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ARS Home » Southeast Area » Baton Rouge, Louisiana » Honey Bee Lab » Research » Research Project #426305

Research Project: Genetics and Breeding in Support of Honey Bee Health

Location: Honey Bee Breeding, Genetics, and Physiology Research

2018 Annual Report

The overall goal of this research is to develop and use genetic resistance or tolerance of honey bees to biotic agents, and to devise management strategies to improve the quality of queen honey bees. This will enhance the economic value of the nation’s honey bees for pollination, honey production, overwintering and hazard resistance. Over the next five years we will focus on interrelated projects with the following objectives: Objective 1: Identify and evaluate traits, strains, and stocks for improved honey bee health, e.g., improved immunity, tolerance, or resistance to Varroa and tracheal mites, the fungi Nosema and chalkbrood, and viruses. Sub-objective 1A: Evaluate the potential for viral resistance. Sub-objective 1B: Establish and characterize genetically and functionally distinct lines that differentially respond to Nosema. Sub-objective 1C: Evaluate the potential of worker brood to suppress reproduction by Varroa. Sub-objective 1D: Determine the effect of Varroa infestations on honey bee behavior. Sub-objective 1E: Evaluate immune and stress related responses in selected honey bee stocks. Objective 2: Characterize genetic and physiological aspects of important traits, strains, and stocks and their interaction with biotic and abiotic stressors. Sub-objective 2A: Characterize the chemical ecology of VSH to aid the development of a practical selection method. Sub-objective 2B: Understand the genetic basis of Ascosphaera apis (chalkbrood) resistance observed in RHB larvae. Sub-objective 2C: Genomic sequencing of multiple stocks of honey bees. Sub-objective 2D: Differential responses to Nosema in honey bees. Objective 3: Conduct traditional and marker-assisted breeding and develop management tools for improved bees. Sub-objective 3A: Develop and use improved methods to evaluate Varroa-resistant phenotypes. Sub-objective 3B: Determine management methods using ionizing radiation to increase honey bee colony fitness. Sub-objective 3C: Evaluate autogrooming as a common resistance mechanism towards tracheal and Varroa mites. Sub-objective 3D: Use traditional and marker-assisted selection programs to improve honey bees with VSH-based resistance. Funding will be used in support of Objective 3: Conduct traditional and marker-assisted breeding and develop management tools for improved bees. [NP305, Component 2, Problem Statements 2A and 2B] Objective 4: Improve knowledge of the biology, physiology, genomics, and behavior of pests (i.e. mites and the small hive beetle) that may be useful for improving their management. Sub-objective 4A: Elucidate the Nosema infection process. Sub-objective 4B: Understand effects of resistance level on Nosema infection across honey bee life stages. Sub-objective 4C: Identify new methods to limit SHB population growth in honey bee colonies.

The health and availability of honey bees (Apis mellifera) have been diminished by multiple biological problems, most notably the parasitic mite Varroa (V.) destructor. Other important threats include fungi (Nosema spp. and chalkbrood), viruses, small hive beetles and tracheal mites. Some of these interact and result in mortality described as colony collapse disorder (CCD). The major goal of this project is to mitigate these threats by finding and selecting for genetically based traits of honey bees that confer resistance to the biological problems. Scientists also will improve management techniques and describe pest biology related to control. Most research targets V. destructor. Efforts that build on our prior work with Varroa-resistant honey bees include using knowledge about Russian Honey Bees to find new traits that enhance resistance, and to improve selection methods. Honey bees with Varroa sensitive hygiene are undergoing traditional and molecular-marker-assisted selection. Marker-assisted selection would be a valuable tool to overcome the difficulty of phenotyping this trait, and increase the capacity to select and breed Varroa-resistant bees. Novel traits for Varroa resistance also are being sought. New selection of honey bees will target Nosema ceranae and Deformed Wing Virus because these pathogens are thought to contribute significantly to colony losses. Research with Nosema, viruses, tracheal mites and chalkbrood will culminate in development of molecular markers for resistance factors. Management research will expand promising preliminary findings about queen and drone health associated with the irradiation of comb in mating colonies The products of this research -- knowledge and technology -- will strengthen integrated pest management strategies for controlling honey bee pests and diseases. This in turn should lead to better profitability for beekeeping and crop pollination.

