POLLINATION AND THE DEVELOPMENT OF ALTERNATIVE CROP POLLINATORS

Goals / Objectives
Objective 1. Deliver improved pollination management systems for alfalfa seed production using the alfalfa leafcutting bee and the alkali bee, and a new pollination management system for orchards using the blue orchard bee. Objective 2. Evaluate pollination efficacies of confined and free-flying populations of select new bee pollinators for small fruit crops and regional forbs in demand for re-vegetation, and develop practical management protocols to encourage adoption of the more promising species as custom pollinators. Objective 3. Develop novel control options and delivery systems for the management of chalkbrood, other diseases, parasites, and predators in commercial-scale pollinator populations. Objective 4. Enhance knowledge of native pollinators by conducting biological surveys, establishing monitoring programs, and expanding knowledge of bee systematics in agricultural and natural systems.

Project Methods
Research is needed to increase the number of bee species used for agricultural pollination. This means improving existing alkali and alfalfa leafcutting bee management systems, delivering the blue orchard bee as a mainstream organic orchard pollinator, assessing the pollinating efficiencies of promising solitary bees for a variety of crops (particularly those not currently well-served by honey bees), and developing programs to maintain and augment the populations necessary to complement and supplement the pollination of crop and native plants. The overarching goal for the 60 months of research outlined in this Project Plan is improved profitability, for U.S. producers of agricultural crops that require cross-pollination, through the diversification of our agricultural pollinator portfolio. In order to achieve this goal, priority will be given to: 1) Delivering improved pollination management systems for alfalfa seed production using the alfalfa leafcutting bee and the alkali bee, and a new pollination management system for orchards using the blue orchard bee; 2) Evaluating pollination efficacies of confined and free-flying populations of select new bee pollinators for small fruit crops and forbs in demand for re-vegetation, and developing practical management protocols to encourage adoption of the more promising species as custom pollinators; 3) Developing novel control options and delivery systems for the management of chalkbrood, other diseases, parasites, and predators in commercial-scale pollinator populations; and, 4) Conducting biological surveys, establishing monitoring programs, and expanding current knowledge of bee systematics in natural and agricultural systems.

Progress 10/01/03 to 09/30/08

Outputs
Progress Report Objectives (from AD-416) Objective 1. Deliver improved pollination management systems for alfalfa seed production using the alfalfa leafcutting bee and the alkali bee, and a new pollination management system for orchards using the blue orchard bee. Objective 2. Evaluate pollination efficacies of confined and free-flying populations of select new bee pollinators for small fruit crops and regional forbs in demand for re-vegetation, and develop practical management protocols to encourage adoption of the more promising species as custom pollinators. Objective 3. Develop novel control options and delivery systems for the management of chalkbrood, other diseases, parasites, and predators in commercial-scale pollinator populations. Objective 4. Enhance knowledge of native pollinators by conducting biological surveys, establishing monitoring programs, and expanding knowledge of bee systematics in agricultural and natural systems. Approach (from AD-416) Research is needed to increase the number of bee species used for agricultural pollination. This means improving existing alkali and alfalfa leafcutting bee management systems, delivering the blue orchard bee as a mainstream organic orchard pollinator, assessing the pollinating efficiencies of promising solitary bees for a variety of crops (particularly those not currently well-served by honey bees), and developing programs to maintain and augment the populations necessary to complement and supplement the pollination of crop and native plants. The overarching goal for the 60 months of research outlined in this Project Plan is improved profitability, for U.S. producers of agricultural crops that require cross-pollination, through the diversification of our agricultural pollinator portfolio. In order to achieve this goal, priority will be given to: 1) Delivering improved pollination management systems for alfalfa seed production using the alfalfa leafcutting bee and the alkali bee, and a new pollination management system for orchards using the blue orchard bee; 2) Evaluating pollination efficacies of confined and free-flying populations of select new bee pollinators for small fruit crops and forbs in demand for re-vegetation, and developing practical management protocols to encourage adoption of the more promising species as custom pollinators; 3) Developing novel control options and delivery systems for the management of chalkbrood, other diseases, parasites, and predators in commercial-scale pollinator populations; and, 4) Conducting biological surveys, establishing monitoring programs, and expanding current knowledge of bee systematics in natural and agricultural systems. Significant Activities that Support Special Target Populations This report serves to document research conducted for project 5428- 21000-010-00D Pollination and the Development of Alternative Crop Pollinators; and National Program 305 Crop Production, Component I. Integrated Sustainable Crop Production Systems Component II. Bees and Pollination, B. Non-Apis Bees. The objective of this project has been to increase pollinating bees available for agriculture. The 5-year project plan concludes this year. During this past year, experiments were conducted to improve the utility of blue orchard bees as pollinators for almonds in an effort to assist the growing pollination demands of this crop in California, during a time when honey bees are facing declines. Management systems for the blue orchard bee were developed for tree fruits in previous years, as part of this project, but California almonds have some special complicating factors such as a lack of availability of other flowers in the orchards for the bees prior to almond bloom, and climate conditions significantly different from the more northern areas where the bees are brought in from. In addition to this research, a new Osmia bee was developed for raspberry pollination that works quite well, and it has been given to producers. Research on chalkbrood control and pollen ball syndrome in the alfalfa leafcutting bee determined that overstocking the bees in alfalfa seed fields increases the incidence of these problems. Knowledge is now needed on what the optimal stocking densities should be, with the goal of avoiding these bee health problems while simultaneously providing sufficient pollination for the crop. Experiments have been initiated to determine these densities. A project to identify the immune response genes for leafcutting bees was also initiated this year, and the genes are being compared to the honey bee genome. This project will lead to better knowledge on how bees combat disease and the genetics associated with disease resistance. Approximately 1000 disease-related EST were developed for this bee under this project. Bee biodiversity surveys for bumble bees were initiated this year, and surveys to quantify the diversity of all bees in natural systems were completed for national parks in the western U.S. These surveys were linked to a national database, and web-based taxonomic keys have been developed and released to assist biologists working on bee biodiversity issues. Technology Transfer Number of New CRADAS: 1 Number of Web Sites managed: 1

