Summary:

This project is directed at enhancing our present understanding of the epidemiology and biogeography of the aster yellows phytoplasma (AYp) with a focus on factors that influence the pathogen’s geographical distribution and spread. Consistent with the objectives originally articulated in the SARE proposal, we developed molecular diagnostic tools to ensure AYp detection was accurate and precise, and characterized the phenology of the aster leafhopper (ALH), at multiple spatial and temporal scales to better understand which habitats surrounding carrot fields have the greatest epidemiological importance as pest or inoculum sources. We found that AYp titer, expressed as Log10 (copies/insect), ranged from 3.53 ( ± 0.07) to 6.26 ( ± 0.11) and increased approximately 100-fold in insects that acquired the AYp. ALH variability among and within years exceeded estimates of variation among farm locations and fields and time covariates explained the largest amount of variation of infectivity. Years in which high ALH abundance occurs co-incidentally with high infectivity can result in 1000-fold greater exposure of the carrot crop to infectious leafhoppers when compared to years in which low leafhopper abundance is co-incident with low infectivity. Similarly, the exposure of the carrot crop to infectious ALHs varies as much as 30-fold throughout the growing season. Thus, without information about insect abundance and infectivity for a specific field, the coincidence of these expected periods of high ALH abundance and infectivity represent a timing interval in which management of the insect could be focused to limit pathogen spread.

Introduction:

In Wisconsin, aster yellows management has focused primarily on controlling the insect vector, the aster leafhopper (ALH), and an AY risk index, known as the aster yellows index (AYI), was developed to describe the maximum allowable numbers of infectious leafhoppers during a discrete time period when plant protection is most needed. The AYI metric is the product of aster leafhopper infectivity, or percent of infectious aster leafhoppers, and the magnitude of the aster leafhopper population. Originally, the AYI was used to make insecticide spray recommendations based on a series of early season leafhopper collections, but following the observations that aster leafhopper abundance and infectivity in and around carrot fields varies, efforts were made to estimate the AYI for specific fields and dates. Contemporary tools (i.e. PCR) are currently used to detect the pathogen in the ALH. However, the relationship between pathogen presence in the vector and the vector’s ability to successfully transmit the pathogen is not known. In turn, many producers avoid risk of pathogen spread by using inexpensive, prophylactic insecticide applications, a management practice that circumvents the utility of the AYI.
Although successful from the perspective of managing insect pests in a cost-effective manner, this approach presents considerable risk, since these insecticides are broad-spectrum compounds with documented mammalian toxicity. The chemicals in this group are also harmful to aquatic organisms, are lipophilic, and in aquatic environments, tend to adsorb to organic sediments. These circumstances have prompted concerns about pyrethroid exposure to non-target areas, especially ecologically sensitive areas such as wetlands, which include the low-land, organic muck soils where the majority of Wisconsin carrot is grown. Thus, it has been our goal to reduce the nearly exclusive reliance on synthetic pyrethroid insecticides for the management of aster yellows in carrot. Specifically, we sought to improve grower adoption of reduced-risk (RR) insecticides by targeting insecticide applications to periods during which AY risk is high thereby reducing the number of applications of the more expensive RR insecticides necessary to control aster yellows improving the cost-efficiency of these newer tools. However, reliable information about ALH abundance and infectivity for specific fields and the coincidence of these expected periods of high insect population sizes and high rates of infectivity were not known.
The primary goal for completing the outlined objectives was to advance our understanding of the epidemiology of aster yellows in Wisconsin towards the development and implementation of a comprehensive disease and pest management plan for carrot. Our approach essentially tried to address the two problems described above by developing the molecular tools to accurately determine the rate of infectious individuals in a field population of ALH and analyzing historical ALH scouting data to identify seasonal trends in factors associated with periods of elevated AY risk.

Project Objectives:

The primary goal for completing this project was to advance our understanding of the epidemiology of aster yellows in Wisconsin towards the development and implementation of a comprehensive disease and pest management plan for carrot. The objectives originally articulated in the SARE proposal were to:
1) Accurately identify of the primary reservoir hosts of AYP in habitats surrounding carrot fields and determine which have the greatest epidemiological importance as potential inoculum sources,
2-A) Compare the genetic structure of the population of AYP isolates collected from reservoir hosts and within affected carrot, and 2-B) determine if AYP variability relates to either disease prevalence or infectivity (virulence) of the pathogen.

Consistent with the overall goals of this project, we have continued to address objectives 1 and 2 in the proposal although greater focus has was placed on the development of molecular diagnostic tools to ensure aster yellows phytoplasma (AYp) detection is accurate and reflected the underlying biology of the aster yellows disease system (Objective 2). Complimentary to objective 1, we continued to examine the phenology of the aster leafhopper (ALH), at multiple spatial and temporal scales to better understand which habitats surrounding carrot fields have the greatest epidemiological importance as pest or inoculum sources in the agricultural landscape.

