Progress 01/01/10 to 12/31/14
Outputs
Target Audience: 1. Beekeepers in Michigan. 2. Beekeepers in the Midwest States, as covered by Heartland Apicultural Society (HAS), which includes MI, OH, IN, IL, KY, AL, TN, MO, and WV. 3. Beekeepers and general public as reached by bees.msu.edu and cyberbee.net, about 130 countries/regions (top level domains). Changes/Problems: We worked in 2007 and 2008 on some experiments but used N. ceranae spores stored under refrigeration. Since then, studies have found that N. ceranae spores were more sensitive to cold storage and that rendered our study less ideal. We had to replicate the same studies using fresh spores (within 24 hours). What opportunities for training and professional development has the project provided? We hosted several visiting scientists, two graduate students and one postdoctoral researcher. How have the results been disseminated to communities of interest? We published three high impact publications in American Bee Journal, Bee Culture and eXtension.org website. Beekeepers found the information highly educational and many discussions were started in various websites. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
We have studied Nosema ceranae and determined that: 1). The large discrepancy between Spanish studies?that bees infected with Nosema ceranae dies?within 7 days after inoculation in Spain but not in North America, was not due to a technical difference. We treated 5 day old bees with carbon dioxide, and then inoculated bees with N. ceranae, similarly to the Spanish studies, but bees did not die as in their studies. We conclude that most likely Spanish bees are genetically different and more susceptible to N. ceranae. 2). We found that workers in colonies experiencing transportation and pollination services showed higher Nosema prevalence (% infected) and abundance (spore loads), compared to the baseline (prior to transportation). A control group (not moved to pollination) did not experience this increase. 3). We found that pollen nutrition exerted such a large effect such that pollen nutrition did not interact with nosema infection as we originally expected. Workers had no pollen survived much worse, regardless if infected with Nosema or not. The effect of nosema on mortality was weaker compared to pollen itself. and 4). we discovered that when bees were inoculated with both species (N. apis and N. ceranae), that their mortality increased significantly compared to those inoculated with a single species (either N. apis or N. ceranae). This suggests that the two species might be attacking different target sites inside honey bee workers.
Publications
- Type:
Journal Articles
Status:
Accepted
Year Published:
2014
Citation:
Meghan Milbrath; Wei-Fong Huang; Leellen Solter; David Tarpy; Frank Lawrence, Zachary Y. Huang. 2014. Comparative
virulence and competition between Nosema apis and Nosema ceranae in honey bees (Apis mellifera) . J. Invert.
Pathology.
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Progress 01/01/13 to 09/30/13
Outputs
Target Audience: Beekeepers and the general public throughout the world accessed my website, http://bees.msu.edu, the Michigan Beekeeper Association website, http://michiganbees.org and eXtension.org web. Colleauges at other institutions are also reached by articles published in referred journals. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided? I attended and presented at the Michigan Beekeepers Association ANR week Program (March 2013), Center of Michigan Beekeepers Meeting (June 2013), Heartland Apicultural Society Conference (July 2013), and Michigan Beekeepers Association Fall Conference (Oct 2013) to interact and educate beekeepers. I also presented at the Science Cafe (Oct 2013) in Ann Arbor to interact and present to the general public. How have the results been disseminated to communities of interest? Through stakeholder meetings and conferences (see above). What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
Under Objective 3 EFFECT OF NOSEMA CERANAE ON HONEY BEES This year we studied whether pollination and its required transportation would lower honey bees resistance and cause their Nosema ceranae prevalence (percent infection) and abundance (spore loads) to increase. We devided 14 colonies into two graoups and one group remained stationary (S) and one group was transported to Fenville for blueberry transportation (T). Both prevalence and abundance increased significantly in the T group compared to the levels prior to transportation, whereas the control group S did not experience such an increase in either parameters. These data are soon to be published as a note in Journal of Apicultural Research (accepted). We tried to study whether Nosema ceranae infection would change the honey bee workers’ thermal preference, hypothesizing that honey bee workers might have ways to detect if they are infected or not and then adjust their temperature preference to reduce either spore germination or spore production. Alternatively, the parasite (Nosema ceranae) migth alter its host behavior to benefit itself (e.g. higher fitness via higher spore production). However we could not find any changes in thermal preferences in bees of various ages by comparing those infected with Nosema ceranae and those not. We replicated the pollen and nosema interaction because last time (2009/2010) we did not use fresh nosema spores since at that time it was not known N. ceranae was sensitive to cold temperature during storage. We used three colonies and subjected newly emerged workers to four treatments: with/without nosema infection and with/without pollen provided. Similar to our previous trials, lack of pollen caused a larger reduction in survival than nosema infections. However, nosema infection also caused a reduction in survival independent of pollen presence or absence. These results suggest the important of pollen nutrition to honey bees and also confirm previous results that N. ceranae itself can cause significant shortening of worker lives.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Goblirsch, M., Z.Y. Huang, M. Spivak. 2013. Physiological and behavioral changes in honey bees (Apis mellifera L.) induced by Nosema ceranae infection: A potential detractor of social resiliency. PLoS ONE 8(3): e58165. doi:10.1371/journal.pone.0058165
Milbrath, M.O., X. Xie, Z.Y. Huang. 2013. Carbon dioxide anesthesia affects mortality of Nosema ceranae infected honey bees. J. Invertebrate Pathology. 144: 42-44.
Zhu, X., Zhou, S, Z.Y. Huang*. 2013. Transportation and pollination service increase abundance and prevalence of Nosema ceranae in honey bees (Apis mellifera). In Press, Journal of Apic Res.
