Location: Invasive Insect Biocontrol & Behavior Laboratory
2016 Annual Report
Objectives
Objective 1: Identify, synthesize, and develop semiochemicals (pheromones, kairomones, plant volatiles, and other attractants and repellants) of significant insect crop pests (such as harlequin bug, bagrada bug, spotted wing drosophila, striped cucumber beetle, kudzu bug) and their natural enemies for use in integrated pest management.
Subobjective 1a: Develop attractant volatiles and trap designs to monitor and manage the harlequin bug.
Subobjective 1b: Assess known stink bug attractants for value in monitoring and/or managing bagrada bug.
Subobjective 1c: Identify plant volatiles that attract squash bugs.
Subobjective 1d: Identify plant and/or insect volatiles for detection and attraction of kudzu bug.
Subobjective 1e: Improve lures for spotted wing drosophila by identification of fruit-based attractants.
Subobjective 1f: Synthesize and test aggregation pheromone in combination with baits and traps for striped cucumber beetle management.
Objective 2: Develop arthropod biological controls for managing key vegetable pests such as stink bugs, squash bug, and cucumber beetles, including integration of natural enemies with other tactics such as microbial control, semiochemicals, and cultural pest controls.
Subobjective 2a: Evaluate the efficacy of the squash bug natural enemy complex for use in pest management and identify the most effective biological control agents.
Subobjective 2b: Assess the value of native egg parasitoids to suppress BMSB.
Objective 3: Advance effective microbial controls for key crop pests such as stink bugs, noctuid moths, and other seedling pests, including discovery of molecular and ecological mechanisms for sustained field reproduction and persistence, using diverse Bacillus thuringiensis (Bt) strains, Chromobacterium spp., baculoviruses, and other entomopathogens.
Subobjective 3a. Discover additional environmental isolates of C. subtsugae and related species, and characterize their insecticidal properties.
Subobjective 3b. Determine ability of Bt strains to persist in common vegetable mulches.
Subobjective 3c. Develop effective baculoviruses for key pest diamondback moth.
Objective 4: Discover naturally occurring biopesticides (such as botanical compounds and/or RNAis) targeting key vegetable pests such as stink bugs and cucumber beetles.
Objective 5: Determine and strengthen the genetic basis of plant defense mechanisms (e.g., pest aversion and resistance) for protection of high-value crops such as cole crops on small farms or gardens against destructive insects, and determine the influence of the microflora of pest insects and/or plants on the plant defense response.
Subobjective 5a: Determine the effect of knocking out or over-expressing the infestation responsive TF StZFP2 in its native species, and in Arabidopsis.
Subobjective 5b: Determine how infestation inducible Q-type C2H2 TFs affect resistance to pest insects in Arabidopsis and B. oleracea.
Subobjective 5c: Determine the influence of plant and insect microflora on the infestation response of Arabidopsis to T. ni.
Approach
The project brings together a research team with diverse expertise for multiple approaches to insect management. The proposed project will focus on control of key insect pests in small farms and urban gardens, both organic and non-organic. Bio-based integrated pest management approaches to be developed will include: (i) discovery and deployment of natural insect attractants and repellents; (ii) conservation and augmentation of beneficial insects including use of their semiochemicals; (iii) pest-specific microbial controls [bacteria (including Bacillus thuringiensis strains and Chromobacterium spp.) and baculoviruses]; and, (iv) crop genetic resistance using molecular-based gene discovery. Research will target insect pests that cause major damage to key crops such as cucurbits and cole crops, although other important crops such as small fruit, beans, and potatoes may receive attention for specific problems. The combination of semiochemical approaches, biological controls, molecular techniques, and crop resistance, will offer a range of non-chemical tactics useful to integrated pest management strategies for major crop pests in urban small farms and gardens.
Progress Report
Objective 1: The harlequin bug aggregation pheromone is a new and important management tool for this key agricultural pest. We identified the pheromone earlier as a mixture of two principal components, both of which are important for attractiveness. To be able to conduct ratio, dose-response, and other field bioassays, we needed sufficient amounts of these pheromone components. Thus, in the current year we developed a new synthetic procedure that provided both components in appreciable yields and allowed scaling-up the pheromone. This new technique was based on an affordable inorganic base used as a catalyst in a key step of the synthesis.
