S1052: The Working Group on Improving Microbial Control of Arthropod Pests
(Multistate Research Project)
S1052: The Working Group on Improving Microbial Control of Arthropod Pests
Duration: 10/01/2012 to 09/30/2017
Statement of Issues and Justification
Broad-spectrum chemical insecticides continue to be the mainstay for control of arthropod pests in most agricultural systems as well as other natural and urban landscapes. While chemical pesticides are capable of rapidly killing various pests, heavy reliance on their use has generated various problems including safety risks due to human poisonings and death, outbreaks of secondary pests normally held in check by natural enemies, environmental contamination, decreases in biodiversity, and insecticide resistance. Thus, there is an urgent need to accelerate the development and implementation of cost-effective, environmentally safe alternatives to chemical pesticides for arthropod control. Changes in pest management programs, such as the reduction in organophosphate use dictated by the Food Quality Protection Act (FQPA), necessitate the development of new management tactics that are environmentally sound and compatible with current production practices. One viable alternative to chemical insecticides is the use of biological control agents. In contrast to chemical insecticides, biological control agents are generally not harmful to humans or the environment, and have minimal or negligible potential to cause resistance or harm non-target organisms. Biological control includes the use of insect predators, parasitoids, and pathogens. The focus of this project is on the development and advancement of entomopathogens for biological pest suppression.
Development of biological control tactics using entomopathogens (i.e., microbial control) is of great importance to US agriculture. The Experiment Station Committee on Organization and Policy (ESCOP), and National Association of State Universities and Land-Grant Colleges (NASULGC) have identified environmental stewardship, including the need to decrease chemical pesticide use, as a primary agricultural challenge in the US. Furthermore, most stakeholder groups developing strategic plans for pest management throughout the U.S. have identified biological control as a major research need, and many have specifically identified use of entomopathogens as a priority. Some examples that list the development of entomopathogens as a priority include pecan, peaches, apples and grapes (http://www.ipmcenters.org/).
Microbial control research and application has already had major impacts on IPM during the past fifty years. The commercialization of Bacillus thuringiensis (Bt) products, including Bt-transgenic plants, is probably the most notable and commercially significant. New discoveries of suitable entomopathogens and advances in their production have facilitated the commercialization of numerous products. However, despite the progress that has been made, entomopathogens still represent an under-developed and under-utilized resource in arthropod pest management. Thus additional research is required to expand the use of entomopathogens in biological control. New scientific tools, including molecular markers, genomics, in vitro production techniques, and others allow for novel discovery, identification, and development of entomopathogens previously overlooked or out of reach. The key challenges that limit microbial control for arthropod pests will be addressed in this project including: enhancing efficacy through strain discovery and improvement, advancing production and delivery, integration with existing management techniques, conservation of endemic entomopathogens, and gaining greater understanding of fundamental entomopathogen biology and ecology to further improve applied pest management. The project objectives will be grouped by agroecosystem (annuals, perennials, and urban/natural systems). The consequences of not doing the proposed research include increased chemical pesticides in the environment (risking the health of humans and other nontargets) and increased crop losses due to endemic and invasive pests.
Given that the research needs indicated above are common to numerous commodities across the US, a cooperative multi-state approach is demanded to provide broad impact solutions that are widely applicable. Entomopathogens and their pest insect hosts are not limited by artificial boundaries. Therefore, tests of efficacy, persistence, safety, resistance management and other parameters must be conducted under different sets of environmental conditions across state lines. Protocols must be developed and standardized for the diverse types of research being proposed. Thus, to be successful in fulfilling the objectives of this project proposal, multi-state cooperative research among State Agricultural Experiment Stations, USDA research groups, and industry is required.
We anticipate that the project will produce substantial benefits for both producer and consumer stakeholder groups. Stakeholders will include farmers, biocontrol producers, the scientific community and the general public. Experiments will be conducted in numerous cropping systems; and, given the broad nature of the research, we anticipate significant knowledge transfer to additional crops during the project period. Foremost, the proposed research will facilitate transition away from the reliance on chemical insecticide usage by providing effective and environmentally-friendly alternatives. Further development of insect pathogens for use in pest management programs will fill vital gaps in pest management caused by removal of broad spectrum chemicals. This is of particular importance in specialty crops that have few remaining pest management options. Furthermore, as new pest assemblages arise, novel microbial control tactics will contribute substantially to the development of innovative integrated pest management programs. Additionally, microbial control is poised to play an important role in transgenic crops (e.g., in resistance management) and in the control of invasive pests. The proposed project will improve quality of life by providing farmers tools to manage arthropod pests without risk of poisoning, and by providing the public with food containing less chemical residues. Economic opportunities will be created by enhancing the biological control industry and improving the productivity of various crops. Finally, fundamental studies conducted within this project will lead to advances in broader scientific disciplines, e.g., ecology, pathology, genetics and molecular biology. The broad and unique expertise represented by this working group will enable us to achieve our ambitious objectives. This project strongly addresses several US agriculture and SAAESD Priority Areas, particularly Priorities 1, 4, and 5, i.e., Developing greater harmony between agriculture and the environment; Establishing an agricultural systems that is highly competitive in the global economy, and Enhanced economic opportunity and quality of life for Americans.
Related, Current and Previous Work
The proposed project is unique. We do not anticipate any significant overlap with other multi-state projects or individual CRIS projects. Ours is the only multistate project that focuses on microbial control of arthropod pests. Several commodity-based projects such as NC205 (corn pests) and S1049 (pecans) include some research on microbial insect control, but these aspects generally constitute minor components of the overall objectives. Also, certain broad-based multistate projects on biological control of arthropods and weeds (e.g., NC1032 and NCERA220) include minimal emphasis on microbial agents. Moreover, the objectives of these other biocontrol projects and the expertise of the groups focus primarily on predators and parasitoids whereas the objectives and expertise in our project is exclusively microbial-based.
A CRIS search (using keywords such as microbial control, insect, entomopathogenic, as well as individual genus names of interest) reveals that the vast majority of research on applied microbial control of arthropod pests in the US is being conducted by our project participants and collaborators. Although some level of microbial control research is being conducted at several institutions outside of our project participation (e.g., University of Maryland, Oklahoma State University, Michigan State University), the specific objectives of these projects do not overlap directly with those of our project. We will continue to encourage all scientists working on microbial control of arthropod pests to join our project as participants or collaborators; we will ensure that these scientists are aware of our project by advertising through professional societies (e.g., Entomological Society of America, Society of Invertebrate Pathology) and other channels.
The proposed project builds strongly upon decades of research on invertebrate pathology and microbial control. The breadth of research on insect pathology and microbial control is reviewed in various texts such as Tanada and Kaya (1993), Vega and Kaya (2012), Grewal et al. (2005), Ekesi and Maniania (2007), Metz (2003), etc. Over the past five years of this project, significant multistate collaborations have been established (or continued) addressing the development of entomopathogens for biological control of arthropod pests. These efforts have resulted in over 450 scientific publications, six patents, and production of educational material for professionals and end users. The following section lists examples of some of the many significant accomplishments the project has made over the past five years. Advancements were made in the discovery, production, formulation and efficacy of microbial control agents used in biological pest suppression. Additionally, significant advances were made in studying the basic biology of microbial control agents. Research conducted under this project has led to the initiation of microbial control, or expanded microbial control, for dozens of arthropod pests in diverse systems. In the proposed project, we will expand microbial control efforts against these targets as well as others. Thus, the accomplishment examples listed below have facilitated expanded and improved use of entomopathogens in insect pest suppression and serve as a basis for future research during the next phase of the project.