Progress Report
This is the final report for project 6050-21000-014-00D, which falls under National Program 305, Component 2 - Bees and Pollination, and it will be replaced with a bridging project pending completion of research review. Progress was made on all four objectives and most of their sub-objectives, despite critical vacancies during a significant portion of the project’s cycle. Progress was made in bidirectional selection of honey bees for response to Deformed wing virus. Bees surviving infection with few symptoms appear to be tolerant of high infection levels, apparently in part due to antiviral immune responses. The heritability of resistance or tolerance to this virus has yet to be determined. Additional research is examining how bees resistant to Deformed wing virus respond to Israeli acute paralysis virus, and the role of reactive oxygen species in viral infection. Research on differential selection of honey bees to Nosema was largely halted after an initial determination of resistance could not be reliably replicated in subsequent generations of selection. Nosema remains a refractory pathogen. A potential trait of honey bee resistance to Varroa mites is an effect that originates from the bee brood on which mites feed; this "brood effect" suppresses reproduction of the mites. We were unable to stabilize this trait over several generations of testing and genetic selection. Work is underway to measure the impacts of Varroa mite parasitism on flight activities, survival and brain chemistry of workers and drones, with the goal of determining magnitude of effects due Varroa directly versus the viruses which the mites transmit. Measuring effects of Varroa on sperm number and viability are ongoing. We determined that Russian honey bees express significant levels of the behavioral trait of Varroa sensitive hygiene, which contributes to enhanced Varroa resistance. Genetic mapping indicated that Varroa sensitive hygiene in Russian honey bees is controlled by the same gene locus found in other bees that express the trait at high levels, but with allelic effects reversed in Russian bees. Progress was made in a subordinate project with partners from Louisiana State University to clarify the dynamics of infection of Deformed wing virus. A 1-year study is underway to compare rates of virus infection, dissemination and transmission in three bee stocks, and to assess the compounding effects of virus prevalence, titers and pathogenicity with Varroa infestation levels. This information will be used to build a model to predict threats from viruses early enough in a season to effectively trigger mite control strategies. Progress was made in the development of a genomic database for commercially important honey bees. Bioinformatics is underway on the sequence information from seven stocks. Resilience of genetically diverse honey bee colonies was found to arise not only from behavioral responses of adult bees but also because individual bees have less variable innate immune responses. Vacancies delayed progress in work to determine the chemical ecology that regulates expression of Varroa sensitive hygiene. We have supplied queens with high expression of the trait to a partner from the University of North Carolina in a subordinate project, and progress was made toward identifying chemical cues related to the behavior. Initial research showed that some parameters of Varroa falling to the bottom of a hive may be an indicator of the mite resistance of a colony, and demonstrated the influence that hygienic behavior has on the rate of falling mites. Vacancies subsequently delayed progress on this research. Laboratory testing showed that in vitro gamma irradiation of microbes completely eliminated the pathogenicity of Nosema, chalkbrood and Deformed wing virus, but had a limited effect on Black queen cell virus and Chronic bee paralysis virus. In the field, the use of irradiated comb in queen mating colonies did not alter the expression of immunity genes in queens. Using gamma irradiated comb in field colonies showed only minor, short-lived effects on pathogen levels in bees emerging from the treated comb, with no benefits to colony productivity. Pol-line honey bees were selectively bred in collaboration with large beekeeping operations during 2008-2014. The bees have mite resistance that is based on Varroa sensitive hygiene. Further breeding currently is being conducted in a public-private partnership in which Pol-line forms much of the founding population. Selection is focused on stabilizing Varroa resistance and improving beekeeping characteristics such as honey production and large colony size, specifically for commercial beekeeping. A refined stock of bees is being targeted for release to industry within about a year. Molecular markers from analysis of quantitative trait loci were developed for bees with chalkbrood resistance and with Varroa resistance based on Varroa sensitive hygiene. However, these markers have not been validated as being useful outside of the original mapping populations of honey bees. This finding indicates the potential difficulty in this approach for finding markers suitable for marker-assisted selection. In an alternative approach for markers related to Varroa sensitive hygiene, several candidate genes with sensory functions were assessed in individual bees and in groups of bees from colonies that had differential expression of the trait. Two genes show potential for further marker development. Also, work is ongoing in which phenotype data and bee samples are provided to a collaborator at the Institut National de la Recherche Agronomique (France) for whole genome sequencing to seek molecular markers associated with Varroa resistance. A comparison of Russian, Italian, Italian X Russian hybrid and commercial outcrossed Russian colonies showed that pure Russian colonies consistently maintained low mite levels and survived the longest while pure Italian colonies had the highest mite numbers and shortest survival. Italian X Russian colonies were intermediate whereas the commercial Russian outcross colonies were inconsistent. Progress was made in several subordinate projects. Studies with collaborators at the University of Sydney (Australia) resulted in a set of single nucleotide polymorphisms which can be used to identify Africanized-bee ancestry in honey bees. Longitudinal field trials with collaborators at Louisiana State University is yielding information about the relative health threats to honey bees in a commercial beekeeping operation; the benefits of mite-resistant stock were apparent under the test conditions. Research in collaboration with partners at the University of Missouri has begun on a search for molecular markers for Varroa sensitive hygiene based on an “eQTL” approach, i.e., a combination of both gene sequence and gene expression data. Research in collaboration with partners at the University of Minnesota has begun to clarify the role of propolis in honey bee immunity and its potential benefits in beekeeping management, and to breed bees with improved health founded on social immunity to parasites and pathogens. Research in conjunction with the ARS Insect Genetics and Biochemistry Research Unit in Fargo, North Dakota, is advancing cryopreservation techniques for both semen and embryos of honey bees. This work supports the USDA National Animal Germplasm Program, a “bee gene bank” that conserves valuable honey bee genetic material. Vacancies and changes in scientific personnel delayed work on whether the trait of autogrooming is effective in helping honey bees resist both Varroa mites and tracheal mites. An assay was developed to detect vegetative and spore life stages of Nosema ceranae using density-gradient centrifugation. The assay is being used to measure changes in Nosema life stages across honey bee developmental stages in field colonies. The reproductive rate of small hive beetles was determined to increase when beetles fed on a diet of brood, pollen and honey, when the temperature was equal to that found in bee colonies, and when male beetles were abundant in a population. Screen bottom boards, used as a tool to manage Varroa mites, did not encourage invasion by small hive beetles. Several endotoxins from Bacillus thuringiensis were screened as potential control materials against small hive beetles, but none were suitably effective to be recommended as a control. In collaboration with partners, genome and transcriptome data for small hive beetles were developed and deposited in a public database; the information is being used as a foundation for further exploration of potential species-specific, molecular-based control measures that may reduce the problems from this hive depredator. Work in subordinate projects increased understanding of other threats to honey bees. A review of the ecology, life history and management of Tropilaelaps mites provided an overview of the threat that these exotic parasites could pose to U.S. honey bees. Novel observations include reproduction despite no phoresy and no mating, and feeding on unsealed bee brood. Studies with partners at Louisiana State University examined the exposure of honey bees to pesticides in a beekeeping operation in the upper Midwest. Pesticide hazards are being analyzed; Varroa mite infestation was a significant factor in colony mortality. Other work on toxicology showed that insecticide sensitivity in honey bees is affected by genetic background and age of bees, and that insecticides used against adult mosquitoes are generally safe for beekeeping when used according to label directions. Work is underway with a U.S. Geological Survey in Lafayette, Louisiana, collaborator to compare sperm quality and sperm count assays, which will be used in the development of an andrology panel useful for understanding not only drone reproductive fitness but also queen health.

