Progress 10/01/14 to 09/30/15
Outputs
Progress Report Objectives (from AD-416): The goal of this project is to develop innovative and sustainable technologies to manage insect pests and their natural enemies to reduce damage to trees, shrubs, and flowering perennial plants in managed and unmanaged urban landscapes. Lepidopteran and coleopteran pests are the principal defoliators in urban landscapes and nearby forests, and constitute 60% of recorded invasive insect species in North America. The research focuses on five of the most destructive lepidopteran and coleopteran pests in urban landscapes: gypsy moth, winter moth, and dogwood borer; and emerald ash borer and Asiatic garden beetle; however, research results are expected to be broadly applicable to other urban landscape insect pests. The research is organized around three objectives: (1) identify and synthesize plant-derived semiochemicals and insect pheromones attractive to pests, and design insect monitoring and management strategies employing them; (2) develop microbial and arthropod natural enemies as biocontrol agents of native and exotic pests of landscape plants; and (3) identify at the molecular level genes and gene pathways critical for success of landscape pests; employ molecular technologies to determine the role of host pathways targeted by entomopathogens; and target these pathways with natural or molecular biopesticides to kill pests. Approach (from AD-416): 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 strategies to combat urban landscape pests. Plant-derived semiochemicals and insect pheromones will be identified, and novel chemical and biological syntheses will be devised to enable their use for monitoring pest populations, disrupting pest reproduction, and luring pests into traps to be exposed to pathogenic microorganisms. Ecological attributes of pathogens underlying virulence, specificity, and accessibility to hosts will be determined. Transcriptome analysis of gypsy moth larvae challenged with various classes of pathogens will expose insect molecular vulnerabilities exploited by virulent pathogens. Naturally occurring pathogens best able to exploit these vulnerabilities will be applied directly to the environment, or placed in traps with species-specific semiochemicals to lure pests into contact with the pathogens to initiate epizootics. Experimental native and exotic assemblages of woody and herbaceous urban landscape plants will be characterized according to the natural enemy resources that they provide, and tested for their ability to support natural enemies and thereby to promote effective biocontrol of urban landscape pests. This Project falls under NP 304 component �Protection of Natural Ecosystems;� and has three objectives. Objective 1: Identify and synthesize plant-derived chemicals and insect pheromones attractive to pests; design pest monitoring and management strategies employing them. Pheromones are urgently needed to control spotted-wing drosophila (SWD), an invasive and highly destructive new orchard pest. Some volatiles derived from fermented apple have been identified in laboratory bioassays as SWD attractants. Field tests are underway to evaluate their attractiveness in real-world situations. These results are of interest to scientists and IPM managers. Brown marmorated stink bug (BMSB) is a devastating orchard pest and a noxious nuisance pest in urban environments. Chemical lures are needed to monitor and control this pest. Earlier, ARS scientists in Beltsville, Maryland, identified two principal pheromone components of the BMSB aggregation pheromone, but our synthetic method provided a low yield for one of them. Efforts during the past year focused on developing a new synthetic procedure to enable scaling-up the two pheromone components in quantities sufficient to conduct ratio, dose-response, and other field bioassays. Through a meticulous literature search a natural amino acid- based catalyst was found that could be used to produce both pheromone components from one reaction in about equal quantities. That catalyst was synthesized from the amino acid (proline) and obtained promising preliminary results, which are of interest to scientists and IPM managers. Objective 2: Develop microbial and arthropod natural enemies as biocontrol agents of landscape plant pests. Novel pathogens are needed to control winter moth (WM), a polyphagous defoliator and fruit pest of deciduous trees and shrubs. ARS scientists in Beltsville, Maryland, developed a new WM cell line from embryonic tissue that is susceptible to infection with WM baculovirus. We also assembled a preliminary draft of the WM baculovirus genome sequence. These tools will assist scientists in developing virus-based control measures for this pest. GypchekTM is a commercial baculovirus formulation used by USDA-Forest Service in the �Slow-the-Spread� program for gypsy moth (GM) control. Previous bioassays with Asian gypsy moth revealed that the baculovirus in GypchekTM did not perform as well as other GM baculovirus strains. A different stock of GypchekTM virus was obtained from a Canadian company and tested against USDA-FS�s GypchekTM in larval bioassays to see if there are differences among GypchekTM stocks. ARS scientists in Beltsville, Maryland, found no consistent differences in insecticidal activity between the two virus stocks. This result is of interest to USDA- FS and other GM control agencies. Larvae in different Asian and European gypsy moth (GM) colonies require different levels of dietary iron for optimal growth and development. To determine if different levels of dietary iron also affect susceptibility to baculovirus infection, ARS scientists in Beltsville, Maryland, set up bioassays with larvae infected and reared on diet differing in dietary iron concentration. No differences in susceptibility to baculovirus infection at different dietary iron levels were found. These results indicate that additional dietary iron has no effect on the results of gypsy moth/baculovirus bioassays. These results are of interest to scientists developing methods for viral control of GM populations. It is generally assumed that native plant species are more desirable than exotic ones for urban ornamental plantings, but little scientific evidence has been gathered to support this notion. ARS