Location: Subtropical Insects and Horticulture Research
2018 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
With the overall goal 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), eight greenhouse chemical efficacy trials were conducted on salvia. In these trials, the compatibility of non-neonic insecticides used in rotation programs to control Bemisia tabaci with the predatory mite, Amblyseius swirskii used in biological control programs was evaluated. Experiments testing flupyradifurone and afidopyropen as a drench application was compatible with the predatory mite, and were found significantly reducing whitefly population during the majority of the study period. Foliar application of spinetoram + sulfoxaflor, pyrifluquinazon and flupyradifurone was effective against whitefly; however, among these only flupyradifurone was found benign on the mite population.
To complement the previous study, additional studies were conducted where the compatibility of insecticides used to control Bemisia tabaci with the parasitic wasp, Eretmocerus eremicus used in biological control programs was evaluated on mint in the greenhouse. Eretmocerus eremicus is among the parasitoids which utilize their prey for both food and site of reproduction resulting in suppression of the pest population on the plant. In this study, whitefly parasitization by wasps was insignificant, and thus parasitized immatures and emerged wasps were not observed. Results showed that the drench application of flupyradifurone, and the parasitic wasps applied alone or in combination can suppress whitefly population = 5 weeks.
The chilli thrips, Scirtothrips dorsalis (S. 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 two years. In Fiscal Year 2018, thrips samples were processed from three counties of Florida (Hillsborough, Orange, and Indian River), two counties in Texas (Fort Bend, and Waller), two counties in California (Los Angeles, and Riverside), and one county in Massachusetts (Barnstable) and New York (Suffolk). Out of the 28 thrips samples processed from different locations in the United States, 19 samples were confirmed as S. dorsalis. The S. dorsalis samples collected from Florida, Texas, and California were South Asia 1 species suggesting this species is more prevalent (dominant species) in the United States compared to another counterpart of S. dorsalis complex reported for the first time from the north eastern United States during 2012. East Asia 1 was only found on hydrangea samples collected from two eastern states (New York and Massachusetts) indicating movement of this species in the neighboring states on hydrangea plants. Identity of S. dorsalis samples collected from blueberry in Hillsborough County, Florida still needs to be verified. Apart from S. dorsalis, eight different thrips species were also identified from the collected samples that includes Frankliniella occidentalis Pergande, Frankliniella schultzei Trybom, Frankliniella bispinosa Morgan, Frankliniella cephalica Crawford, Echinothrips americanus Morgan, Neohydatothrips burungae (Hood) Pseudothrips inaequalis (Beach) and Thrips hawaiiensis Morgan. Additional thrips samples from ornamental nurseries in Alabama, North Carolina, Louisiana and Connecticut are expected during 2018.
To determine the host range of the dominant member of chilli thrips, Scirtothrips dorsalis complex, South Asia 1, we conducted four greenhouse trials, where we evaluated 25 plant taxa (21 different species within 13 families and 11 plant orders) among ornamentals, landscape plant, herb and flowering shrubs as feeding and/or reproductive hosts of this pest. Among all the tested plant taxa, 16 were true hosts (reproductive plus feeding) and one was just feeding hosts (thrips will feed on these but avoid laying eggs). We also found eight new true hosts of chilli thrips which were not earlier reported as the reproductive hosts of this pest in the literature.
Native to the Neotropical region, Aleurotrachelus trachoides Back, commonly known as the solanum or pepper whitefly, is an emerging pest of pepper and many other horticultural crops in the United States. Feeding by this pest can cause stress to the plant by removing nutrients and water and the sheer numbers of whitefly can bring down plants and cause death. It has been in the United States for more than five decades as an intermittent pest of pepper although, until recently, was never considered a key pest of economic importance. However, in the past few years, records of its spread and damage have been reported from private residences, nurseries and vegetable growers (including tomato) throughout Florida. Aleurotrachelus trachoides is not a known vector of any plant damaging viruses, but to reassure tomato growers that this whitefly does not have the ability to acquire and transmit tomato yellow leaf curl virus, experiments are currently in progress.
