Progress 08/15/07 to 04/14/12

Outputs
OUTPUTS: We have completed this research on assessing the status, population genetic structure and pathogen prevalence of target bumble bee species in the U.S. This research led to worldwide interest in the status of bumble bees in the U.S. and is the only national study of its kind. Throughout the project we gave presentations at national meetings and symposia on our conclusions from the research. We completed all of the major goals (and more) of our proposed research and disseminated the results in top journals (6 reviewed journals, one international report, multiple abstract proceedings), including PNAS (Cameron et al. 2011, 100 citations in the last year), Molecular Ecology and Journal of Insect Pathology. We completed an 80,000+ specimen database of museum records assembled and used in our study to determine historical distributions of target bumble bee species across the U.S. The database is available on GBIF and on the PI's personal website. We developed a survey protocol that will be used in subsequent studies on the status of bumble bees. We developed a guide to the bumble bees of Illinois and Missouri as a teaching tool for citizen scientists and for visitors to the St. Louis Zoo. We helped to develop the website BeeSpotter, a citizen-scientist reporting site for spottings of bumble bees through Illinois. We trained a postdoc who went on to a faculty position at the Univ. of Alabama and continues to study the population genetics of bumble bees in the U.S., we trained two graduate students who received their masters from research on the project, and multiple undergraduates in the methods of the research. We participated in outreach throughout, including presentations about the importance of wild pollinators at National Pollinator Week, to Master Naturalists, Master Gardeners and farm extension groups. PARTICIPANTS: Dr. Jeffrey Lozier, a postdoc in the PI's lab, recruited specifically to work on this research, completed his tenure in the Cameron lab and accepted a faculty position at the University of Alabama, beginning Fall 2011. Dr. Lozier will continue working on bumble bee conservation genetics as part of his career trajectory, a direction he developed after coming to the PI's lab. Thus he represents a new generation of bumble bee specialist, bringing valuable skills in population genetics with which to address pollinator decline in the U.S. Co-PI Strange has trained a graduate student, Jonathan Koch, in bumble bee decline research, including databasing, ecological modeling, rearing colonies for experimental studies, and population genetics. Koch, who completed his masters in 2010, is currently completing research for his PhD thesis, which focuses on species boundaries and changing distributions of certain western bumble bee species. Co-PI Solter trained a masters student, Nils Cordes in bumble bee pathogen screening. PI Cameron and co-PI Strange trained more than a dozen undergraduate student and three high school students assistants during the project, all of whom contributed to the monitoring and databasing of target bumble bee species in the eastern and western U.S. Co-PI Griswold helped to organize the database of the western target species addressed in the project. TARGET AUDIENCES: The primary target audience is the international community of scientists and conservation groups interested in global decline of wild pollinators. This research has succeeded in providing a framework for understanding the severity of bumble bee decline in the U.S. and elucidated two potential causal factors (high Nosema bombi prevalence and low genetic diversity) of the decline. This study is cited whenever comparative statements about global decline are made. The study led to an international meeting to consider issues and policy associated with bumble bee decline worldwide, and led to the formation of the IUCN Bumble Bee Specialist Group. PI Cameron is the scientific advisor to this group and chairs a special committee on bumble bee conservation genetics, which includes Jeffrey Lozier, a principal contributor to the NIFA study. The conservation genetics committee has completed a Joint Statement (http://www.iucn.org/about/work/programmes/ species/who_we_are/ssc_specialist_groups_and_red_list_authorities_dir ectory/ invertebrates/bumblebee_specialist_group/commercial_bumblebee_policy_ statement/) concerning the views on commercial movement of bumble bee colonies worldwide; this statement can be used for policy decisions internationally. PI Cameron, co-PI Strange, postdoc Lozier and PhD student Koch have participated in numerous outreach and educational activities with Master Naturalists, local farmers and extension personnel (Utah). The goals of these activities are to inform both the agricultural and local public of the seriousness of pollinator decline and to provide them with tools that will allow them to be part of the solution rather than the problem. One of PI Cameron's graduate students (Isaac Stewart), who made a career shift to teaching high school biology (and in the process was awarded a national Teaching Fellowship in Biology from the prestigious Knowles Foundation), has transferred his knowledge and skills in pollinator research to the classroom, including working with his students to create a native prairie site near his school for attracting bumble bees and other wild pollinators. This site assists in teaching students the importance of maintaining native plant and pollinator biodiversity and in the importance of native bees for plant pollination. PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
The results of our study generated a groundswell of interest by the media (radio, TV, internet blogs and articles), which reported our results worldwide. This has led to an awareness of the possible causes of bumble bee decline in the U.S. and increased interest in and research on bumble bee pathogens, particularly the fungal pathogen Nosema bombi. We achieved insights into bumble bee decline in the U.S. by developing a historical museum record database, which we used for ecological niche modeling of each target species across its historical range. This database has already been used in multiple studies by other research groups in the U.S. and Canada, and to publications by those groups. Our research activities overall led to the first nationwide analysis and subsequent conclusion that multiple species of bumble bees have significantly declined in range and abundance, the impact of which stimulated the first international meeting to address bumble bee decline in North America and potential mitigating policies and procedures. This meeting included all the stakeholders, including government agencies, the commercial bumble bee industry, NGOs and academic scientists. This resulted in organization of an international Bumble Bee Specialist Group, under the umbrella of the IUCN. The results of our research supported the local and regional observations of several previous studies that had suggested bumble bee populations were declining. Our research also revealed two significant correlations associated with declining populations: 1) higher prevalences of the microsporidian pathogen, Nosema bombi, and 2) lower levels of genetic diversity. These observations led to an ongoing USDA-funded project (PI Cameron, coPI's J.D. Lozier and R. Thorp) to test the hypothesis that Nosema bombi found in U.S. bumble bees is an invasive European strain. The project also helped to support the development of a field guide to the bumble bees of Illinois and Missouri, which has been disseminated to more than 1,000 individuals at bumble bee workshops and the St. Louis Zoo.

