Research Project: Exotic Whitefly Pests of Vegetables and Ornamental Plants

Location: Subtropical Insects and Horticulture Research

2019 Annual Report

Objectives
Objective 1: Investigate biological control and ecological interactions of whiteflies with their natural enemies using banker plant systems to promote environmentally sound control in vegetable and ornamental crops. Sub-objective 1b: Determine the compatibility of insecticide regimes with beneficial insects and natural enemies used in banker plant systems. Objective 2: Investigate structural, physiological, molecular and chemical aspects of the whitefly feeding process and identify inhibitor strategies/molecules such as but not limited to feeding disruptors and peptide inhibitors of disease transmission than can be used in the development of novel interdiction strategies envisioned to work either through production of transgenic plants or application of chemical treatments that block feeding/disease transmission. Sub-objective 2a: Develop transgenic tobacco expressing enzymatic inhibitors of whitefly salivary sheath formation test for resistance to Bemisia tabaci feeding. Sub-objective 2b: Test application of discovered small molecule inhibitors of sheath formation for their effect on whitefly feeding on tomato. Sub-objective 2c: Conduct proteomic analysis of salivary exudates to identify salivary sheath structural and biosynthetic proteins. Objective 3: Use molecular strategies to develop disease resistant banker plants to support large numbers of whitefly populations for production of biocontrol agents for use in the greenhouse.

Approach
Research will focus on constructing a nonflowering papaya banker plant through biotechnology that is resistant to both papaya ringspot virus and powdery mildew. Strategically timed insecticide applications including neonicotinoids in the rotation regimes will be evaluated against MED (Q-biotype whitefly) for whitefly efficacy and compatibility with two of the natural enemies used in our banker plant systems: the predatory mite, Amblyseius swirskii, and the whitefly parasitoid, Encarsia sophia. Development and testing molecular inhibitors of whitefly feeding processes with specific emphasis on the processes that must occur for the whitefly to develop a successful feeding event. Two main objectives include: 1) Continued characterization of the whitefly salivary sheath biosynthesis and composition. We already have basic compositional data and some knowledge on structural arrangement. Molecular, biochemical and structural analyses will continue to identify key biosynthetic enzymes and sheath structural components. 2) Evaluation of inhibitors of sheath formation as control agents. This evaluation will be performed using artificial diet assays and development of transgenic tobacco expressing inhibitors and conducting bioassays where the whitefly feed on the artificial diet or the transgenic plants.

