Project : USDA ARS

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Research Project: New Technologies and Strategies for Managing Emerging Insect Pests and Insect Transmitted Pathogens of Potatoes

Location: Temperate Tree Fruit and Vegetable Research

2022 Annual Report

Objectives
Objective 1: Develop new tools and approaches for examining landscape-scale movement by Hemipteran vectors and plant pathogens between non-crop plant species and potato fields. • Sub-objective 1A: Identify weedy plant sources of infective potato psyllids and beet leafhoppers entering potato fields of the Columbia Basin growing region. • Sub-objective 1B: Characterize genetic variation in beet leafhopper populations across geographic areas and between host species within the Columbia Basin and use these data to evaluate host-linked dispersal of leafhoppers into potato fields. Objective 2: Describe the biology of Hemipteran vectors of potato pathogens in crop and non-crop habitats, including reproduction and development, feeding ecology, chemical ecology, seasonal phenology, interactions with natural enemies, and transmission/acquisition of pathogens. • Sub-objective 2A: Characterize beet leafhopper feeding behavior and stylet penetration activities to examine acquisition and inoculation of BLTVA across host species. • Sub-objective 2B: Identify predator species important in reducing densities of potato psyllid and beet leafhopper in stands of weedy host plants. Objective 3: Develop new or improved integrated management strategies to control emerging insect pests and insect-transmitted pathogens of potatoes. • Sub-objective 3A: Produce a “risk matrix” that ranks non-crop weedy plants as to importance as sources of infective potato psyllid or beet leafhopper arriving in potato fields and forward those rankings to the potato industry.

Approach
Sub-objective 1A: Identify weed sources of psyllids and leafhoppers. Approach: Molecular gut content analysis will be used to identify plant DNA in insects and define their feeding histories. Both species will be collected as they enter potato fields. Specimens will be tested with PCR for presence of plant pathogens. Presence of a specific plant in insect guts and correlated presence of pathogen DNA will be evidence the plant is a source of infective insects. Contingencies: Plant DNA that cannot be identified to species based on representation in the NCBI database will be identified to genus. Sub-objective 1B: Characterize genetic variation in beet leafhopper populations across regions and host species. Approach: NextRAD sequencing will be used to identify genetically-defined leafhopper subpopulations collected from potato fields and weed hosts. Genetic differentiation among regions and plants will be assessed by analysis of molecular variance. Contingencies: NextRAD sequencing is time intensive which may make it difficult to evaluate all regions and host-sources. We will supplement NextRAD data as needed by analysis of the CO1 gene. Sub-objective 2A: Characterize beet leafhopper feeding behavior to examine acquisition and inoculation of plant pathogens. Approach: Electropentagraphy (EPG) technology to be used to examine how leafhopper feeding behavior affects pathogen acquisition and inoculation in cultivated and weedy hosts. We will record the time required to begin a feeding event and time spent in an event for three behaviors: xylem ingestion, phloem salivation, and phloem ingestion. Probability of pathogen acquisition and inoculation will be evaluated as a function of these time durations. Contingencies: If we encounter difficulties with the EPG assays, we will consult the literature on EPG work with other leafhoppers. Sub-objective 2B: Identify predator species that attack potato psyllid and beet leafhopper in non-crop habitats. Approach: Molecular gut content analysis will be used to identify predators feeding on potato psyllid and beet leafhoppers in non-crop habitats. Insects for molecular assay will be extracted from plant samples in Berlese funnels. The COI gene will be PCR amplified to detect psyllid or leafhopper DNA. We will identify which predatory taxa most readily attack potato psyllid and beet leafhopper by comparing presence vs absence of prey DNA across predator specimens. Contingencies: No difficulties in completing this work is anticipated. Sub-objective 3A: Produce a risk matrix that ranks weedy hosts by importance as sources of infective psyllids and leafhoppers. Approach: Rankings will color-code each plant species according to risk (red, yellow, or green). Host plants color-coded red will be those found to be sources of vectors and pathogens, and to be common in the study region. Rankings will be made available to growers at research meetings and publication in industry newsletter. We will include suggestions of how risk rankings can be used to assist IPM programs through monitoring of at-risk fields or by eradication of high-risk species. Contingencies: No difficulties in completing this work is anticipated.