Impacts
(N/A)

Publications

Alston, D.G., Tepedino, V.J., Bradley, B.A., Toler, T.R., Griswold, T.L. 2007. Effects of the insecticide phosmet on solitary bee foraging and nesting in orchards of Capitol Reef National Park, Utah (U.S.A.). Environmental Entomology. 36(4): 811-816
Ascher, J.S., Engel, M.S., Griswold, T.L. 2006. A new subgenus and species of Oxaea from Ecuador (Hymenoptera: Andrenidae). Polskie Pismo Entomologiczne 75(4): 539-552
Cane, J.H. 2008. Acta horticulturae an effective, manageable bee for pollination of rubus cane fruits, osmia aglaia. Acta Horticulturae. (ISHS) 777: 459-464
Cane, J.H. 2008. A native ground nesting bee, Nomia melanderi, sustainably managed to pollinate alfalfa across an intensively agricultural landscape. Apidologie. 39: 315-323
Cane, J.H., Griswold, T.L., Parker, F.D. 2007. Substrates and Materials Used for Nesting by North American Osmia bees (Apiformes: Megachilidae). Annuals of the Entomological Society of America. 100(3):350-358(9)
Griswold, T.L., Hanson, P.E., Alvos Dos Santos, I. 2006. Capitulo 18. Apoidea: Las Abejas. In: I. Gauld & P. Hanson, Hymenoptera de la Region Neotropical. American Entomological Institute.
Gonzalez, V.H., Griswold, T.L. 2007. A review of the North and Central American Megachile subgenus Argyropile subgenus Argyropile mitchell (Hymenoptera: Megachilidas). Zootaxa. 1461:68
Guedot, C., Bosch, J., Kemp, W.P. 2007. Effect of three dimension and color contrast patterns on nest localization performance of two solitary bees (hymenoptera: megachilidae). Journal of Kansas Entomological Society. 80:2:90-104.
Huntzinger, C., James, R.R., Bosch, J., Kemp, W.P. 2008. Fungicide Tests on Adult Alfalfa Leafcutting Bees Megachile rotundata (F.) (Hymenoptera:Megachilidae). Journal of Economic Entomology. 101(3): 660- 667
Pitts Singer, T., Bosch, J., Kemp, W.P., Trostle, G.E. 2008. Field use of an incubation box for improved emergence timing of Osmia lignaria populations used for orchard pollination. Apidologie. 39: 235-246
Pitts Singer, T., Espelie, K.E. 2007. Nest demographics and foraging behavior of apterostigma collare emery (hymenoptera, formicidae)provide evidence of colony independence. Insectes Sociaux. 54: 310-318
Pitts Singer, T., James, R.R. 2008. Do weather conditions correlate with findings in failed, provision-filled nest cells of Megachile rotundata (Hymenoptera:Megachilidae) in western North America?. Journal of Economic Entomology. 101(3): 674-685
Strange, J.P., Cicciarelli, R.P., Calderone, N.W. 2008. What's in that Package? An Evaluation of Quality of Package Honey Bee Apis mellifera L. (Hymenoptera:Apidae) shipments in the U.S.. Journal of Economic Entomology. 101(3): 668-673
Tepedino, V.J., Toler, T.R., Bradley, B.A., Hawk, J.L., Griswold, T.L. 2007. Pollination biology of a disjunct population of the endangered sandhills endemic Penstemon haydenii S. Wats. (Scrophulariaceae) in Wyoming, USA. Journal Of Plant Ecology. 193:1:59-69
Cane, J.H. 2008. Pollinating Bees Crucial to Farming Wildflower Seed for U. S. Habitat Restoration. Book Chapter. Bee Pollination in Agricultural Ecosystems. Oxford University Press
James, R.R. 2008. The Problem of Disease When Domesticating Bees. Book Chapter. Bee Pollination in Agricultural Ecosystems. Oxford University Press.
James, R.R., Pitts Singer, T. 2008. The Future of Agricultural Pollination. Book Chapter. Bee Pollination in Agricultural Ecosystems. Oxford University Press
Pitts Singer, T. 2008. Past and Present Management of Alfalfa Bees. Book Chapter. Bee Pollination in Agricultural Ecosystems. Oxford University Press
Pitts Singer, T., James, R.R. 2008. Bees in Nature and on the Farm. Book Chapter. Bee Pollination in Agricultural Ecosystems. Oxford University Press
James, R.R., Pitts Singer, T. 2008. Bee Pollination in Agricultural Eco- Systems. Complete Book. Oxford University Press.

