2006 Annual Report 1.What major problem or issue is being resolved and how are you resolving it (summarize project aims and objectives)? How serious is the problem? Why does it matter? The research to be undertaken falls within National Program 305 (Crop Production) Component 3 Section B (Bee Management and Pollination). Specifically, the project addresses the goals to:. 1)identify nutritional deficiencies that contribute to colony decline during pollination and develop/improve supplemental feeds, and. 2)determine factors that promote interbreeding and colony takeovers by African bees. Developing new methods to control Varroa mites falls within the goals of Bees and Pollination Section A. (Pest Management). More honey bee colonies are needed now by U.S. agriculture than ever before to pollinate crops. However, fewer colonies are available. Modern beekeeping practices exert extraordinary stress on honey bees because of overcrowded apiaries, frequent movement of colonies for pollination, and challenges from diseases and parasitic mites. Also facing the U.S. Beekeeping Industry is the problem of Africanization of European honey bees. Colonies from across the U.S. are moved to pollinate more than $15 billion worth of crops each year. Some enter areas with feral African populations (e.g., California for fruit and nut pollination or Florida for citrus). If colonies are moved into regions with African honey bees, they could become Africanized. When colonies are returned to their home apiaries, Africanized bees could be spread to non-Africanized regions of the U.S. The project has three specific objectives:. 1)create an artificial diet that beekeepers can feed to colonies during times when flowering plants are unavailable or when colonies are being transported for pollination;. 2)develop methods to control Varroa population growth in honey bee colonies by identifying new miticides and also by altering cues used by Varroa during host finding and reproduction, and. 3)develop methods to introduce European queens into Africanized colonies and prevent the invasion of European colonies by swarms of African bees. A minor project that we are conducting in cooperation with the USDA, APHIS, PPQ, CPHST, Soil Inhabiting Pest Laboratory, Gulfport, MS is to test the effects of fire ant repellents on honey bee colony foraging behavior and mortality. Fire ants are a major problem on bee equipment and can be spread when colonies are moved between states for pollination. In addition to testing repellents, we also are developing bait stations that can be used for inspection of honey bee colonies while in transit. The device will enable inspectors to determine quickly whether fire ants are on equipment being moved from a fire ant infested area to one that is free of fire ants. Attaining our Objectives will benefit beekeepers, queen breeders, and growers producing crops that require honey bee pollination for seed/fruit set. An artificial diet will help insure that colonies remain populous when flowering plants are not in bloom and while hives are transported for pollination. New methods to control Varroa will reduce the dependency on chemical miticides. Methods to prevent the Africanization of managed honey bee colonies will insure that manageable colonies are available for pollination, and possibly slow the spread of Africanized bees. Scientists will benefit from the information we will generate on honey bee nutrition, behavior and population interactions between European and African honey bees and advances in Varroa mite host-parasite behavior and population control. 2.List by year the currently approved milestones (indicators of research progress) FY 2005 Sub-Objective 1.1:Develop Patty Diet 1. Determine components needed for patty diet and test formulations for texture and stability. 2. Collect pollen from colonies to make SPP for feeding studies. 3. Identify phagostimulants in pollen. 4. Write manuscript on nutritional needs of honey bees. Sub-Objective 1.2: Objective 1.2 to state: Identify methods to incorporate miticides into liquid and solid artificial diets to control Varroa mites. 1. Begin choice tests with essential oils in the diet at various concentrations using adult worker bees. Sub-Objective 2.1: Identify volatile cues from honey bee hosts, Varroa, and brood cells. 1. Start colonies of various resistant and susceptible lines. Release Varroa into colonies. 2. Begin collecting volatiles from larvae and pupae. 3. Write manuscript on volatile profiles from resistant and tolerant bee lines. Sub-Objective 3.1: Determine effects of usurpation on worker-queen interactions. 1. Establish European colonies. 2. Begin filming worker-queen interactions as usurpations occur. 3. Refine sampling procedures as needed. 4. Collect data from films. Sub-Objective 3.2: Characterize pheromone profiles of invading bees. 1. Sample volatiles from swarm and host colony bees. 2. Adjust sampling procedures if needed. 3. Begin compound identification. 