Accomplishments are presented in relation to prior s-1024 Subproject objectives:
SUBPROJECT 1: Discovery of entomopathogens and their integration and safety in pest management programs for major acreage crops.
Implementing biological agents to control insect pests affecting major acreage crops is limited by the ecology and economics associated with large scale production of these select crops. Nonetheless, the development of Bacillus thuringiensis (Bt) as a biological pesticide and its eventual use in genetically engineered crops, serves as an example of the tremendous impact resulting from researching biological control agents for pest control. Beginning with research on production, application and efficacy of Bt as a biological pesticide, continued research has resulted in the transformation of the seed production industry that provides significant pest control based on a biological agent.
Wide-spread use of Bt crops has raised concerns about the development of insect resistance to Bt toxins by target pests. Levels of resistance and mechanisms of resistance have been documented (Gahan et al. 2005; Moar and McCollum 2006; Anilkumar et al. 2008a, b) and generally support continued use of this pest control strategy combined with mandated refuge requirements, areas planted to non-Bt varieties of crops. Additionally, entomopathogenic nematodes were found to magnify fitness costs of Bt resistance (e.g., in pink bollworm, Pectinophora gossypiella), and thus entomopathogenic nematode applications in non-Bt refuges could delay resistance to Bt cotton (Gassmann et al 2008, 2009).
Additionally, discovery of new technologies continue to improve the potential of developing entomopathogens as biological control agents including: fermentation techniques that produce novel fungal structures (microsclerotia) for application to soil (Jackson and Jaronski 2009; Jackson et al. 2010), formulations containing natural products shown to protect microbes from environmental degradation (Salamouny et al. 2009), new pathogens (Nielsen and Hajek 2005; Lacey and Neven 2006) and isolates of known pathogens (Leland et al. 2005) with better virulence to native and exotic insect pests, and integration of pathogens into cropping systems (Leland et al. 2005; Campbell et al. 2006).
SUBPROJECT 2: Discovery of entomopathogens and their integration and safety in pest management programs for ornamental, vegetable, fruit and nut crops.
Recent research on a variety of microbial control agents has demonstrated that high levels of inundative control can be achieved for various economically important insect pests, e.g., 80-90% reductions in pecan weevil, Curculio caryae, and peachtree borer, Synanthedon exitiosa, by applications of entomopathogenic nematodes (EPN) Steinernema carpocapsae (Cottrell and Shapiro-Ilan 2006; Shapiro-Ilan et al. 2006); 76-100% control of the white grub, Phyllophaga georgiana, following greenhouse applications of the nematode S. scarabaei (Koppenhöfer et al. 2008); > 90% control of plum curculio, Conotrachelus nenuphar, larvae and the citrus root weevil, Diaprepes abbreviatus, using the nematode S. riobrave (Alston et al. 2005; Duncan et al. 2007; Jenkins et al. 2007, 2008; Shapiro-Ilan et al. 2008a); 86-96% reductions in potato tuber moth, Phthorimaea operculella, populations due to application of potato tuber moth granulovirus (Arthurs et al. 2008b, c), and high levels of black vine weevil, Otiorhynchus sulcatus, suppression using EPNs and entomopathogenic fungi (EPF) (Bruck et al. 2005). As a result of this research, commercial control of these pests has been initiated or enhanced using microbial agents. Additionally, recently discovered microbial agents offer novel potential for use in biological control programs. This includes a newly discovered fungus, Hirsutella homalodiscae nom. prov., which infects all stages of the glassy-winged sharpshooter, Homalodisca coagulate (Boucias et al. 2007); new isolates of the fungi Hirsutella citriformis and Isaria fumosorosea associated with Asian citrus psyllid, and a novel microsporidium of the black vine weevil, Otiorhynchus sulcatus (Bruck et al. 2008).
Improvements in production of entomopathogens were also made including advances in in vivo production of entomopathogenic nematodes (resulting in patented technology that has been adopted by the biocontrol industry) (Shapiro-Ilan et al. 2007, 2008b). Novel formulation technology was developed to improve biocontrol efficacy by facilitating delivery or protecting entomopathogens from harmful UV radiation or other environmental stressors (Arthurs et al. 2006, 2008a; Lacey et al. 2006, 2010; Shapiro-Ilan et al. 2010).
Significant advances were made in understanding the basic biology and ecology of entomopathogens, and could lead to improved biocontrol utility. The dynamics of group movement and infection behavior by entomopathogenic nematodes were elucidated (Fushing et al. 2008), and the behavioral responses of entomopathogenic nematodes to electrical fields (Shapiro-Ilan et al. 2009a) and to the volatile cues produced by nematophagous fungi (El-Borai et al. 2011) explored. Additionally, quantitative realtime PCR (qPCR) assays were developed to detect and quantify entomopathogenic nematode species that are naturally distributed in Florida citrus orchards (Campos-Herrera et al. 2009, 2010). The qPCR assay was more effective than standard baiting methods for detecting nematode species composition in population mixtures and offers great potential for studying their ecology. Furthermore, novel virulence factors were identified in B. bassiana (Xu et al. 2008, 2009).
SUBPROJECT 3: Discovery of entomopathogens and their integration and safety in pest management programs for urban and natural landscapes.
Evaluations of different entomopathogen isolates and formulations suggest significant potential for use in perennial systems. Efficacy was demonstrated in using EPNs for oriental beetle in turf. EPFs were tested against Asian longhorned beetle in deciduous forests, grasshoppers, and Mormon crickets on rangeland. A new gypsy moth virus and two species of microsporidian were tested against gypsy moth. Also, the nematode Deladenus siricidicola is in development to use against recent invasions of Sirex noctilio. New methods of applications included a gel formulation for EPNs that increases the nematodes persistence in non-protected areas and using cloth bands wrapped around tree trunks for EPFs. Ecological relationships impacting entomopathogens were studied, e.g., Entomopthora maimaiga continues to cause infection in very low density gypsy moth, but has not caused an epizootic in several years, and studies of physical and chemical properties of soil were shown to interact in a predictable way with EPN efficacy. New entomopathogenic nematode species have been discovered in the Sonoran desert and semi desert areas (discovery in climactic extremes such as these may offer potential for enhanced environmental tolerance in the pathogens).
New pathogen relationships were also found for beneficial insects (e.g., pollinators). Evaluation of Bombus spp. populations for presence of Nosema bombi was completed in 2010. Bombus populations were also evaluated for the protozoan, Crithidia bombi. Combinations of viruses and microsporidia were investigated for their role in colony collapse disorder (results are currently being analyzed). Evaluation of the progression of Nosema apis and N. ceranae disease in individual host insects showed that infection only occurs in midgut tissues.
SUBPROJECT 4: Discovery of entomopathogens and their integration and safety in pest management programs for medical, veterinary, and structural pests.