scientists in Beltsville, Maryland, set up a residential-scale experiment to determine whether urban landscapes composed of native plants have more insect natural enemies, and hence better biological control of insect pests, than those comprising exotic plants. Brown marmorated stink bug egg masses were attached to leaves of trees and shrubs in these experimental landscapes, and found statistically significantly more spiders resting near egg masses in the native than in the exotic plantings, thus supporting the hypothesis. These results are of interest to scientists, gardeners, and urban planners. Objective 3: Identify gene pathways critical for success of landscape pests; use molecular methods to identify host pathways targeted by insect pathogens; target them with natural or molecular bio-insecticides. Gypsy moth (GM) is a voracious and rapidly spreading insect pest of the U.S. urban landscape. ARS scientists in Beltsville, Maryland, are pursuing novel approaches to GM control. With colleagues at the University of Maryland, College Park, we developed genetically engineered plants that will allow us to test a gene that is expressed at high levels after insect attack begins. Also, we have tested the promoter for poplar gene pot167 and found it to be strongly wound-induced, making it a useful part of an RNAi construct, since it is known to be strongly responsive to infestation (Lawrence et al. Biotechnology Letters [2006] 28:1493�1501). A plant producing large amounts of the anti-GM agent only as needed is advantageous since it would be present only during insect attack, thereby reducing the amount of the transgene in the environment. Another advantage is that this new genetic material has not been patented and is freely available for public use. Ultimately new genetic tree stocks may be available that resist gypsy moth attack. This research is of interest to plant breeders and landscapers. RNA interference (RNAi) for brown marmorated stink bug (BMSB) control. RNAi deactivates or silences specific genes to disable a target organism. Utilizing our published BMSB, ARS scientists in Beltsville, Maryland, identified specific molecular gene targets for an RNAi strategy against BMSB, a devastating exotic and invasive landscape pest. Delivery methods to enable ingestion of RNAi by BMSB were investigated and used to evaluate RNAi efficacy in vivo. Accomplishments 01 Comparative transcriptomes for brown marmorated stink bug generated. Halyomorpha halys (St�l) (Heteroptera: Pentatomidae), the brown marmorated stink bug (BMSB), is an invasive insect native to Asia that has emerged as an important major insect pest in the United States. BMSB is a polyphagous piercing/sucking feeder. It poses considerable ecological and economic threat to specialty crops (apples, stone and pome fruits, grapes, ornamental plants, vegetables, seed crops) as well staple crops (soybean and corn); it is also an indoor nuisance pest. ARS scientists in Beltsville, Maryland, determined the total complement of active genes - the transcriptome - of different BMSB life stages and both sexes of the adult stage. Comparison of these transcriptomes provided the basis for identifying a wide variety of differentially expressed transcripts and a reliable source of BMSB genes involved in key physiological processes. Distinct transcripts were identified that were differentially regulated or induced in specific BMSB life stages, were associated with particular life stages or sexes, or were associated with highly specific biological systems. These represent candidate BMSB-specific gene targets that may be candidates for disruption by molecular biopesticide technologies that could cause negative impacts on the insect. These results will be of interest to biopesticide companies developing new insect control technologies.
Impacts
(N/A)
Publications
- Khrimian, A., Fay, H.A., Guzman, F., Chauhan, K.R., Moore, C., Aldrich, J. R. 2012. Pheromone of the banana-spotting bug, amblypelta lutescens lutescens Distant (Hemiptera: Coreidae): identification, synthesis and field bioassay. Psyche. Article ID 536149, pgs.1-8.
- Khrimian, A., Zhang, A., Weber, D.C., Ho, H., Aldrich, J., Vermillion, K., Siegler, M., Shirali, S., Guzman, F., Leskey, T.C. 2014. Discovery of the Aggregation Pheromone of the Brown Marmorated Stink Bug (Halyomorpha halys) through the Creation of Stereoisomeric Libraries of 1-Bisabolen-3-ols. Journal of Natural Products. 77:1708-1717.
- Abraham, J., Zhang, A., Abubeker, S.U., Angeli, S., Rodriquez-Saona, C. 2015. Behavioral and antennal responses of spotted wing drosophila, drosophila suzukii, to volatiles from fruit extracts. Environmental Entomology. 44(2):356-367.
- Carrillo-Tripp, J., Krueger, E.N., Harrison, R.L., Toth, A.L., Miller, W., Bonning, B.C. 2014. Lymantria dispar iflavirus 1 (LdIV1), a new model to study iflaviral persistence in lepidopterans. Journal of General Virology. 95(10):2285-2296.
- Hock, V., Chouinard, G., Lucas, E., Cormier, D., Leskey, T.C., Zhang, A. 2015. Behavioral responses of plum curculio (Coleoptera: Curculionidae) to different enantiomer concentrations and blends of the synthetic aggregation pheromone grandisoic acid. Journal of Economic Entomology. 108(2):549-558.
- Li, H., Pan, X., Chen, L., Wang, D., Zhang, A., Wang, M. 2015. Expression profile and ligand-binding characterization of odorant-binding protein 2 in Batocera horsfieldi (Hope). Journal of Applied Entomology. 139(5):361- 369.
- Khrimian, A., Shirali, S., Vermillion, K., Siegler, M.A., Guzman, F., Chauhan, K.R., Aldrich, J.R., Weber, D.C. 2014. Determination of the stereochemistry of the aggregation pheromone of harlequin bug, Murgantia histrionica. Journal of Chemical Ecology. 40:1260-1268.
- Nakai, M., Harrison, R.L., Uchida, H., Ukuda, R., Hikihara, S., Ishii, K., Kunimi, Y. 2015. Isolation of an adoxophyes orana granulovirus (AdorGV) occlusion body morphology mutant: Biological activity, genome sequence, and relationship to other isolates of AdorGV. Journal of General Virology. 96(4):904-914.
- Simmons, A.M., Weber, D.C., Payton, M.E., Hu, J.S., Greenstone, M.H. 2015. Do heteropterans have longer molecular prey detectability half-lives than other predators? A test with Geocoris punctipes (Geocoridae) and Orius insidiosus (Anthocoridae). Journal of Entomological Science. 50(2):99-105.