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Progress 01/01/12 to 12/31/12
Outputs
OUTPUTS: EFFECT OF NOSEMA CERANAE ON HONEY BEES Last summer (2011) we had two trials examining the effect of nosema infection honey bee foraging and survival. This summer (2012) we replicated the experiment in two more nucs. In 2 out of 3 trials, bees with mixed infections foraged earlier than either control bees or those with pure infections. In all three trails, bees with mixed infections had lower survival than bees in the other treatment categories. These data indicate that effects of nosema infection my be dependent on co-infection and other factors. Because results in North America and elsewhere cannot repeat Higes' early results that N. ceranae was more virulent than N. apis that N. ceranae kills bees within 10 days post infection, we examined the joint and separate effects of CO2 anesthesia and nosema infection using a 2x2 design. Of the bees not anesthetized with CO2, those that were infected with nosema survived significantly shorter than uninfected bees (P < 0.01). But these that were anesthetized with CO2, the differences in survivalship between infected and uninfected bees disappeared (P > 0.05). Interestingly, we observed significantly higher spore counts in bees that did not undergo CO2 anesthesia (t=7.53, df=138.67, P < 0.01). We anticipated that this was a function of a longer survival time, and thus longer multiplication time for nosema. However, we observed a higher mean spore count consistently for bees that died at all ages. These data not only aid in the understanding of nosema related mortality, but also provide useful insight on experimental techniques that can be used to understand discrepancies in previous work. HUNTING FOR GENES IMPORTANT FOR VARROA MITE SURVIVAL AND REPRODUCTION USING RNAi We have shown last year that using RNA interference (RNAi), some genes affected mite reproduction and others affected mite survival. This year we examined three more genes: Rps13, RPL11 and Rpp0, all of them affected mite reproduction. Daughterless (Da) and Pts26.4 affected mite survival. Understanding genes important in mite survival or reproduction will help us devise novel method for varroa mite control. PARTICIPANTS: Xianbing Xie, Postdoctoral Fellow Meghan Milbrat, Postdoctoral Fellow Samuel Slagell, summer assistant Emily Crouse, HSHSP Fellow TARGET AUDIENCES: Beekeepers and the general public through out the world accessed my website,http://bees.msu.edu, the Michigan Beekeeper Association website, http://michiganbees.org and eXtension.org web. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
Honey bees play a critical role in US and Michigan agriculture by providing pollination services to the majority of fruits and vegetables. Honey bees are attacked by a plethora of diseases and pests that threaten their health. Beekeepers are gaining more knowledge in pest and disease control as a result of our research and extension efforts. Results of our study were jointly published in the American Bee Journal and Bee Culture, the two most important trade journals for beekeepers in the U.S. The same materials were published electronically on eXtension.org and Michiganbees.org. Requests are coming in for the use and instructions of the new technology for varroa mite control, Mitezapper. Our goal is to sustain or increase honey bee population by helping beekeepers better manage diseases and pests.
Publications
- Ahn, K., X. Xie, J. Riddle, J. Pettis, Z.Y. Huang. 2012. Effects of Long distance transportation on honey bee physiology. Psyche, doi:10.1155/2012/193029
- Huang, Z.Y. 2012. Pollen nutrition affects honey bee stress resistance. Terrestrial Arthropod Reviews 5: 175-189 (Invited review).
- Huang, Z.Y. 2012. Varroa mite reproductive biology. American Bee Journal 152: 981-985. Also published on Bee Culture.
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Progress 01/01/11 to 12/31/11
Outputs
OUTPUTS: EFFECT OF NOSEMA CERANAE ON HONEY BEES Last summer we discovered that mixed infections of two species of Nosema, N. apis and N. ceranae caused significantly higher mortality in caged bees and that N. ceranae was not more virulent than N. apis. This year we determined whether this is also true in colony settings. We compared single species infection by Nosema apis (50,000 spores/bee) and Nosema ceranae (same dose), and one mixed infection (25,000 spores of each species) and a negative control (no spores). The experiment was repeated three times. In all cases, mixed infected bees lived shorter lives than single species infections and also foraged earlier than them. We will determine the ratio of the two species using real time PCR (rtPCR) during the winter. rtPCR results from last year`s caged bees showed that regardless of the initial proportion (18%, 50%, or 82%) of N. pis, it always out-competed N. ceranae inside individual workers. HUNTING FOR GENES IN VARROA USING RNAi We have shown previously that using our microinjection protocol, mite survival 48 h p.i. was 78.8% (N = 41 mites), and a 77% efficiency gene knock-down efficiency was achieved. In this study we tried to determine the effect on mites of injecting double stranded RNA (dsRNA) of three genes: ribosome protein L8(RPL8), proteasome 26S subunit (Pros26.4) and ribosome protein S13(RPS13), all of which have shown to play roles in survival or reproduction in other tick species. RPL8 affected mite reproduction while Pts26.4 and RPS13 affected mite survival. INTERACTION OF SUBLETHAL LEVELS OF PESTICIDES WITH NOSEMA INFECTION We tested 4 pesticides by exposing bees to them during larval stage and then feeding them Nosema ceranae spores at the adult stage. The pesticides tested included fluvalinate, coumaphos, imidacloprid and clorothalonil. Most pesticides, even though showing no significantly increased mortality at the larval stage (therefore truly exposed at sublethal doses), still showed increased mortality, even when not fed with nosema. Nosem infection in most cases did not cause higher mortality due to prior exposure to pesticides. This is good news in the sense that pesticides did not exasperate the effect of nosema infection. PARTICIPANTS: Undergraduate Research Assistants: Mike Jessup, David Huang. Postdoctoral Fellows, Samina Qamer, Xianbing Xie. Graduate students (MSc):Matthew Lundquist, Julie Adams. Program for Undergraduate Research in the Life-sciences: Austin Dreyer, Justin Merkel, Kevin Baker. High School Students: Henry Pointon, Alice Jiang. TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
Honey bees play a critical role in US agriculture by providing pollination services to the majority of fruits and vegetables. Honey bees are attacked by a plethora of diseases and pests that threaten their health. Beekeepers are gaining more knowledge in pest and disease control as a result of our research and extension efforts. Requests are coming in for the use and instructions of the new technology for varroa mite control, Mitezapper. Our goal is to sustain or increase honey bee population by helping beekeepers better manage diseases and pests.
Publications
- Huang, Z.Y. 2011. Effects of Nosema on Honey Bee Behavior and Physiology. American Bee Journal 151: 871-874. Also published on Bee Culture, 138: 21-24.