Substantial advances were made on developing entomopathogens for control of important Diptera pests. The Musca domestica salivary gland hypertrophy virus (MdSGHV) that infects and sterilizes adult house flies and stable flies was characterized. Research on this Hytrosavirus group has established a framework to incorporate MdSGHV into bait for fly management, and to develop novel RNAi technologies targeting both host and viral genes via an oral delivery system. To develop more effective mosquito control, efficacy and resistance management studies on two recombinant larvicidal stains of B. thuringiensis subsp. israelensis (Bti) were conducted. The Bti/BsBin strain produces the B. sphaericus binary (Bin) toxin in combination with the four major endotoxins of Bti, and was nine-fold more effective against Anopheles gambiae than the strains of Bti and B. sphaericus used in current commercial products. Selection studies with the Bti/BsBin recombinant show no resistance development after nine generations in An. gambiae and similar findings have occurred with Culex quinquefasciatus. An expressed sequence tag (EST) survey of the mosquito parasite Edhazardia aedis indicated E. aedis seems to lack the multi-gene transcripts present in other microsporidia, and in addition, the first case of transcription of a transposable element in microsporidia was documented. Complete genome sequencing for 3 diverse species of microsporidia from mosquitoes is part of a project to sequence a total of 12 microsporidian species which will provide unique insight into the core genes responsible for the specialized intracellular lifestyle of these pathogens of important disease vectors. The baculovirus OcsoNPV (=AesoNPV) was active against a variety of important mosquito vectors and pests. New information on structural proteins of a baculovirus that kills Culex mosquitoes opens new possibilities for understanding host-viral interactions.
Surveys for pathogens in invasive ants have yielded novel viruses, fungi, and nematodes that infect these invasives. Three viruses were discovered from the red imported fire ant, Solenopsis invicta, with one virus associated with dramatic colony declines. Three new fungi were identified and infective nematodes are being described from the European fire ant, Myrmica rubra. Pathogenicity was confirmed for the newly isolated Hirsutella sp. fungus. The multistate - USDA-ARS Area wide Pest Management Project for the suppression of fire ants, which included the utilization of the microsporidium Kneallhazia (=Thelohania) solenopsae was concluded. Potential benefit associated with the fire ant reduction from the pathogen was estimated to be over $300 million annually. Several species and strains of nematodes were capable of infecting and killing structural pests such as the bark scorpion, Centruroides exilicauda and the desert subterranean termite, Heterotermes aureus. A Beauveria species was isolated from black-legged ticks, Ixodes scapularis. Transposon based transformation vectors for genetic manipulation of endosymbiotic rickettsiae of ticks were developed.
Advance the role of entomopathogens in annual crops (= row crops, vegetables etc).
Advance the role of entomopathogens in perennial crops (= orchards, small fruits, forage, etc
Advance the role of entomopathogens in natural and urban landscapes (= Med/vet, turf, ornamental, urban, forests).
The overall goal of the project is to improve and expand the use of entomopathogens in biological arthropod pest suppression in the following cropping systems. The goal in each objective is enhanced safety, sustainability, and productivity in US agriculture, forest, and urban systems. Additionally, each Subproject will address the following approaches to enhance microbial control efficacy:
Entomopathogen discovery and characterization, including biology and ecology.
Entomopathogen production, formulation, application methods, and integration into management systems.
MethodsGeneral Methods & Approach Across Sub-Projects: Building upon the research accomplishments from the previous project, research will target discovery of new entomopathogens, improving production and formulation technology, and enhanced implementation of microbial control in IPM systems. Research will also elucidate the basic biology and ecology of entomopathogens. The basic scientific approaches will be organized among three Subprojects and will be similar as outlined in the General Methods section. This process will facilitate the development of useful protocols that are transferrable among cropping systems. All Subprojects will include substantial participation and collaboration among state universities, USDA-ARS and other USDA agencies, industry, and extension personnel. The Project Chair and Subproject Chairs will then coordinate efforts (from shared protocols to reporting) among all project participants to tie together the collaborative efforts and leverage findings into improved IPM strategies with broad impact. Significant research efforts will also be directed toward the use of entomopathogens to manage current pests as well as those recently introduced invasive species in North America. Efforts to discover new entomopathogen strains and species will include the most recent techniques for surveying, isolating, identifying and examining pathogenicity of nematodes, bacteria, viruses, and fungi (including Microsporidia). Foreign exploration and implementation of new biopesticides will adhere to all APHIS and EPA guidelines. Nematodes and Hypocreales fungi will be isolated by baiting soil samples with sentinel insects, as well as surveying live insects, and grown on appropriate media or recycled in vivo for testing. Surveys for viruses and microsporidia will be conducted by collecting diseased individuals in the field and, if necessary, stressing the pests to induce epizootics of latent infections. New entomopathogens will be identified using morphological and molecular techniques. Virulence and pathogenicity of selected entomopathogens will be determined in laboratory assays against a range of appropriate target pest insects. Entomopathogens showing potential for commercial development will be tested in semi-field and field evaluations to determine efficacy potential. Entomopathogen production and formulation technology will be advanced through various approaches. Improvements to in vitro pathogen production (e.g. fungi or nematodes) will be based on optimization of media and bioreactor parameters. Improvement in in vivo production will be based on mechanization. Novel technologies will reduce costs and provide ample quantities of high quality microbial agents. Formulation research will develop of grower-adoptable materials, improve protection from environmental stresses, and extend shelf-life. Novel formulations will be compared with standards targeting appropriate economic pests. Enhanced implementation of microbial control in diverse systems will be achieved in a multi-faceted approach. Entomopathogen strain improvement techniques will be implemented to enhance efficacy. Efficacy will also be improved by developing novel application techniques and optimizing parameters such as application rates and timing. Efforts will include conservation, classical introduction, and inoculative or inundative approaches to biocontrol. Research will also be directed toward understanding fundamental entomopathogen biology and ecology. These studies will provide novel insights that can be leveraged to improve biocontrol efficacy. Ecological studies will focus on epizootiology (using molecular and non-molecular approaches), microhabitat preferences, as well as infection dynamics. Genetic studies will include new projects on sequencing and annotation of entomopathogen strains and species, as well as elucidation of beneficial trait changes during culturing. The following projects and experiments divided by Subproject represent critical research needs within specific commodities that will lead to major advances in safe and effective pest management. Methods for Sub-objectives within each Subproject: Subproject I: Advance the role of entomopathogens in annual crops Sub-objective 1: Development of Metarhizium spp. for control of soil-borne insect pest of row crops. The fungal pathogens, Metarhizium spp., kill a wide range of arthropod pests; most of which feed on aerial portions of crop plants. The sugar beet root maggot (SBRM), Tetanops myapaeformis, is a key pest of sugar beet roots and is susceptible to fungal pathogens under laboratory conditions (Jaronski 2006). Unfortunately, control of soil pests under field conditions has been relatively ineffective partially due to the inability to adequately apply