Progress 02/28/13 to 02/27/14
Outputs Target Audience: The target audience for this research includes academic researchers in the fields of entomology and toxicology as well as beekeepers and others in the agricultural sector interested in managed pollination. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided? This project has contributed to the training of one postdoctoral student (Wenfu Mao), two graduate students (Catherine Dana, Ling-Hsiu Liao) and several undergraduates who assisted in gathering and compiling data. How have the results been disseminated to communities of interest? Scientific findings were disseminated in refereed journals aimed at the research community. In terms of public audiences, activities included: June 12, 2013, interview on honey chemistry for digital magazine The Column, LCGC; June 14, 2013, interview for digital magazine Vetstreet.com on bee health; “Applied Bee-nomics,” Congressional Hill Lunch-n-Learn seminar, June 16, 2013; Stakeholder meeting, “Open dialogue—research priorities, translating research from hive to honey and other products”, American Farm Bureau, CropLife America, June 16, 2013; Kathleen Dunn call-in show, “Honeybee health,” Wisconsin Public Radio, June 19, 2013; "Pollination fascination—pollinator genomics”, UI Pollinatarium, June 30, 2013; New technologies for U.S. apiculture, Suzanne Zionts and Stephanie Becker, Al Jazeera America, August 5, 2013; Alexandra Sifferlin, Time.com, entomophagy and food security query, August 20, 2013; Neonicotinoids and pest control operators, Ann Nagro, Pest Control Technology magazine, August 26, 2013; Interview and podcast with Adrian Smith, aodpod.com, Origins, September 4, 2013; Interview with Seth Shostak, “Disappearing bees,” Big Picture Science radio and podcast, September 4, 2013; Interview, “Is there a bee crisis?” Richard Wills, Pittsburg Tribune-Review, September 13, 2013; George S. Saul Public Lecture, Middlebury College, Middlebury VT, September 19, 2013;“How the honey bee genome project is revolutionizing pollinator research” AgMasters Conference, Champaign, IL December 2, 2013; Interview with Joel Brownstein, LiveScience, on tobacco hornworm defense; interview with Susan Milius, Science News, on tobacco hornworm defense, December 30, 2013; Interview with Paul Wood, Champaign-Urbana News Gazette, powder-post beetles in Spurlock, “Exhibiting an intolerance for pests,” January 18, 2014; Interview with Voice of Russia, Bees and Pesticides, London, UK, January 24, 2014; Interview with Seth Borenstein, Associated Press, “Zom-bees”, January 29, 2014; Pesticides, WILL AM Focus 580, February 18, 2014; Interview with Seth Borenstein, Associated Press, disease associations between honey bees and bumble bees, February 19, 2014. George S. Saul Lecture, Department of Biology, Middlebury College, Middlebury VT, September 20, 2013; “Predicting the future of entomology—lessons from the past,” First Annual Richards-Hodson Lecture, Department of Entomology, University of Minnesota, St. Paul, MN, October 24, 2013; “Bees and pesticides and gypsy moths,” Annual Gypsy Moth Review, Evanston, IL, November 5, 2013; Entomological Society of America Annual Meeting, Austin, TX: “Does the honey bee risk cup runneth over?” P-IE Section Symposium: Beyond the LC50: Advancements in Toxicological Research on Pollinators, November 10, 2013; “Impacts of landscape-scale conversion to bioenergy crops on microlepidopteran diversity: Biofuels and LBMs”, P-IE Section Symposium: Biofuel Cropping Systems: Connecting Beneficial Arthropods, Ecosystem Services, and Landscape Effects, November 12, 2013; “Farming out pharmacology: Symbiont detoxification of phytochemicals",P-IE Section Symposium: The Effect of Microbes on Insect-Plant Interactions, November 12, 2013; You are what you eat: honey and the honey bee", P-IE Section Symposium: Connecting Research, Outreach and Regulatory efforts to Protect Honey Bee Health, November 12, 2013. “Honeybee cytochrome P450s: how a 6-million-year-old genome copes with pesticide-intensive modern agriculture” at “Impact of Pesticides on Bee Health” meeting, Society of Experimental Biology/Biochemical Society/British Ecological Society, London, UK, January 22-24, 2014; Friday Seminar Programme, John Innes Plant Research Center, Norwich, UK, March 24, 2014; Rochester Community and Technical College Environmental Science Program, Bees in Crisis: Colony Collapse, Honey Laundering and Other Problems Bee-Setting American Apiculture, Rochester, MN, April 8, 2014; Storer Lectures, Department of Entomology, University of California at Davis, Davis, CA, May 20-21, 2014; Back Yard Beekeepers, Weston CT May 27, 2014. What do you plan to do during the next reporting period to accomplish the goals? Upcoming presentations are scheduled with beekeeping groups in Maryland and Illinois during summer 2014 and scientific presentations will be made at the November 2014 Entomological Society of America meeting.