Publications

• Lozier, J. D. and Cameron, S. A. 2009. Comparative genetic analyses of historical and contemporary collections highlight contrasting demographic histories for the bumble bees Bombus pensylvanicus and B. impatiens in Illinois. Molecular Ecology 18: 1875-1886.
• Grixti, J. C., Wong, L. T. Cameron, S. A. and Favret. C. 2009. Decline of bumble bees (Bombus) in the North American Midwest. Biological Conservation 142: 75-84.
• Cordes, N., Huang, W.-F., Strange, J. P., Cameron, S. A., Lozier, J. D. and Solter, L. F. 2012. Interspecific geographic distribution and variation of the pathogens Nosema bombi and Crithidia species in United States bumble bee populations. Journal of Invertebrate Pathology 109:209-216.

Progress 08/15/10 to 08/14/11

Outputs
OUTPUTS: During the last year (08/15/2010-08/14/2011) we completed the analyses and wrote multiple papers (see pubs) and gave multiple presentations on the overall results of the research project. We have completed all of the major goals of our proposed research and disseminated the results in top journals, including PNAS and Molecular Ecology. We organized a conference on bumble bee decline and disseminated the results in a large report. We gave presentations at national meetings, and conducted outreach activities, including National Pollinator Week 2011. We completed our 78,000 specimen database of museum records, used in our study to determine historical distributions of target bumble bee species across the U.S. PARTICIPANTS: Dr. Jeffrey Lozier, a postdoc in the the PI's lab recruited specifically to work on this research, completed his tenure in the Cameron lab and accepted a faculty position at the University of Alabama, beginning Fall 2011. Dr. Lozier will continue working on bumble bee conservation genetics as part of his career trajectory, a direction he developed after coming to the PI's lab. Thus he represents a new generation of bumble bee specialist, bringing valuable skills in population genetics with which to address pollinator decline in the U.S. Co-PI Strange has trained a graduate student, Jonathan Koch, in bumble bee decline research, including databasing, ecological modeling, rearing colonies for experimental studies, and population genetics. Koch, who completed his masters in 2010, is currently completing research for his PhD thesis, which focuses on species boundaries and changing distributions of certain western bumble bee species. PI Cameron and co-PI Strange continued training student field assistants during the summer of 2010, including undergraduates and two high school students, who contributed to ongoing monitoring of target bumble bee species in the eastern and western U.S. TARGET AUDIENCES: PI Cameron, co-PI Strange, postdoc Lozier and PhD student Koch have participated in numerous outreach and educational activities with Master Naturalists, local farmers and Extension personnel (Utah). The goals of these activities are to inform both the agricultural and local public of the seriousness of pollinator decline and to provide them with tools that will allow them to be part of the solution rather than the problem. One of PI Cameron's graduate students (Isaac Stewart), who made a career shift to teaching high school biology (and in the process was awarded a national prestigious fellowship, the Knowles Foundation Teaching Fellowship in Biology), has transferred his knowledge and skills in pollinator research to the classroom, including working with his students to create a native prairie site near his school for attracting bumble bees and other pollinators. This site assists in teaching students the importance of maintaining native plant and pollinator biodiversity and in the importance of native bees for plant pollination. PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
The results of our study generated a groundswell of interest by the media (radio, TV, internet blogs and articles), which reported our results worldwide. We achieved insights into bumble bee decline in the U.S. through our development of a historical database, which we used for ecological niche modeling of each target species across its historical range. Our research activities overall led to the first nationwide analysis and subsequent conclusion that multiple species of bumble bees have significantly declined in range and abundance. The results supported the local and regional observations of several previous studies suggesting bumble bee populations were declining. Our research also revealed two significant correlations associated with declining populations: 1) higher prevalences of the microsporidian pathogen, Nosema bombi, and 2) lower levels of genetic diversity. These observations led to an ongoing USDA-funded project (PI Cameron, coPI's J.D. Lozier and R. Thorp) to test the hypothesis that Nosema bombi found in U.S. bumble bees is an invasive European strain.