fungal conidia to target soil pests and the corresponding high costs of the treatments (Campbell et al. 2006). New fermentation technology has the ability to economically produce fungal microsclerotia (Jaronski and Jackson, 2008, Jackson and Jaronski 2009), which can be formulated as granules and applied to the soil where infective conidia are subsequently produced in situ. Fermentation studies will optimize economical production and microsclerotia fitness. Formulated samples and application techniques will be evaluated in laboratory and field studies to verify conidia production after application for efficacy. Although SBRM is an initial target, this technology may be evaluated against soil pests of other crops, such as corn rootworm, root weevils, and wireworms. Subproject I Collaborating Institutions: North Dakota State University, Fargo, ND; USDA-ARS-Peoria, IL; USDA-ARS-Sidney, MT; USDA-ARS-Wapato, WA. Subproject II: Advance the role of entomopathogens in perennial crops Sub-objective 1: Development of improved microbial control measures for grasshoppers and Mormon crickets on rangeland and improved pastures. Grasshoppers and Mormon crickets continue to impact forage production in the 17 western states (2.4 million acres were treated with chemical pesticides in 2010). Several hundred domestic entomopathogenic fungi (EPFs, obtained from the USDA-ARS Entomopathogenic Fungal Collection; curator (Richard Humber: firstname.lastname@example.org) will be evaluated (using criteria of ecologically relevant thermal tolerances, mass production potential, as well as efficacy) as candidate mycoacridicides, and permitting is being pursued for the field testing of two commercial, nonindigenous, mycoinsecticides based on M. acridum as possible substitutes. Based on the recent discovery of locust-active Bt in China, molecular approaches will be used to screen the extensive USDA-ARS Bt collection for others. In addition, ecological and physiological factors will be screened for effects on pathogen immune defenses of Mormon crickets and grasshoppers. Sub-objective 2: Develop an integrated pest management program for fungus gnats (Bradysia spp.) in greenhouses utilizing predators and entomopathogens. The effectiveness of two registered EPFs, M. anisopliae (Met52) and Beauveria bassiana (GHA) and the recently commercially available predatory rove beetle, Atheta coriaria, will be evaluated against adult and larval fungus gnats. Their effectiveness will be assessed on seedling flats in small cages with the following 6 treatments: control; M. anisopliae granules; B. bassiana granules; rove beetles; M. anisopliae granules and rove beetles; B. bassiana granules and rove beetles. Emergence of fungus gnat adults will be monitored beginning two weeks after infestation for 4 weeks using yellow sticky cards. The most effective treatment from small scale cage studies will be compared to an untreated control and Gnatrol® (Valent USA Corporation, Walnut Creek CA) as a standard in greenhouses with fungus gnat infestations. Sub-objective 3: Development of improved control measures for key pests of pecan and peaches using entomopathogenic nematodes. Key southeastern US orchard pests include the peachtree borer (PB), Synanthedon exitiosa, lesser peachtree borer (LPB), S. pictipes, and plum curculio (PC), Conotrachelus nenuphar in peach, and the pecan weevil (PW), Curculio caryae in pecan. Entomopathogenic nematodes (EPN), e.g., Steinernema carpocapsae or S. riobrave, cause high levels of mortality in these pests under field conditions (Shapiro-Ilan et al. 2008a, 2009b). However, application and delivery methods need optimization. In small field plots as well as commercial scale peach and pecan orchards, methods of EPN application will be compared (e.g., injection, boom sprayer, trunk spray) for control of PB and PW. Additionally, a diverse array of formulations will be tested for optimizing LPB control. Lab screening will identify the optimum EPN for use against PC, and an integrated multi-stage program for control of this pest will ensue. The impact of the biocontrol approach on pest control and natural enemy abundance will be measured in comparison with conventional insecticide sprays. In related studies, beneficial trait stability of EPNs will be characterized (Bai et al. 2005, Adhikari et al. 2009, Chaston et al. 2011). Sub-objective 4: Analysis of pathogens associated with Asian citrus pysllid (ACP) and development of control measures. The ACP, Diaphorina citri, is an invasive pest of citrus. This pest vectors phloem-inhabiting bacteria thought to be responsible for huanglongbing disease (HLB, citrus greening). Field surveys by Hall et al. (2011) demonstrated that the incidence of the mycopathogen Hirsutella citriformis (Meyer et al. 2007) in fall/winter of mycosed adults may exceed 75%. Various materials derived from H. citriformis will be evaluated against ACP. Unformulated material from both mono- and biphasic production lots will be assayed against ACP in the laboratory, greenhouse and field. Current mycoinsecticides based on a North American Isaria fumosorosea isolate and other fungal species which offer potential for selective control of ACP (Arthurs et al. 2011). Significant progress on application methods, formulation additives, and timing with respect to climatic effects, are needed to maximize the potential of this technology. In greenhouse and small plot trials, mycoinsecticides and common insecticides will be evaluated against ACP and beneficial arthropods on citrus. Sub-objective 5: Development of improved control measures for white grubs in highbush blueberries and cranberries using entomopathogenic nematodes. The root-feeding larvae of scarab beetles are major blueberry and cranberry pests in the US. In New Jersey, the oriental beetle, Anomala (=Exomala) orientalis is a major pest of highbush blueberries and the May/June beetle Phyllophaga georgiana is a common and often severe pest of cranberries. In previous studies the EPN, Steinernema scarabaei, controlled both scarabs, whereas other scarab-pathogenic nematodes such as Heterorhabditis bacteriophora were less effective (Polavarapu et al. 2007; Koppenhöfer et al. 2008). All EPN species are less effective in the acidic blueberry/cranberry soils than in typical agricultural soils (Koppenhöfer and Fuzy 2006). Several EPN species isolated from New Jersey blueberry fields were scarab-pathogenic in preliminary trials. These species will be evaluated for virulence/efficacy against scarabs in blueberry and cranberry fields. Isolation of other species/strains adapted to these crop environments should continue, followed by virulence/ efficacy testing. Effective and well-adapted species/strains will be characterized and evaluated for suppression of scarab pests. Sub-objective 6: Habitat manipulation to enhance biological control of soil pests in nurseries. Ornamental plants and vegetables produced in nurseries are high-value specialty crops. Nursery crops are plagued by many insect pests, some of which spend part of their life cycle in soil. Larval stages of root weevils, scarab beetles, and fungus gnats can damage a plant by feeding on roots, introducing pathogens, limiting water and nutrient uptake and possibly killing the plant. Current pest management programs include insecticide applications and/or biocontrol using EPNs and EPFs. Nurseries rely on soilless media composed of sphagnum peat moss, coconut coir, sawdust, bark, sand, perlite, vermiculite, and other similar materials customized and mixed at the nursery. Certain components of soilless media may interfere or enhance biocontrol strategies. Standard commercial and modified (experimental) blends of soilless media will be evaluated to optimize the habitat for biocontrol agents of root-feeding pests. Subproject II Collaborating Institutions: Brigham Young University, UT; Department of Plant Industry, Gainesville, FL; Koppert Biological Systems; OR State University; Rutgers University, NJ; University of CA, Davis; University of FL; University of GA; USDA APHIS CPHST Phoenix AZ; USDA ARS Beltsville MD; USDA-ARS-Byron, GA; USDA-ARS Corvallis, OR; USDA ARS Fort Pierce, FL; USDA ARS-Kearneysville, WV; USDA ARS-Peoria, IL; USDA ARS- Sidney MT, University of AZ, Utah State University, Subproject III. Advance the role of entomopathogens in natural and urban landscapes Sub-objective 1. Identification and management of microsporidian pathogens of beneficial beetles used for classical biological control of hemlock woolly adelgid (HWA) in forests. The HWA has devastated relic hemlock in eastern North America. Predatory beetles have been imported from Asia and from western North American hemlock stands for use as biological