- Leniaud, L., E. Darrouzet, K. Ahn, F. Dedeine, Z.Y. Huang, A.-G. Bagneres. 2011. Ontogenic potentialities of the worker caste in two sympatric subterranean termites. Evolution and Development. 13: 138-148 DOI: 10.1111/j.1525-142X.2011.00464.x
- Luo C.W., Z. Y. Huang, X.M. Chen, K. Li, Y. Chen, Y. Y. Sun. 2011. Contribution of diurnal and nocturnal insects to the pollination of Jatropha curcas in Southwest China. J. Economic Entomology. 104: 149-154 DOI: 10.1603/EC10265
- Shi, Y. Y., Z.Y. Huang, Z.J. Zeng, Z.L. Wang, X.B. Wu, W.Y. Yan. 2011. Diet and cell size both affect queen-worker differentiation through DNA methylation in honey bees (Apis mellifera, Apidae). PLoS ONE 6(4): e18808. doi:10.1371/journal.pone.0018808
- Slone, J. D., T.A. Stout, Z.Y. Huang, S.S. Schneider, 2011. Characteristics of honey bee drones that receive vibration signals from workers. Behavioral Ecology and Sociobiology. DOI 10.1007/s00040-011-0195-5
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Progress 01/01/10 to 12/31/10
Outputs
OUTPUTS: EFFECT OF NOSEMA CERANAE ON HONEY BEES We compared single species infection by Nosema apis (30,000 spores/bee) and Nosema ceranae (same dose), and various mixed doses (25:5, 20:10, 15:15, 10:20, and 5:25 ratio, for the two species, each total of 30,000 spores). The results from survival analysis suggest two important things. 1. N. apis and N. ceranae do not affect survival of caged bees differently and 2. Mixed infections are more deadly than single species infections in caged bees. It is so far unclear whether or not there is competition between N. ceranae and N. apis from the survival curves. If one spore species out competed the other species, the survival curves should not have been different amongst the mixed and the single species infections. It is more likely that the two spore species work together and therefore become more deadly. We are waiting for results of quantitative rtPCR to estimate the proportion of each species. TESTING OF VARIOUS PYTHROIDS ON VARROA MITES We tested 5 different pyrethroid insecticides on varroa mites. These were bifenthrin, bioresmethrin, cyfluthrin, fenpropathrin and fluvalinate. We found mites were resistant to fluvalinate (LD 50 of 886 ng), but were more sensitive, or highly sensitive to other chemicals. One chemical had a LD50 of 0.31 ng. This is almost three thousand times more sensitive than to fluvalinate. Understanding the mechanisms of the differential sensitivity will help use develop new mitecides for varroa control. HONEY BEE LEARNING AFFECTED BY INSECTICIDES We studied the learning and memory of honey bees using a "relay" assay. Bees were released 1 km away from their hive and then released at 2 km in the same direction. Bees usually will return at a higher rate from this location than a control group which was released 1 km away in a different direction. This was turned a relay effect because bees were thought to perform reconaince flights trying to find their hive and during this process they also remember the landscape. We used this assay to study subtle effects of pesticide synergism on learning and memory. We used sublethal doses of both fluvalinate (50 ng/bee) and imidacloprid (0.2 ng/bee), and a mixed group (25 fluvalinate + 0.1 ng imidacloprid /bee) to treat the bees before their first release (1 km), returned bees were recaptured and were released at 2 km in the same direction. While bees returned at similar rates in all four groups (3 treatment and an untreated group), the mixed group showed significant reduction for the second return (relay assay). These results suggest that pesticide interaction can affect honey bee learning or memory, almost most likely learning in this assay. These results might shed light to the colony collapse disorder where bees mysteriously disappear from their hives. PARTICIPANTS: Toan van Tran, Visiting Scientist, MSU Entomology Joseph Riddle, Undergraduate Research Assistant, MSU Entomology Mike Jessup, Undergraduate Research Assistant, MSU Entomology R.W.K. Punchihewa, visiting scientist, MSU Entomology Matthew Lundquist, Graduate student (MSc), MSU Entomology Ali Nemeth, MSU High School Honors Science Program Jill Donowich, MSU High School Honors Science Program TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
Honey bees play a critical role in US agriculture by providing pollination services to the majority of fruits and vegetables. Honey bees are attacked by a plethora of diseases and pests that threaten their health. Beekeepers are gaining more knowledge in pest and disease control as a result of our research and extension efforts. Requests are coming in for the use and instructions of the new technology for varroa mite control, Mitezapper. Our goal is to sustain or increase honey bee population by helping beekeepers better manage diseases and pests.
Publications
- Chen, Y.P., Z.Y. Huang. 2010. Nosema ceranae, a newly identified pathogen of Apis mellifera in the U.S. and Asia. Apidologie 41: 364-374 (Invited review)
- Huang, Z.Y., K. Ahn, J. Riddle, J. Pettis. 2010. Effect of transportation on honey bee physiology. Proceedings of the American Bee Research Conference, American Bee Journal 150: 502-503. Abstract
- Huang, Z.Y. 2010. Honey bee nutrition. American Bee Journal 150 (8): 773-776 (also published on Bee Culture 138 (9):22-25, and eXtension.org: http://www.extension.org/pages/Honey_Bee_Nutrition).
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Progress 01/01/09 to 12/31/09
Outputs
OUTPUTS: EFFECT OF MIGRATION ON HONEY BEE PHYSIOLOGY We studied (as a third replicate, with the two replicates from summer of 2008) how migration affected honey bee physiology. Bees were transported around about 800 miles per day for 3 days but returned to E. Lansing each night. Control bees stayed stationary. One group of bees experienced the migration as young bees (4-6 days old) and one cohort as old bees (14-16 day old bees). We then sampled aged marked bees for their juvenile hormone titer, total protein in the head, and size of hypopharyngeal glands. Size of hypopharyngeal glands showed consistent differences between the stationary and migrated bees, in both age groups. Juvenile hormone titers and head proteins did not show consistent differences. ALTERNATIVE CONTROL METHOD FOR VARROA MITE With the hard work from Mr. John Sinanis (Mite Zapper LLC), and more field testing in my lab and by Prof. Tugrul Giray (University of Puerto Rico), the designs for Mitezapper control box and frame were both finalized in 2009. One thousand pieces will be produced and distributed March/April of 2010. EFFECT OF NOSEMA CERANAE ON HONEY BEES We tested the effect of the emerging pathogen Nosema ceranae on honey bee longevity in combination of pollen nutrition. We failed to find an interaction between the absence/presence of pollen and infection/no infection of Nosema. We also compared the virulence of both nosema species and did not find a difference between N. apis and N. ceranae. HONEY BEE ORIENTATION We studied the ontogeny of honey bee orientation in relation to worker age. Workers showed increased return rate at greater distances when they aged. We also studied the "relay landscape learning" in foragers whereby bees released at a site close to a previously released site show increased homing rate compared to the control (a site more than 1 km away form the previous release site). These results have implications for honey bee learning and can lead insights to the causes of Colony Collapse Disorder. PARTICIPANTS: Kiheung Ahn, Visiting Scientist, MSU Entomology Toan van Tran, Visiting Scientist, MSU Entomology Joseph Riddle, Undergraduate Research Assistant, MSU Entomology Melissa Huang, Undergraduate Research Assistant, MSU Entomology Neil Kondamuri, MSU High School Honors Science Program Joanne Shin, MSU High School Honors Science Program John Sinanis, Mite Zapper LLC, Michigan Tugrul Giray, University of Puerto Rico TARGET AUDIENCES: Beekeepers, fruit and vegetable growers. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
Honey bees play a critical role in US agriculture by providing pollination services to the majority of fruits and vegetables. Honey bees are attached by a plethora of diseases and pests that threaten their health. Beekeepers are gaining more knowledge in pest and disease control as a result of our research and extension efforts. Requests are coming in for the use and instructions of the new technology, Mitezapper. Our goal is to sustain or increase honey bee population by helping beekeepers better manage diseases and pests.