Publications

• Cameron, S.A., Lozier, J.D., Strange, J.P., Koch, J.B., Cordes, N., Solter, L.F. and Griswold, T.L. 2011. Patterns of widespread decline in North American bumble bees. Proc. Natl. Acad. Sci. USA 108: 662-667.
• Nature News and Views; Brown, M.J F. 2011. The trouble with bumblebees. Nature 469:169-170.
• Lozier, J.D., Strange, J.P., Stewart, I. and Cameron, S.A. 2011. Patterns of range-wide genetic variation in six North American bumble bee (Apidae: Bombus) species. Molecular Ecology 20:4870-4888.
• Cordes, N., Huang, W.-F., Strange, J.P., Cameron, S.A., Lozier, J.D. and Solter, L.F. 2011. Multi-species host distributions of the bumble bee pathogens, Nosema bombi and Crithidia bombi, in United States Bombus populations. Journal of Invertebrate Pathology (In Press).
• Kissinger, C.N., Cameron, S.A., Thorp, R.W., White, B. and Solter, L.F. 2011. Survey of bumble bee (Bombus) pathogens and parasites in Illinois and selected areas of northern California and southern Oregon. Journal of Invertebrate Pathology 107:220-224.
• Cameron, S., Jepsen, S., Spevak, E., Strange, J., Vaughan, M., Engler, J. and Byers, O. (eds.). CBSG. 2011. North American Bumble Bee Species Conservation Planning Workshop Final Report. IUCN/SSC Conservation Breeding Specialist Group: Apple Valley, MN. An electronic version of this report can be downloaded at www.cbsg.org.
• Cameron, S.A., Lozier, J.D., Strange, J., Koch, J., Cordes, N., Solter, L. and Griswold, T. 2010. Status of bumble bee decline in the U.S. IUCN bumble bee conservation workshop, St. Louis (Disseminated Online).
• Cameron, S.A. and Lozier, J.D. 2010. Bumble bee decline across North America. Conservation and Ecological Applications of Native Pollinators symposium, Penn State. (Abstract).
• Lozier, J.D. and Cameron, S.A. 2010. Bumble bee decline in North America. Ecological Society of America Annual Meeting (Abstract).