control agents; two species have been released. Six species of microsporidia have been isolated from four beetle species in laboratory colonies including the two released species and two that remain in quarantine. Three microsporidian species have destroyed colonies and aborted release programs (Solter et al. 2011, in press). Beetles in mass reared colonies, field collected beetles, and beetles recovered from release sites will be monitored to determine the phylogenetic identity of the pathogens and whether they infect multiple beetle species in the field. Recommendations to eliminate microsporidia from mass rearing facilities and future releases of beetles will be provided to collaborating institutions. Sub-objective 2. Immune responses of gypsy moth (GM), Lymantria dispar, to naturally occurring pathogens and parasites. The GM and its associated pathogens and parasites will serve as a model system for studies of comparative immune response in Lepidoptera. Host proteomic responses to two species of microsporidia, Endoreticulatus schubergi (midgut pathogen), Vairimorpha disparis (fat body pathogen), and to LdMNPV (DNA virus), Ld cypovirus (RNA virus), and Bt, all orally administered, and Glypapanteles (parasitoid) administered by stinging, will be compared using 2-D gel electrophoresis. Host proteins of interest will be identified by mass spectometry, and differential responses to various pathogens determined. The goal is to provide a better understanding of lepidopteran immune response to naturally occurring pathogens, an understudied area of research (Beckage, 2008). The data will provide information about pathogen virulence and host susceptibility at the level of post-genomic host response. Sub-objective 3: Evaluation of high temperatures on Entomophaga maimaiga for control of gypsy moth (GM), Lymantria dispar. The GM is an invasive forest defoliator presently established in North America and is spreading west and south. Two entomopathogens provide natural control of GM: the fungus, E. maimaiga, and the NPV, Lymantria dispar. Laboratory studies demonstrated high temperatures (30oC) can cure GM larvae of E. maimaiga infections (Hajek et al. 1990). E. maimaiga will be exposed to such temperatures as GM spreads further south in a warmer climate. Tests are underway to determine if infected larvae sun-bask to cure themselves of infections. Bioassays will expose GM larvae infected through protoplast injection and conidial showering to high temperatures for various time intervals. The results will be used to model the effect of E. maimaiga in different areas of North America. Sub-objective 4: Development of strategies for applying Metarhizium brunneum for control of Asian longhorned beetles (ALHB). The invasive ALHB kills many species of trees and is presently the target of USDA-APHIS eradication efforts. Only tree removal and tree injection with imidacloprid are effective. Additional methods for control are needed. Non-woven fiber bands containing entomopathogenic fungi are used for longhorn beetle control in Japan and China. Evaluations of control strategies and delivery methods for M. brunneum targeting ALHB are underway in US (Peng et al. 2011). There is no industry in North America interested in making these bands (Shanley et al. 2009). Alternative substrates for making bands to improve conidial deposition on beetles, oil formulations, and application strategies will be evaluated through conidial counts, beetle longevity, reproduction and behavior after exposure, with and without the addition of semiochemical attractants. Bands will be evaluated for fungus longevity in field environments. Sub-objective 5: Management of invasive thrips in ornamental plants using entomopathogenic fungi. Several species of thrips (Thysanoptera: Thripidae) are among the most serious pests of ornamental plants in southeastern USA. Nursery growers and landscape mangers require biorational alternatives to protect the limited effective insecticides often applied on a calendar basis. Newly available mycoinsecticides can provide effective control of thrips in greenhouses and nurseries under certain circumstances. Further research is needed to optimize methods of application and delivery (e.g. testing formulation, ULV application and efficacy soil versus foliage applications). The impact of integrated thrips management programs including mycoinsecticides also needs to be tested against additional thrips species. Sub-objective 6: Improving control measures for white grubs, annual bluegrass weevil, and Lepidoptera pests in turfgrass using EPNs. A complex of white grubs (Coleoptera: Scarabaeidae) are the most important pests of turfgrass in the eastern US. The EPN, S. scarabaei, can provide excellent control of white grubs, but is difficult to mass-produce; while commercial EPN species are often less effective (Koppenhöfer et al. 2006). Additionally, commercial EPNs have provided highly variable control of the annual bluegrass weevil, Listronotus maculicollis, a severe pest of fine turf in the eastern US (McGraw et al. 2010). Several other EPNs, especially S. carpocapsae, while very effective against the black cutworm, Agrotis ipsilon, on golf course greens and tees, do not always provide satisfactory control (Ebssa and Koppenhöfer 2011). New EPN species/strains will be evaluated against these pests in split applications, species combinations, and potentially synergistic combinations of nematodes with other control agents to improve control of these pests. Promising combinations will be tested for long-term pest suppression. Sub-objective 7: Analysis of Hytrosavirus dynamics in Dipterans. Salivary gland hypertrophy viruses (SGHVs) are unique DNA viruses reported from insects within three dipteran genera including the house fly Musca domestica (Lietze et al. 2011a). These viruses induce salivary gland enlargement (hypertrophy) and inhibit reproduction by suppressing vitellogenesis and disrupting mating behavior. The basis for sterility, mechanisms underlying SGHV transmission and maintenance in field populations are unknown (Lietze et al. 2011b). Research on M. domestica-specific SGHV will target the physiology causing the virus-induced sterility and the mechanisms regulating the transmission of this virus in field populations. These results will be used to model virus-host interactions. Sub-objective 8: Gene homology/identity survey for pathogens of invasive ants to discover and implement potential biological control agents. Nylanaderia pubens, unofficially called the Rasberry, Caribbean, or hairy, crazy ant, is an invasive from the Caribbean or Central/South America that has established in several Gulf Coast states. Extreme proliferations of N. pubens populations inundate urban and natural landscapes and their sheer numbers result in incessant complaints, electrical malfunctions, crop damage, and loss of commercial bee hives. Following methods used to discover viruses in fire ants (Oi and Valles 2009); a pathogen survey utilizing gene homology will be conducted among Florida N. pubens populations to identify pathogens. Survey, identification/characterization of biocontrol agents (B. bassiana, EPNs) and efficacy tests also will be directed toward the invasive, stinging European fire ant, Myrmica rubra, which has become pestiferous in Maine. Sub-objective 9. Improving control measures for public health and structural pests using entomopathogenic nematodes. Several public health and structural pests are susceptible to EPNs, e.g., medically important scorpions Centruroides sculpturatus (Scorpiones: Buthidae), subterranean termites Heterotermes aureus (Isoptera: Rhinotermitidae) (Gouge & Snyder 2005; Yu et al. 2010). Additionally, there are a number currently under investigation, e.g. bed bugs. To expand upon previous findings, lab-based efficacy tests will assess S. carpocapsae and S. riobrave against juvenile C. sculpturatus. Lab- and home-based efficacy tests will be done on baseboard harborage areas, assessing S. feltiae and S. riobrave against bed bugsmin a BSL 2 facility. Subproject III Collaborating Institutions: Clemson Univ., SC; INHS/Univ. of IL; PA State University; Novozymes Biologicals Inc. Salem, VA; NJ Dept. of Agriculture; Rutgers University, NJ; State University of NY; University of AZ; University of FL; University of GA; University of ME, USDA APHIS; USDA-ARS-Beltsville, MD; USDA-ARS, Gainesville, FL; USDA-ARS Maricopa, AZ; USDA-FS-Morgantown; USDA-FS-Hamden, VA Tech. University, Vineland Research and Innovation Center, Vineland Station, Canada.
Measurement of Progress and Results
- New microbial control tools including novel or improved species and strains and superior methods of production and delivery.