1. Identification of Africanized ancestry in honey bees. Africanized bees remain a concern in some areas, and are a major regulatory issues when honey bees are moved between countries. Methods to analyze the degree of Africanization are outdated or expensive. ARS researchers at Baton Rouge, Louisiana, and collaborators developed a technique that is accurate and also reduces the time and cost of screening samples. The analysis can be used for reliable determination of Africanized heritage of bees imported from countries like the United States (where Africanized bees are present) into countries like Australia (where Africanized bees are absent).

2. Genetic stability of a honey bee breeding population tracked with genetic markers. As a companion to the commercial release of the Russian honey bees, a genetic stock identification assay was developed to distinguish Russian honey bees from other U.S. commercial bees. An ARS researcher at Baton Rouge, Louisiana, used genotype data to assess the genetic stability of the Russian honey bee breeding population. The genetic population structure in 2016 remained similar to that first characterized in 2010. The successful application of the genetic stock identification assay in a commercial breeding program demonstrates the utility and stability of such technology to contribute to and monitor the genetic integrity of a breeding stock of an insect species.

Review Publications
Kirrane, M.J., De Guzman, L.I., Frake, A.M., Rinderer, T.E., Whelan, P.M. 2018. Evaluations of the removal of Varroa destructor in Russian honey bee colonies that display different levels of Varroa Sensitive Hygienic activities. Journal of Insect Behavior.
Simone-Finstrom, M., Tarpy, D.R. 2018. Honey bee queens do not count mates to assess their mating success. Journal of Insect Behavior. 31(2):200-209. doi: 10.1007/s10905-018-9671-3.
De Guzman, L.I., Phokasem, P., Khongphinitbunjong, K., Frake, A.M., Chantawannakul, P. 2018. Successful reproduction of unmated Tropilaelaps mercedesae and its implication on mite population growth in Apis mellifera colonies. Journal of Invertebrate Pathology. 153:35-37.
De Guzman, L.I., Frake, A.M., Simone-Finstrom, M. 2017. Comparative flight activities and pathogen load of two stocks of honey bees reared in gamma-irradiated combs. Insects. 8, 127. doi:10.3390/insects8040127
Simone-Finstrom, M., Aronstein, K.A., Goblirsch, M., Rinkevich Jr, F.D., De Guzman, L.I. 2018. Gamma irradiation inactivates honey bee fungal, microsporidian, and viral pathogens and parasites. Journal of Invertebrate Pathology. 153:57-64.
Chapman, N.C., Bourgeois, A.L., Beaman, G.D., Lim, J., Harpur, B.A., Zayed, A., Allsopp, M.H., Rinderer, T.E., Oldroyd, B.P. 2017. An abbreviated SNP panel for ancestry assignment of honeybees (Apis mellifera). Apidologie. doi:10.1007/s13592-017-0522-6.
Bourgeois, A.L., Beaman, G.D. 2017. Tracking the genetic stability of a honeybee breeding program with genetic markers. Journal of Economic Entomology. 110(4):1419-1423.