Progress 08/15/09 to 08/14/10

Outputs
OUTPUTS: MIDWESTERN AND EASTERN REGIONAL SUMMARY OF RESEARCH: 1. DATABASING: Total records of target species (B. pensylvanicus, B. impatiens, B. bimaculatus, B. terricola, B. affinis) in the database compiled by the Eastern team = 41,192. We have completed our midwestern/eastern databasing effort. Number of institutions represented in the database = 25. 2. FIELD SURVEYS: Completed field surveys in 27 states, 112 sites. A total of 5,294 bees surveyed during summer 2009 for distribution and abundance; a total 8,674 specimens have been surveyed to date over the last two summer seasons. Additional field trips to sites in Ohio, Pennsylvania, Minnesota, and Wisconsin were conducted in 2010 specifically to look for B. penyslvanicus, B. terricola, and B. affinis (7 sites surveyed with standardized methods applied to all project surveys). 3. PATHOGEN SCREENING AND SUSCEPTIBILITY EXPERIMENTS: Midwestern and Eastern species: Screened 3,000 bees for pathogens (total to date 5,500). Western bumble bees: 2,300 specimens screened (total to date 4,348). SUSCEPTIBILITY STUDIES: Repetition of individual inoculations of adult bumble bees of two different species (B. occidentalis, B. huntii) with N. bombi spores. Colonies were developed from Spring-reared bumble bee queens in the Strange lab in Utah. Observations conducted in Utah for a month following inoculations. 4. GENOTYPING: (11 loci) target species collected from across the midwestern, southern, southeastern and eastern regions. Totals for all genotyping: B. pensylvanicus = 455 individuals. B. bimaculatus = 534 individuals. B impatiens = 666 individuals. Genotyping of eastern species has been completed. PARTICIPANTS: Nothing significant to report during this reporting period. TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
We have completed all data summarization for the project and have a paper in review on our major findings. We provided evidence that four species have significantly reduced geographic ranges compared to historical ranges of the last century, they also have significantly reduced relative abundances. These reductions are associated with higher infection levels of the microsporidium pathogen Nosema bombi, as well as reduced genetic diversity in their populations. We are careful not to mix cause and effect in these decline patterns. Whether the correlations we have found are actually causes of the observed declines has yet to be established. Of the four species that have undergone population declines, three belong to the subgenus Bombus sensu stricto (B. occidentalis, B. terricola and B. affinis) and one is from the subgenus Thoracobombus (B. pensylvanicus). The remaining four target species (B. vosnesenskii, B. bifarius, B. impatiens and B. bimaculatus) were classified as stable, having undergone no observable population declines across their native ranges in the U.S. Besides submission of a manuscript for publication, we have reported these major findings in several venues over the last year, including talks at the Penn State Pollinator Conference in July 2010 and the Ecological Society of America meetings (July 2010). We will present the findings also at the USDA workshop in San Diego in Dec 2010. In addition to the paper currently in review which summarizes the major results written in a relatively brief format for a high-impact journal, we are also working on two additional publications, one on the genetics and one on the pathogen work. We expect to release our specimen database shortly, as soon as our first paper is in press. This year we have again confirmed the presence of Bombus affinis in Illinois. This species was thought to be pretty much gone a couple of years ago, but it appears to exist in pockets in the Midwest.

Publications

• Marlin, J. and Cameron, S. 2009. Are wild bee pollinator populations declining Change in the Heartland Issue 1:8 ref: http://hdl.handle.net/2142/16449.