Publications
- 1. Johnson, R.M., Z.Y. Huang, M.R. Berenbaum. 2009. Role of detoxification in Varroa destructor (Acari: Parasitidae) tolerance of the miticide tau-fluvalinate. International J Acarology, In press
- 2. Maria Navajas, D.L. Anderson, J. Clement, Z.Y. Huang, T. Zhou, L.I. de Guzman, Y. Le Conte. 2009. New Asian types of Varroa destructor represent new threats for world apiculture. Apidologie, In press
- 3. Wegener, J., Z.Y. Huang, M.W. Lorenz, K. Bienefeld. 2009. Regulation of hypopharyngeal gland activity and oogenesis in honey bee (Apis mellifera) workers. Journal of Insect Physiology 55: 716-25
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Progress 01/01/08 to 12/31/08
Outputs
OUTPUTS: VARROA MITE BIOLOGY Many studies have shown that Varroa destructor prefer nurses, however all studies were performed under laboratory settings. In addition, we do not fully understand why varroa mites prefer nurses. For example, do nurses have different nutrition so that mites reproduce better after feeing on them Or is it simply because nurses provide the best access to brood stages that are ready to be capped so that mites can invade their host easier We studied Varroa destructor host preference in experimental colonies using Apis mellifera. We found that the host preference is nurses > newly emerged bees > foragers, with nurses being the most preferred host. To determine why mites prefer nurses, we conducted a second experiment. We reared varroa mites on the three different types of hosts for three days and introduced them into newly capped (<6 hrs) worker brood. Brood was incubated at 34 C and 50% relative humidity. After 9 days we examined the reproductive status and number of offspring of each mite. We discovered that mites reproduced the best when nurses were used as their phoretic host, followed by foragers and then newly emerged bees. We now finally understand that varroa mites prefer nurses as phoretic hosts because they give them the highest reproductive potential, possibly due to physiological factors in their hemolymph. ALTERNATIVE CONTROL METHOD FOR VARROA MITE: I continue to work closely with Mite Zapper LLC (Detroit, MI) and we designed a new temperature box to lower the cost. We tested various temperature and time combination to obtain the shortest time and highest possible temperature without melting the beeswax. The product should be in the market May/June 2009. USE OF ESSENTIAL OIL FOR NOSEMA CONTROL: We tested three types (thymol, origanum and clove oil, either neat, or in starch encapsulated form). Bees infected with nosema lived significantly shorter than those not infected with nosema, however, treating with essential oils did not improve survival in the host honey bees. The used essential oils did not affect the mortality of bees which were not fed with nosema, or their food consumption. Some essential oils such as thymol, starch-encapsulated origanum oil, and neat origanum oil showed a negative impact on the nosema infected bees. Survivorship of infected bees treated with starch-encapsulated thymol in sugar syrup (26%+-4) and starch-encapsulated origanum oil in pollen patty (38%+-4) were higher compared to the control group (19%+-3) but these differences were not significant. EFFECT OF NOSEMA AND FOOD TYPE ON HONEY BEE LONGEVITY: We repeated for another season whether high fructose corn syrup can lower honey bees immune system, making them more susceptible to diseases and pests. We inoculated workers with Nosema apis and fed them either beet sugar or high fructose corn syrup. While nosema did not have a significant impact on worker longevity, corn syrup did have a significant negative impact on worker longevity. PARTICIPANTS: Zachary Huang, P.I. Yulun Fu, graduate student Kiheung Ahn, visiting scientist Xianbing Xie, visiting scientist Paitoon Seanbualuang, visiting scientist Gorden Wardell, collaborator, SAFE LLC, AZ Joseph Riddle, undergraduate assistant TARGET AUDIENCES: Scientists and students at other universities were reached through invited seminars. Beekeepers and fruit growers (cherry, apple, blueberry etc) and the general public were reached through seminars at state, regional and national extension and outreach meetings. Anyone, anywhere, with a computer could also reach the website cyberbee.msu.edu for information related to honey bees. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
The honey bee plays a critical role in US agriculture by providing pollination to fruits and vegetables. The honey bee is attacked by many diseases and pests that threaten the beekeeping industry world wide. Understanding varroa mite biology will help us find weak links in its behavior or physiology and design better control strategies. Understanding how nosema affect honey bees will also help us come up with better management for this disease.
