Research Project: Urban Landscape Integrated Pest Management

Location: Invasive Insect Biocontrol & Behavior Laboratory

2019 Annual Report

Objectives
OBJECTIVE 1: Develop semiochemicals (pheromones, kairomones including plant volatiles, and other attractants and repellents) of significant landscape pests (such as brown marmorated stink bug and exotic scarabaeids) and their natural enemies (e.g., parasitoids and spiders) for monitoring and management. Sub-objective 1.A: Identify and synthesize volatiles and secretions from ornamental landscape pests or their plant hosts that are important in host- or mate-finding behavior, as components for biologically based control strategies. Sub-objective 1.B: Develop effective strategies for using synthesized semiochemicals to reduce plant damage due to ornamental landscape insect pests. OBJECTIVE 2: Develop and genomically characterize effective microbial and arthropod natural enemies as biocontrol agents, including bacteria (e.g., Bacillus thuringiensis [Bt]; Chromobacterium spp.), baculoviruses, and other entomopathogens, against insect pests of landscape plants (such as European and Asian gypsy moth, brown marmorated stink bug, and winter moth). Sub-objective 2.A: Discover additional environmental isolates of C. subtsugae and related species, and characterize their insecticidal properties. Sub-objective 2.B: Make phylogenetic and pathogenic comparisons of soil and phylloplane populations of Bt. Sub-objective 2.C: Evaluate and develop viral pathogens as potential control agents for selected lepidopteran pests in unmanaged urban forest landscapes. Sub-objective 2.D: Compare the floral and extrafloral resources of native and exotic urban landscape plant species for their ability to support natural enemies. Sub-objective 2.E: Use molecular gut-content analysis to identify brown marmorated stink bug egg predators. OBJECTIVE 3: Employ new technologies to characterize genomes and transcriptomes, characterize gene expression patterns, and develop RNAi-based molecular biopesticides for control of insect pests of landscape plants, especially brown marmorated stink bug and gypsy moth. Sub-objective 3.A: Characterize and generate draft reference genome for brown marmorated stink bug (BMSB) and gypsy moth. Sub-objective 3.B. Develop RNAi-based microbial biopesticides targeting brown marmorated stink bug (BMSB) and European and Asian gypsy moth.

Approach
This highly interdisciplinary project combines concepts and methodologies from analytical and synthetic organic chemistry, insect chemical behavior, insect pest-pathogen genomics, and microbial and insect ecology to develop multiple strategies for managing key insect pests of urban landscapes, orchards, and surrounding forest remnants. Integrated pest management approaches to be developed will include: (1) discovery and deployment of insect- and plant-derived attractants and repellents; (2) conservation of parasitic and predatory insects and spiders through selection of host plants for their ability to support natural enemies and thereby to promote effective biocontrol of pests; (3) pest-specific microbial controls including bacteria (e.g., Bacillus thuringiensis strains and Chromobacterium spp.) and baculoviruses; and, (4) plant genetic resistance using molecular-based gene discovery. Research will target insect pests that cause major damage to woody plants, including key landscape and forest genera, as well as fruit-bearing trees. 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 for major pests of urban landscapes and associated orchard and forest ecosystems.