4. Write manuscript on differences in volatile from colonies depending upon their state. FY 2006 Sub-Objective 1.1: Develop patty diet. 1. Begin feeding trials with different formulations of diet to determine those most palatable to bees. 2. Conduct brood survival, longevity, and hemolymph protein analyses on colonies fed artificial diets. 3. Include compounds that are phagostimulants in diet mixtures to determine if they increase palatability. 4. Write manuscript on factors that stimulate and deter feeding. Sub-Objective 1.2: Objective 1.2 to state: Identify methods to incorporate miticides into liquid and solid artificial diets to control Varroa mites. 1. Track oils fed in the liquid diet to adult bees, nurse bees, and larvae. 2. Measure mite cell infestation and reproductive rates. 3. Begin choice tests with essential oils in the patty diet at various concentrations using adult worker bees. Sub-Objective: 2.1: Identify volatile cues from honey bee hosts, Varroa, and brood cells. 1. Continue collection of volatiles from larvae, mites, and cells. 2. Test for seasonal differences in volatiles from samples. 3. Begin identification of compounds. 4. Write manuscript on seasonal changes in chemical profiles of bees and mites. Sub-Objective 3.1: Determine effects of usurpation on worker-queen interactions. 1. Continue filming and data collection of worker-queen interactions during usurpation events. 2. Write manuscript on nest usurpation behaviors. Sub-Objective 3.2: Characterize pheromone profiles of invading bees. 1. Continue sampling volatiles from swarm and host colony bees. 2. Continue compound identification. FY 2007 Sub-Objective 1.1: Develop Patty Diet. 1. Add phagostimulants to the diet to determine if it causes bees to consume more diet. 2. Determine if and how long colonies can rear brood when fed exclusively on the patty diet. 3. Conduct brood survival, longevity, and hemolymph protein analyses on colonies fed artificial diets. 4. Write manuscript on the effects of diet on brood rearing. Sub-Objective 1.2: Objective 1.2 to state: Identify methods to incorporate miticides into liquid and solid artificial diets to control Varroa mites. 1. Continue to measure cell infestation and reproductive rates of mites. 2. Write manuscript on the effects of essential oils fed I diets on mite reproduction. Sub-Objective 2.1: Identify volatile cues from honey bee hosts, Varroa, and brood cells. 1. Continue identification of compounds. Sub-Objective 2.2: Evaluate reproductive rates of mites when exposed to selected compounds isolated from Objective 2.1. 1. Begin behavior bioassays to identify compounds that influence mite reproduction. 2. Write manuscript on effects of compounds on mite reproductive behaviors. Sub-Objective 3.1: Determine effects of usurpation on worker-queen interactions. 1. If necessary, continue filming and data collection. 2. Write manuscripts of worker-queen interactions during usurpation events. Sub-Objective 3.2: Characterize pheromone profiles of invading bees. 1. Continue sampling volatiles from swarm and host colony bees. 2. Continue compound identification. 3. Begin analysis to determine if volatiles from swarm workers change as usurpation progresses. FY 2008 Sub-Objective 1.1: Develop Patty Diet. 1. Continue tests to determine if and how l9ong colonies can reear brood when fed exclusively o the patty diet containing phagostimulants. 2. Conduct tests on longevity of adult workers fed the patty diet while as larvae and as adults. 3. Apply for possible patent for diet formulation. Sub-Objective 1.2: Objective 1.2 to state: Identify methods to incorporate miticides into liquid and solid artificial diets to control Varroa mites. 1. Determine minimum amounts of compounds needed to stimulate feeding. Sub-Objective 2.1: Identify volatile cues from honey bee hosts, Varroa, and brood cells. 1. Continue compound identification. Sub-Objective 2.2: Evaluate reproductive rates of mites when exposed to selected compounds isolated from Objective 2.1. 1. Continue bioassays with single compounds and combinations. 2. Write manuscript on effects of compounds (Various amounts and ratios) on mite reproduction. Sub-Objective 3.2: Characterize pheromone profiles of invading bees. 1. Continue analysis of the changes in volatiles. 2. Continue compound identification. 3. Write manuscripts on the role of volatiles in the usurpation of honey bee colonies. FY 2009 Sub-Objective 1.1: Develop Patty Diet. 1. Continue longevity tests. 2. Begin testing of patty diets in commercial beekeeping operations. 3. Write manuscripts on feeding patty diet to honey bees. Sub-Objective 2.2: Evaluate reproductive rates of mites when exposed to selected compounds isolated from Objective 2.1. 1. Continue bioassays with single compounds and combinations. 2. Write manuscripts on influence of various volatile compounds on mite reproduction rates. 