Progress Report
The overall goal is to improve existing management strategies for Bemisia tabaci (B. tabaci) (MED or biotype Q) whitefly and stewardship of neonicotinoid class of insecticide (dinotefuran - grower standard). Currently, six non-neonicotinoid insecticide rotation regimes, based on efficacy of the high rate compatibility studies with biological control agents, were evaluated for whitefly control. Rotation regimes consisted of an application every 21 days (times 3) with an insecticide strategically placed in a rotation so that the mode of action was never replicated and the drench method of application was only used once in any rotation regime to manage insecticide resistance development. Rotation regimes with only non-neonicotinoid insecticides at the lowest label rate (abamectin, afidopyropen, cyantraniliprole, flupyradifurone pyrifluquinazon) overall provided good to excellent control of MED whitefly season-long compared to the untreated control. Drenches did not perform as well at the low rate compared to the high rate evaluated in previous studies. In the event the neonicotinoid class of insecticides are pulled from the market, growers are being provided with effective alternatives for whitefly management that are compatible with biological control agents and can be used at the lowest label rates applied as foliar sprays to minimize the overall cost of the rotation. In collaboration with University in Florida, ARS researchers are developing barcodes for predatory mites. Traditional taxonomy positively identified cohorts of each predatory mite population available from commercial biocontrol companies or occurring naturally using key morphological traits under stereomicroscope slide mount. After mite populations were validated for purity of species, DNA was isolated and genomic DNA libraries were constructed for 23 predatory mite populations representing 7 species: Phytoseiulus persimilis (5), Neoseiulus californicus (4), Neoseiulus cucumeris (4), Neoseiulus fallacis (2), Amblyseius swirskii (4), Amblyseius andersoni (3), Amblydromalus limonicus (1). Librairies have been sequenced using Illumina sequencing (PacBio sequencing was performed on one sample for development of a more complete reference scaffold) resulting in large volumes of raw sequencing data. Genome assembly is complete for 15 of the 23 libraries. Initial analysis of phylogenetic relationships among or between species using a reference population with the highest quality sequence library for each species and genes suitable for phylogeny construction has already identified taxonomic ambiguities among the populations and genera that are being evaluated is currently being conducted. Once all genomes are assembled, whole genome comparisons will be initiated so intra-population diversity can be evaluated to screen for potential cryptic species issues and purity of commercially available samples and for identification of diagnostic genome regions within and among species to develop and validate reliable Polymerase chain reaction (PCR) barcode strategies for quick evaluation of predator mite samples. The chilli thrips, Scirtothrips dorsalis, is a cryptic species complex (group of morphologically indistinguishable species) of at least nine distinct species, two (South Asia 1 and East Asia 1) of which exist in the United States. To determine their distribution range and find the dominant member of this thrips complex in the United States, a nationwide survey program is underway for the past four years. In FY19, we processed 17 samples (114 total thrips: 93 adults/21 larvae) from Georgia (3), Tennessee (4), Rhode Island (1) and Florida (9). Unforturnately thrips larvae are difficult to identify using either morphological or molecular techniques. Roses were the primary host plant sampled except in Georgia where thrips were sampled from Dystylium ‘Emerald Heights’, Aucuba ‘Goldstrike’ and blueberry and Florida where one sample was from Hemp. Five of the samples did not contain any Scirtothrips dorsalis and were mixtures of other thrips species including Thrips tabaci, Scolothrips sp., Frankliniella bispinosa, Selenothrips rubrocinctus, Aeolothripidas sp. and possible Franklinothrips. All adult thrips samples that were sequenced were South Asia 1 which is the predominant cryptic species to date. ARS researchers in collaboration with University in Florida and University in New York as part of a FARMBILL grant conducted a biological control study to compare the ability of Phytoseiulis macropilis and Neoseiulus californicus to manage the Glover mite (Tetranychus gloveri). Both predatory mites managed this mite but at the end of the experiment Chilli thrips populations were severely damaging the controls plants (Tetranychus gloveri and no predators) and the plants treated with P. macropilis. The plants treated with Neoseilus californicus had very little damage and almost no Chilli thrips. The mean number of immature thrips per 10 sampled leaves were 30, 1.6 and 41.1 for the untreated control, Neoseilus californicus and Phytoseiulis macropilis treatments, respectively. Based on these preliminary results, Neoseilus californicus is an excellent predator of chilli thrips and glover mite. ARS researchers in collaboration with APHIS, Florida Department of Agriculture and Consumer Services and the University of Florida initiated research on the phantasma scale, Fiorinia phantasma Cockerell & Robinson (Hemiptera: Diaspididae). Phantasma scale is a significant pest of at least 24 families and 44 genera of nursery and ornamental plants, particularly palms, as well as several fruit crops. The primary purpose of this FARMBILL grant is to support research for enhanced mitigation capabilities against the invasive, newly established armored scale by determining its current spread in south Florida as well as its potential plant host range, by improving our ability to properly identify this pest to species through the production of a taxonomic key, and by identifying effective biological and chemical control options. ARS researchers in an international collaboration with Brazilian scientists are evaluating new biological molecules for the control of the Bemisia whitefly. Together they demonstrated the effectiveness of a plant-derived peptide that is specifically toxic to insects and constructed transgenic plants expressing this peptide. Evaluation of these plants demonstrated that they induced mortality to several Lepidopteran insects that are economically important phytophagous pests in crops. Analysis of transgenic tobacco plants expressing this peptide are on-going. To support this work we have advanced two methods of delivering peptides directly to plant: 1) a direct vascular tissue infusion device was demonstrated to systemically delivery peptides to citrus trees where it induced a greater than 90% reduction in a phloem-limited bacterium after one week from a single application. This device is now being tested in other crop systems including tomato and potato; 2) The ARS scientists have developed a method of localized plant tissue transformation, where a specific tissue within the plant is transformed to produce desired molecules (peptides). In such plants only the modified tissue is transformed and we have demonstrated rapid systemic activity of peptides produced in this transformed “biofactory” tissue. This system is now available for use in evaluating potential biocontrol peptides of any type.

Accomplishments
1. Georgia survey of Bemisia tabaci. Bemisia tabaci (Gennaduis) is a large complex of cryptic species whose members are particularly invasive pests of hundreds of economically important commodities including cotton, vegetables and ornamental crops. In 2016, MED (Mediterranean; biotype Q) was detected in the field and residential landscapes for the first time in the United States. In 2018, a new state detection on poinsettia from a commercial Wisconsin greenhouse brought the total positive MED states to 27 indicating that MED is still expanding both its geographical and environmental ranges in the United States. With high whitefly populations that were difficult to control and new viruses in high dollar agricultural crops appearing across the southeast in 2016, growers were very concerned if MED had also moved into the field in Georgia. In collaboration with Universities of Georgia and Florida, ARS scientists in Ft. Pierce, Florida, conducted a whitefly survey in Georgia of agricultural ecosystems from field, greenhouse and nursery plants. Seventy samples were collected from 19 crops across 23 counties in Georgia with the bulk of the samples taken in 2016 and 2017. Overall, cotton was the most heavily sampled commodity (27) followed by seven samples of bell or ornamental pepper and six samples each of poinsettia, peanut, and squash. Other crops sampled included verbena, lantana, cucumber, soybean, cowpea, corn, snap bean, zucchini, kale, tomato, sweet potato, eggplant, cantaloupe, and mum. MED whitefly was detected on verbena in 2011 and poinsettia in 2012 at commercial greenhouses. All MED whitefly sequenced were identical and considered to have an Eastern Mediterranean origin. Only MEAM1 (Middle Eastern Asia Minor 1; biotype B) was detected in all the field grown commodities sampled in Georgia regardless of the year. This survey serves as a baseline for Georgia in the event that MED whitefly are eventually detected in the field.