- Vanhove, W., Vanhoudt, N., Bhanu, K., Damme, P., Abubeker, S.U., Zhang, A. 2015. Geometric isomers of sex pheromone components do not affect attractancy of Conopomorpha cramerella in cocoa plantations. Journal of Applied Entomology. DOI:10.1111/jen.12212.
- Harrison, R.L., Rowley, D.L. 2015. Complete genome sequence of the strain of Lymantria dispar multiple nucleopolyhedrovirus found in the gypsy moth biopesticide, Virin-ENSh.. Genome Announcements. 3(1):e0147-14.
- Sparks, M., Shelby, K., Kuhar, D.J., Gundersen, D.E. 2014. Transcriptome of the invasive brown marmorated stink bug, halyomorpha halys (st�l) (heteroptera: pentatomidae). PLoS One. 9(11_:e111646. DOI: 10.1371/journal. pone.0111646.
- Gammon, D.B., Duraffour, S., Hehnly, H., Rozelle, D.K., Sharma, R., Sparks, M., West, C.C., Andrei, G., Connor, J.H., Conte, J., Doxsey, S., Gundersen, D.E., Marshall, W.L., Silverman, N., Mello, C.C. 2014. A single vertebrate DNA virus protein disarms invertebrate immunity to RNA virus infection. Cell Host and Microbe. eLife 25:3. doi: 10.7554/eLife.02910.
- Laumann, R., Borges, M., Aldrich, J.R., Khrimian, A., Blassioli Moraes, M. 2012. Attractant pheromone of the neotropical species Neomegalotomus parvus (Westwood)(Heteroptera: Alydidae). Psyche. 1(1):27-35.
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Progress 10/01/13 to 09/30/14
Outputs
Progress Report Objectives (from AD-416): The goal of this project is to develop innovative and sustainable technologies to manage insect pests and their natural enemies to reduce damage to trees, shrubs, and flowering perennial plants in managed and unmanaged urban landscapes. Lepidopteran and coleopteran pests are the principal defoliators in urban landscapes and nearby forests, and constitute 60% of recorded invasive insect species in North America. The research focuses on five of the most destructive lepidopteran and coleopteran pests in urban landscapes: gypsy moth, winter moth, and dogwood borer; and emerald ash borer and Asiatic garden beetle; however, research results are expected to be broadly applicable to other urban landscape insect pests. The research is organized around three objectives: (1) identify and synthesize plant-derived semiochemicals and insect pheromones attractive to pests, and design insect monitoring and management strategies employing them; (2) develop microbial and arthropod natural enemies as biocontrol agents of native and exotic pests of landscape plants; and (3) identify at the molecular level genes and gene pathways critical for success of landscape pests; employ molecular technologies to determine the role of host pathways targeted by entomopathogens; and target these pathways with natural or molecular biopesticides to kill pests. Approach (from AD-416): 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 strategies to combat urban landscape pests. Plant-derived semiochemicals and insect pheromones will be identified, and novel chemical and biological syntheses will be devised to enable their use for monitoring pest populations, disrupting pest reproduction, and luring pests into traps to be exposed to pathogenic microorganisms. Ecological attributes of pathogens underlying virulence, specificity, and accessibility to hosts will be determined. Transcriptome analysis of gypsy moth larvae challenged with various classes of pathogens will expose insect molecular vulnerabilities exploited by virulent pathogens. Naturally occurring pathogens best able to exploit these vulnerabilities will be applied directly to the environment, or placed in traps with species-specific semiochemicals to lure pests into contact with the pathogens to initiate epizootics. Experimental native and exotic assemblages of woody and herbaceous urban landscape plants will be characterized according to the natural enemy resources that they provide, and tested for their ability to support natural enemies and thereby to promote effective biocontrol of urban landscape pests. This Project falls under NP 304 component �Protection of Natural Ecosystems;� it has three objectives. Objective 1: Identify and synthesize plant-derived chemicals and insect pheromones attractive to pests; design pest monitoring and management strategies employing them. A gene encoding a protein that is potentially involved in dogwood borer pheromone biosynthesis was isolated from the pheromone glands of female adult dogwood borer moths. Two alleles of this gene were inserted into insect virus vectors for production of the encoded proteins in insect cells. Experiments are underway to produce the proteins and evaluate their enzymatic activity. Objective 2: Develop microbial and arthropod natural enemies as biocontrol agents of landscape plant pests. A new virus was identified in a cell line derived from the gypsy moth. This virus was also detected in other gypsy moth cell lines and in gypsy moth eggs and tissues dissected from gypsy larvae and adults. Infectious particles of this virus were used to successfully infect a gypsy moth cell line that did not already contain the virus. A new winter moth cell line that potentially can be used for winter moth virus infection and propagation was developed from embryonic tissue. This cell line has undergone ten passages in culture, and is being adapted to a variety of different culture conditions and cell media. Objective 3: Identify gene pathways critical for success of landscape pests; use molecular methods to identify host pathways targeted by insect pathogens; target them with natural or molecular bio-insecticides. Gypsy moth is a voracious and rapidly spreading insect pest of the nation�s urban landscape. Scientists at USDA-ARS are pursuing novel approaches to control this pest. With scientists at the University of Maryland, College Park, they have developed genetically engineered plants that will allow them to test a gene that is only expressed after insect attack begins. A plant producing an anti-gypsy moth agent as needed would