Progress 08/15/08 to 08/14/09

Outputs
OUTPUTS: Midwestern and Eastern Regional Summary of Research: 1. Databasing total target species (B. pensylvanicus, B. impatiens, B. bimaculatus, B. terricola, B. affinis) in database to date = 25,930 plus 16,000+ specimens from the Illinois Natural History Survey (total records = approx. 42,000). Number of institutions represented in the database: 25. We are 90% complete with our midwestern/eastern databasing effort, with two institutions remaining: the second half of the Cornell collection (approx. 500 specimens) and the Archbold Biological Station, FL. 2. Field Surveys: Completed: Field surveys in 27 states, 112 sites. A total of 5,294 bees surveyed during summer 2009 for distribution and abundance; a total 8,674 specimens have been surveyed to date over the last two summer seasons. 3. Pathogen screening and susceptibility experiments: Midwestern and Eastern bumble bee surveys: screened 2,400 bees for pathogens. Western bumble bees: 1,500 specimens screened (500 remain to be screened). Susceptibility studies: Individual inoculation experiments of adult bumble bees were conducted with with Nosema bombi spores for 4 different species (B. occidentalis, B. huntii, B. impatiens, B. bimaculatus). Two full late season B. impatiens colonies were also fed with N. bombi spores over the course of a week and observed for a month following inoculations. 4. Genotyping (11 loci) target species collected from across the midwestern, southern, southeastern and eastern regions. B. pensylvanicus = 455 individuals. B. bimaculatus = 534 individuals. B impatiens = 437 individuals. We have only B. impatiens left to complete, with 80% of the genotyping completed for this last species, which we expect to complete by 10 January, 2010. Western Summary 1. Databasing: B. occidentalis = 4,454. B. vosnesenskii = 5,279. B. bifarius =6,763. Total = 16,496. Number of institutions represented in the database is 16. Additional large databases have also provided databases of verified specimen data: Yale University (B. occidentalis = 754, B. vosnesenskii = 191, B. bifarius = 1,007; Total = 1,952), American Museum of Natural History (B. occidentalis = 340, B. vosnesenskii = 191, B. bifarius = 1,041; Total = 1,572) and the University of Kansas (B. occidentalis = 403, B. vosnesenskii = 358, B. bifarius = 802; Total = 1,563). By including these databases, total of digitized specimens = 21,583. We are still working on historic records of B. vosnesenskii (75% complete) and B. bifarius (75%) at the Cal Acad Sci; UC Berkley; UC Davis; UC Riverside; USNM. 2. Field Surveys: Completed: Field surveys in 10 states. Surveyed 311 localities, including 137 new locations (2009). 4,462 bees were surveyed in 2009 for abundance and distribution. 3. Pathogen Study: Dissected and sent >1,500 guts to Illinois for analysis in 2009. 4. Genotyping Completed: B. occidentalis. Genotyped extant pops (total of ~100 bees). B. vosnesenskii. Genotyped 900 bees from approximately 30 populations throughout the entire range. We have a minimum of 30 B. vosnesenskii per site. B. bifarius Extracted DNA from and genotyped 670 bees with approximately 600 remaining to process. Expect to finish genotyping by January 15, 2010. PARTICIPANTS: Besides the PIs working on the project, the follwing students were hired for the summer 2009 surveys, in addition to my ongoing postdoc on the project, Dr. J.D. Lozier: -Isaac Stewart, grad student in the Biology Teaching Program. Responsibilities: Organize collecting permits, organize field surveys and lead field surveys through the southern and midwestern U.S. (returning from last season) -Paul Karnstedt, undergrad student in teaching biology program, hired for field surveys and databasing. -Juraj Cech, grad student in Entomology, U Illinois, hired for field surveys and databasing. -Whitney Stewart, middle school biology teacher, returned from last season to help conduct field surveys. -Scott Czarnik, completed undergraduate and hired Feb-Aug 2009 to assist with databasing and assist with field surveys through the southern and eastern U.S. Each of these young assistants on the project has been trained in bumble bee identification, field survey protocols, floral resource identification, and in databasing museum specimen records, including georeferencing. In sum, this experience can be incorporated into the biology teaching programs of three of our