2011 Annual Report
1a.Objectives (from AD-416)
The central theme of this project is to reduce the impacts of pests and pathogens on honey bees using approaches ranging from field experiments to controlled pathology experiments and modern genetic and genomic applications. Specific objectives are to.
1)Improve screening and management methods used by beekeepers to minimize losses due to Varroa mites and other stress factors, focusing on queen supersedure, worker longevity, and catastrophic losses such as colony collapse disorder,.
2)Measure the individual and combined impacts of key honey bee disease agents including Varroa, viruses, Nosema, and the American foulbrood bacterium under field, cage, and laboratory conditions, .
3)Define the resistance mechanisms of bees toward pathogens, especially bacteria and viruses, focusing on individual and group defenses as a means of providing candidate traits for breeding programs,.
4)Develop and improve collection, culture and expression systems for continuous production of disease-causing pathogens in order to provide ready source experimental material and disease reproduction models for in vitro and in vivo assessments of pathogenesis and host-pathogen interactions of honey bees and 5)Determine the roles of aging and stress on honey bee worker and queen longevity in order to improve overwinter survival and minimize losses to CCD.
1b.Approach (from AD-416)
Developing and adult bees will be exposed to incidental pesticides, and to acaricides used to control Varroa mites, in order to determine the vulnerabilities of bees to these chemicals. A central goal will be to determine and validate methods for remediating honey bee comb containing potentially dangerous levels of chemicals or pathogens. Impacts of two species of Nosema on queen supersedure rates, worker mortality, and colony declines will be studied using controlled cage experiments and field treatments with the Nosema control fumagillin. These experiments will be followed by microscopic and genetic tests of Nosema loads, and tests of honey bee immune responses and resistance to Nosema. Activity levels of honey bee immune genes and genes related to chemical stress can be indicators of resistance mechanisms present in some bee lines, and can help test the impacts on bees of specific management techniques. Resistance to American foulbrood disease will be determined by screening lines of bees that survive controlled infection to the bacterial cause of this disease. In addition, new techniques for silencing honey bee and/or bacterial genes will be used to determine new avenues for controlling this important disease. Work on viral pathogens of bees will focus on developing controlled genetic assays for diverse viral species in bees, determining specific virulence factors in these viruses, and determining the efficacy of gene silencing and other resistance mechanisms used by honey bees to resist viral disease. Viral research will also focus on transmission mechanisms of viruses, in anticipation of determining the most economical means for reducing the impacts of direct or indirect (e.g., Varroa mite) transmission of bee viruses. A genome sequencing project for the critical honey bee pest Varroa destructor will be used to identify and validate targets for mite control, define mechanisms of mite orientation and reproduction, disrupt the ability of mites to transmit viruses, determine novel microbial control agents for this parasite, and genetic information for novel mite controls. New methods will be developed to collect, transport, purify and diagnose honey bee pathogens from the field using genetic techniques. Experimental systems for propagating and maintaining viruses and other pathogens will be used to assess virulence and host-pathogen interactions. The aging process in workers bees will be examined by exploring the physiological parameters that define long-lived bees. Specifically, research into the genetics of longevity will be undertaken along with studies using specific stressors, pesticides and resource availability to determine their role in worker life expectancy. Genetic and experimental approaches will be used to determine the impacts of pesticide exposure on the virulence and spread of honey bee pathogens. Colony level experiments will build on the work with individual bees and explore the role of the above stressors on colony overwintering and the production of long-lived winter bees, a key to understanding colony collapse disorder (CCD) as most colonies die from CCD in the fall and winter.
Honey bee colonies are threatened by numerous parasites, pathogens, and pests. The industry is also impacted by nutritional and chemical stresses placed on honey bees. In the past few years, domesticated honey bee colonies have suffered alarming and enigmatic losses, a syndrome labeled Colony Collapse Disorder (CCD). Specific focus areas in 2011 included experimental analyses of the biology and impacts of several pathogens implicated in Colony Collapse Disorder, and experiments seeking to determine the effects of nutrition and chemical exposure on pathogen and parasites. As a representative RNA virus, Israeli acute paralysis virus was studied, and it was determined that Varroa mites transmitted this virus, and that nutritional status of bees had an effect on the ability of this virus to replicate. The effects of the important gut parasite Nosema ceranae on bees were measured in controlled settings and, while this parasite seems to have a minimal effect on bees in clean environments, there is some suggestion of negative interactions with other bee threats.
Viruses transmitted to bees by parasitic mites are able to pass major barriers in the bee defense system, leading to higher impacts. Along with acting as vectors for viruses, these mites are known to cause extensive damages to bee health, through physiological effects on developing bees. A main advance for 2011 was the publication of a draft genome sequence for the parasitic mite Varroa destructor. The genome draft, while incomplete, showed many elements of the mite immune system and the proteins available to mites to recognize bee hosts and develop as parasites. Several microbes were also found in association with mites, including a virus and a bacterial species that are possible candidates for controlling for these mites.