Progress Report
Objective 1. Develop semiochemicals of significant landscape pests and natural enemies for monitoring and management. ARS scientists in Beltsville, Maryland, continued to evaluate methyl benzoate and its analogs as insecticides, with efficacy against bed bugs, mosquitoes, thrips, nematodes, and some stored product insect pests demonstrated under laboratory conditions. ARS scientists in Beltsville, Maryland, and Newark, Delaware, collaborated in identifying volatiles emitted by brown marmorated stink bug (BMSB) that could be used as attractants or repellents for the parasitic wasp Trissolcus japonicus. Two compounds, tridecane, and (E)-2-decenal, in the BMSB volatiles emission were behaviorally active against this known egg parasitoid. While the former chemical seemed to attract the egg parasitoid, the latter displayed a repellent activity when tested in laboratory bioassay. Objective 2 Develop microbial and arthropod natural enemies as biocontrol agents of native and exotic pests of landscape plants. In collaboration with Brazilian researchers, ARS scientists in Beltsville, Maryland, identified variants of the brown marmorated stink bug iflavirus in the transcriptome data of three unrelated South American stinkbug species. The stinkbug iflaviruses shared 84% to 99% genome sequence identity and appear to be members of the same iflavirus species. Virus-like particles resembling those of iflaviruses were extracted from the South American stink bugs. Between 40% and 90% of the transcriptome sequencing reads derived from the antennae of the South American stinkbugs mapped to the iflavirus genome sequence, indicating that the iflavirus replicated to a very high level in these insects without causing any obvious pathology. ARS scientists in Beltsville, Maryland, in collaboration with scientists at Virginia Commonwealth University, examined Cotesia congregata (Hymenoptera: Braconidae) for their host-parasitoid interactions in suppression of herbivorous insect pest populations, specifically investigating the process of diversification among sympatric populations that differed in host and host foodplant usage. Through analysis of two incipient species on different food sources and comparison of relative expression in vivo of certain C. congregata bracovirus (CcBV) genes among these parental and hybrid crosses, it was shown that C. congregata parasitoid is composed of two sympatric incipient species that can utilize multiple host species rather than of several host-associated races or cryptic species. ARS researchers in Beltsville, Maryland, continued to characterize new species of insecticidal Chromobacterium, comparing biochemical and genetic features of two new species isolated from ponds in Maryland and Florida. The new species were found to have activity against seedcorn maggot. In addition, a strain of Bacillus wiedmannii was discovered to produce parasporal crystals like the insect pathogen Bacillus thuringiensis (Bt). While not as toxic to gypsy moth larvae as Bt kurstaki, B. wiedmannii caused significant mortality compared with the non-pathogenic Bt finitimus. Objective 3. Employ new technologies to characterize genomes and transcriptomes, characterize gene expression patterns, and develop RNAi-based molecular biopesticides for control of insect pests of landscape plants, especially brown marmorated stink bug and gypsy moths. ARS researchers in Beltsville, Maryland, investigated the effect of orally administered RNAi on the efficacy of gypsy moth baculovirus against North American, Chinese, and Russian populations of gypsy moth. Double stranded RNA interference triggers for four different gypsy moth genes were fed to newly hatched larvae alone, or in combination with the virus, and compared with the effect of the virus alone. No consistent effects were observed on larval growth or virus-induced mortality. ARS scientists in Beltsville, Maryland, in collaboration with scientists in Natural Resources Canada, Université Laval, and McGill University, prepared a draft genome assembly and annotation for the Asian gypsy moth subspecies, Lymantria dispar asiatica and L. dispar japonica. In addition, this working group is considering ways to incorporate a draft assembly of the European gypsy moth, prepared by ARS scientists, into a comparative genomic study involving the Asian subspecies mentioned above, as well as a genome for the Ld652Y cell line derived from European gypsy moth ovary tissue. ARS scientists in Beltsville, Maryland, and Stoneville, Missippi, have collaboratively prepared a transcriptome for Solenopsis invicta, the red imported fire ant, and identified genes differentially expressed between larval and pupal stages. The genes identified in the study are candidate targets for RNAi-mediated biocontrol efforts.

Accomplishments
1. Discovery of a new insecticidal bacterium. ARS scientists in Beltsville, Maryland, have isolated and characterized a previously unknown species of bacteria that kill insects. The new bacterium, Chromobacterium phragmitis, was found in water and sediment samples collected from low salinity tidal marshes along the Potomac and James rivers in Maryland and Virginia. When grown in liquid cultures, C. phragmitis was found to produce compounds that are orally toxic to larvae of gypsy moth, diamondback moth, seedcorn maggot and red four beetle. The new bacterium has the potential to be developed into a commercial organic insecticide. A patent application covering the use of C. phragmitis for insect control has been filed.