assistants. All the assistants have been introduced to the scientific method, and have learned to use tools important in ecology and entomology, including quantitative methods in field surveying, the use of GPS devices, the correct preservation of insect specimens and the pinning of voucher material for museum deposit, and the keeping of scientific field notes. The bumble bee pathogen screening aspect of the project was carried this summer in co-PI Solter's lab, with lab assistance from Nils Cordes and two other undergrad/grad student assistants. The assistants learned to distinguish Nosema bombi spores, which required learning the anatomy of the bees, the microscope techniques, and slide mounting material. Our western collaborators (directed by Dr. Jamie Strange) included the following assistants through the year: -Jonathon Koch (Graduate Research Assistant) worked 20 hr per week through 2009, for a total of 1,000 hours. - Three undergraduates worked a total of 600 hours on the project. - Joyce Knoblett (520 hours) and Harold Ikerd (250 hours) (USDA technicians) contributed a combined 770 hours to the project, including field sampling, specimen curation, genotyping, and data analysis. Co-PI Strange contributed 400 hours to the project and Co-PI Griswold contributed 100 hours. TARGET AUDIENCES: PI Cameron, co-PI Strange and postdoctoral associate, J. Lozier have been targeting political action groups in an attempt to educate them about the bumble bee decline issue as we have discovered new information. They are in need of accurate information on the status of the bees and on possible causal factors in decline. To date, there have been no national studies to address either of these questions. Although we cannot give out quantitative data until we have published papers relevant to our findings, we have been able to give them some information on collecting protocols and on our perspectives of the decline as we understand it from our summer field research and databasing. This assists them by reducing the amount of speculative information they offer the public. We all attended a workshop of Bombus experts held in Washington, DC last June, organized by the U.S. Geological Survey and the North American Pollinator Protection Campaign (NAPPC). The goals of the workshop were to discuss and decide on a possible national collection protocol for bumble bees. We managed to convey our view that we are already engaged in a national survey effort, that our protocols are appropriate, that we have targeted scientific questions to answer, and that we do not need at this time lots of independent efforts to collect bees. We are engaged in a major conservation effort, and our methods minimize the sacrifice of live bees surveyed in the field. We feel that we can no longer justify massive surveys that simply kill and preserve bees without careful thought as to what the data will be used for. We believe we were successful in communicating this message. PI Cameron and co-PI Strange were asked to participate as co-chairs of an NAPPC Task Force on bumble bees, which we accepted. The first meeting was held in October, which Strange attended and moderated. Several short-term projects were organized at the meeting for completion within the year. The meeting brought together interested individuals from govt. agencies, NGOs, and universities to discuss pollinator decline and possible mitigation. PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
We are in the midst of completing all data summarization for the project. We expect to report the major findings of the research in scientific papers to be submitted in winter 2010. We will be able to summarize the current status of all target specimens, release our constructed databases, and draw conclusions about the association/correlation between apparently threatened species and Nosema bombi. At this time we can conclude that we have confirmed the presence of at least 2 target eastern species that were thought to be seriously disappearing: B. affinis (thought to be nearly extinct across its range) has been found in several locations in Illinois and Wisconsin; B. terricola was found in fairly large numbers along the Maine mid-coastal region. Based on western surveys in 7 National Parks, we can confirm the presence of B. occidentalis, a threatened western species, in (or near) 5 of the parks within the historic species range.