Cooperators

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  • Dr. Russell Groves

Research

Materials and methods:

A combination of molecular and analytical tools were developed to complete the outlined objectives. Full descriptions of these methods have been published or are currently in press (see publication list below). Interested parties can contact us for reprints and details. In brief we:

  • Developed a qPCR assay for the quantification of the aster yellows phytoplasma (AYp) in its insect vector, Macrosteles quadrilineatus, and characterized AYp growth pattern and titer within the leafhopper for approximately 9 days after acquisition.
    Completed a detailed analysis of the capabilities and limitations of the qPCR assay that examined the variation of qPCR calibration curves, defined limits of detection of the assay and modeled AYp growth in the insect vector.

    Described an analysis that used a multi-year, multi-location observational pest scouting data set to obtain information about the scale at which ecological factors were contributing to the variability of leafhopper abundance and infectivity occurred.
    Deduced seasonal “windows” of elevated risk for spread of the aster yellows phytoplasma to susceptible crops.
    Demonstrated the utility of long-term data sets for improving our understanding of the spatial and temporal patterns of variation of insect abundance and infectivity.
    Discussed the use of generalized linear mixed models and generalized additive mixed models for the analysis of long-term ecological data.

Research results and discussion:

Again, full descriptions of our results have been published or are currently in press (see publication list below). Interested parties can contact us for reprints and details. In brief we found:

  • Average AYp titer, expressed as Log10 (copies/insect), ranged from 3.53 (±0.07) to 6.26 (±0.11) at 1 and 7 days after the aquisition access period.
    AYp titers per insect and relative to an ALH chromosomal reference gene, cp6 wingless (cp6), increased approximately 100-fold in insects that acquired the AYp.
    High quantification cycle values obtained for ALHs not exposed to an AYp-infected plant were interpreted as background and used to define a limit of detection for the qPCR assay.
    Aster leafhopper variability among and within years (39% of the total variation) exceeded estimates of variation among farm locations and fields (7%).
    Time covariates explained the largest amount of variation of aster leafhopper infectivity (50%).
    Leafhopper abundance has been decreasing since 2001 and reached a minimum in 2010.
    The average seasonal pattern indicated that periods of above average abundance occurred between 11 June and 1 August.
    Annual infectivity appears to oscillate around an average value of 2% and seasonal periods of above average infectivity occur between 19 May and 15 July.
Participation Summary

Educational & Outreach Activities

Participation Summary

Education/outreach description:

Peer-reviewed publications:

Frost, K.E., Esker, P.D., Van Haren R., Kotolski, L., and R.L. Groves. 2012. Seasonal trends of Macrosteles quadrilineatus (Hemiptera: Cicadellidae) abundance and aster yellows phytoplasma infectivity in Wisconsin carrot fields. Journal of Environmental Entomology (in Review).

Frost, K.E., Esker, P.D., Van Haren R., Kotolski, L., and R.L. Groves. 2012. Factors influencing aster leafhopper, Macrosteles quadrilineatus (Hemiptera: Cicadellidae), abundance and aster yellows phytoplasma infectivity in Wisconsin carrot fields. Journal of Environmental Entomology (in Review).

Frost, K.E., Willis, D.K., and R.L. Groves. 2011. Detection and variability of aster yellows phytoplasma titer in its insect vector, Macrosteles quadrilineatus (Hemiptera: Cicadellidae). Journal of Economic Entomology 104(6):1800-1815.

Abstracts/Presentations:

Frost, K.E., Willis, D.K. and R. L. Groves. 2010. Variation in aster yellows phytoplasma (Candidatus Phytoplasma asteris) titer in its insect vector, Macrosteles quadrilineatus. Entomological Society of America Annual Meeting. Dec. 9-15, 2010, San Diego, CA.

Frost, K. E., Groves, C.L., and Russell L. Groves. 2009. Refining the aster yellows index in Wisconsin: developing sustainable control tactics for susceptible vegetable crops [abstract]. Phytopathology 99(6S):S38.

Frost, K. E., Willis, D.K., Groves, C.L., and Russell L. Groves. 2009. Using real-time PCR to quantify aster yellows phytoplasma in its insect vector; relationship of infectivity to transmissibility in the aster leafhopper, Macrosteles quadrilineatus [abstract]. Phytopathology 99(6S):S38.

Extension Proceedings:

Rogers, P.M., Stevenson, W.R., Wyman, J.A., Frost K.E. and R.L. Groves. 2011. IPM Perspectives for Carrot Foliar Diseases in Wisconsin. University of Wisconsin – Extension, publication A3953.