2. First report of Bemisia tabaci MED (Q biotype) in the Dominican Republic. The first reports of Bemisia tabaci in the Dominican Republic (DR) date back to 1975 when the New World (NW) member of the complex (biotype A) caused severe damage in beans in the Southwestern provinces primarily due to transmission of bean golden mosaic virus. In 1994, MEAM1 (Middle Eastern Asia Minor 1, biotype B) was detected and quickly moved across the country spreading plant viruses and causing physiological disorders. Recent reports of insecticide resistance and increasing whitefly populations in tomatoes indicated Mediterranean (MED or biotype Q) whitefly may be present. ARS scientists in Ft. Pierce, Florida, and University of Florida in collaboration with the Instituto Dominicano de Investigaciones Agropecuarios y Forestales (IDIAF) conducted a whitefly survey to determine the distribution and composition of Bemisia tabaci cryptic species populations in the Dominican Republic. Sixteen samples were collected from 11 crops across 11 Provinces in the DR. Provinces included Azua, Distrito Nacional (2), Espaillat, Independencia, La Vega, Monseñor Nouel, San Juan (2), Santiago (2), Santo Domingo, Samaná, and Peravia (3). Host plants sampled included squash, eggplant, Asian spiderflower, mint, tomato, bean, tobacco, sesame, spurge, turkey berry, and Mexican prickly poppy. MED was detected for the first time in the DR from two different Provinces (Santo Domingo and Santiago) and host plants (tomato and tobacco) and environments (greenhouse and open field). MEAM1 was detected in all but one sample which was 100% MED. NW was detected twice on eggplant in different geographical regions and once on Mexican prickly poppy in a native habitat, but always with MEAM1. Widespread difficulties managing whitefly populations efficiently make additional confirmatory cryptic species detections necessary to determine the extent of the MED invasion in the country, especially where control failures have been reported.

3. A new method of delivering systemically active peptides to the vascular tissue of plants has been developed and demonstrated in tomato. This method allows the engineering of specific plant tissue that grows as a “biofactory” producing desired peptides that are secreted into the plant vascular system. ARS scientists in Ft. Pierce, Florida, have demonstrated systemic activity of peptides produced by localized biofactory tissue with this method. Using this strategy, only the “biofactory” tissue is transformed while the rest of the plant is non-transgenic. Systemic activity of biofactory-produced peptides can be visualized in as early as 3 weeks after initiation and provides a rapid way of screening for in planta bioactive peptide activity. It would also be a rapid way to apply transgenic technology to emerging problems in diverse crop genotypes without having to produce whole transgenic plants.

Review Publications
Taravati, S., Mannion, C., McKenzie, C.L., Osborne, L. 2018. Oviposition preference of rugose spiraling whitefly (Hemiptera:Aleyrodidae) on five host plant species. Florida Entomologist. 101(4):611-616. https://doi.org/10.1653/024.101.0423.
Taravati, S., Mannion, C., Osborne, L., McKenzie, C.L. 2019. Lethal and sublethal effects of selected systemic and contact insecticides on the whitefly predator, Nephaspis oculata (Coleoptera: Coccinellidae), in a tri-trophic system. Journal of Economic Entomology. 112(2):543-548. https://doi.org/10.1093/jee/toy364.
Avery, P.B., Kumar, V., Skvarch, E.A., Mannion, C.M., Powell, C.A., McKenzie, C.L., Osborne, L.S. 2019. An ecological assessment of Isaria fumosorosea compared to a neonicotinoid treatment for regulating invasive ficus whitefly. The Journal of Fungi. 5:36
Kumar, V., Kakkar, G., Palmer, C., Myers, W., McKenzie, C.L., Osborne, L.S. 2019. Chilli thrips, Scirtothrips dorsalis - small player with big damage. Acta Horticulturae. 1232:247-251. https://doi.org/10.17660/ActaHortic.2019.1232.36.
Sahayaraj, K., Subash, N., Allingham, R.W., Kumar, V., Avery, P., Mehra, L., McKenzie, C.L., Osborne, L.S. 2018. Lethal and sub-lethal effects of three microbial biocontrol agents on Spodoptera litura and its natural predator Rhynocoris kumarii. Insects. 9(3):101. https://doi.org/10.3390/insects9030101.
Sahavaraj, K., Kumar, V., Banu, N., Avery, P., Radhika, A., McKenzie, C., Osborne, L. 2018. Predation potential of Rhynocoris marginatus (Hemiptera: Reduviidae) against three mealybug species of agricultural importance. Applied Entomology and Zoology. https://doi.org/10.1007/s13355-018-0576-6.