Progress Report
In support of Objective 1, Sub-objective 1A, three years of sampling data for beet leafhopper on non-crop hosts were compiled and analyzed to identify seasonal changes in host use by beet leafhopper. Leafhoppers and plants were tested for presence of Phytoplasma trifolia and beet curly top virus using polymerase chain reaction (PCR). Molecular gut content analysis is being used to identify plants that the captured beet leafhopper adults had previously fed upon. Nymphal leafhoppers captured over the three-year sampling period were sorted by morphology, and PCR primers were designed to help identify beet leafhopper nymphs from indistinguishable nymphs of other species. To identify non-crop sources of infective potato psyllids, gut content analysis was performed on psyllid adults with and without the zebra chip pathogen that were captured on sticky cards placed in potato field in 2021. In support of Sub-objective 1B, beet leafhopper adults have been collected for DNA extraction and Rapid Application Development (RAD)-seq assays during March–September of 2021. Leafhoppers were collected from 20 locations within the Columbia Basin of Washington (from potato fields, sugar beet fields, coriander seed fields, and stands of kochia, Russian thistle, flixweed, and tumble mustard), two locations in southern Idaho (from a potato field, a sugar beet field, and a stand of kochia), and several locations within the Klamath Basin of Oregon (from potato fields). DNA from specimens is currently being extracted. Results suggest a lack of host-associated differentiation among beet leafhoppers collected off different host-plants species; however, there is some differentiation between Idaho and Columbia Basin leafhoppers. In support of Sub-objective 2A, two beet leafhopper colonies have been established and are currently being maintained. Beet Leafhopper-Transmitted Virescence Agent (BLTVA)-infected plants are also currently maintained in the greenhouse and BLTVA-infected potato tubers in storage. Due to the difficulties of keeping plants alive long enough for BLTVA titers to build while infested with beet leafhoppers, we are maintaining the insects and pathogen-infected plants separately. When needed for experimental assays, we will allow beet leafhoppers to feed on and acquire BLTVA from an infected plant. Additionally, work is underway to establish BLTVA in tissue culture. This would serve an additional source from which beet leafhoppers could acquire BLTVA prior to experimental manipulation. In support of Sub-objective 2B, predatory arthropods were collected from weedy host plants of potato psyllid at 10 locations in the potato growing region in the state of Washington. Molecular techniques were used to look for the presence of potato psyllid DNA in guts of the specimens. DNA of potato psyllid was detected in several taxa including minute pirate bug, ladybeetles, spiders, and, surprisingly, extensively in the guts of a whirligig mite (Anystis), a taxon which has received almost no attention as a predator of psyllids in general. These results confirm that the molecular approach developed in this sub-objective is a viable way of identifying predators of potato psyllid under field conditions and has unexpectedly identified Anystis as one of the more important species of predators that attacks potato psyllid in natural field conditions. In support of Sub-objective 3A, stands of matrimony vine were sampled for presence of potato psyllids from early March until late May. Data were used to improve the psyllid prediction model. Results were also provided to extension specialists and presented to growers in the Washington State University Potato Pest Alerts.

Accomplishments
1. Identification of a psyllid associated with a new Liberibacter and discovery of the psyllid’s plant host. A new genetic type of the pathogen (Liberibacter) that causes zebra chip in potato was discovered by an ARS scientist located in Prosser, Washington, in a common but unidentified species of psyllid in Washington State. ARS scientists in Wapato and Prosser, Washington, consulted taxonomic literature from Europe and North America, examined morphological traits of the psyllids, and used molecular analysis to identify the psyllids as a species of Aphalara. This is the first mention worldwide that Aphalara may harbor Liberibacter. The plant species listed in the literature as host of the psyllid is not known to be present in the study region and was therefore dismissed as a source of the psyllid and as a possible reservoir of the pathogen. Instead, sampling of plant species closely related to the listed host led us to identify an introduced species of knotweed as the host plant of the psyllid in Washington State. Research is now underway to determine whether the weed is also a reservoir of the new Liberibacter haplotype.

2. Newly discovered virus infects potato psyllid. Potato psyllid is a pest of potato as a vector of the pathogen that causes zebra chip disease. New methods that are safe for consumers and the environment are needed to effectively control potato psyllid and zebra chip disease. Researchers at the University of Idaho, along with ARS researchers in Wapato, Washington, discovered a new virus named Bactericera cockerelli picorna-like virus (BcPLV) that infects potato psyllids. BcPLV, along with a related virus that infects the Asian citrus psyllid, form a new genus which the researchers provisionally named Psylloidivirus. This discovery will allow researchers to test whether BcPLV provides biological control of potato psyllid to prevent zebra chip disease and reduce the need for chemical insecticides.

U.S. DEPARTMENT OF AGRICULTURE

Temperate Tree Fruit and Vegetable Research: Wapato, WA