- Extension-related presentations and workshops and scientific symposia.
- Scientific publications including refereed manuscripts and books and book chapters, videos and CDs on new developed techniques.
- Annual reporting will include the summary of research achievements, publication lists, tabulation of extension related activities (including videos, web-based resources, factsheets and presentations) related to this project.
- The project will maintain a website which, in addition to the Annual Reports and meeting information, will publish the specific recommendations from workshops.
- Output 6. Tabulated hits at the project and related websites will be used as outcome indicators.
Outcomes or Projected Impacts
- Reduced impact on non-target organisms and the environment, and protection of human health due to decreased use of chemical pesticides.
- Economic benefits to farmers using microbial control based on higher virulence in biocontrol products or enhanced efficiency in production and delivery.
- Increased food security due to the development of alternative pest management tools that be applied to increase productivity and protect against new threats (e.g., invasive pests).
- Expansion and increased profitability in the biocontrol industry.
Milestones(0): Milestones Attachment.
Projected ParticipationView Appendix E: Participation
This project will provide educational material for: a) Entomology colleagues via symposia at national meetings, and refereed research publications, books and book chapters; and b) Extension personnel and growers via participation in annual project meetings, trade shows, field days, presentations, trade journal articles and factsheets, and production of published and web-based resources on the use of entomopathogens in pest management. Products from this project have applications for both conventional and organic producers, resource managers (e.g. forests), and urban clientele. Organic growers in particular have typically been under-served by research and extension activities, which have tended to focus on development of chemically-based pest management programs. Additionally, outreach will also extend to 1890 institutions as some of the project participants have ties to these colleges (e.g., through adjunct professorships) and populations sectors they serve. Today, research and outreach efforts emphasize increasing the implementation of IPM strategies on farms, forests and urban landscapes, with biological controls forming the first line of defense against pests. This project enhances our ability to achieve this goal on a broad range of agricultural commodities and other managed ecosystems.
Organization: Officers include Chair, Vice-Chair, Member-at-Large, and Secretary. Each position runs for a two-year term, at which time a new Member-at-Large and a new Secretary are elected at the annual meeting, the retiring the Member-at-Large replaces the Vice-Chair, and retiring Vice-Chair replaces the Chair. The Chair is charge of running all aspects of project, the Vice-Chair and Member-at-Large assist the Chair and serve in his/her place when the Chair is unavailable. The Chair can appoint other ad-hoc officers as needed. The Secretarys primary responsibility is to compile annual reports and meeting minutes.
Chair: David Shapiro-Ilan, USDA-ARS, Byron, GA.
Vice-Chair: Denny Bruck, USDA-ARS, Corvallis, OR.
Member-at-Large: Steve Arthurs, University of Florida, Apopka.
Secretary: Robert Behle, USDA-ARS, Peoria, IL.
Subproject Chairs: I: Mark Boetel (ND State U) and Robert Behle (USDA-ARS-Peoria, IL).
II: Edwin Lewis (U of CA, Davis).
III. David Oi (USDA-ARS-Gainesville, FL) and Steven Arthurs (U of FL).
Adhikari, B. N., L. Chin-Yo, B. Xiaodong, T. A. Ciche, P. S. Grewal, A. R. Dillman, J. M. Chaston, D. I. Shapiro-Ilan, A. L. Bilgrami, R. Gaugler, P. W. Sternberg, and B. J. Adams. 2009. Transcriptional profiling of trait deterioration in the insect pathogenic nematode Heterorhabditis bacteriophora. BMC Genomics. 10: 609 doi:10.1186/1471-2164-10-609.
Alston, D.G., D. E. N. Rangel, L. A. Lacey, H. G. Golez, J. J. Kim and D.W. Roberts. 2005. Evaluation of novel fungus and nematode isolates for control of Conotrachelus nenuphar (Coleoptera: Curculionidae) larvae. Biol. Contr. 35: 163171.
Anilkumar, K. J., A. Rodrigo-Simón, J. Ferré, M. Pusztai-Carey, S. Sivasupramaniam, and W. J. Moar. 2008a. Production and characterization of Bacillus thuringiensis Cry1Ac-resistant cotton bollworm, Helicoverpa zea (Boddie) Appl. Environ. Microbiol. 74: 462-469.
Anilkumar, K. J., M. Pusztai-Carey, and W. J. Moar. 2008b. Fitness costs associated with Cry1Ac-resistant Helicoverpa zea (Lepidoptera: Noctuidae): a factor countering selection for resistance to Bt cotton? J. Econ. Entomol. 101: 1421-1431.
Arthurs, S. P., L. A. Lacey, and R.W. Behle. 2006. Evaluation of spray-dried lignin-based formulations and adjuvants as ultraviolet light protectants for the granulovirus of the codling moth, Cydia pomonella (L). J. Invertebr. Pathol. 93: 8895.
Arthurs, S. P., L. A. Lacey and R. W. Behle. 2008a. Formulation for improved activity of codling moth granulovirus: evaluation of environmental/UV protectants. Biocontrol Sci. Technol. 18: 639-663.
Arthurs, S. P., L. A. Lacey and F. de la Rosa. 2008b. Evaluation of a granulovirus (PoGV) and Bacillus thuringiensis subsp. kurstaki for control of the potato tuberworm in stored tubers. J. Econ. Entomol. 101: 1540-1546.
Arthurs, S. P., L. A. Lacey, J. N. Pruneda and S. Rondon. 2008c. Semi-field evaluation of a granulovirus and Bacillus thuringiensis ssp. kurstaki for season-long control of the potato tuber moth, Phthorimaea operculella. Entomol. Exp. Appl. 129: 276285.
Arthurs, S, Avery, P. B., and Stauderman, K. 2011. Evaluation of low risk insecticides for Asian citrus psyllid on jasmine, 2010. Arthropod Management Tests (in press)
Bai, C., D. I. Shapiro-Ilan, R. Gaugler, and K. R. Hopper. 2005. Stabilization of beneficial traits in Heterorhabditis bacteriophora through creation of inbred lines. Biol. Contr. 32: 220-227.
Beckage, N.E. 2008, Insect Immunology. Elsevier, NY.
Boucias, D. G., D. W. Scharf, S. E. Breaux, D. H. Purcell, and R. F. Mizell. 2007. Studies on the fungi associated with the glassy-winged sharpshooter Homalodisca coagulata with emphasis on a new species Hirsutella homalodisca. BioControl 52: 231-258.
Bruck, D. J., L. F. Solter, and A. Lake. 2008. Effects of a novel microsporidium on the black vine weevil, Otiorhynchus sulcatus (F.) (Coleoptera: Curculionidae). J. Invertebr. Pathol. 98: 351-355.
Bruck, D. J., D. I. Shapiro-Ilan and E. E. Lewis. 2005. Evaluation of application technologies of entomopathogenic nematodes for control of the black vine weevil, Otiorhynchus sulcatus. J. Econ. Entomol. 98: 1884-1889.
Campbell, L. G., M. A. Boetel, N. B. Jonason, S. T. Jaronski, and L. J. Smith. 2006. Grower-adoptable formulations of the entomopathogenic fungus Metarhizium anisopliae (Ascomycota: Hypocreales) for sugarbeet root maggot (Diptera: Ulidiidae) management. Environ. Entomol. 35: 986-991.