Publications
- Huang, Z.Y. (2008). Website design and contents for http://cyberbee.msu.edu,over 700 photos, and over 40 pages of information related to honey bees. Over 6 million hits during 2008
- Huang, Z.Y. 2008. Teaching beekeeping in Nepal, Michigan Beekeepers Association Newsletter, July, pp 6-7
- Huang, Z.Y., 2008. Swarm led by a virgin queen, Michigan Beekeepers Association Newsletter, June, pp 4-5
- Huang, Z.Y., W. Pett. 2008. Working with your beekeeper for optimal fruit pollination, Michigan Farm News, April 15, 2008, page 20-21
- Huang, Z.Y., W. Pett. 2008. Working with your beekeeper for optimal fruit pollination, MSU Fruit Crop Advisory Team Alert Newsletter http://www.ipm.msu.edu/cat08fruit/ f03-18-08.htm#1
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Progress 01/01/07 to 12/31/07
Outputs
OUTPUTS: VARROA MITE BIOLOGY (RESISTANCE): In collaboration with Ke Dong, we isolated 30 full-length cDNA clones from a varroa mite field population in Michigan. Only one of these clones, VmNav-1, generates sodium currents when expressed in oocytes. Interestingly, VmNav-1 is highly resistant to fluvalinate and deltamethrin. Compared with the previously published sequence, there are 33 amino acid changes in VmNav-1 including the methionine (M) to isoleucine (I) mutation at amino acid position 1991 in IVS6, which is one of the four resistance-associated mutations previously identified. So far we have examined a total of 28 amino acid changes including the M to I mutation in IVS6 using site-directed mutagenesis and functional assay in oocytes. None of them are responsible for the pyrethroid resistance in VmNav-1. Completion of the analysis of the remaining five amino acid changes should reveal the residue that is responsible for the pyrethroid resistance in VmNav-1. ALTERNATIVE CONTROL
METHOD FOR VARROA MITE: We have studied whether essential oils can be used to regulate varroa mite reproduction. We found little or no effect on mite reproduction of thymol and origanum oil (both oils in neat, starch encapsulated, or beta- cyclodextrin encapsulated forms) and clove oil (neat only). The lowest reproduction rate occurred in 0.0006mg beta-cyclodextrin encapsulated origanum oil, which was only 8% lower than the control (84%). All three oils showed some repellency to varroa mites when brood cells were opened to exam the invasion rate after applying the essential oils. The Mitezapper is being developed by Mite Zapper LLC in Detroit and 50 copies of a working prototype is produced and will be used for beta test early 2008. Effect of Nosema and food type on honey bee longevity We tested the hypothesis that high fructose corn syrup can lower honey bees immune system, making them more susceptible to diseases and pests. We inoculated workers with Nosema apis and fed them either
beet sugar or high fructose corn syrup. While nosema did not have a significant impact on worker longevity, corn syrup did have a significant negative impact on worker longevity.
PARTICIPANTS: Huang, Zachary Dong, Ke Fu, Yulun Ahn, Kiheung
TARGET AUDIENCES: Targe audiences include anyone with a computer who can access internet to access cyberbee.msu.edu). Beekeepers and fruit growers (cherry, apple, blueberry etc) and the general public were also reached through seminars at state, regional and national extension and outreach meetings.
Impacts
The honey bee plays a critical role in US agriculture by providing pollination to most fruits and vegetables. The honey bee is attacked by many diseases and pests that threaten the beekeeping industry world wide. Understanding varroa mite biology will help us find weak links in its behavior or physiology and design better control strategies.
Publications
- Huang, Z.Y. (2007). Honey bee colony collapse disorder and vegetable growers. MSUVegetable IPM Crop Advisory Team Alert Newsletter. http://www.ipm. msu.edu/cat07veg/v04-18-07.htm#2
- Huang, Z.Y. (2007). As a beekeeper you should know about CCD. Midwest Beekeeping, 1(2): 8.
- Duan, J.J, M. Marvier, J. Huesing, G. Dively, Z.Y. Huang. (2008). A meta-analysis of effects of Bt crops on honey bees (Hymenoptera: Apidae). PLOS ONE, 3(1): e1415. doi:10.1371/journal.pone.0001415
- Huang, Z.Y. (2007). Website design and contents for http://cyberbee.msu.edu, over 700 photos, and over 40 pages of information related to honey bees. 5.7 million hits during 2007
- Huang, Z.Y., W. Pett. (2007). "Working with your beekeeper for optimal pollination of fruits and vegetables," Proceedings of Great Lakes Fruit, Vegetable and Farm Market Expo. http://www.glexpo.com/abstracts/2007abstracts/pollination_2007.pdf
- Huang, Z.Y. (2007). Information on colony collapse disorder, cyberbee.msu.edu. http://cyberbee.net/ccd.shtml
- Huang, Z.Y., R. Isaacs. (2007). Using bees for pollination of fruit crops. MSU Fruit Crop Advisory Team Alert Newsletter. http://www.ipm.msu.edu/cat07fruit/f03-27-07.htm#1
- Huang, Z.Y. (2007). The recent honey bee crisis and its implications for Michigan fruit growers. MSU Fruit Crop Advisory Team Alert Newsletter. http://www.ipm.msu.edu/cat07fruit/ f03-27-07.htm#2
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Progress 01/01/06 to 12/31/06
Outputs
VARROA MITE BIOLOGY (RESISTANCE): In collaboration with Ke Dong, we have recently successfully isolated and expressed one varroa mite sodium channel in Xenopus oocytes. However, the expressed sodium channel was extremely resistant to pyrethroids. This was unexpected because the full-length sodium channel clone was isolated from a field population that showed no pyrethroid resistance. Previous studies have shown that due to alternative splicing and/or RNA editing, even pyrethroid-susceptible insects can carry pyrethroid-resistant sodium channel variants. It is possible that this is a naturally occurring resistant variant. Sequence analysis revealed that none of the previously identified four mutations (F758L, L826P, I982V and M1055I) were present in this clone but there are 25 scattered amino acid changes. We are currently determining which amino acid change is responsible for the insensitivity to pyrethroid of this clone. ALTERNATIVE CONTROL METHOD FOR VARROA MITE: We
have conducted another summer of field study of using the Mitezappers for mite control. We saw significant suppression of mite population by using the Mitezapper 4-5 times in each colony throughout the summer. Most of the colonies showed mite population level below economic thread (30 mite drop per day) when treated with the Mitezappers, but untreated colonies had mite populations requiring chemical treatment. Together with Mite Zapper, LLC, we are also working on the development of more efficient Mitezapper prototypes. EPIDEMIOLOGY OF NOSEMA APIS: we conducted a fourth year field experiment to track the nosema spore levels in honey and beeswax and correlate these spore levels with rates of worker infection. This is our last year and we expect to publish a paper with four years of results during 2007.
Impacts
The honey bee plays a critical role in US agriculture by providing pollination to fruits and vegetables. The honey bee is attacked by many diseases and pests that threaten the beekeeping industry world wide. Understanding varroa mite biology will help us find weak links in its behavior or physiology and design better control strategies. Understanding nosema epidemiology also helps us come up with better management for this disease.