be advantageous since the product would be present only during insect attack reducing the amount found in the environment. Another advantage to this work is that this new genetic material has not been patented and is freely available for public use. Ultimately new genetic tree stocks may be available that resist gypsy moth attack. RNA interference (RNAi) deactivates specific genes to disable a target organism. To identify molecular targets for an RNAi strategy against Brown Marmorated Stink Bug (BMSB), a devastating exotic and invasive landscape pest, the total complement of active genes - the transcriptome - in different BMSB life stages and both sexes of adult, and selected candidate differentially expressed genes as candidate BMSB-specific RNAi targets were identified. For gypsy moth (GM), prior year analyses of transcriptomes were used to identify differentially expressed genes as candidate GM-specific RNAi targets. Double-stranded RNAs (DsRNAs) were tested individually for efficacy in causing distress to live caterpillars. Significant Activities that Support Special Target Populations: Inner-city minority populations depend upon green spaces for their appreciation of the natural world; also, they receive relatively little exposure to agriculture and agricultural research. Under Objective 2 of this project, a residential-scale experiment on the role of exotic vs. native plants in determining the efficacy of biological control of insect pests was sited at the U.S. National Arboretum in Northeast Washington, D. C. Through professional signage and day-to-day interactions with ARS scientists, local residents are introduced to the need for and principles of insect biocontrol, experimental design, and hypothesis testing, and are encouraged to think about their own roles in fostering the sustainability of urban landscape plantings. Local school children take field trips to the Student Discovery Garden on the campus of the Beltsville Agricultural Research Center, where a microcosm of this experiment is set up. Here they interact with ARS scientists, and are stimulated to ask questions about the role of landscape plants and plant-insect interactions in urban environments. Accomplishments 01 Biological and genomic characterization of gypsy moth virus isolates with superior insecticidal activity against Asian gypsy moth. Gypsy moth (GM) is the most serious woody landscape pest in the U.S. Two foreign sub-species of GM, collectively referred to as Asian gypsy moth, are potentially invasive strains that threaten to cause more extensive damage to landscapes in the U.S. due to their wider plant host range and greater capacity to disseminate to distant areas with a suitable habitat. A GM biopesticide, Gypchek, developed from a GM virus isolated in Connecticut, has been used to control GM outbreaks in the U.S. and Canada. To identify isolates of this GM virus with greater activity against Asian gypsy moth, GM virus isolates from Japan, Korea, and Russia were tested along with Gypchek in bioassays against Asian gypsy moth strains from Japan, Siberia, far east Russia, and Lithuania, as well as GM strains from Greece and Connecticut, USA. The Japan, Korea, and Russia virus isolates exhibited 2- to 6-fold greater activity than Gypchek against the Asian gypsy moth strains at the lower virus dose range. Genome sequences of these viruses revealed genetic differences that correlated with their geographic origins. These new GM virus isolates can be developed as biopesticides to control outbreaks of Asian gypsy moth in the U.S., especially in ecologically sensitive areas, and their genome sequences can be used in EPA registrations of the viruses as biopesticides.
Impacts
(N/A)
Publications
- Sparks, M., Gundersen, D.E., Harrison, R.L. 2013. Complete genome sequence of a novel iflavirus from the transcriptome of Halyomorpha halys, the brown marmorated stink bug.. Genome Announcements. 1(6):e00910-13.
- Hock, V., Chouinard, G., Lucas, E., Cormier, D., Leskey, T.C., Wright, S.E. , Zhang, A., Pichette, A. 2014. Establishing abiotic and biotic factors necessary for reliable male pheromone production and attraction to pheromones by female plum curculios Conotrachelus nenuphar (Coleoptera: Curculionidae). The Canadian Entomologist. DOI.org/10.4039/tce.2014.1.
- Baldwin, R.L., Zhang, A., Fultz, S., Abubeker, S.U., Harris, C., Connor, E. E., Van Hekken, D.L. 2014. Hot Topic: Brown marmorated stink bug odor compounds do not transfer into milk by feeding bug-contaminated corn silage to lactating dairy cattle. Journal of Dairy Science. 97:1877-1884.
- Lin, F., Ye, J., Wan, H., Zhang, A., Zhao, B. 2013. Host deception: Predaceous fungus, esteya vermicola, entices pine wood nematode by mimicking the scent of its host pine for nutrient. PLoS One. 8(8):1-9.
- Li, H., Zhang, A., Wang, M., Zhang, G. 2014. Construction and analysis of cDNA libraries from the antennae of Batocera horsfieldi and expression pattern of putative odorant binding proteins. Journal of Insect Science. 14(57):1-15.
- Greenstone, M.H., Tillman, P.G., Hu, J.S. 2014. Predation of the newly invasive pest Megacopta cribraria (F.) (Hemiptera: Plataspidae) in soybean habitats adjacent to cotton by a complex of predators. Journal of Economic Entomology. 107:947-954.
- Rijal, J., Zhang, A., Bergh, C. 2013. Behavioral response of grape root borer (Lepidopetera: Sesiidae) neonates to grape root volatiles. Environmental Entomology. 42(6):1338-1347.
- Frank, D., Zhang, A., Wright, S.E., Walgenbach, J., Bergh, C.J., Leskey, T. C. 2014. Effect of a pheromone antagonist-based disruption blend on dogwood borer (Lepidoptera: Sesiidae) mate-finding and infestation in a commercial apple orchard. Journal of Entomological Science. 49(1):44-45.
- Hoover, K., Keena, M., Nehme, M., Wang, S., Meng, P., Zhang, A. 2014. Sex- specific trail pheromone mediates complex mate finding behavior in Anoplophora glabripennis (Coleoptera: Cerambycidae). Journal of Chemical Ecology. 40(2):169-180.