Progress 10/01/06 to 09/30/07

Outputs
Progress Report Objectives (from AD-416) Objective 1. Deliver improved pollination management systems for alfalfa seed production using the alfalfa leafcutting bee and the alkali bee, and a new pollination management system for orchards using the blue orchard bee. Objective 2. Evaluate pollination efficacies of confined and free-flying populations of select new bee pollinators for small fruit crops and regional forbs in demand for re-vegetation, and develop practical management protocols to encourage adoption of the more promising species as custom pollinators. Objective 3. Develop novel control options and delivery systems for the management of chalkbrood, other diseases, parasites, and predators in commercial-scale pollinator populations. Objective 4. Enhance knowledge of native pollinators by conducting biological surveys, establishing monitoring programs, and expanding knowledge of bee systematics in agricultural and natural systems. Approach (from AD-416) Research is needed to increase the number of bee species used for agricultural pollination. This means improving existing alkali and alfalfa leafcutting bee management systems, delivering the blue orchard bee as a mainstream organic orchard pollinator, assessing the pollinating efficiencies of promising solitary bees for a variety of crops (particularly those not currently well-served by honey bees), and developing programs to maintain and augment the populations necessary to complement and supplement the pollination of crop and native plants. The overarching goal for the 60 months of research outlined in this Project Plan is improved profitability, for U.S. producers of agricultural crops that require cross-pollination, through the diversification of our agricultural pollinator portfolio. In order to achieve this goal, priority will be given to: 1) Delivering improved pollination management systems for alfalfa seed production using the alfalfa leafcutting bee and the alkali bee, and a new pollination management system for orchards using the blue orchard bee; 2) Evaluating pollination efficacies of confined and free-flying populations of select new bee pollinators for small fruit crops and forbs in demand for re-vegetation, and developing practical management protocols to encourage adoption of the more promising species as custom pollinators; 3) Developing novel control options and delivery systems for the management of chalkbrood, other diseases, parasites, and predators in commercial-scale pollinator populations; and, 4) Conducting biological surveys, establishing monitoring programs, and expanding current knowledge of bee systematics in natural and agricultural systems. Accomplishments Native Pollinator Effective for Alfalfa Seed Pollination. A large-scale, multi-year field experiment on the population dynamics of the alkali bee was completed in cooperation with Washington State growers. This bee is a native, ground nesting bee used for alfalfa seed production. We determined that 3 million nesting females from one nesting bed can readily pollinate 160 acres of alfalfa, yielding 1000 lbs clean seed per acre. Flowers were pollinated in the first 6 hours of the day. Seven years of sustained population growth across the watershed resulted in 16 million nesting females. Impact: This project has demonstrated that the alkali bee can be a sufficient pollinator, alone, for alfalfa seed production. It has now become the primary pollinator for the region of the study, pollinating several million pounds of alfalfa seed annually at a fraction of the management cost for alfalfa leaf-cutting bees. This research was conducted under National Program 305 Crop Production, and directly contributes to research Component 6.0 Bees and Pollination, ARS Strategic Plan Goal 1.2 Contribute to the Efficiency of Agricultural Production Systems. Blue Orchard Bee Incubation Box Improves Emergence in Orchard. Blue orchard bees are excellent pollinators for tree crops such as almonds, cherries, apples and pears, and a relatively small number of bees are needed for these crops. However, fruit trees bloom early in the spring when temperatures can sometimes be cool enough to inhibit bee emergence from the winter cocoon stage and reduce