Impacts What was accomplished under these goals?
Our specific objectives and our progress to date are: [1] To identify inducers and inhibitors of cytochrome P450 detoxification gene transcription in naturally occurring (nectar, pollen) and processed (honey, beebread) components of the honey bee diet. With respect to this objective, we identified four constituents of honey (p-coumaric acid, pinocembrin, pinobanksin and pinobanksin 5-methyl ether) that upregulate detoxification and immunity genes in both larval and adult bees; [2] To determine how in-hive and agricultural pesticides induce or inhibit P450 detoxification. With respect to this objective we determined that CYP9Q enzymes are involved in detoxification of acaricides and that tau-fluvalinate and coumaphos compete for access to the catalytic site and synergize each other; [3] To identify specific P450 enzymes that metabolize pesticides and phytochemicals. With respect to this objective we identified CYP6AS genes as encoding enzymes that metabolize flavonoids in honey and beebread/pollen and CYP9Q genes that encode enzymes that metabolize naturally occurring flavonoids as well as in-hive and agricultural pesticides; and [4] To test whether diet (honey, sucrose, high fructose corn syrup) influences transcription of P450 genes and toxin tolerance. With respect to this objective, we found that induction by p-coumaric acid enhances pesticide detoxification and tolerance and that consumption of honey enhances tolerance of aflatoxin B1 relative to consumption of high fructose corn syrup.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Schuler, M.A. and M.R. Berenbaum. Structure and function of cytochrome P450s in insect adaptation to natural and synthetic toxins: Insights gained from molecular modeling. Journal of Chemical Ecology 39: 1232-1245.
- Type:
Journal Articles
Status:
Awaiting Publication
Year Published:
2014
Citation:
Berenbaum, M.R. Bees in crisis: Colony collapse, honey laundering, and other problems bee-setting American apiculture. Proc. Am. Phil. Soc.
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Mao, W., M.A. Schuler and M.R. Berenbaum. Honey constituents upregulate detoxification and immunity genes in the western honey bee Apis mellifera. Proceedings of the National Academy of Sciences 110: 8842-8846.
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Progress 03/01/11 to 02/28/12
Outputs OUTPUTS: Objectives of this project (and experimental approaches) are: 1. Identify inducers and inhibitors of P450 gene transcription in natural substances (nectar, pollen) and hive products (honey, beebread, propolis) using quantitative real-time PCR followed by GC-MS/LC-MS to identify specific inducers/inhibitors. 2. Identify in-hive and agricultural pesticides that induce or inhibit P450 gene transcription (using qPCR). 3. Determine substrates for enzymes encoded by upregulated genes using baculovirus-mediated expression and molecular modeling. 4. Test whether inducers from hive products incorporated can hence enhances toxin tolerance using pesticide LD50 bioassays with and without inducers. Dissemination of these findings has taken place via radio interviews and personal presentations, including: "Honey and health," Illinois State Beekeepers Association, Champaign, IL June 25, 2011; WILL AM 580 Afternoon Magazine, Pollinatarium programs, August 5, 2011; UI Pollinatarium, Colony Collapse Disorder update, August 7, 2011; WILL AM 580 Focus 580, Bees in the news; ABC News bee stings August 26 2011; Chicago Tribune CCD September 1, 2011; Big Ten Housing Conference, "Honey and housing," Urbana, IL, October 25, 2011; Honey chemistry, Peggy Notebaert Museum, October 29, 2011; "History of honey," Garfield Park Conservatory Beekeeping Workshop, Garfield Park Conservatory, Chicago, IL, November 19, 2011; Fourth Dimension Fourth Presbyterian Church, Chicago, IL January 13, 2012; Southern Illinois Audubon Society, Carbondale, IL, January 27, 2012; interview for Naomi Klein January 31, 2012; "Beespotter," MSTE Lunch, Champaign, IL, February 17, 2012; "Bees and pesticides," Missouri State Beekeepers Association, St. Louis, MO, March 9, 2012; Bees and pesticides, UIUC Beekeeping Short Course, Urbana, IL, March 31, 2012; and "Honey chemistry," Backyard Beekeepers Association, Weston, CT, May 29, 2012. PARTICIPANTS: Wenfu Mao is a postdoctoral associate on this project learning and applying methods of molecular genetic analysis to bee toxicology. Catherine Dana is a graduate student learning how to conduct bioassays of behavioral responses of honey bees to pesticides. Katherine Noble is a graduate student learning how to conduct bioassays of behavioral responses of honey bees to p-coumaric acid and other