- Greenstone, M.H., Peyton, M.E., Weber, D.C., Hu, J.S., Simmons, A.M. 2013. The detectability half-life in predator-prey research: what it is, why we need it, how to measure it, and what it�s good for. Molecular Ecology. DOI:10.1111/mec.12552.
- Harrison, R.L., Keena, M.A., Rowley, D.L. 2014. Classification, genetic variation and pathogenicity of Lymantria dispar nucleopolyhedrovirus isolates from Asia, Europe, and North America. Journal of Invertebrate Pathology. 116:27-35.
- Blackburn, M.B., Martin, P.A., Kuhar, D.J., Farrar, R.R., Gundersen, D.E. 2014. Crystalliferous Bacillus cereus group bacteria from a Maryland hardwood forest are dominated by psychrotolerant strains. MicrobiologyOpen. DOI:10.1002/mbo3.189.
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Progress 10/01/12 to 09/30/13
Outputs
Progress Report Objectives (from AD-416): The goal of this project is to develop innovative and sustainable technologies to manage insect pests and their natural enemies to reduce damage to trees, shrubs, and flowering perennial plants in managed and unmanaged urban landscapes. Lepidopteran and coleopteran pests are the principal defoliators in urban landscapes and nearby forests, and constitute 60% of recorded invasive insect species in North America. The research focuses on five of the most destructive lepidopteran and coleopteran pests in urban landscapes: gypsy moth, winter moth, and dogwood borer; and emerald ash borer and Asiatic garden beetle; however, research results are expected to be broadly applicable to other urban landscape insect pests. The research is organized around three objectives: (1) identify and synthesize plant-derived semiochemicals and insect pheromones attractive to pests, and design insect monitoring and management strategies employing them; (2) develop microbial and arthropod natural enemies as biocontrol agents of native and exotic pests of landscape plants; and (3) identify at the molecular level genes and gene pathways critical for success of landscape pests; employ molecular technologies to determine the role of host pathways targeted by entomopathogens; and target these pathways with natural or molecular biopesticides to kill pests. Approach (from AD-416): 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 strategies to combat urban landscape pests. Plant-derived semiochemicals and insect pheromones will be identified, and novel chemical and biological syntheses will be devised to enable their use for monitoring pest populations, disrupting pest reproduction, and luring pests into traps to be exposed to pathogenic microorganisms. Ecological attributes of pathogens underlying virulence, specificity, and accessibility to hosts will be determined. Transcriptome analysis of gypsy moth larvae challenged with various classes of pathogens will expose insect molecular vulnerabilities exploited by virulent pathogens. Naturally occurring pathogens best able to exploit these vulnerabilities will be applied directly to the environment, or placed in traps with species-specific semiochemicals to lure pests into contact with the pathogens to initiate epizootics. Experimental native and exotic assemblages of woody and herbaceous urban landscape plants will be characterized according to the natural enemy resources that they provide, and tested for their ability to support natural enemies and thereby to promote effective biocontrol of urban landscape pests. Objective 1: Identify and synthesize plant-derived chemicals and insect pheromones attractive to pests; design pest monitoring and management strategies employing them. Two approaches to producing 7-epi-sesquithujene (7ES), a long-distance attractant for the emerald ash borer (EAB). The Asian invasive EAB threatens all native ash species. First, we made insect virus vectors that carry the TPS4 gene, which directs synthesis of 7ES, but they did not produce TPS4 in insect cells. Though we successfully introduced alternative non-viral vectors into insect cells, the cells became contaminated with fungus, indicating a need for new methods. We then developed a synthesis for 7ES using commercially available chemicals. 7ES was not an effective EAB lure in field traps, either alone or in combination with (3Z)-hexenol, a generalized insect attractant. We plan further field tests of 7ES as an EAB attractant. Objective 2: Develop microbial and arthropod natural enemies as biocontrol agents of landscape plant pests Virulent new gypsy moth (GM) nucleopolyhedrovirus (NPV) isolates. GM is the most serious landscape pest in the US, and new biocontrol agents are needed. NPVs from Asia and Massachusetts (MA) were tested for toxicity against GM caterpillars along with Gypchek, the NPV product used by the Forest Service. MA and Russian isolates were about twice as toxic as Gypchek. Genetic analysis of these and several other isolates found that the Asian isolates are separate and distinct from North American and European isolates. Asian isolates appear to be more toxic against Asian GM than Gypchek. Native parasitoids are more diverse in native than exotic urban landscapes. Native natural enemies are expected to be more effective biocontrol agents on native than on exotic plants. In a residential-scale experiment, we found that parasitic wasps are more abundant, and comprise more species, in habitats dominated by native than by exotic plants. Objective 3: Identify gene pathways critical for success of landscape pests; use molecular methods to identify host pathways targeted by insect pathogens; target them with natural or molecular bio-insecticides. RNA interference (RNAi) for brown marmorated stink bug (BMSB) and gypsy moth (GM) control. RNAi deactivates specific genes to disable a target organism. To identify molecular targets for an RNAi strategy against BMSB, a devastating exotic and invasive landscape pest, we determined the total complement of active genes - the transcriptome - in different BMSB stages and both sexes. In the process we recovered the complete gene sequence of a novel virus that may be useful as a BMSB biocontrol agent. For GM, we analyzed transcriptomes to compare active genes in tissues of healthy GM caterpillars, versus those infected with the bacterium Bacillus thuringiensis, the fungus Varimorpha dispar, or a second bacterium. Differences in levels of gene transcription (copying from DNA to RNA) in healthy