Traditional taxonomy for mite identification is becoming a lost art and expertise is extremely difficult to acquire. In collaboration with University of Florida, ARS researchers are developing barcodes for predatory mites that are commercially available as well as those native to Florida. Traditional taxonomy is being used to identify mites morphologically and then directly correlate those populations with complementary (cDNA) library construction for deep sequencing data mining for identification of the best diagnostic genes for identification. Cryptic species analysis and intra and inter species variation for selected genes will be validated. Benefits to stakeholders include new tools to identify new predators and also help identify predators that are working in commercial settings, especially when growers are using multiple species and sources.
Rugose spiraling whitefly (RSW) is an exotic invasive species which had an outbreak in south Florida that became economically damaging to numerous plant species in many urban areas. Oviposition preference was determined for the following five known plant hosts: coconut, gumbo limbo, avocado, black olive v. shady lady and giant white bird of paradise (GWBP). Gumbo limbo was the most preferred and also had the highest number of eggs per spiral. GWBP was the least preferred host plant species but no statistically significant difference was found among the survival of RSW on different host plant species tested. Also, no significant correlation was found between the leaf area and number of eggs on each host species.
Nephaspis oculata is a whitefly beetle predator which has been reported feeding on several whitefly species including rugose spiraling whitefly (RSW) in south Florida. The management of RSW has been heavy reliant on the use of insecticides which may negatively impact biological control agents. We studied the effect of bifenthrin (spray) and imidacloprid (drench) application on survival, fecundity, and behavior of Nephaspis oculata in the laboratory. Generally, beetles avoided bifenthrin treatments and did not survive as long as imidacloprid treatments. The results from this study shows that systemic imidacloprid is not repellent to Nephaspis oculata and does not significantly affect mortality of adult beetles in the tri-trophic system tested. Therefore, using systemic imidacloprid and Nephaspis oculata for controlling RSW might be compatible or at least not significantly incompatible. Future studies in the field may help us to have a better understanding about such interactions in the landscape.
Work on discovery of new molecules for the control of the Bemisia whitefly was advanced through international collaboration with scientists in Brazil. Together we have 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. This research has led to the production of transgenic tobacco plants expressing this peptide that are now being used for feeding studies with Bemisia tabaci whiteflies. To support this work we have developed a method to deliver peptides directly into the phloem tissue and we are evaluating this strategy for delivery of the peptide to crop plants on which Bemisia tabaci is a pest.
Because isolation and analysis of psyllid salivary sheaths was easier than that for the whitefly and because we previously demonstrated that the same sheath inhibitors worked for both insects, ARS researchers used the psyllid system as a surrogate for the whitefly. We have identified and purified a single highly abundant approximately 2.8kD peptide in salivary secretions of the psyllid and we hypothesis that this peptide is an important structural component of the salivary sheaths. Sequencing of this peptide is being done through a collaboration with a research molecular biologist, USDA-Agricultural Research Service, Ithaca, New York. This discovery has led to development of a research program to identify inhibitors of one of these proteins/enzymes.
Accomplishments
1. Pre-shipping treatments of biopesticides mitigates trans-boundary movement of whitefly. Bemisia tabaci (B. tabaci) cryptic species complex represents a major threat to several important crops including vegetables, cereals, fruits, and ornamentals worldwide. One important mode of its dispersal is through the trans-boundary (domestic, international) movement of infested plant materials. To prevent the spread of this invasive whitefly pest on inter-state ornamental shipments, ARS researchers at Ft. Pierce, Florida, in collaboration with researchers from a university in Florida tested the efficacy of biopesticides through dip treatments of plant material pre-shipping. In several experiments, clean Mentha sp. (Lamiaceae) mint plants were exposed to adults of B. tabaci for 1 or 6 days. Adults were removed from plants and eggs, and immature stages were treated with one or a combination of the following: entomopathogenic fungi Beauveria bassiana (BotaniGard®), Isaria fumosorosea (Preferal®), the surfactant Natur'l Oil™, and soap (Publix®). After treatment application, plants were placed in commercial cardboard boxes and shipped from Apopka, Florida to Fort Pierce, Florida. Assessments for the treatment efficacies were conducted at 3, 7, and 14 days post arrival of the shipment. Fungal entomopathogens; B. bassiana, I. fumosorosea, and the surfactant Natur'l Oil™, showed significant reduction in the whitefly population compared to the untreated control (P < 0.05). Significantly higher corrected mortality of the whitefly populations was observed when Beauveria bassiana was applied alone (81%) or in combination with Natur'l Oil™ (86%), as well as Isaria fumosorosea applied in combination with Natur'l Oil™ (77%). Our results showed that pre-shipping dip treatment applications of either Natur'l Oil™ mixed with Beauveria bassiana or Isaria fumosorosea or Beauveria bassiana applied alone could help mitigate the inter-state spread of whitefly on ornamental shipments.