Frost, K.E., Van Haren, R. and R. Groves. Historical variation in leafhopper abundance and aster yellows phytoplasma infectivity: assessing periods of elevated risk. [proceedings] Wisconsin Potato and Vegetable Growers Association, Inc. Annual Grower Meeting & Convention. February 1-3, 2011, Stevens Point, WI.

Frost, K.E. and R. Groves. Overwintering non-crop sources of aster yellows phytoplasma. [proceedings] Wisconsin Potato and Vegetable Growers Association, Inc. Annual Grower Meeting & Convention. February 2-4, 2010, Stevens Point, WI.

Trade Journal Publications:

Frost, K., Gevens, A., Van Haren, R, Miller, P., and R. Groves. 2012. Refining Pest Management Programs in Processing Carrots: New Technologies for an Old Problem. In Carrot Country. Columbia Publishing & Design, Yakima, WA. Summer 2012, p. 14-15.

Szendrei, Z., Frost, K., and R. Groves. 2010. Aster Yellows can distort carrot growth. In Vegetable Grower News [online]. Great American Media Services, Sparta, MI. March 2012.

Groves R. and Frost, K.E. 2010. Towards an Improved Understanding of the Aster Yellows Phytoplasma. In The Badger Common’tater. Wisconsin Potato and Vegetable Growers Association, Inc. July 2010.

Szendrei, Z., Frost, K., and R. Groves. 2010. Aster leafhopper index: what is it and how do you use it? In Carrot Country. Columbia Publishing & Design. Summer 2010.

Project Outcomes

Project outcomes:

The purpose of this work has been to increase the sustainability of carrot crop production in Wisconsin by improving production practices. Sustainability goals included reduction of foliar-applied pesticides to decrease cost, environmental impact, and exposure to farm workers. It also sought to enhance the economic return experienced by growers by lowering input costs while maintaining the yield and quality of the carrot crop.

Our work has contributed to the growing body of research of phytoplasma replication in their insect host by describing the AYp growth pattern and titer variation among individual ALH. A primary contribution of our work was to demonstrate qPCR as a reliable and accurate method for measuring AYp titer in ALHs and detecting differences in AYp titers among insect individuals which is consistent with objective 2 of the original grant proposal.
In our work, we have also examined aster leafhopper phenonlogy in Wisconsin to produced new information about the occurrence of seasonal trends in factors associated with elevated risk for AYp spread. A primary contribution of our research in this area has been to demonstrate the utility long-term data sets for improving the understanding of the spatial and temporal patterns of variation of insect abundance and infectivity. However, we also have highlighted and discussed the use of contemporary statistical methodologies including generalized linear mixed models and generalized additive mixed models for the analysis of these types of long-term ecological data sets. These data are currently being used as a benchmark to further refine current pest control strategies, reduce the number of pesticide applications and promote the use of reduced risk, less broad spectrum insecticides for the control of aster yellows. To date, we have deduced seasonal “windows” of elevated risk for spread of the aster yellows phytoplasma to susceptible crops and proposed changed
Manuscripts detailing this work are either published or in review as described below.

Project outcomes to date have been consistent with project goals of developing a multidisciplinary approach for the management of AYP in the carrot producing regions of Wisconsin and are complimentary to a core goal of the NCR-SARE program of enhancing environmental quality through a reduction of insecticide inputs.

Economic Analysis

As part of our continued work, we are examining the operational, technological, and economic feasibility of our proposed revision to the insecticide program currently used for the control of aster yellows in the Wisconsin carrot crop. Specifically, we are comparing the response of the aster leafhopper, Aster yellows disease symptoms, and yield and quality, of the carrot crop associated with the proposed revisions to the AY management program when compared to the current insecticide program. We are also using historical pest scouting data to compare the cost structures associated with the different management programs, including the costs associated with the new insecticide products and application technologies. Additionally, the historical pest scouting data is being used to examine the value of having management options. For example, the ability to choose the most cost-effective program for a given year can minimize plant protection costs in the long term.

Farmer Adoption

Not currently applicable.

Recommendations:

Areas needing additional study

Improvements to AYp detection will continue to be important to AY management. An accurate estimate of the rate of infectious aster leafhoppers in a population is needed to accurately and quickly prescribe an insecticide application for protection of the carrot crop. Research is needed to refine the molecular detection methodologies so that site-specific insecticide recommendations can be prescribed with greater accuracy.

Advancing predictive tools to address the sporadic occurrence of AY ‘risk intervals’, can help to minimize costs associated with unwarranted pesticide applications and reduce yield losses due to advanced preparation for aster leaf hopper infestations. Continued research on the impact of weather on aster leafhopper development can begin to identify geographically defined regions in the Midwest where winged insects are present and simulated air parcel trajectories can be used to develop models for predicting aster leafhopper dispersal and deposition in the environment.

Any opinions, findings, conclusions, or recommendations expressed in this publication are those of the author(s) and do not necessarily reflect the view of the U.S. Department of Agriculture or SARE.