An effective tool for detection of Spotted wing drosophila (SWD), an highly damaging invasive fruit fly species which has spread rapidly across the United States, is urgently needed for management of this pest. After performing a series of tests on apple juice volatile aroma, a ternary blend has been identified as the most effective attractant for trapping SWD under laboratory conditions. The activity was confirmed by field tests. In addition, another volatile compound, methyl benzoate (MB), from apple juice was found to exhibit significant acute toxicity against the invasive species, brown marmorated stink bug (BMSB), gypsy moth (GM), SWD, as well as diamondback moth (DBM) and tobacco hornworm (THW).
Striped cucumber beetle is a perennial problem with growers of squash, pumpkin, cucumbers, and melons. Its aggregation pheromone was previously discovered but quantities available were not sufficient for field testing. To remedy this need, a facile synthesis was developed producing gram quantities of diastereomeric mixture of the pheromone. We concluded field studies during the 2015 season demonstrating the attractiveness of this synthetic preparation to adult cucumber beetles, and the patent was recently published.
Objective 2: Native parasitoid impact on brown marmorated stink bug (BMSB) sentinel egg masses was measured in four crops (bell pepper, tomato, squash, and soybean), in landscaped experimental plots comprised of either native or exotic plant species, and in wooded habitats in Maryland and D.C. The parasitoid species composition and parasitism rate varied in different habitats. Parasitism rates ranged from 12.4% in vegetable crops to 3.7% in landscaped plots.
Preliminary bioassays of 65 diamondback moth granulovirus isolates from China, Taiwan, and Japan revealed that 25 of the isolates possessed insecticidal activity against diamondback moth larvae. PCR and DNA sequence analysis of diagnostic genes from these 25 samples revealed that they are all isolates of the betabaculovirus species Plutella xylostella granulovirus.
Objective 3: New Chromobacterium isolates were obtained from a variety of aquatic habitats. Bioassays of newly collected isolates that have been identified indicate that within a Chromobacterium species, toxicity to particular insects is relatively constant. Biochemical phenotypes, growth characteristics, and genome level comparisons of two new Chromobacterium species were investigated to determine which features are useful in distinguishing them from other members of the Chromobacterium. A number of genes were investigated for their utility in predicting phylogenetic relationships among Chromobacterium, as 16S rRNA genes are not always reliable.
Objective 4: Transcriptome sequence data for Murgantia histrionica, the harlequin bug, was assembled, annotated and quantitatively analyzed. Genes exhibiting differential expression across sexes and developmental stages were identified, and primers have been designed for further inquiry using molecular approaches. Preparation of biological replicates to enable molecular validation efforts is currently in process. These results will assist in the identification of gene targets of potential use in insect biocontrol programs.
Objective 5: Arabidopsis transgenics expressing the StZFP2 gene were tested for resistance to diamondback moth using bioassays. Unlike their increased resistance to cabbage looper, evidence indicates that the transgenics do not have increased resistance to diamondback moth. The goal of this project is to determine how StZFP2 affects the plant’s resistance to insects. Genetically modified potato lines were produced in which the level of expression of StZFP2 was increased or decreased. We are currently analyzing genes induced by mechanical wounding, which induces genes associated with defense but is more rapidly performed and reproducible than insect infestation, and is thus a more reliable way to see how changes in StZFP2 levels alter expression of other genes. StZFP2 is produced rapidly after wounding, peaking in expression within 20 minutes. In addition, we are testing whether we can predict the function of StZFP2-like transcription factors by the features of their protein sequence; 27 StZFP2-like genes were identified in potato, and five of these were found to have expression patterns similar to StZFP2 upon wounding. Further studies will determine whether any of these newly discovered transcription factors also play a role in the defense against chewing insect pests.
Accomplishments
1. Demonstration of an important native biocontrol for key pest, squash bug. ARS researchers in Beltsville, Maryland, have demonstrated that the egg parasitoid Gryon pennsylvanicum is an important biological control agent of squash bug, a key pest of squash, pumpkins, melon, and watermelon. A two-year study determined that Gryon was able to efficiently track wild squash bug eggs throughout the season. The rate of parasitism on wild egg masses peaked at the end of July at over seventy-two percent. From the end of July until the first week of September, less than 20 percent of squash bug nymphs hatched successfully from wild eggs. These results indicate that Gryon has the potential to be an effective biological control agent and play an important role in the suppression of squash bug populations.