Publications
- Liu, Z., J. Tan, Z.Y. Huang, K. Dong. 2006. Effect of a fluvalinate-resistance-associated sodium channel mutation from varroa mites on cockroach sodium channel sensitivity to fluvalinate, a pyrethroid insecticide. Insect Biochemistry and Molecular Biology 36: 885-889.
- Cho, S., Z.Y. Huang, D.R. Green, D.R. Smith, J. Zhang. 2006. Evolution of the complementary sex-determination gene of honey bees: balancing selection and trans-species polymorphisms. Genome Research16: 1366-1375.
- Huang, Z.Y., J. Zhao, L. Zhou, Y. Qin.. 2006. Electronic monitoring of feeding behavior of Varroa mites on honey bees. Journal of Apicultural Research 45(3): 157-158.
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Progress 01/01/05 to 12/31/05
Outputs
VARROA MITE BIOLOGY (RESISTANCE): We have tested the previously identified four mutations (F758L, L826P, I982V and M1055I) using cochroach sodium channels and found one mutation that caused a 10 fold change of resistance to fluvalinate. We also have successfully expressed the mite sodium channel in Xenopus eggs. Pharmacological study of mite sodium channel is finally underway. ALTERNATIVE CONTROL METHOD FOR VARROA MITE: We have conducted a second year field study of using mitezappers for mite control. It appears that to increase efficiency of the mitezapper, we need to use the zappers early in the spring (May/June). EPIDEMIOLOGY OF NOSEMA APIS: we conducted a third year field experiment to track the nosema spore levels in honey and beeswax and correlate these spore levels with rates of worker infection. We are still counting the nosema spores in samples. SWARM BIOLOGY: we studied how colony conditions can affect levels of juvenile hormone, a hormone (JH) that plays
important roles in division of labor. We discovered that crowded conditions that leads to swarming also reduced JH levels, suggesting that workers in pre-swarming colonies have delayed behavioral development, resulting later ages of foraging.
Impacts
Our long term goal is to understand the mode of action of disease pathogens and the effect of pests on honey bees workers, so that the knowledge can be used for better pest and disease management. The mitezapper patent is in negotiation phase with potential buyers. The sodium channel work might lead to new types of pesticides if we have comparative data of differential sodium channel sensitivities of these from bees and mites. Knowledge on Nosema apis will lead to its better control.
Publications
- Zeng, Z., Z.Y. Huang, Y. Qin and H. Pang.. 2005. Hemolymph juvenile hormone titers in worker honey bees under normal and pre-swarming conditions. Journal of Economical Entomology 98: 274-278.
- Tan, J., Z. Liu, R. Wang, Z. Y. Huang, A.C. Chen, M. Gurevitz, K. Dong. 2005. Identification of amino acid residues that are critical for pyrethroid binding to an insect sodium channel. Molecular Pharmacology 67: 513-522.
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Progress 01/01/04 to 12/31/04
Outputs
VARROA MITE BIOLOGY (RESISTANCE): we have developed a PCR assay for mutations that might be associated with varroa mite resistance to pyrethroid insecticides. We also conducted a field survey sampling mites from 6 beekeepers throughout the state. The mite samples are waiting for PCR tests to determine the frequencies of the various mutations. We also tested the feasibility of using an Insect Feeding Monitor to electronically monitor varroa mite feeding behavior. ALTERNATIVE CONTROL METHOD FOR VARROA MITE: We have tested the mitezappers in 2 locations and the efficacy arranged from 28% to 86% (mean=51.5%). We also obtained more data of thermal kinetic data, comparing bees of various physiological status (nurses, foragers, winter bees) bees to if they have different thermal sensitivity. Foragers and winter bees showed no difference in thermal tolerance, but nurses are more tolerant than both foragers and winter bees. We also participated in a field trial to determine
the effectiveness of various semiochemicals in deterring or attracting varroa mites, causing them to fall to a sticky trap on the hive bottom. EPIDEMIOLOGY OF NOSEMA APIS: we repeated the tracking of Nosema spores in beeswax, honey and related these to the rate of infection in honey bees. The highest correlation is found between spore levels in honey and rate of worker infection (r=0.77, n=12 colonies).
Impacts
Our long term goal is to understand the mode of action of disease pathogens and the effect of pests on honey bees workers, so that the knowledge can be used for better pest and disease management. Already one novel idea for varroa mite control has been patented by MSU. Our sodium channel work might lead to new types of pesticides if we have comparative data of differential sodium channel sensitivities of these from bees and mites. Knowledge on Nosema apis will lead to its better control.
Publications
- Huang, Z.Y., Hanley, A.V., Pett, W., Langenberger, M., Duan, J.J. 2004. Field and semi-field evaluation of impacts of transgenic canola pollen on survival and development of worker honey bees, Journal of Economic Entomology 97: 1517-1523
- Zhou, T., Anderson, D.L., Huang, Z.Y., Huang, S. Yao, J., Ken,T., Zhang, Q. 2004. Identification of Varroa mites infesting Apis cerana and Apis mellifera in China. Apidologie 35: 645-654
- Zhou, T., Anderson, D., Huang, Z.Y., Huang, S., Yao, J., Tan, K., Zhang, Q. 2004. Absence of Varroa jacobsoni in mites infesting Apis cerana and Apis mellifera in China. In (Eds E.N. Camaya, C.R. Cervencia) Proceedings of the 7th Asian Apicultural Association Conference and 10th Beenet Symposium and Technofora, pp 187-192
- Huang, Z.Y., Lin, R. 2004. JH titers, biosynthesis and metabolism in honey bee workers infected by a microsporidia parasite, Nosema apis. Eighth International Conference on the Juvenile Hormones, Scientific Program Abstracts, p12, Abstract
- Qin, Y., Huang, Z.Y. 2004. Dynamics of Nosema spores in honey, beeswax and their relationship to worker infection. Proceedings of the American Bee Research Conference, American Bee Journal 144: 406-407, Abstract
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Progress 01/01/03 to 12/31/03
Outputs
VARROA MITE RESISTANCE MECHANISM: We have tested several full-length cDNA clones of the varroa mite sodium channel gene for the functional expression of the varroa mite sodium channel in Xenopus oocyte. However none of the clones showed any current in expressed channels. ALTERNATIVE CONTROL METHOD FOR VARROA MITE: We have determined the thermal death kinetics for both varroa mites and the honey bees. These information are crucial to design a circuit that would kill mites but not harm honey bees. We also determined the thermal capacity of honey bee drone pupae. These data are allowing us to design the third version of the heating circuit that will be used in colonies next spring. EPIDEMIOLOGY OF NOSEMA APIS: we tracked nosema spore loads in honey and beeswax in three groups of colonies. One group had old equipment and regular queens, one old equipment and hygienic queens, and one group of regular queens and new equipment. Samples were taken monthly from May to October.