and infected tissues revealed ten candidate GM-specific RNAi targets. These are being tested for efficacy in live caterpillars. Significant Activities that Support Special Target Populations: Inner-city minority populations depend upon green spaces for their appreciation of the natural world; also, they receive relatively little exposure to agriculture and agricultural research. Under Objective 2 of this project, a residential-scale experiment on the role of exotic vs. native plants in determining the efficacy of biological control of insect pests was sited at the U.S. National Arboretum in Northeast Washington, D. C. Through professional signage and day-to-day interactions with ARS scientists, local residents are introduced to the need for and principles of insect biocontrol, experimental design, and hypothesis testing, and are encouraged to think about their own roles in fostering the sustainability of urban landscape plantings. Local school children take field trips to the Student Discovery Garden on the campus of the Beltsville Agricultural Research Center, where a microcosm of this experiment is set up. Here they interact with ARS scientists, and are stimulated to ask questions about the role of landscape plants and plant-insect interactions in urban environments. Accomplishments 01 Comparison of healthy versus Bacillus thuringiensis (Bt)- infected gypsy moth genetic components revealed important genes that could be involved in resistance development or biological control. Gypsy moth (GM) is the most serious woody landscape pest in the U.S. Nevertheless, almost no genetic information existed for GM, even though this information could help scientists devise new biological control strategies and understand development of resistance to biocontrol and other management strategies. We determined the total genetic components, called the �transcriptome�, produced in midgut tissue in healthy and in Bacillus thuringiensis (Bt)-infected GM caterpillars. High- throughput approaches were used to assemble and analyze gene expression (activity) profiles. The genes exhibiting increased or reduced activity in response to Bt infection were primarily associated with functions of digestion; immune response; development; or binding (acid-, lipid-, protein-, or nucleic acid-binding). This is the first study using large- scale sequencing technologies to specifically investigate how GM responds to a bacterial infection challenge. The results highlight important genes that could be involved in biopesticide resistance development, or could serve as targets for new biopesticides in biologically-based control strategies of this devastating pest.
Impacts
(N/A)
Publications
- Blackburn, M.B., Martin, P.A., Kuhar, D.J., Farrar, R.R., Gundersen, D.E. 2013. Phylogenetic distribution of phenotypic traits in bacillus thuringiensis analyzed by multilocus sequence typing . PLoS One. 8(6) :e66061.
- Gundersen, D.E., Dupuy, C., Huguet, E., Drezen, J. 2013. Polydnaviruses: Roles in insect pathology and applications. Biocontrol Science and Technology. 23:1-61.
- Ryall, K.L., Silk, P.J., Mayo, P., Crook, D., Khrimian, A., Cosse, A.A., Sweeney, J., Scarr, T. 2012. Attraction of agrilus planipennis fairmaire (Coleoptera: Buprestidae) to a volatile pheromone: effects of release rate, host volatile and trap placement. Environmental Entomology. 41(3):648-656.
- Brockerhoff, E., Suckling, D., Kimberley, M., Richardson, B., Coker, G., Gous, S., Cowan, D., Lance, D., Strand, T., Zhang, A. 2012. Mating disruption by aerial application of sex pheromone against the invasive light brown apple moth and implications for the management of biological invasions. Meeting Proceedings. 42(3):558-563.
- Hansen, J., Klingeman, W., Moulton, J., Oliver, J., Windham, M., Zhang, A., Trigiano, R.2012. Molecular phylogeny of Synanthedonini (Lepidoptera: Sesiidae) using cytochrome oxidase I with comparison of morphological variation among some members justifying their generic reclassification. Annals of the Entomological Society of America. 42(3):558-563.
- Sparks, M., Blackburn, M.B., Kuhar, D.J., Gundersen, D.E. 2013. Transcriptome of the lymantria dispar (gypsy moth) larval midgut and its response to infection by bacillus thuringiensis. PLoS One. 8(5):e61190.
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Progress 10/01/11 to 09/30/12
Outputs
Progress Report Objectives (from AD-416): The goal of this project is to develop innovative and sustainable technologies to manage insect pests and their natural enemies to reduce damage to trees, shrubs, and flowering perennial plants in managed and unmanaged urban landscapes. Lepidopteran and coleopteran pests are the principal defoliators in urban landscapes and nearby forests, and constitute 60% of recorded invasive insect species in North America. The research focuses on five of the most destructive lepidopteran and coleopteran pests in urban landscapes: gypsy moth, winter moth, and dogwood borer; and emerald ash borer and Asiatic garden beetle; however, research results are expected to be broadly applicable to other urban landscape insect pests. The research is organized around three objectives: (1) identify and synthesize plant-derived semiochemicals and insect pheromones attractive to pests, and design insect monitoring and management strategies employing them; (2) develop microbial and arthropod natural enemies as biocontrol agents of native and exotic pests of landscape plants; and (3) identify at the molecular level genes and gene pathways critical for success of landscape pests; employ molecular technologies to determine the role of host pathways targeted by entomopathogens; and target these pathways with natural or molecular biopesticides to kill pests. Approach (from AD-416): 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 strategies to combat urban landscape pests. Plant-derived semiochemicals and insect pheromones will be identified, and novel chemical and biological syntheses will be devised to enable their use for monitoring pest populations, disrupting pest reproduction, and luring pests into traps to be exposed to pathogenic microorganisms. Ecological attributes of pathogens underlying virulence, specificity, and accessibility to hosts will be determined. Transcriptome analysis of gypsy moth larvae challenged with various classes of pathogens will expose insect molecular