Publications

• Grixti, J.C., Cameron, S.A. and Favret, C. 2008. Decline of bumble bees (Bombus) in the North American Midwest. Biological Conservation, 142:75-84. doi:10.1016/j.biocon.2008.09.027.
• Lozier, J.D. and Cameron, S.A. 2009. Comparative genetic analyses of historical and contemporary collections highlight contrasting demographic histories for the bumble bees Bombus pensylvanicus and B. impatiens in Illinois. Molecular Ecology, 18:1875-1886. doi:10.1111/j.1365-294X.2009.04160.x.
• Strange J.P., Knoblett J. and Griswold, T. 2008. DNA amplification from pin-mounted bumble bees (Bombus) in a museum collection: Effects of fragment size and specimen age on successful PCR. Apidologie, 40:134-139. DOI: 10.1051/apido/2008070.
• Marlin, J. and Cameron, S.A. 2010. Are wild bumble bee pollinators declining Change and the Heartland, University of Illinois Agroecology and Sustainable Agriculture Program for the Environmental Change Institute, Univ. of Illinois. (In Press).

Progress 08/15/07 to 08/14/08

Outputs
OUTPUTS: Between June and September 2008 we conducted field surveys of bumble bee diversity and abundance throughout the Midwestern Region, TX, AR, and OK. Our surveys also included the eastern portion of the western states of CO, WY, SD and ND. Overall we surveyed a total of 86 sites across 15 states. In 2008 Co-PIs Griswold and Strange led sampling of western bumble bee populations at 93 sites in ten states within the historic range of B. occidentalis. A total of 15 sites yielded specimens of B. occidentalis. A total of 3300 bees were surveyed and of those, 2465 were dissected and the guts preserved for the Nosema bombi survey. - Screening of Nosema and Crithidia pathogens in bumble bees surveyed from the Western and Midwestern regions has begun. Midguts have been examined for the presence of Nosema or Crithidia in a total of 965 of 1695 individuals collected in the Midwestern states and 518 individuals in the western states. - In our examination of genetic diversity and population structure in the Midwestern Region, we have performed preliminary analyses of microsatellite diversity and locus optimization and have developed 11 loci that amplify well in all three target species. A subset of these loci amplified in 40 year-old museum specimens from the IL Natural History Survey, which enabled us to conduct a preliminary microsatellite analysis comparing historical IL B. impatiens (8 loci) and B. pensylvanicus (9 loci) populations (3 populations each sampled between 1966 and 1970) to collections made in the same regions during the current survey. -We are continuing to database specimen records obtained from entomology collections at university museums to clarify the historical distributions of target Bombus species. To date, we have entered and georeferenced 20,994 specimen records consisting of 33 Bombus species from 15 museum collections. To predict the likely future geographic distributions of target species, we have used georeferenced museum data to implement ecological niche models using the computer program MAXENT. Co-PIs Strange and Griswold have generated preliminary historic range maps for B. occidentalis, B. vosnesenskii and B. huntii based upon labeled specimens deposited in federal and university entomological collections. They are currently adding data to these projects. -Seminars/Workshops: Co-PI Strange: May 2008 "Native bumble bees in the garden: backyard conservation" Workshop presenter for Utah Master Gardener Annual Workshop, Brigham City, UT; April 2008 "Bombus in the West: range contractions and population declines" Washington State University, Dept of Entomology, Pullman, WA. -Co-PI Griswold: Presentation on bumblebee conservation by collaborator Robbin Thorp at the September 2008 Bee Course (22 international participants) with following discussion involving co-PI Strange. Cameron, Solter, Strange and Griswold presented a poster at the CSREES workshop in Dec. 2007; postdoc Lozier presented a talk at the Evolution meetings June 2008 on the results of ecological niche modeling, predicting future Bombus species distributions. PARTICIPANTS: PI Cameron's lab: During the first year of the project, Dr. Jeffrey Lozier was hired as a postdoc to work on the population genetic portion of the research. Nils Cordes was recruited to work on the pathogens and has been trained in the methods of DNA sequencing in the Cameron lab. Three summer field assistants were hired, who were invaluable for the wide-ranging surveys conducted in summer 2008. These assistants included Isaac Stewart, a graduate student in the Biological Sciences teaching program, Whitney Stewart, a high school biology teacher, Beth Peregrine, an undergraduate with interests in entomology. Jennifer Grixti, who recently received a Masters degree centered around solitary bee biodiversity in North America, was hired to do the databasing. All of these individuals were new to bumble bee research when they arrived in the PI's lab, and subsequently have learned to identify bumble bees in the field and laboratory, to collect them, and to preserve them for various purposes. Dr. Lozier has learned how to extract Bombus DNA for genotyping populations and has developed microsatellite primers for genotyping. Co-PI Solter's lab: Solter and Cameron recruited Nils Cordes, a graduate student at the PhD level, who has focused on the pathogen portion of the project. Co-PI Solter has trained Cordes in pathogen identification, isolation and storage, and in DNA isolation. She also organized, convened and hosted an international workshop on Microsporidia. Co-PIs Strange and Griswold: During the first year, they hired and advised graduate student, Jonathan Koch, and an undergraduate assistant, Leah Waldner. Koch has participated in field surveys of western Bombus, spending 30 days traveling for field collections. In conjunction with Waldner, he has dissected over 800 Bombus specimens, removing gut tissue for pathogen analysis. He has databased records of field sampling sites and bee species identifications. Waldner assists Koch with data entry and has been entering historic specimen data into the database. Waldner has assisted with field collections totaling 10 days in the field. She works with Koch to complete the dissections of bees and assists Co-PI Strange with DNA extraction and amplification. CO-PI Strange coordinates the input of historic data and field sampling for the project. He has spent over 35 days field sampling in summer 2008 in addition to providing his technician (not funded by the grant) to the