Our data showed that while new equipment initially showed low spore loads in both wax and honey, their levels later became indistinguishable from the other two treatments. Colonies headed by hygienic queens also did not show lower spore levels in most samplings.
Impacts
Our long term goal is to understand the mode of action of disease pathogens and the effect of pests on honey bees workers, so that the knowledge can be used for better pest and disease management. Already one novel idea for varroa mite control has been patented by MSU. Our sodium channel work might lead to new types of pesticides if we have comparative data of differential sodium channel sensitivities of these from bees and mites. Knowledge on Nosema apis will lead to its better control.
Publications
- Wang, R., Z.Y. Huang, K. Dong. 2003. Molecular characterization of an arachnid sodium channel gene from the varroa mite (Varroa destructor). Insect Biochemistry and Molecular Biology 33: 733-739
- Hanley, A.V., Z.Y. Huang, W. Pett. 2003. Effect of transgenic Bt corn pollen on larval development of honey bee (Apis mellifera L.) and greater wax moth (Galleria mellonella L.) Journal of Apicultural Research 42: 77-81
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Progress 01/01/02 to 12/31/02
Outputs
VARROA MITE RESISTANCE MECHANISM: Because functional expression of arachnid sodium channels has not been established yet, we introduced the varroa mite pyrethroid resistance-associated mutations into the German cockroach sodium channel gene using site-direct mutagenesis. The mutant channels were expressed in Xenopus oocytes and channel sensitivity to three pyrethroids including fluvalinate was examined. Unexpectedly, three of the four fluvalinate resistance-associated mutations, L826P, I982V and M1055I did not alter the cockroach sodium channel sensitivity to pyrethroids, suggesting that these mutations may not be involved in pyrethroid resistance. However, it is equally possible that effects of these mutations can only be detected in the native varroa mite sodium channel because of the low (less than 50%) sequence identity between the cockroach and varroa mite sodium channel proteins. The cockroach sodium channel backbone sequence may not allow the varroa mite
mutations to exert effects on channel interaction with pyrethroids. This is further supported by the observation that, when the fourth mutation, F758L, was introduced into the cockroach sodium channel, no sodium current was detected. This mutation may have altered the folding, assembly in the membrane, or the ion selectivity of the cockroach sodium channel. We therefore could not use the cockroach sodium channel to assess the effect of the fourth mutation on cockroach sodium channel sensitivity to pyrethroids. Establishment of a mite sodium channel expression system is therefore crucial for further progress of this project. Toward this goal we have isolated several full-length cDNA clones of the varroa mite sodium channel gene for the functional expression of the varroa mite sodium channel in Xenopus oocyte. ALTERNATIVE CONTROL METHOD FOR VARROA MITE: We have been working with Dr. Tom Mase in Engineering to develop a plastic foundation with wires embedded, we are making progress but
the prototype was not ready during summer of 2002. We therefore hand-wired 50 drone frames with wax foundation. We tried to test the best time for killing mites using these wax-foundation prototype mitezappers. While the wax foundation alone (without combs built or pupae inside) melted in about 4-8 seconds, we discovered that with drone pupae in every cell, the same portable battery did not have enough power to heat the comb even after 5 minutes. A regular car battery, on the other hand, burned all the wires immediately, without any harm to the bees or mites. Clearly we will need some current regulation mechanisms for the wires. MODE OF ACTION OF NOSEMA APIS: We tested the hypothesis that Nosema apis might reproduce differently in different behavioral castes of worker honey bees. Nosema spores were fed individually to newly emerged workers, 8- day old nurses or foragers of unknown ages. Bees were then introduced into a nursery colony for 8 days before the total number of nosema spores
were determined in their mid-guts. In three separate trials (9 different colonies) we did not find any significant differences in nosema spores among workers of different behavioral castes.
Impacts
Our long term goal is to understand the mode of action of disease pathogens and the effect of pests on honey bees workers, so that the knowledge can be used for better pest and disease management. Already one novel idea for varroa mite control has been patented by MSU. Our sodium channel work might lead to new types of pesticides if we have comparative data of differential sodium channel sensitivities of these from bees and mites. Knowledge on Nosema apis will provide new information for its management and control.
Publications
- Wang, R., K. Dong, Z.Y. Huang, P.J. Elzen & J. Pettis. 2002. Resistance Mechanism of Varroa jacobsoni to Fluvalinate: Altered Sodium Channel? In (eds: E.H. Erickson, R.E. Page Jr, and A.A. Hanna) Proceedings of the 2nd International Conference on Africanized Honey Bees and Bee Mites. pp 173-176
- Wang R., Liu Z., Dong K., Elzen P. J., Pettis J., Huang Z. Y. (2002) Novel mutations in a sodium channel gene are associated with fluvalinate resistance in the varroa mite, Varroa destroctor. J. Apicultural Research. 40:17-25.