4a.List the single most significant research accomplishment during FY 2006. We are the first to isolate and identify volatile compounds specific to honey bees. Four of the compounds were present only in queens and not in workers. One of these four compounds, identified as E-(-ocimene, was expressed fully only in established mated queens and may be a signal of queen egg-laying activity. The research falls within National Program 305 (Crop Production) Component 3 Section B (Bee Management and Pollination, Goal 2: determine factors that promote interbreeding and colony takeovers by African bees. A major problem in keeping European colonies in Africanized areas is colony takeovers by African bee swarms and difficulties with acceptance of European queens in Africanized colonies. Identifying compounds associated with laying queens will enable us to develop tools to protect European colonies that are in the process of being requeened, from invasion by swarms of African bees. Those tools might also be useful in increasing the acceptance rate of European queens into Africanized colonies. 4b.List other significant research accomplishment(s), if any. New methods to control Varroa mites: The research falls within National Program 305 (Crop Production) Component 3-Section A. (Pest Management). New methods to control Varroa mites in honey bee colonies must be developed, because the mites are resistant to compounds that are currently registered for control. Oxalic acid (OA) and a new miticide called Sucrocide were tested by Dr. Diana Sammataro in collaboration with investigators in Italy to determine their effectiveness in reducing Varroa populations. New delivery systems for the miticide, 2-heptanone, also were tested by Dr. DeGrandi-Hoffman. We concluded that Sucrocide is not effective when applied at recommended rate, but OA and 2-heptanone prototypes are effective at reducing mite populations. Dr. Sammataro also added various emulsified essential oils to the liquid protein diet formulated by our laboratory and when it was fed to bees, Varroa reproductive rates declined. These finding will serve as a basis for developing new products for beekeepers to control Varroa mites. Preventing European Colony invasions by Swarms of African Bees: The research falls within National Program 305 (Crop Production) Component 3 Section B (Bee Management and Pollination). Specifically, the project addresses Goal 2: determine factors that promote interbreeding and colony takeovers by African bees. Africanization of European honey bee colonies will reduce the number of colonies available for pollination. One way that colonies become Africanized is by invasion (i.e., usurpation) of European colonies by swarm of African bees. Drs. DeGrandi-Hoffman and Gilley developed methods for sampling nest odor and demonstrated that queen loss in European colonies affects the composition of volatile compounds emanating from the hive. Differences in nest odor between a queenright and queenless colony might be detected by usurpation swarms and be a basis for their selection of queenless colonies. Identifying the compounds that usurpation swarms use to find queenless European colonies can serve as a basis for developing tools to prevent colony usurpation by altering the odors of queenless colonies. 4c.List significant activities that support special target populations. None. 4d.Progress report. none. 5.Describe the major accomplishments to date and their predicted or actual impact. All accomplishment is to achieve goals of National Program 305 (Crop Production) Component 3 Section B (Bee Management and Pollination). Under Goal 1: Identify nutritional deficiencies that contribute to colony decline during pollination and develop/improve supplemental feeds, we formulated a liquid protein diet for honey bees that enables colonies to rear brood and produce worker bees with the same adult longevity as those fed a diet of natural pollens. This is the first liquid diet to be developed for honey bees. The diet is an improvement over current commercially available supplements in that bees can rear brood over extended periods (i.e., more than 2 months) when fed exclusively on the diet. A patent for the diet was filed as were the licensing agreements with our CRADA partner and contracts with a commercial manufacturer. A solid patty form of the diet is being developed. The diet will be used by beekeepers and queen breeders to feed colonies during times when pollen is not available. The accomplishment will benefit both the beekeeping industry and growers of crops requiring honey bee pollination for seed and fruit or nut set. The Varroa mite is the most important pest of honey bees and there are no reliably effective products available for controlling this pest. We have identified new methods to control Varroa mites in honey bee colonies using non-petroleum based miticides. These accomplishments address National Program 305 (Crop Production) Component 3 Section A. (Pest Management). The new miticides we have identified are either produced by honey bees (2-heptanone) or