Campos-Herrera, R., L. W. Duncan, R. J. Stuart, F. El-Borai, and C. Gutierrez. 2009. Entomopathogenic Nematode Ecology and Biological Control in Florida Citrus Orchards. In, Integrated Management of Arthropod Pests and Insect Borne Diseases, A. Ciancio & K. G. Mukerji, eds. Springer Science + Business Media B.V., p. 97-126.
CamposHerrera, R., E. Johnson, F. K. ElBorai, R. J. Stuart, J. H. Graham, and L. W. Duncan. 2010. Long-term stability of entomopathogenic nematode spatial patterns measured by sentinel insects and real-time PCR assays. Ann. Appl. Biol. 158: 55-68.
Chaston J. M., A. R. Dillman, D. I. Shapiro-Ilan, A. L. Bilgrami, R. Gaugler, K. R. Hopper, and B. J. Adams. 2011. Outcrossing and crossbreeding recovers deteriorated traits in laboratory cultured Steinernema carpocapsae nematodes. Int. J. Parasitol. 41: 801809.
Cottrell, T. E. and D. I. Shapiro-Ilan. 2006. Susceptibility of the peachtree borer, Synanthedon exitiosa, to Steinernema carpocapsae and Steinernema riobrave in laboratory and field trials. J. Invertebr. Pathol. 92: 85-88.
Duncan, L. W., J. H. Graham, J. Zellers, D. Bright, D. C. Dunn, and F.E. El-Borai. 2007. Food web responses to augmentation biological control using entomopathogenic nematodes in bare and composted-manure amended soil. J. Nematol. 39: 176189.
Ebssa, L., and A. M. Koppenhöfer. 2011. Efficacy and persistence of entomopathogenic nematodes for black cutworm control in turfgrass. Biocontrol Sci. Technol. 21: 779-796.
Ekesi, S., and K. N. Maniania. 2007. Use of Entomopathogenic Fungi in Biological Pest Management. Signpost, Kerala, India.
ElBorai, F. E., R. CamposHerrera, R. J. Stuart and L. W. Duncan. 2011. Substrate modulation, group effects and the behavioral responses of entomopathogenic nematodes to nematophagous fungi. J. Invertebr. Pathol. 106: 347356.
Fushing, H., D. I. Shapiro-Ilan, J. F. Campbell, and E. Lewis. 2008. State-space based mass event-history model I: many decision-making agents with one target. Ann. Appl. Stat. 2: 1503-1522.
Gahan, L., Y. T. Ma, M. L. MacGregor Coble, F. Gould, W. J. Moar, and D. G. Heckel. 2005. Genetic basis of resistance to Cry1Ac and Cry2A in Heliothis virescens. J. Econ. Entomol. 98: 1357-1368.
Gassmann A. J., J. A. Fabrick, M. Sisterson, E. R. Hannon, S. P. Stock, Y. Carrière, and B. E. Tabashnik. 2009. Effects of pink bollworm resistance to Bacillus thuringiensis on phenoloxidase activity and susceptibility to entomopathogenic nematodes. J. Econ. Entomol. 102: 1224-1232.
Gassmann A. J., S. P. Stock, M. Sisterson, Y. Carrière, and B. E. Tabashnik. 2008. Synergism between entomopathogenic nematodes and Bt crops: integrating biological control and resistance management. J. Applied Ecol. 45: 957-966.
Gouge, D. H., and J. L. Snyder. 2005. Parasitism of bark scorpions Centruroides exilicauda (Scorpiones: Buthidae) by entomopathogenic nematodes (Rhabditida: Steinernematidae; Heterorhabditidae). J. Econ. Entomol. 98: 1486-1493.
Grewal, P. S., R-U Ehlers, and D. I. Shapiro-Ilan. 2005 Nematodes as Biocontrol Agents. CABI Publishing, Wallingford.
Hajek, A. E., R. I. Carruthers, and R. S. Soper. 1990. Temperature and moisture relations of sporulation and germination by Entomophaga maimaiga (Zygomycetes: Entomophthoraceae), a fungal pathogen of Lymantria dispar (Lepidoptera: Lymantriidae). Environ. Entomol. 19: 85-90.
Hall, D. G., M. Hentz, G. Meyer, J. M. and D. G Boucias. 2011. Observations on the entomopathogenic fungus Hirsutella citriformis attacking adult Diaphorina citri (Hemiptera: Psyllid) in a managed citrus grove. Biol. Contr. (submitted).
Jackson, M. A., C. A. Dunlap, and S. T. Jaronski. 2010. Ecological considerations in producing and formulating fungal entomopathogens for use in insect biocontrol. Biocontrol 55: 129-145.
Jackson, M. A., and S. T. Jaronski. 2009. Production of microsclerotia of the fungal entomopathogen Metarhizium anisopliae and their potential for use as a biocontrol agent for soil-inhabiting insects. Mycological Res. 113: 842-850.
Jaronski, S. 2006. Fungus controls sugar beet insect pest. Industrial Bioprocessing 28: 2-3.
Jaronski, S.T. and M.A. Jackson. 2008. Efficacy of Metarhizium anisopliae microsclerotial granules. Biocont. Sci. Technol. 18: 849-863.
Jenkins, D. A., D. I. Shapiro-Ilan, and R. Goenanga. 2007. Virulence of entomopathogenic nematodes against Diaprepes abbreviatus in an oxisol. Fla. Entomo. 90: 401-403.
Jenkins, D. A., D. I. Shapiro-Ilan, and R. Goenaga. 2008. Efficacy of entomopathogenic nematodes versus Diaprepes abbreviatus (Coleoptera: Curculionidae) larvae in a high clay content oxisol soil: greenhouse trials with potted Litchi chinensis. Fla. Entomol. 91: 75-78.
Koppenhöfer, A. M., and E. M. Fuzy. 2006. Effect of soil type on infectivity and persistence of the entomopathogenic nematodes Steinernema scarabaei, Steinernema glaseri, Heterorhabditis zealandica, and Heterorhabditis bacteriophora. J. Invertebr. Pathol. 92: 11-22.
Koppenhöfer, A. M., P. S. Grewal, and E. M. Fuzy. 2006. Virulence of the entomopathogenic nematodes Heterorhabditis bacteriophora, H. zealandica, and Steinernema scarabaei against five white grub species (Coleoptera: Scarabaeidae) of economic importance in turfgrass in North America. Biol. Contr. 38: 397-404.
Koppenhöfer, A. M., C. R. Rodriguez-Saona, S. Polavarapu, and R. J. Holdcraft. 2008. Entomopathogenic nematodes for control of Phyllophaga georgiana (Coleoptera: Scarabaeidae) in cranberries. Biocontrol Sci. Technol. 18: 21-31.
Lacey, L. A. and L. G. Neven. 2006. The potential of the fungus, Muscodor albus as a microbial control agent of potato tuber moth (Lepidoptera: Gelechiidae) in stored potatoes. J. Invertebr. Pathol. 91: 195-198.
Lacey, L. A., S. P. Arthurs, T. R. Unruh, H. Headrick and R. Fritts, Jr. 2006. Entomopathogenic nematodes for control of codling moth (Lepidoptera: Tortricidae) in apple and pear orchards: effect of nematode species and seasonal temperatures, adjuvants, application equipment and post-application irrigation. Biol. Contr. 37: 214223.