2. Horticultural oils compatible with entompathogenic fungi for Asian Citrus Psyllid Control. Horticultural oils are an important component of integrated management programs of several phytophagous arthropods and pathogens affecting fruit, ornamentals and vegetables in greenhouse and field production systems. Although effective against the target pest, their incompatibility with biological control agents can compromise efforts to develop eco-friendly management programs for important agricultural pests. ARS researchers at Ft. Pierce, Florida, in collaboration with University of Florida assessed the in-vitro effect of selected refined petroleum oils used in citrus and other horticultural crops with a biopesticide containing the entomopathogenic fungi, Isaria fumosorosea (PFR-97) under laboratory conditions. Leaf disk bioassays were used to evaluate the combined efficacy of petroleum oils and PFR-97 against the Asian citrus psyllid, Diaphorina citri, a major pest of citrus in the United States. All five petroleum oil treatments were compatible with PFR-97 blastospores, as none of them were found to affect colony-forming units and radial fungal growth measured up to 12 days post-inoculation. All mixed treatments performed better than PFR-97 alone against Diaphorina citri. Results indicated that horticultural oils in combination with PFR-97 could offer citrus growers an alternative treatment for integrating into their current management programs while battling against Diaphorina citri in citrus production systems. Due to their eco-friendly, broad-spectrum effect, it could provide control against various citrus pests, while also encouraging the retention of effective chemistries for a longer period in the marketplace. However promising, these combination treatments need to be tested further with PFR-97 under grove conditions to confirm their field efficacy.
3. New insect-toxic peptide (Jaburetox from Jack bean) shown to be effective at controlling insects when expressed in transgenic plants. Currently, the only commercial transgenic method for insect control involves the use of bacterial toxins called Bacillus thuringiensis (BT) endotoxin. Because of this limited diversity of control strategies, insect resistance to BT is becoming a major concern. Therefore new gene products that provide effective insect control are needed. Furthermore, no BT-toxin has been identified that works against hemipterans. Jaburetox is a plant derived insect toxin that has shown activity against some hemipterans. Therefore in collaboration with researchers from Brazil, ARS researchers at Ft. Pierce, Florida, have evaluated activity of this peptide in laboratory bioassays and in transgenic plants and demonstrated high insect mortality and developmental disruption in pest insects. They have now produced transgenic tobacco to study Jaburetox’s effect on Bemisa tabaci. These experiments are currently underway and demonstration of their effectiveness will create a new strategy in the fight against this severe crop pest.
Review Publications
Aristizabal, L., Avery, P., Kumar, V., Caldwell, J., McKenzie, C.L., Osborne, L.S. 2018. Mitigating trans-boundary movement of Bemisia tabaci (Hemiptera: Aleyrodidae) on Mentha sp by pre-shipping treaments of biopesticides. Crop Protection. 107:71-78.
Kumar, V., Avery, P., Ahmed, J., Cave, R., McKenzie, C.L., Osborne, L.S. 2017. Compatibility and efficacy of Isaria fumosorosea with horticultural oils for mitigation of the Asian citrus psyllid, Diaphorina citri (Hemiptera: Liviidae). Insects. 8:119. doi.org/10.3390/insects.8040119. https://doi.org/10.3390/insects.8040119
Kumar, V., Kakkar, G., Seal, D.R., McKenzie, C.L., Osborne, L.S. 2017. Evaluation of insecticides for curative, preventative and rotational use on Scirtothrips dorsalis South Asia 1 (Thysanoptera: Thripidae). Florida Entomologist. 100(3):634-646.
Kumar, V., Francis, A., Avery, P., McKenzie, C.L., Osborne, L.S. 2018. Assessing compatibility of Isaria fumosorosea and buprofezin for mitigation of Aleurodicus rugioperculatus (Hemiptera: Aleyrodidae) - an invasive pest in the Florida landscape. Journal of Economic Entomology. 111(3):1069-1079.