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Progress 01/01/01 to 12/31/01
Outputs
VARROA MITE RESISTANCE MECHANISM: To investigate the molecular mechanism of resistance to fluvalinate in varroa mites, we have cloned and sequenced a large cDNA fragment corresponding to segment 3 of domain II (IIS3) to segment 6 of domain IV (IVS6) of a para-homologous sodium channel gene (VmNa) from susceptible and resistant varroa mite populations. The deduced amino acid sequence from this cDNA shares 71%, 60%, and 50% identity with the corresponding region of the Para-homologous protein of the Southern cattle tick Boophilus microplus, Drosophila melanogaster Para, and rat brain type II sodium channel a-subunit, respectively. Sequence analysis revealed that four amino acid changes, F758L in IIIS6, P826L in the linker connecting domains III and IV, I982V in IVS5 and M1055I in IV6, were correlated with fluvalinate resistance in both Florida and Michigan populations. Interestingly, the kdr or super-kdr (which confers much higher level resistance than kdr) mutations
previously identified in insects was not detected in the resistant varroa mites. These results support the emerging notion that distinct sodium channel gene mutations are selected in different insects and arachnids in response to pyrethroid selection. ALTERNATIVE CONTROL METHOD FOR VARROA MITE: Last year I conceived a non-chemical method for mite control. The Mitezapper uses electricity embedded in drone combs to generate heat and kill the mites. This year we developed a much better prototype of Mitezapper and now it can heat up in 5-8 seconds instead of the 5-8 minutes in last year's model. We will conduct a field test this year to evaluate the usefulness of this method for controlling mites in 60 colonies. MODE OF ACTION OF NOSEMA APIS: We conducted experiments to determine whether juvenile hormone (JH) is involved in the earlier foraging of nosema-infected workers. Newly emerged workers from a naturally mated queen, or from queens artificially-inseminated with semen from single
drones were fed with 450,000 nosema spores per bee in 1ul sugar syrup, and introduced into normal colonies. Rates of JH biosynthesis in infected workers and control bees were compared when bees were 4-14 days old to determine if infected workers produce JH at higher rates than control bees before foraging started. Infected workers had almost 2 times the rates of JH biosynthesis compared to control bees when bees were 6-8 days old in 4 out of 4 colonies. We studied the possibility that JH or JH like chemical(s) were produced by Nosema to affect age of first foraging in workers. We performed surgery (allatectomy) to remove the corpora allata, the only source that produce JH. Allatectomized bees were fed with Nosema spores or sugar syrup only as control. Both groups of bees were introduced into a colony. Behavioral observations for the onset of foraging were made for the two groups of bees. In two out of two colonies, workers without corpora allata foraged the same time. These results
suggest that nosema infection causes earlier foraging in workers, probably through chemicals stimulates host corpora allata to produce more JH. Without host corpora allata, the behavioral differences disappear.
Impacts
Our long term goal is to understand the mode of action of disease pathogens and the effect of pests on honey bees workers, so that the knowledge can be used for better pest and disease management. Our sodium channel work might lead to new types of pesticides if we have comparative data of differential sodium channel sensitivities of these from bees and mites. The non-chemical method for mite control has received much media attention and may prove to be highly effective in preventing the damages caused by mites to bees.
Publications
- 1. Huang, Z. Y. & R. Wang. 1999. Nosema disease and division of labor in honey bees. Proceedings of XXXVI Congress of Apimondia, Vancouver, Canada, p 102, Abstract
- 2. Zhou, T., J. Yao, S.X. Huang & Z.Y. Huang. 2001. Larger cell size reduces varroa mite reproduction. Proceedings of the American Bee Research Conference, American Bee Journal 141: 895-896
- 3. Huang, Z.Y. 2001. Mitezapper: a new device for varroa mite control. Proceedings of Apimondia Congress,Durban, South Africa, p. 54
- 4. Huang, Z.Y. & H. Lin. 2001. The small hive beetle - a new pest of honey bees in the United States. Newsletter of the Michigan Entomological Society 46: 5, 8
- 5. Huang, Z.Y. 2001. MiteZapper - a new and effective method for varroa mite control. American Bee Journal. 141: 730-732
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Progress 01/01/00 to 12/31/00
Outputs
Objective 1 & 2. We conducted an extensive survey for mite resistance against Apistan throughout the state. We tested a total of 451 colonies for mite prevalence and among these 165 were further tested for mite resistance using a field assay. Mites showed an average mortality of 95% when exposed to fluvalinate, indicating no resistance to fluvalinate in mites from these sampled colonies. Objective 3. To determine if sodium channel mutations are responsible for fluvalinate resistance, we cloned and sequenced a full-length sodium channel a subunit cDNA (VMSC) from the Michigan mite population. The deduced protein sequence of VMSC has one of the smallest coding regions of known sodium channel with four domains, having a coding region of 2284 amino acids, and a calculated molecular mass of 256.67 Kd. Comparison of the deduced amino acid sequence obtained from five (2 susceptible, 3 resistant) mite populations revealed 7 amino acid differences. However the usual kdr or
super-kdr mutation was not detected in any of the three resistant mite populations. Objective 4. We transferred mites from European bees to Asian worker brood and found that the mites reproduced on worker brood just as much as drone brood. We could not find any mites on Asian bees, after surveying more than 40 colonie. We also found that mites do not reproduce on worker brood as well, if the workers are reared in large drone cells (by accident we found that queens would lay worker eggs in drone cells in early fall). This "cell-size" effect is very interesting because it reduced the mite reproduction . Objective 5. We finally selected colonies that would respond to nosema infection by becoming foragers early. After selection, 100% of colonies would respond to nosema infection, compared to 50% prior to selection. This suggests there is a genetic variation in workers responding to nosema infections. Intriguingly, we also found a significantly higher "missing" rate in nosema-infected bees
-- these bees did not die inside the colony, but seems to have disappeared after one or more trips of foraging. Measurement of juvenile hormone biosynthesis indicated that bees infected with nosema have a higher rate of hormone production. However, bees with their hormonal sources removed (corpora allata removal surgery) still responded to nosema infection by foraging and "disappearing" earlier. These results suggests that nosema may affect worker bee behavior via many different pathways. Objective 6. A survey was conducted July to August, this was done together with the survey for varroa mite resistance. We contacted 15 beekeepers who move bees to other states and inspected 11 of these. We concentrated our first year effort on the migratory beekeepers because it is more likely that they bring in beetles from the southern states. Indeed, we found beetles in 2 of the checked colonies of a beekeeper. A separate beekeeper, who purchased his package bees from Georgia, also
found a few beetles and larvae in the pollen trap of one of his colonies. Samples were sent in and beetles were positively identified.
Impacts
(N/A)
Publications
- Wang, R. D. Ke, P.J. Elzen, J. Pettis, Z.Y. Huang. 2000. Altered sodium channel as a possible resistance mechanism of Varroa jacobsoni to fluvalinate. American Bee Journal. 140: 474. Abstract.
- Schmidt-Bailey, J., D. Westervelt, Z.Y. Huang. 2000. The Spartan Beetle Buster: testing of a new method for surveying the small hive beetle. American Bee Journal. 140: 907. Abstract.
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