Lacey, L. A., D. I. Shapiro-Ilan and G. M. Glenn. 2010. Post-application of anti-desiccant agents improves efficacy of entomopathogenic nematodes in formulated host cadavers or aqueous suspension against diapausing codling moth larvae (Lepidoptera: Tortricidae). Biocontrol Sci. Technol. 20: 909-921.
Lietze, V-U., A. M. Abd-Alla, M. J. Vreysen, C. J. Geden, C. J., and D. G. Boucias.. 2011a. Salivary gland hypertrophy viruses (SGHVs): a novel group of insect pathogenic viruses. Ann. Rev. Entomol. 56: 6380.
Lietze, V-U, C. J. Geden, M. Doyle, and D. G. Boucias. 2011b. Disease dynamics and persistence of MdSGHV-infections in laboratory house fly (Musca domestica) populations. Appl. Environ. Microbiol. (under review).
Leland, J. E., M. R. McGuire, J. A. Grace, S. T. Jaronski, M. Ulloa, Y. H. Park, and R. D. Plattner. 2005. Strain selection of a fungal entomopathogen, Beauveria bassiana, for control of plant bugs (Lygus spp.) (Heteroptera: Miridae). Biol. Contr. 35: 104-114.
McGraw, B. A., R. Cowles, P. J. Vittum, and A. M. Koppenhöfer. 2010. Field evaluation of entomopathogenic nematodes for the biological control of the annual bluegrass weevil, Listronotus maculicollis (Coleoptera: Curculionidae) in golf course turfgrass. Biocontrol Sci. Technol. 20: 149-163.
Metz, M. 2003. Bacillus thuringiensis: A Cornerstone of Modern Agriculture. Haworth Press, New York.
Meyer, J. M., M. A. Hoy, and D. G. Boucias. 2007. Morphological and molecular characterization of a Hirsutella species infecting the Asian citrus psyllid, Diaphorina citri Kuwayama (Homoptera: Psyllidae), in Florida. J. Invertebr. Pathol. 95: 101-109.
Moar, W. J., and R. C. McCollum. 2006. Bt formulated products: should there be more concern about resistance development with the introduction of Bt transgenic plants? GMOs in Integrated Plant Production. Ecological Impact of Genetically Modified Organisms. IOBC wprs Bulletin. 29: 99-102.
Nielsen, C., and A. E. Hajek. 2005. Control of invasive soybean aphid, Aphis glycines (Hemiptera: Aphididae) populations by existing natural enemies in New York State. Environ. Entomol. 34: 1036-1047.
Oi, D. H., and S. M. Valles. 2009. Fire ant control with entomopathogens in the USA, pp. 237-257. In A. E. Hajek, T. R. Glare and M. O'Callaghan [eds.], Use of microbes for control and eradication of invasive arthropods. Springer Science + Business Media B.V.
Peng, F., S. Gardescu, and A. E. Hajek. 2011. Transmission of Metarhizium brunneum conidia between male and female Anoplophora glabripennis adults. BioControl 56: 771780.
Polavarapu, S., A. M. Koppenhöfer, J. D. Barry, R. J. Holdcraft, and E. M. Fuzy. 2007. Entomopathogenic nematodes and neonicotinoids for remedial control of oriental beetle, Anomala orientalis (Coleoptera: Scarabaeidae), in highbush blueberry. Crop Protect. 26: 1266-1271.
Salamouny, S. E., M. Shapiro, K. S. Ling, and B. M. Shepard. 2009. Black tea and lignin as ultraviolet protectants for the beet armyworm nucleopolyhedrovirus 1. J. Entomol. Sci. 44: 50-58.
Shanley, R. P., J. Leland, M. Keena, M.M. Wheeler, A. E. Hajek. 2009. Evaluating the virulence and longevity of non-woven fiber bands impregnated with Metarhizium anisopliae against the Asian longhorned beetle, Anoplophora glabripennis (Coleoptera: Cerambycidae). Biol. Contr. 50: 94-102.
Shapiro-Ilan, D. I., T. E. Cottrell, I. Brown, I., W. A. Gardner, R. K. Hubbard, and B. W. Wood. 2006. Effect of soil moisture and a surfactant on entomopathogenic nematode suppression of the pecan weevil, Curculio caryae. J. Nematol. 38: 474-482.
Shapiro-Ilan, D. I., J. F. Campbell, E. E. Lewis, J. M. Elkon, and D. B. Kim-Shapiro. 2009a. Directional movement of parasitic nematodes in response to electrical current. J. Invertebr. Pathol. 100: 134-137.
Shapiro-Ilan, D. I., T. E. Cottrell, R. F. Mizell III, D. L. Horton, and J. Davis. 2009b. A novel approach to biological control with entomopathogenic nematodes: Prophylactic control of the peachtree borer, Synanthedon exitiosa. Biol. Contr. 48: 259-263.
Shapiro-Ilan, D. I., T. E. Cottrell, R. F. Mizell, D. L. Horton, B. Behle, and C. Dunlap. 2010. Efficacy of Steinernema carpocapsae for control of the lesser peachtree borer, Synanthedon pictipes: Improved aboveground suppression with a novel gel application. Biol. Contr. 54: 23-28.
Shapiro-Ilan, D. I. W. A. Gardner, J. R. Fuxa, and B. W. Wood. 2007. US Patent 7,241,612. Materials and Methods for Control of Insects Such as Pecan Weevils.
Shapiro-Ilan, D. I., Mizell, R. F., Cottrell, T. E. and Horton, D. L. 2008a. Control of plum curculio, Conotrachelus nenuphar with entomopathogenic nematodes: effects of application timing, alternate host plant, and nematode strain. Biol. Control 44: 207-215.
Shapiro-Ilan, D.I., W. L. Tedders, and E. E. Lewis. 2008b. US Patent 7,374,773. Application of entomopathogenic nematode-infected cadavers from hard-bodied arthropods for insect suppression.
Solter, L. F., W. F. Huang, and B. Onken. 2011. Microsporidian Disease in Predatory Beetles. In Implementation and Status of Biological Control of the Hemlock Woolly Adelgid [Onken, B., Ed.] USDA Forest Service Technical Report.
Tanada, Y., and H. K. Kaya. 1993. Insect Pathology. Academic Press, San Diego.
Vega F., and H. K. Kaya. 2012. Insect Pathology (2nd Edition). Elsevier, San Diego. In Press.
Xu, Y., R. Orozco, E M. Kithsiri, P. Espinosa-Artiles, L. Gunatilaka, S. P. Stock, and I. Molnár. 2009. Biosynthesis of the cyclooligomer depsipeptide bassianolide, an insecticidal virulence factor of Beauveria bassiana. Fungal Genetics Biol. 46: 353-364.
Xu, Y., R. Orozco, E M. Kithsri-Wijeratne, P. A. A Gunatilaka, S. P. Stock, and I. Molnár. 2008. Biosynthesis of the cyclooligomer depsipeptide beauvericin, a virulence factor of the entomopathogenic fungus Beauveria bassiana. Chemistry and Biology 15: 898-907.
Yu, H., D. H. Gouge, and D. I. Shapiro-Ilan. 2010. A novel strain of Steinernema riobrave (Rhabditida: Steinernematidae) possesses superior virulence to subterranean termites (Isoptera: Rhinotermitidae). J. Nematol. 42: 91-95.