vulnerabilities exploited by virulent pathogens. Naturally occurring pathogens best able to exploit these vulnerabilities will be applied directly to the environment, or placed in traps with species-specific semiochemicals to lure pests into contact with the pathogens to initiate epizootics. Experimental native and exotic assemblages of woody and herbaceous urban landscape plants will be characterized according to the natural enemy resources that they provide, and tested for their ability to support natural enemies and thereby to promote effective biocontrol of urban landscape pests. Significant progress was made in achieving three objectives towards our overall goal. To address our goal to �Identify and synthesize plant- derived semiochemicals and insect pheromones attractive to pests� effective long-distance volatile attractants were made by several strategies for an emerald ash borer (EAB) monitoring trapping system. EAB invasive pest threatens native ash species. We developed an in vitro system to produce a plant volatile that stimulates EAB antennae. To produce an active enzyme, we constructed recombinant baculoviruses for expression in insect cells, and cloned a fruit fly gene for another enzyme that synthesizes a precursor line that produced the attractant in insect cells. We also synthesized 7ES directly from commercially available starting material. To address our goal to �Develop microbial and arthropod natural enemies as biocontrol agents of insect pests of landscape plants� we characterized baculovirus (NPV) isolates in the IIBBL virus collection at molecular and genome sequence levels for use against gypsy moth (GM), the most serious U.S. landscape pest. Isolates from Korea, Japan, Russia, Spain, and Massachusetts were selected for characterization by complete genome sequencing. A complete draft of the Massachusetts genome and partial drafts of genomes for other isolates were produced. To develop an alternative to winter moth larvae for evaluating NPVs for WM control, we initiated a cell line from WM embryos from dissected and homogenized eggs. We also identified a diverse group of non-crystal-forming Bacilli from forest soil that carried insecticidal toxin genes like those in Photorhabdus and Yersinia, but distinct from other bacteria. Multi-locus sequence typing analysis of B. thuringiensis populations (Bts) revealed great diversity even among close phylogenetic relatives. The majority of Bts belonged to few major phylogenetic groups. The distribution of crystal-forming Bacillus species from hardwood forests revealed a new group of mild pathogens related to B. weihenstephanensis which grow at low temperature. Sampling at different elevations revealed that these were dominant crystal-forming Bacilli on slopes and hilltops, but absent from floodplains. Floodplain soils were dominated by Bacilli belonging to an undescribed group producing trapezoidal crystals, which were occasionally toxic to GM. To address our goal to �Identify genes and gene pathways critical for success of pests; employ molecular technologies to determine the role of host pathways targeted by entomopathogens; and target these pathways to kill pests� we conducted studies and analyzed transcriptome profiles to identify expressed genes in GM cell line Ld652Y and healthy 3rd instar larvae for both midgut and hemocyte tissues. Tissues were separately infected with Bt and other microbial pathogens, and parasitized by braconid parasitoid G. flavicoxis. Transcriptome profiles were generated for each to assess differences in levels of gene transcription between healthy and pathogen-infected tissues and to identify target genetic pathways. This will allow us to determine molecular vulnerabilities of GM to pathogens and parasitoids. Accomplishments 01 Gypsy moth (cell line IPBL-Ld652Y) total expressed gene profile determin Almost no genetic information existed for gypsy moth (GM) even though this information could help scientists devise new biological control strategies. GM cells were analyzed to determine the total genetic components (called the �transcriptome�) produced by the cells. More than 14,000 transcripts were identified for GM. Numerous virus genetic components associated with the GM cells were also discovered, as well as several candidate genetic components of interest for developing into biological control methods.
Impacts
(N/A)
Publications
- Dupuy, C., Gundersen, D.E., Cusson, M. 2012. Genomics and Replication of Polydnaviruses. In: Beckage, N.E., Drezen, J.M., editors. Parasitoid Viruses: Symbionts and Pathogens. London, England: Elsevier. p. 47-61.
- Sparks, M., Gundersen, D.E. 2011. The Lymantria dispar IPLB-Ld652Y cell line transcriptome comprises diverse virus-associated transcripts. Viruses. 3(11): 2339-2350.
- Bergh, J.C., Zhang, A., Meyer, J.R., Kim, D. 2011. Response of grape root borer (lepidoptera: sesiidae) neonates to root extracts from vitaceae species and rootstocks. Environmental Entomology. 40:880-888.
- Cha, D., Linn, C., Teal, P.E., Zhang, A., Roelofs, W., Loeb, G. 2011. Eaves-dropping on plant volatiles by a specialist moth: significance of ratio and concentration. PLoS One. 6(2):e17033.
- Crook, D., Khrimian, A., Cosse, A.A., Frazer, I., Mastro, V. 2012. Influence of trap color and host volatiles on capture of the emerald ash borer, Agrilus planipennis (Coleoptera: Buprestidae). Journal of Environmental Entomology. 105:429-437.
- Loeb, G., Cha, D., Hesler, S., Linn, C., Zhang, A., Teal, P.E., Roelofs, W. 2011. Monitoring grape berry moth (Paralobesia vitianna: Lepidoptera) in commercial vineyards using a host plant based synthetic lure. Environmental Entomology. 4(6):1511-1522.
- Martin, P.A., Mongeon, P., Gundersen, D.E., Blackburn, M.B. 2011. Recovery of Bacillus thuringiensis and insect toxic related strains from forest soil. Applied and Environmental Microbiology. 2(7):285-291.
- Tobin, P.C., Zhang, A., Onufrieva, K., Leonard, D.S. 2011. A field evaluation of the effect of temperature on the release of disparlure from a pheromone-baited trapping system used to monitor the gypsy moth (Lepidotera: Lymantriidae). Journal of Economic Entomology. 104(4):1265- 1271.
- Frank, D., Zhang, A., Leskey, T.C., Bergh, J. 2011. Electrophysiological response of female dogwood borer (lepidoptera: sesiidae) to apple volatile compounds. Journal of Entomological Science. 46(3):204-215.