2. Development of attract and kill strategy for cocoa pod borer pest management. The cocoa pod borer (CPB) is a major threat to cocoa production in South-East Asia. Crop losses to farmers can be up to 80%. Until now, management of CPB has heavily relied on pesticide applications, which is neither environmentally sustainable nor economically effective. An ARS scientist in Beltsville, Maryland, has developed an effective attract & kill method using the CPB sex pheromone as the attractant, and cypermethrin as the pesticide, to control CPB populations in cocoa plantations. The attract and kill technology uses 6 to 12 thousand times less attractant than mass trapping or mating disruption technologies, respectively, and 300 times less pesticide than traditional chemical spray treatments. It has great potential to be an efficient, safe, and environmentally friendly alternative in CPB control programs.

Review Publications
Sparks, M., Bottner-Parker, K.D., Gundersen, D.E., Lee, I. 2018. Draft genome sequence of the New Jersey aster yellows strain of ‘Candidatus Phytoplasma asteris’. PLoS One. https://doi.org/10.1371/journal.pone.0192379.
Blackburn, M.B., Farrar, R.R., Sparks, M., Kuhar, D.J., Mowery, J.D., Mitchell, A.D., Gundersen, D.E. 2019. Chromobacterium phragmitis sp. nov., an insecticidal bacterium isolated from estuarine marshes. International Journal of Systematic and Evolutionary Microbiology. https://doi.org/10.1099/ijsem.0.003508.
Bredlau, J., Kuhar, D.J., Gundersen, D.E., Kester, K. 2019. The parasitic wasp, Cotesia congregata, consists of two incipient species isolated by asymmetric reproductive incompatibility and hybrid inability to overcome host defenses. Frontiers in Ecology and Evolution. https://doi.org/10.3389/fevo.2019.00187.
Farrar, R.R., Gundersen, D.E., Kuhar, D.J., Blackburn, M.B. 2018. Insecticidal activity of Chromobacterium vaccinii. Journal of Entomological Science. 53:339-346.
Feng, Y., Bruton, R.G., Park, A.L., Zhang, A. 2018. Identification of attractive blend for spotted wing drosophila, Drosophila suzukii from apple juice. Journal of Pest Science. https://doi.org/10.1007/s10340-018-1006-9.
Harrison, R.L., Herniou, E.A., Jehle, J.A., Theilmann, D.A., Burand, J.P., Becnel, J.J., Krell, P.J., Van Oers, M.M., Mowery, J.D., Bauchan, G.R. 2018. ICTV virus taxonomy profile: Baculoviridae. Journal of General Virology. 99:1185-1186.
Lancaster, J., Khrimian, A., Young, S., Lehner, B., Luck, K., Wallingford, A., Ghosh, S.B., Zerbe, P., Marek, P., Sparks, M., Tittiger, C., Kollner, T.G., Weber, D.C., Gundersen, D.E., Kuhar, T., Tholl, D. 2018. De novo formation of an aggregation pheromone precursor by an isoprenyl diphosphate synthase-related terpene synthase in the harlequin bug. Proceedings of the National Academy of Sciences. https://doi.org/10.1073/pnas.1800008115.
Lancaster, J., Lehner, B., Khrimian, A., Muchlinsky, A., Luck, K., Kollner, T., Weber, D.C., Gundersen, D.E., Tholl, D. 2018. An IDS type sesquiterpene synthase produces the pheromone precursor (Z)-a-bisabolene in Nezara viridula. Journal of Chemical Ecology. https://doi.org/10.1007/s10886-018-1019-0.
Vanhove, W., Zhang, A., Osman, H.I., Vanhoudt, N., Van Damme, P. 2019. Development of an attract-and-kill strategy for Cocoa Pod Borer (Conopomorpha cramerella Snellen) control. Pest Management Science. https://doi.org/10.1080/09670874.2019.1578003.
Zhang, A., Feng, Y., Larson, N., Feldlaufer, M.F. 2018. Eucalyptol detected by aeration from the eggs of the common bed bug (Hemiptera: cimicidae). Journal of Economic Entomology. https://doi.org/10.1093/jee/toy385.
Farrar, R.R., Gundersen, D.E., Kuhar, D.J., Blackburn, M.B. 2018. Insecticidal activity of a recently described bacterium, Chromobacterium sphagni. Journal of Entomological Science. 53:333-338.