2. Development of spotted winged drosophila attractant. ARS researchers in Beltsville, Maryland, have developed an effective lure for spotted winged drosophila (SWD), a destructive invasive fruit fly native to southeast Asia that is currently spreading across the U.S. Laboratory experiments led to the formulation of four volatile compounds from apple juice that were attractive to the fly under field conditions. Early detection of SWD will allow growers to apply timely pest management interventions and avoid unnecessary applications of insecticides.
Urban agriculture is increasingly considered a key component for nutrition, employment, and well-being of urban populations which are often underserved by agricultural research and extension programs. ARS scientists are cooperating with University of the District of Columbia (UDC) College of Agriculture, Urban Sustainability & Environmental Sciences to mentor student projects addressing urban agricultural needs for their ambitious program by UDC’s Center for Urban Agriculture & Gardening Education with urban farms in each of the nine wards of the District. One of these projects involves the harlequin bug, a threat to cole crops and related nutritious vegetables. This project will determine the importance of existing biological controls, mainly parasitoids that attack the egg masses of this key pest, at several sites in DC, as well as UDC’s Firebird Farm and ARS fields in Beltsville, Maryland. Based on this research we will learn the best mix of non-chemical controls, including biological, cultural, and behavioral (pheromone-based) tactics that are most applicable in different urban farms and gardens.
Review Publications
Prieto, N., Swift, M., Summerfelt, S., Juarez, M., Zijlstra, R., Aalhus, J. 2015. A feasibility study on the use of near infrared spectroscopy for the authentication of depurated salmon fillets. Journal of Food Analytical Methods. 8:2660-2664.
Cornelius, M.L., Buffington, M.L., Talamas, E.J., Gates, M.W. 2016. Parasitism and predation rates on sentinel and naturally occurring egg masses of the squash bug (Hemiptera: Coreidae) in Maryland. Environmental Entomology. 45(2) 367-375.
Harris, C., Abubeker, S.U., Yu, M., Leskey, T.C., Zhang, A. 2015. Semiochemical production and laboratory behavior response of the brown marmorated stink bug, Halyomorpha Halys. Journal of Insect Physiology. 10(1):e0140876.
Herlihy, M.V., Talamas, E.J., Weber, D.C. 2016. Attack and success of native and exotic parasitoids on eggs of Halyomorpha halys in three Maryland habitats. PLoS One. 11(3):e0150275.
Khrimian, A., Shirali, S., Guzman, F. 2015. Absolute configurations of zingiberenols isolated from ginger (Zingiber officinale) rhizomes. Journal of Natural Products. 78(1)3071-3074.
Mahadwar, G., Chauhan, K.R., Bhagavathy, G., Murphy, C.F., Smith, A.D., Bhagwat, A.A. 2015. Swarm motility of Salmonella enterica serovar Typhimurium is inhibited by compounds from fruit peel extracts. Letters in Applied Microbiology. 60:334-340.
Wada-Katsumata, A., Zurek, L., Nalyanya, G., Roelofs, W.L., Zhang, A., Schal, C. 2015. Gut bacteria mediate aggregation in the German cockroach. Proceedings of the National Academy of Sciences. 112(51):15678-15683.
Zeng, F., Zhao, Z., Yan, M., Zhou, W., Zhang, A., Lu, Z., Wang, M. 2015. Identification and comparative expression profiles of chemoreception genes revealed from major chemoreception organs of the rice leaf folder, Cnaphalocrocis medinalis (Lepidoptera: Pyralidae). PLoS One. 10(12):e0144267.
Zhang, Q., Zhou, G., Hoover, D.R., Michaelson, N.J., Bryant, P., Margaryan, A., Chauhan, K.R., Aldrich, J., Schneidmiller, R. 2015. (1R,2S,5R,8R)-Iridodial and Z,E-nepetalactol: first long-range 4 chemical attractants for antlions (Neuroptera: Myrmeleontidae). Frontiers in Ecology and the Environment. 2:80.
Zhang, Z., Gao, Q., Liu, L., Zhang, A., Zhang, B., Luo, J., Chen, L., Wang, M., Lei, C. 2015. Sex pheromone of the mirid bug, Adelphocoris suturalis. Journal of Chemical Ecology. 26(1):25-31.