- Greenstone, M.H., Weber, D.C., Hu, J.S., Coudron, T.A., Payton, M.E. 2012. Removing external DNA contamination from arthropod predators destined for molecular gut-content analysis. Molecular Ecology Resources. 12(3):464-469.
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Progress 10/01/10 to 09/30/11
Outputs
Progress Report Objectives (from AD-416) The goal of this project is to develop innovative and sustainable technologies to manage insect pests and their natural enemies to reduce damage to trees, shrubs, and flowering perennial plants in managed and unmanaged urban landscapes. Lepidopteran and coleopteran pests are the principal defoliators in urban landscapes and nearby forests, and constitute 60% of recorded invasive insect species in North America. The research focuses on five of the most destructive lepidopteran and coleopteran pests in urban landscapes: gypsy moth, winter moth, and dogwood borer; and emerald ash borer and Asiatic garden beetle; however, research results are expected to be broadly applicable to other urban landscape insect pests. The research is organized around three objectives: (1) identify and synthesize plant-derived semiochemicals and insect pheromones attractive to pests, and design insect monitoring and management strategies employing them; (2) develop microbial and arthropod natural enemies as biocontrol agents of native and exotic pests of landscape plants; and (3) identify at the molecular level genes and gene pathways critical for success of landscape pests; employ molecular technologies to determine the role of host pathways targeted by entomopathogens; and target these pathways with natural or molecular biopesticides to kill pests. Approach (from AD-416) 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 strategies to combat urban landscape pests. Plant-derived semiochemicals and insect pheromones will be identified, and novel chemical and biological syntheses will be devised to enable their use for monitoring pest populations, disrupting pest reproduction, and luring pests into traps to be exposed to pathogenic microorganisms. Ecological attributes of pathogens underlying virulence, specificity, and accessibility to hosts will be determined. Transcriptome analysis of gypsy moth larvae challenged with various classes of pathogens will expose insect molecular vulnerabilities exploited by virulent pathogens. Naturally occurring pathogens best able to exploit these vulnerabilities will be applied directly to the environment, or placed in traps with species-specific semiochemicals to lure pests into contact with the pathogens to initiate epizootics. Experimental native and exotic assemblages of woody and herbaceous urban landscape plants will be characterized according to the natural enemy resources that they provide, and tested for their ability to support natural enemies and thereby to promote effective biocontrol of urban landscape pests. IIBBL scientists addressed the first project objective, �Identify and synthesize plant-derived semiochemicals and insect pheromones attractive to pests� by developing a conventional chemical synthesis for 7-epi- sesquithujene, a plant volatile attractive to emerald ash borer (EAB), Agrilus planipennis. EAB is an invasive Asian pest that threatens all native ash (Fraxinus) species. Effective long-distance volatile attractants have been needed to develop a trapping system for this pest and as a monitoring tool. In a parallel effort to produce this compound in an in-vitro cell culture system, we succeeded in producing an enzyme essential for its synthesis, the terpene synthase TPS4, in bacteria. IIBBL scientists addressed the second project objective, �Develop microbial and arthropod natural enemies as biocontrol agents of insect pests of landscape plants� by determining the genetic relatedness of soil bacterium Bacillus thuringiensis (Bt) bacterial strains chosen to include diverse combinations of biochemical traits, using multi-locus sequence typing. The soil bacterium Bt has been an important biocontrol agent for moth pests and determining the habitat-specificities of different varieties was crucial for the selection of Bts with improved persistence in urban landscapes. Results indicated that most soil Bts belong to a relatively few dominant groups. IIBBL scientists addressed the third project objective, �Identify genes and gene pathways critical for success of pests; employ molecular technologies to determine the role of host pathways targeted by entomopathogens; and target these pathways to kill pests� through conducting new transcriptomic studies of caterpillar responses to pathogen infection. New biological controls are needed for the gypsy moth (Lymantria dispar), the most serious woody landscape pest in the U.S. In order to determine molecular vulnerabilities of gypsy moth to pathogens and parasites, we initiated transcriptome studies to identify expressed genes in a cultured cell line and in specific larval tissues. Larval tissues were also infected with the microbial pathogen Bt or parasitized by the braconid parasitoid wasp Glyptapanteles indiensis to evaluate the pest gene differential regulation. We also demonstrated the ability of DNA from G. indiensis parasitoid wasp polydnavirus to integrate into DNA of gypsy moth larvae. Accomplishments 01 Absolute configuration and synthesis of 7-epi-sesquithujene. Emerald as borer (EAB), Agrilus planipennis, is an invasive Asian pest that threate all native ash tree (Fraxinus) species. One promising candidate attracta for this pest, the plant volatile 7-epi-sesquithujene, gives positive neurophysiological responses in olfactory pathways of both male and fema EAB. We determined the spatial arrangement of atoms of 7-epi- sesquithujene, which is necessary for its effective and economical synthesis. The attractiveness of the synthetic product to EAB is being studied under field conditions. This will enable use of effective long- distance volatile attractants in a trapping system as a monitoring tool for EAB.
Impacts
(N/A)
Publications
- Blackburn, M.B., Martin, P.A., Kuhar, D.J., Farrar, R.R., Gundersen, D.E. 2011. The occurrence of Photorhabdus-like toxin complexes in Bacillus thuringiensis. PLoS One. 6(3):e18122.
- Khrimian, A., Cosse, A.A., Crook, D.J. 2011. Absolute configuration of 7- epi-sesquithujene. Journal of Natural Products. 74(6):1414-1420.
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