constituents of honey, beebread and propolis. Ling Hsiu Liao is a graduate student studying how honey processing changes the phytochemistry of honey and how esterase activity contributes to xenobiotic detoxification in honey bees. Mark Demkovich is a graduate student learning how to express honey bee P450s in a heterologous system in order to assay possible synergistic interactions among pesticides. Brendan Colon is an undergraduate who is learning how to model molecular structure of xenobiotic-metabolizing cytochrome P450s in honey bees to determine mechanisms of synergistic interactions. TARGET AUDIENCES: Target audiences include insect toxicologists, physiologists, and sociobiologists (at meetings of professional societies), beekeepers and others in the apicultural community (through meetings of beekeeping societies), and the general public (through a variety of public meetings, presentations, and the like) PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts We used several approaches to ascertain effects of dietary toxins on bee susceptibility to synthetic and natural xenobiotics, including the miticides tau-fluvalinate and coumaphos, the agricultural pesticide imidacloprid, and the naturally occurring mycotoxin aflatoxin. We administered potential inducers of cytochrome P450 enzymes to investigate how detoxification is regulated. The drug phenobarbital induces P450s in many insects, yet in tau-fluvalinate bioassays no induction occurred in bees fed phenobarbital. Similarly, no P450 induction, as measured by tau-fluvalinate tolerance, occurred in bees fed xanthotoxin, salicylic acid, or indole-3-carbinol, all of which induce P450s in other insects. Only quercetin, a common pollen and honey constituent, reduced tau-fluvalinate toxicity. Specialty microarrays containing all bee P450s revealed no change in detoxicative gene expression in guts of phenobarbital-treated bees. However, Northern blot analyses of guts of bees fed extracts of honey, pollen and propolis showed elevated expression of three CYP6AS P450s. In bioassays, diet did not influence tau-fluvalinate or imidacloprid toxicity but aflatoxin toxicity was higher in bees consuming sucrose or high-fructose corn syrup than in bees consuming honey. These results suggest that regulation of honey bee P450s is tuned to chemicals occurring naturally in the hive environment and that, in terms of toxicological capacity, a diet of sugar is not equivalent to a diet of honey. We also used high pressure liquid chromatography, MS-MS, and bioassays to identify several honey constituents that act as inducers of detoxification gene transcription. These include pinobanksin, pinobanksin 5 methyl ether, pinocembrin, and p-coumaric acid; whereas the first three likely arise from propolis, p-coumaric acid is a universal constituent of pollen and is likely present in all honey and bee bread. Via RNAseq analysis, we demonstrated that p-coumaric acid specifically upregulates detoxification genes and select antimicrobial genes. As a major component of pollen grains, p-coumaric acid is ubiquitous in the natural diet of honeybees and may function as a nutraceutical regulating immune and detoxification processes. This finding confirms that the widespread apicultural use of honey substitutes, including high fructose corn syrup, may compromise the ability of honeybees to resist pesticides and pathogens and contribute to colony losses. As well, this finding raises uncertainties as to the applicability of previous studies of pesticide toxicity which were conducted with sucrose, rather than honey, as the delivery medium 3. Studies of behavioral responses to p-coumaric acid in addition to bioassays of p-coumaric acid impacts on pesticide toxicity 3. Additional bioassays of herbicides, insecticides and fungicides found in wax comb, bee bodies, or beebread are being conducted to determine whether these compounds interact synergistically. In addition to bioassays, molecular models are under construction to determine if these compounds interfere with each other in the catalytic site of the principal detoxificative enzymes (CYP9Q).
Publications
- Niu, G., Johnson, R.M. and Berenbaum, M.R. 2011. Toxicity of mycotoxins to honeybees and its amelioration by propolis. Apidologie: 10.1051/apido/2010039
- Johnson, R.M., Mao, W., Pollock, H.S., Niu, G., Schuler, M.A. and Berenbaum, M.R. 2012. Ecologically appropriate xenobiotics induce cytochrome P450s in Apis mellifera. PLoS ONE 7(2): e31051. doi:10.1371/journal.pone.0031051.
- Mao, W., Schuler, M.A. and Berenbaum, M.R. 2012. Honey bees rely on honey constituents to upregulate detoxification and immunity genes. For submission to Science.
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