are food additives (plant acids from hops and plant essential oils). We have documented that 2-heptanone is a potent miticide and are developing a delivery system in collaboration with the USDA-ARS Bioproducts Chemistry and Engineering Research Unit in Albany, California to dispense it in colonies. A U.S. patent has been filed for the compound. We also have identified several plant-based compounds that are byproducts from the processing of hops. We have established a CRADA with John Haas Inc. to develop formulations and delivery systems for these compounds so that they can be used in hives to control Varroa. We also have identified an inert compound as a potential mite control that does not affect bee behaviors. This work is in collaboration with CRADA partner, Pima Research LLC. We have developed techniques to collect volatile compounds from all lifestages of worker and drone honey bees. With these techniques, we have obtained chemical profiles from mite resistant and non-resistant bee lines. We are now documenting the presence/absence of specific volatile compounds from the bee lines and are identifying them. Those compounds that influence Varroa host finding and reproduction will become the basis for our novel mite controls in the next few years. The beekeeping industry will benefit greatly from our research as will growers of crops requiring honey bees. Furthermore, since we have identified several methods to control Varroa, beekeepers can use the products that will be developed in rotation to avoid mite resistance to any one compound. In our research on determining factors that promote interbreeding and colony takeovers by African bees (National Program 305 (Crop Production) Component 3 Section B (Bee Management and Pollination)) we have documented that differences in volatile compounds particularly E-'-ocimene affect the acceptance rates of queens in honey bee colonies. We also have documented that Africanized honey bee colonies have lower acceptance rates of queens compared with European colonies. This research is the first step in solving the problem of Africanization of managed honey bee colonies. Identifying compounds associated with queen egg laying strength will enable us to develop tools to improve the acceptance rate of European queens into Africanized honey bee colonies so that the Africanization process can be reversed. The research also will lead to tools to protect European colonies from invasion by African swarms particularly when the colony is in the process of being requeened and is particularly vulnerable to invasion. The beekeeping industry will benefit greatly from our research as will growers of crops requiring honey bees. 6.What science and/or technologies have been transferred and to whom? When is the science and/or technology likely to become available to the end-user (industry, farmer, other scientists)? What are the constraints, if known, to the adoption and durability of the technology products? Through a series of articles, we have reported to beekeepers how to identify the early stages of Africanization in colonies, how to prevent Africanization, and what to do if colonies become Africanized. The papers were the result of research on how African honey bees displace European populations and behaviors associated with the hybridization of European and African bees. Research on control of Varroa mites was presented to the following groups: January 2006: National meetings for the American Honey Producers, Houston, TX; and the American Beekeeping Federation, Louisville, KY. February 2006: North Carolina State Beekeepers Association, Asheville, NC March 2006: Kansas and Missouri Beekeepers, Overland Park, KS; Michigan State University ANR Week, E. Lansing MI; 24th Annual Science and Math Conference for 7-12th grade women in S. Arizona, University of Arizona, Tucson, AZ (panel). April 2006. Ulster Bee Keeper's Association, Greenmount, N. Ireland August 2006. International Acarology Congress, Amsterdam, Netherlands. November 2006. Empire State Beekeepers, Syracuse, NY and S.E. New England Beekeepers Assembly, Hamden, CT. 7.List your most important publications in the popular press and presentations to organizations and articles written about your work. (NOTE: List your peer reviewed publications below). DeGrandi-Hoffman, G., Tarpy, D., Chambers, M., Schneider, S., Smith, D. The African Honey Bee Has Arrived – So Where Do We Go From Here? Bee Culture 134 (June): 24-32. 2006. Loper, G.M., Sammataro, D., Finley, J., Cole, J. Feral honey bees in Southern Arizona, 10 years after Varroa infestation. Am. Bee J. 146:(6): 521-524. 2006. Sammataro D. Mites of the Honey Bee. Bee Craft 88:14-19. 2006. Schneider, S.S., DeGrandi-Hoffman, G., Smith, D. The African Bee: A case study of biological invasion. Bee Culture 134 (April): 21-25. 2006. Schneider, S.S., DeGrandi-Hoffman, G., Smith, D. The Displacement of European Honey Bees by African Bees in the New World. Bee Culture 134 (May): 25-30. 2006.