Thomson, M., Sahavaraj, K., Kumar, V., Avery, P.B., McKenzie, C.L., Osborne, L.S. 2017. Mass rearing and augmentative biological control evaluation of Rhynocoris fuscipes (Hemiptera: Reduviidae) against multiple pests of cotton. Pest Management Science. 73(8):1743-1752.
Kumar, V., Dickey, A.M., Seal, D.R., Shatters, R.G., Osborne, L.S., McKenzie, C.L. 2017. Unexpected high intragenomic variation in two of three major pest thrips species does not affect ribosomal transcribed spacer 2(ITS2) utility for thrips identification. International Journal of Molecular Sciences. 18(10):2100.
Stover, E.W., Shatters, R.G., Gruber, B., Kumar, Moore, G.A. 2016. Influence of photoperiod duration and phloem disruption through scoring on growth, disease symptoms and bacterial titer in citrus graft-inoculated with Candidatus Liberibacter asiaticus. HortScience. 51:1215-1219.
Ammar, E.D., Hall, D.G., Shatters, R.G. 2017. Ultrastructure of the salivary glands, alimentary canal and bacteria-like organisms in the Asian citrus psyllid, vector of citrus huanglongbing-disease bacteria. Journal of Microscopy and Ultrastructure. 5(1):9-20.
Zhang, S., Shi, Q., Albrecht, U., Shatters, R.G., Stange Jr, R.R., McCollum, T.G., Zhang, S., Fan, C., Stover, E.W. 2017. Comparative transcriptome analysis during early fruit development between three seedy citrus genotypes and their seedless mutants. Horticulture Research. 4:17041.
Hall, D.G., Borovsky, D., Chauhan, K.R., Shatters, R.G. 2018. An evaluation of mosquito repellents and essential plant oils as deterrents of Asian citrus psyllid. Crop Protection. 108:87-94.
Borovsky, D., Nauewelaers, S., Powell, C., Shatters, R.G. 2017. Cloning, genetic engineering and characterization of TMOF expressed in Saccharomyces cerevisiae to control larval mosquitoes. Journal of Insect Physiology. 106(2):134-146. https://doi.org/10.1016/j.jinsphys.01.008.
Mann, M., Fattah-Hosseini, S., Ammar, E., Strange, R., Warrick, E., Sturgeon, K., Shatters, R.G., Heck, M.L. 2018. Diaphorina citri nymphs are resistant to morphological changes induced by “Candidatus Liberibacter asiaticus” in midgut epithelial cells. Infection and Immunity. 86(4):e00889-17. https://doi.org/10.1128/IAI.00889-17.
Kruse, A., Fattah-Hosseini, S., Saha, S., Johnson, R., Warwick, E., Sturgeon, K., Mueller, L., Maccoss, M.J., Shatters, R.G., Cilia, M. 2017. Combining 'omics and microscopy to visualize interactions between the Asian citrus psyllid vector and the Huanglongbing pathogen Candidatus Liberibacter asiaticus in the insect gut. PLoS One. https://doi.org/10.1371/journal.pone.0179531.
Saha, S., Hosmani, P.S., Billalobos-Ayala, K., Miller, S., Shippy, T., Flores, M., Rosendale, A., Cordola, C., Bell, T., Mann, H., DeAvila, G., DeAvila, D., Moore, Z., Buller, K., Ciolkevich, K., Nandyal, S., Mahoney, R., Van Voorhis, J., Dunlevy, M., Farrow, D., Hunter, D., Morgan, T., Shore, K., Guzman, V., Izsak, A., Dixon, D.E., Cridge, A., Cano, L., Cao, X., Jiang, H., Leng, N., Johnson, S., Cantarel, B.L., Richards, S., English, A., Shatters, R.G., Childers, C., Chen, M., Cilia, M., Mueller, L., Munoz-Torres, M., Nelson, D., Poelchau, M.F., Benoit, J., Wiersma-Koch, Brown, S.J. 2017. Improved annotation of the insect vector of citrus greening disease: biocuration by a diverse genomics community. bioRxiv. https://doi.org/10.1093/database/bax032.