Controlling the key parasite Varroa destructor through genomics. Novel controls based on genomics will provide for strong control of parasitic mites while reducing the use of chemicals. A genome draft sequence was published for this mite, revealing potential weak points in mite biology (defensive proteins, and proteins used in chemical mitigation) and candidates for novel controls such as RNA interference (RNAi), a method for knocking down specific pest proteins. The description of mite candidate genes allowed the initiation worldwide of RNAi efforts for this parasite, ending in late 2010 with the first successful demonstration of RNAi activity in Varroa. Microbes identified in this screen have also been screened across bees and mites as plausible controls for Varroa. Companies producing products for bee health will use these results to expand the mite control tools available to beekeepers.
Tools development for tracking and understanding CCD. Efforts to improve bee health have suffered from an inability to accurately assess disease caused by viruses and other pathogens. BRL scientists improved current methods to collect field honey bees populations and ship for genetic analyses, stabilize and extract RNA, conduct high-throughput genetic screens for viruses and other pests, collect embryos from established colonies, and carry out controlled experiments on adult bees using sterile cups. These methods are being used in national surveys in the U.S. in order to establish cell lines and other genetic techniques, and to better determine interactive effects.
Nosema ceranae impacts. Controlled experiments documented interactions between Nosema parasites and other pathogens in causing CCD. While multifactorial analyses did not point to Nosema as an integral part of CCD, this parasite was confirmed as being present wherever CCD was found, and at higher levels than in past surveys. In addition, it became clear that Nosema ceranae has recently spread worldwide and is a permanent part of bee populations on all continents with beekeeping.
Insect Immunity. Honey bees combat American foulbrood, viruses, and Nosema with immunity proteins found across the insects. A BRL scientist determined immune gene protein variation across bees and ants, an essential step in determining which proteins are responsive to novel viruses, bacteria, and fungi. This information is being used in programs to test specific immune genes for breeding purposes.
Movement of the small hive beetle worldwide. Genetic analyses revealed the origins of two new populations of the small hive beetle, Aethina tumida, an important parasite of bees. This beetle has left subharan Africa at least twice (and likely three times), and continues to spread even to countries which do no accept bee importations form outside (e.g., Australia). Along with reducing impacts of this pest across the world, these results have immediate impact in trade decisions and regulations.
Causes and treatment for Colony Collapse Disorder. BRL scientists determined likely factors in CCD, including RNA viruses, chemical exposure, and the gut parasite Nosema ceranae. These efforts confirmed the view that CCD is a multifaceted problem, with both biological and abiotic causes, and that hive management offered one possible route for mitigating CCD. Understanding how biological causes like viruses interact with other bee threats will improve the ability of beekeepers to manage their bees.
Viljakainen, L., Evans, J.D., Hasslemann, M., Rueppel, O., Tingek, S., Pamilo, P. 2009. Rapid evolution of immune proteins in social insects. Molecular Biology and Evolution. 26:1791-1801.
Evans, J.D., Chen, Y., Diprisco, G., Pettis, J.S., Williams, V.P. 2009. Bee cups: Single-use cages for honey bee experiments. Journal of Apicultural Research. 48(4):300-302.
Frazier, M., Muli, E., Conklin, T., Schmehl, D., Torto, B., Frazier, J., Tumlinson, J., Evans, J.D., Raina, S. 2010. A Scientific note on Varroa mites found in East Africa; Threat or Opportunity. Apidologie. 41:463-465.
Lounsberry, Z.T., Spiewok, S., Pernal, S., Sonstegard, T.S., Hood, M.W., Pettis, J.S., Neumann, P., Evans, J.D. 2010. Chasing your honey: Worldwide diaspora of the small hive beetle, a parasite of honey bee colonies. Annals of the Entomological Society of America. 104:671-677.
Evans, J.D., Boncristiani Jr., H.F., Chen, Y. 2010. Scientific note on mass collection and hatching of honey bee embryos. Apidologie. 41:654-656.
Cornman, R.S., Schatz, M.C., Johnston, S.J., Chen, Y., Pettis, J.S., Hunt, G., Bourgeois, A.L., Elsik, C., Anderson, D., Grozinger, C.M., Evans, J.D. 2010. Genomic survey of the ectoparasitic mite Varroa destructor, a major pest of the honey bee Apis mellifera. Biomed Central (BMC) Genomics. 11:602.
Dainat, B., Evans, J.D., Chen, Y., Neumann, P. 2011. Sampling and RNA quality for successful diagnostics using quantitative PCR. Journal of Pest Science. 174:150-152.