project for over 30 days of field sampling, museum data collection and dissections. Co-PI Griswold contributed 7 days of field sampling and his technician clocked an additional 20 days of field sampling, specimen curation and dissections. Both Co-PIs have partnered with the Xerces Society for Invertebrate Conservation to gather historic data and to produce historic range maps for B. occidentalis and other Bombus species. Field collections in CA and OR have been coordinated with Dr. Robbin Thorp (UC-Davis), who has his own funding sources yet shares data and sampling sites. Field collections were coordinated with Dr. Graham Pyke (Australian Museum) at the Rocky Mountain Biological Laboratory, CO. TARGET AUDIENCES: PI Cameron ran a workshop on bee identification for National Pollinator Week. The focus was bees of Illinois, with an emphasis on bumble bees. This was held through the University of Illinois and the Champaign Public Library, and was succesful in bringing in a broad array of individuals from the community, plus a few students. The workshop was assisted by two graduate students from Cameron's laboratory (Heather Hines and Claus Rasmussen), both experts on bees, and Dr. James B. Whitfield. Participants were provided with pictoral guides for identification of IL bumble bees, floral guides to bee gardening, and as much information on biology as possible during the several hous. Cameron and graduate student, H. Hines, created a pictoral key with photographs and graphics for use by the St. Louis Zoo and by interested individuals in IL and Missouri. This has been helpful for training students in identification of bumble bees in the field, as the key points to the major distinguishing features of the different species, some of which are difficult to tell apart because of similarity in color pattern. Co-PI Solter organized, convened and hosted an international workshop on Microsporidia. This brought scientists and students together internationally for several days to discuss key problem areas and possible solutions. Co-PIs Strange and Griswold partnered with the Xerces Society for Invertebrate Conservation to gather historic data and to produce historic range maps for B. occidentalis and other Bombus species. They also had two meetings with UT State Dept. of Food and Agriculture personnel to discuss the impact of Japanese beetle trapping on Bombus populations with special focus on trap placement to reduce the number of spring queens caught. We used museum data gathered for this grant (timing of queen flight in the spring for all species we studied in UT) to help develop our recommendations. UDFA agreed to place traps later in the spring and retrieve them earlier in the fall in areas where B. occidentalis is known from historic data. PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
- From the Bombus species diversity and distibution surveys of > 4,000 bees in the Midwestern region we found that B. impatiens was the most numerous species collected (787), followed by B. griseocollis (780), B. bimaculatus (390), B. auricomus (208), and B. pensylvanicus (206). Nosema screening is in the earliest stages but summary results suggest that in some sites Nosema may have a high prevalence within one species but the overall prevalence in most populations is low. Based on initial sequencing results, the only microsporidian species isolated thus far is Nosema bombi. Thus far, the incidence of Crithidia has been lower than that of Nosema. - Results from the historical analysis of genetic diversity suggest that modern B. pensylvanicus populations exhibit a small but consistent reduction in heterozygosity relative to the historical populations (averages +/- s.d. = 0.572 +/- 0.000 versus 0.609 +/- 0.002). The same was not true for B. impatiens, which showed an increase in heterozygosity relative to historical levels (0.666 +/- 0.029 versus 0.622 +/- 0.012). For B. pensylvanicus, population structure was low and insignificant in historical populations (FST = 0.008) but was stronger and highly significant for modern populations (FST = 0.030), suggesting a decline in population connectivity and gene flow over the last 40 generations. FST levels for B. impatiens did not differ from zero for either set of populations, suggesting that populations of this species remain highly connected. -Graduate student Nils Cordes has been trained to identify multiple bumble bee pathogens, including Nosema and Crithidia. He has been trained in DNA sequencing procedures and primer development as a means to eventually reveal the origin of Nosema bombi in North America. We are expecting that the Nosema ceranae genome will assist in our efforts to identify variable N. bombi genes for insights into its geographic origins. -We have trained multiple undergraduates, assistants, graduate students and a postdoc to survey, collect and identify North American bumble bees, and have developed (collaboration with Paul Williams of NHM, London) a pictoral key to Bombus species of IL and MO and color diagrams of species color patterns for the western and Midwestern regions. We have helped to develop the BeeSpotter website for bee identification by citizen scientists. - Two meetings with UT State Dept. of Food and Agriculture personnel to discuss the impact of Japanese beetle trapping on Bombus populations with special focus on trap placement to reduce the number of spring queens caught. We used museum data gathered for this grant (timing of queen flight in the spring for all species we studied in UT) to help develop our recommendations. UDFA agreed to place traps later in the spring and retrieve them earlier in the fall in areas where B. occidentalis is known from historic data.

Publications

• Grixti, J.C., Wong, L.T., Cameron, S.A. and Favret, C. 2008. Decline of bumble bees (Bombus) in the North American Midwest. Biological Conservation (in press).
• Lozier J.D. and Cameron S.A. 2009. Genetic diversity in bumble bees from the past and present: microsatellite analyses of natural history collection specimens provide insights into the recent histories of Bombus pensylvanicus and Bombus impatiens populations in Illinois (in prep.)
• Strange J.P., Knoblett J. and T. Griswold. 2008. DNA amplification from pin-mounted bumble bees (Bombus) in a museum collection: effects of fragment size and specimen age on successful PCR. Apidologie (in press).
• Williams, P.H., Cameron, S.A., Hines, H.M., Cederberg, B. and Rasmont, P. 2008. A simplified subgeneric classification of the bumblebees (genus Bombus). Apidologie 39:46-74.