1a. Objectives (from AD-416):
1. Prevent invasions of chilli thrips and whiteflies by developing genetic tools to track/predict/monitor and ultimately aid in controlling movement from likely sources of infestation and suppress established populations with biologically-based and environmentally-sound techniques. 1a. Determine the global sources of invasive chilli thrips, Scirtothrips dorsalis and describe the global genetic diversity of this pest species so that effective international and national barriers can be erected to further introductions. 1b. Investigate biological control and ecological interactions of chilli thrips and whiteflies with their natural enemies in order to promote their environmentally sound control in vegetable and ornamental crops 2. Investigate structural, physiological, molecular and chemical aspects of the whitefly feeding process and identify inhibitor strategies/molecules that 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. 2a. Characterize the method of polymerization that occurs when whitefly saliva is secreted and solidifies during insect probing for the plant vascular tissue. 2b. Determine the structural composition of salivary sheaths and the source of the precursors. 2c. Identify small molecule inhibitors of the whitefly feeding process using the artificial diet system as a bioassay.
1b. Approach (from AD-416):
Research will focus on establishing a DNA barcoding reference data set for S. dorsalis that will allow construction of a robust global molecular phylogeny for this species using both mitochondrial cytochrome oxidase I (mtCOI) and nuclear genes to be identified. A commercially available predatory mite (Amblyseius swirskii) will be evaluated for sustainable control of chilli thrips, whiteflies and other key pests in ornamental and vegetable crops using banker plants and ‘predator-in-first’ tactics. Molecular and biochemical methods will be used to develop a model describing whitefly salivary sheath formation and to identify inhibitors of this process that can be evaluated for use in control strategies.
3. Progress Report:
India is thought to be the likely geographical origin of chilli thrips, Scirtothrips dorsalis. To help determine the phylogenetic placement of this species within the genus thirty variants of a mitochondrial gene from chilli thrips collected in India were sequenced. Primers were then developed that improved the success rate for amplifying and sequencing this gene from a greater diversity of chilli thrips samples globally. Chilli thrips mitochondrial haplotypes from Japan are more than 14% divergent from other chilli thrips samples and are consistent with findings from morphometric analysis of five geographically distinct populations. Eleven varieties of hot pepper and 17 varieties of sweet pepper were screened for their ability to sustain the predatory mite, Amblyseius swirskii, with/without pollen. Five hot peppers sustained an average of 8-12.7 mites per seedling for 70 d post release. Among 17 selected sweet pepper varieties, 8 maintained the most mites and 9 varieties effectively sustained mites when supplemented with pollen. Overall, four varieties maintained the higheset predatory mite populations for at least 45 d averaging 5.5 mites per plant. Functional response studies showed that the predatory mite, Amblyseius swirskii, reared on different diets (peach, cattail, ornamental pepper pollen, and other prey) was equally effective against chilli thrips nymphs. Among these diets, the maximum daily predation was 35-43 nymphs of chilli thrips/day by an A. swirskii female. The findings improve the ability to rear predatory mites for biological control against multiple pests in vegetable and ornamental production. Tomato yellow leaf curl virus and its vector, the sweetpotato whitefly, threaten the economic value of greenhouse tomato production. We demonstrated the use of Papaya plants could be used as a banker plant to conserve wasps that, in turn control this whitefly. Our lab also showed that ‘Maradol’ papaya is not a host for tomato yellow leaf-curl virus strengthening the case for growers to use the papaya banker plant system for whitefly control. We have successfully developed an optimized strategy to use Flinders Technology Associates DNA storage card technology to archive DNA from arthropod samples using a new quick-boil method that recovers DNA from the card in five minutes for less than $0.01 per reaction. Research on B. tabaci saliary sheaths was problematic due to inability to obtain pure sheaths. Complications in the isolation procedure led to extended research on new method of purification. A novel method for B. tabaci salivary sheath isolation was developed using soluble membranes through which whitefly pierces and produces a salivary sheath that remained attached to the membranes. Large quantities of pure salivary sheaths were obtained using this method in combination with a filtration strategy. This method worked for numerous plant feeding hemipteran insects and allows isolation of large quanties of salavary sheaths per compositional structural and biosynthetic analysis. Initial salivary sheath composition analysis revealed a structure primarily composed of a specific carbohydrate polymer with protein crosslinks.
1. North American Bemisia biotypes after the Q invasion. The appearance of biotype Q in the U.S. in 2004 was associated with reports from ornamental growers of increasing problems in controlling whitefly infestations. As part of an APHIS-coordinated multi-state, -industry, -commodity, -agency and -institutional Q biotype Task Force initiative, a coordinated whole-country survey was led by ARS researchers in Fort Pierce, FL and conducted across North America from Jan 2005 to Dec 2011 and included Bermuda and Hawaii (islands of Hawaii and Oahu), Canada and Mexico. Our findings show: 1) The New World biotype was detected in Texas and represents the first report of this biotype in the U.S. since its rapid displacement in the late 1980s by biotype B; 2) Biotype Q was detected in 23 U.S. states and Mexico, with first reports of biotype Q in Canada and Bermuda; 3) Biotype Q was found in protected commercial horticultural greenhouse plantings of forty-five percent of all collections of ornamental and herb plants and a single tomato transplant collection, but never in open field agriculture; 4) Genetic markers identified three distinct haplotypes of biotype Q whiteflies supporting the inference of independent invasions from at least three different sources; 5) Although we provide data of rarely occurring B-Q hybrids, these showed no evidence of persistence; and 6) Our results suggest that, unlike other countries where the Q biotype has invaded field crops, in the U.S. rapid detection and implementation of improved control strategies targeting the Q biotype has prevented the establishment of this pest beyond greenhouse production.
2. Population diversity of North American Bemisia tabaci. During the course of tracking continental spread of Bemisia tabaci biotype Q, two genetic markers were used to discriminate between invasive biotypes B and Q and native biotype NW. ARS researchers in Fort Pierce, Florida have completed an intensive sequencing effort at one of these markers that reveals possible hybrids between biotypes were not hybrids at all. This is important because B and Q possess different sets of high-fitness traits so a hybrid of this highly polyphagous pest would be particularly problematic to affected growers. A suite of seven genetic markers has also been used to track continental spread of biotype Q. Population genetics analyses of the US population conducted by ARS researchers in Fort Pierce, Florida indicates that it is highly structured, with as many as nine different sub-populations with limited gene flow between them. This is consistent with previous findings indicating the probability of multiple invasions into the U.S.
3. Molecular diagnostics for field implementation. A single temperature DNA amplification technique is being pursued as a means to discriminate between two invasive whiteflies, Bemisia tabaci biotypes B and Q, in high-value greenhouse ornamental crops. At present growers and other stakeholders send whiteflies to forensics labs for “biotyping” in order to manage pesticide resistance and to track the continental spread of biotype Q. ARS researchers in Fort Pierce, Florida have developed an assay that amplifies target DNA from biotype B whiteflies in 30 minutes with results based on color change in the reaction tube. This assay is much faster and easier to carry out and interpret than other biotyping assays. The combination of speed and simplicity make this assay more amenable to deployment in the greenhouse for real-time pest management decisions. Once additional barriers to field deployment are overcome, this assay will provide this real-time data to ornamentals growers and other stakeholders. Furthermore, once proof-of-concept is established, this DNA amplification technique is amenable to help solve other pest/pathogen diagnostics problems in agriculture.
Dickey, A.. Osborne, L.S., McKenzie, C. 2012. Papaya is not a host for tomato yellow leaf-curl virus. Florida Entomologist. 95:211-213. Available: http://dx.doi.org/10.1653/024.095.0136