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:
A portion of mitochondrial gene from chilli thrips was sequenced from populations in four Florida counties feeding on ten hosts as well as populations in China, Japan, India, Israel, Thailand, and Singapore and uncovered more than 80 distinct haplotypes. There is 14% sequence divergence between Japan and U.S. populations, 14% sequence divergence between China and U.S. populations, and 7% sequence divergence between Japan and China populations of chilli thrips. The common Florida haplotype is identical in sequence to samples from Thailand and Singapore. Next-generation DNA sequencing of the invasive chilli thrips is being conducted and more than 1 billion nucleotides of the chilli thrips metagenome have been sequenced. Morphological and molecular identification techniques were coupled to develop a quick, reliable and simple diagnostic method for thrips. Prior to the DNA extraction of chilli thrips specimens, larvae and adults were subjected to traditional morphological identification using high resolution scanning electron microscopy (SEM) and then gold/palladium sputter coated thrips specimens were processed for polymerase chain reaction assay for molecular identification. Sequence results of two molecular markers of individual larva and adult thrips were in agreement with the taxonomic identification conducted using SEM and each result confirmed the other technique. Two commercial bell pepper varieties were evaluated for their ability to sustain predatory mites (Amblyseius swirskii) under no pollen or prey conditions at trial initiation in greenhouse experiments. Pepper seedlings were infested with two rates of predatory mites (20 or 40 mites/) before transplanting to pots. Predatory mites were able to survive, develop and oviposit in all mite treatments and mites preferred top over lower leaves in both pepper varieties. Natural infestation of whiteflies occurred later in the experiment and the number of whitefly eggs, nymphs and adults were significantly higher in the untreated control compared to all mite infested treatments. Preliminary results indicate predator in first approach is viable under greenhouse conditions. Bacterial communities in 7 species of the whitefly Bemisia tabaci complex were investigated by pyrosequencing bacterial 16S rRNA gene amplicons. Representatives of just 9 bacterial genera were identified, with 7 previously described vertically-transmitted taxa accounting for >99.9-100% of the reads in each sample.The abundance of the bacterial symbionts in the most common species in the United States was elevated in insect cultures bearing begomoviruses (tomato mottle virus and tomato yellow leaf curl virus) relative to a non-viruliferous culture, and varied significantly with rearing plant. Specific proteins were identified that block de novo whitefly salivary sheath formation and degrade existing sheaths. An invention disclosure was submitted for this and accepted by the patent review board. The patent is now being written. Efficacy of soap oils and entomopathogenic fungi are being evaluated as potential biopesticides to prevent the spread of whitefly on ornamental shipment from nurseries in Florida to other states.
1. Controlling Bemisia whitefly feeding through the use of inhibitors that block salivary sheath formation. Discovery of specific proteins (single gene products) that inhibit whitefly (Bemisia species) salivary sheath formation and degrade preformed sheaths. Researchers at the USDA, ARS, U.S. Horticultural Research Laboratory in Fort Pierce, Florida showed that specific proteins can block salivary sheath formation by whitefly and present this discovery as a novel method of control of controlling whiteflies as well as many plant-feeding Piercing-sucking hemiptera (including aphids, mealybugs, scales, leafhoppers, planthoppers, psyllids etc.). This work is being advanced using transgenic plant strategies to produce plants that prevent whitefly sheath production and thus inhibit feeding.
2. North American Bemisia Biotypes After the Q Invasion. The Bemisia tabaci cryptic species complex of whiteflies contains two invasive groups, often called biotype B or the Mid-East Asia Minor 1 group and biotype Q or the Mediterranean group. In the U.S. biotype B occurs both in outdoor plantings and in the greenhouse, but biotype Q is only found in greenhouses. To determine the genetic structure and number of distinct populations of each species, and to estimate the origin of biotype Q within the U.S., ARS researchers in Fort Pierce, Florida carried out population genetic analyses on both cryptic species using microsatellite markers. The major findings of the study are: 1) biotype Q contains a greater number of genetically different populations in U.S. than biotype B despite having been established for only a few years, 2) biotype Q consists of two major genetic lineages, one originating in the eastern mediterranean region and one originating in the western mediterranean region, 3) biotype Q with origins in both the eastern and western Mediterranean are found throughout the continental U.S. and eastern Mediterran biotype Q is present in Hawaii, and 4) biotype B in the U.S. contains two populations from greenhouses (one in Arizona and one in New York) that are genetically different from one another and from all other biotype B in the U.S. The results suggest that biotype B was introduced into the U.S. on at least three occasions and rapidly spread throughout the U.S., showing no discernable differentiation across 7,000 kilometers. The results further suggest that there is an enhanced role of greenhouses in promoting and maintaining genetic differentiation in both cryptic species.
3. Intragenomic variations of three economically important thrips species. Correct identification is a fundamental step in the development of sound management practices against a pest. Thus, it is important to select an appropriate marker gene (region) for the molecular characterization of the target pest species. ARS researchers in Fort Pierce, Florida compared intra- and intergenomic variability in the sequence of the two most widely used genes (mtCO1 and ITS2) for thrips identification. Relatively low variation in mtCO1 and variant paralogous haplotype was observed in chilli thrips, melon thrips and western flower thrips suggesting this gene was suitable for taxonomic characterization of the three major economic thrips species. However, extensive inter- and intragenomic variations were observed in the ITS2 gene of both chilli and melon thrips. Although the possible reason for ITS2 variation of these thrips species could not be traced, it can be concluded that the ITS2 copies in these thrips do not evolve in concert based on the degree of intra- and intergenomic variability. Thus, this marker region is not phylogenetically informative for population studies, and should not be used for species-specific PCR identification of the thrips species studied.
Xiao, Y., Avery, P.B., Chen, J., McKenzie, C., Osborne, L. 2012. Ornamental pepper as banker plants for establishment of Amblyseius swirskii (Acari: Phytoseiidae) for biological control of multiple pests in greenhouse vegetable production. Biological Control. 63:279-286. Available: http://dx.doi.org/10.1016/j.biocontrol.2012.09.007.