1a. Objectives (from AD-416):
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.
1b. 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 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.
3. Progress Report:
Objective 1. Develop semiochemicals of significant landscape pests and natural enemies for monitoring and management. The two component aggregation pheromone of the brown marmorated stinkbug is an essential tool for managing this important agricultural and nuisance pest. The existing chemical synthesis provides a satisfactory yield for the major but not the minor component. Thus, we focused on the development of a new synthetic procedure that provides both components in appreciable yields and allows scaling-up of the pheromone in quantities sufficient to conduct ratio, dose-response, and other field bioassays. A careful review of the chemical literature revealed a natural amino acid-based catalyst that was used to produce similar quantities of both pheromone components in one reaction. A second procedure that we developed was based on a less costly inorganic base which was ultimately used for the scale up reaction. Due to the rapid spread of the invasive fruit fly species, spotted wing drosophila (SWD), across the United States, an efficient SWD detection tool 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. Objective 2. Develop microbial and arthropod natural enemies as biocontrol agents of native and exotic pests of landscape plants. A draft sequence of the genome of a new gypsy moth baculovirus isolate obtained from larvae collected near Harbin, China has been completed. Clonal isolates derived from the gypsy moth baculovirus field isolate 3029 have been generated and characterized by PCR and sequencing of variable regions within the isolate 3029 genome. New Chromobacterium isolates were obtained from a variety of aquatic habitats. Descriptions for two new species of insecticidal Chromobacterium were developed, using a combination of genetic and phenotypic characteristics. Assembly and annotation of Chromobacterium subtsugae-infected gypsy moth transcripts was performed to identify host genes potentially involved in Lepidopteran immune response to Gram-negative bacteria challenge. Results will shed light on the mechanisms of establishment and progression of disease in the insect host, potentially helping to identify ways to enhance the insecticidal potential of the bacterium. 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 scientists in Beltsville, Maryland, made progress in implementing 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 transriptomes, specific molecular gene targets were identified for an RNAi strategy against BMSB, a devastating exotic and invasive landscape pest. Delivery methods to enable ingestion of RNAi- inducing dsRNAs by BMSB were developed and used to evaluate RNAi efficacy in vivo. The ability to deliver RNAi-based molecular biopesticides to BMSB or other invasive insect pests through feeding will be essential to their use as biocontrol agents. ARS scientists in Beltsville initiated an international i5K working group effort to assemble and annotate the 1.2 Gb BMSB genome sequence, with eleven global university and government research institutions participating. This group has already discovered that, consistent with its generalist dietary habits, BMSB contains the largest number of gustatory receptors that has been observed among sequenced arthropods to date. The working group is currently annotating and cataloging a variety of opsin genes and identifying genes expressed in a sex-specific manner. The results could lead to development of new means for BMSB biocontrol, including RNAi-based methods.
1. RNA interference (RNAi)-based molecular biopesticides effective against European gypsy moth (Lymantria dispar). Gypsy moth is a voracious and rapidly spreading insect pest that defoliates millions of acres of hardwood forest annually, and often plagues the nation’s urban greenscapes. Using their recently published gypsy moth transcriptomes, ARS scientists in Beltsville, Maryland, used recently developed technologies to create RNA interference (RNAi)-based molecular biopesticides that target specific genes that are crucial to the survival of the moth. Several novel dsRNAs were specifically designed to deactivate or silence these genes, negatively impacting the normal physiology of the insect. These double-stranded RNAs were delivered by feeding and tested individually for efficacy in causing distress to live caterpillars, or in causing reduction of egg mass in adult females. These new and highly-specific molecular biopesticide technologies will be useful tools for controlling gypsy moth pests, negatively impacting the gypsy moth, while sparing non-target insects.
2. RNA interference (RNAi)-based molecular biopesticides effective against brown marmorated stink bug (BMSB), Halyomorpha halys (Stål) (Heteroptera: Pentatomidae). The BMSB, an invasive insect native to Asia, has rapidly emerged as a major insect pest in the United States. Well known to the public as an irritating indoor nuisance pest, BMSB is a polyphagous piercing/sucking feeder that 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). ARS scientists in Beltsville, Maryland, specifically designed novel double-stranded RNAs, based on their recently published BMSB transcriptomes, to deactivate or silence specific genes critical to BMSB. When fed to BMSB, these novel RNA interference (RNAi)-based molecular biopesticides caused detrimental effects such as inhibition of development or reduced fecundity. These new and highly-specific molecular biopesticide technologies will be useful tools for controlling BMSB pests through causing negative impact to the target insects without affecting non-target insects.
5. 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. A microcosm of this experiment has been set up at the Student Discovery Garden on the campus of the Beltsville Agricultural Research Center. School children and teachers from the D.C. - Baltimore metropolitan area take field trips to the Discovery Garden, where they interact with ARS scientists and are stimulated to ask questions about the role of landscape plants and plant-insect interactions in urban environments.
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