blue orchard bee activity. We designed and field-tested a new outdoor incubation box and compared emergence rates of bees incubated in the boxes with those incubated under standard conditions and determined that the boxes facilitated faster bee emergence without increasing mortality. Thus, we have developed an incubation box that is an effective tool for shortening emergence periods and, therefore, improving management of these bees when weather is variable and unpredictable. Impact: A patent application for the box has been filed, and a customer has expressed interest in licensing the patent once it is available. This research was conducted under National Program 305 Crop Production, and directly contributes to research Component 6.0 Bees and Pollination, ARS Strategic Plan Goal 1.2 Contribute to the Efficiency of Agricultural Production Systems. Restoration of the Nation's Rangelands. Federal, state and private land managers in the western U.S. are critically in need of affordable native plant seed for revegetation of rangeland, and other wildlands, due to the destruction of native plant communities from invasive weeds (e.g. cheat grass), natural disasters (e.g. wildfires), and destructive human activities. Farm production methods for wildflower seed could generate restoration seed at affordable prices; however, in order to produce seed on farms, pollinators for these plants are needed. We determined that the majority of desired wildflower species require bees for pollination, but some are satisfactorily pollinated with conventional pollinators (honey bees or alfalfa leafcutting bees). We also identified two native pollinators that work well for some of the other flowers, and we have successfully developed methods to propagate these bees using shelters and nesting substrates. Impact: This research is necessary for a small cadre of seed growers, but the economic impact of this small market is overshadowed by the millions of acres of western rangelands that will benefit by having their degraded plant communities restored. The research reported here is being conducted under National Program 305 Crop Production, and directly contributes to research Component 6.0 Bees and Pollination. Research activities are further related to ARS Strategic Plan Goal 2 Support Increased Economic Opportunities and Improved Quality of Life in Rural America, and Goal 5 Protection and Enhance the Nation�s Natural Resource Base and Environment, Objective 5.1 Provide Science Based Knowledge and Education to Improve the Management of Forest, Rangelands, and Pastures. Old Nest Components Attract Female Alfalfa Leafcutting Bees and Blue Orchard Bees. These two solitary bees are used by farmers for pollination of alfalfa seed and tree fruit crops, respectively, and it has long been recognized by the growers that these bees prefer to nest in or near previously used nesting materials. We found that volatile compounds emanating from leaf pieces used by the alfalfa leafcutting bees to line their nests, and feces attached to the cast-off cocoons were attractive to nesting leafcutting bees, but other nesting materials were not. For the blue orchard bee, volatile compounds from caste-off cocoons and the paper straws we use to line the nest cavities were the attractive materials. We also found that, for these two bee species, actively nesting bees excrete chemicals that they use to mark and identify their own nests. Impact: Understanding that solitary bees are attracted to particular chemical components for nesting is the first step in developing attractants that could be manufactured and applied to nests to encourage the bees to nest where we want them to, and to reduce absconding of economically important bees. This research was conducted under National Program 305 Crop Production, and directly contributes to Component 6.0 Bees and Pollination, and ARS Strategic Plan Goal 1 Enhance Economic Opportunities for Agricultural Producers, Objective 1.2 Contribute to the Efficiency of Agricultural Production Systems. Development of Bumble Bee Genetic Markers to Evaluate Population Health. Increasingly, questions arise regarding whether our native pollinators are declining, particularly our native bumble bee populations; however, little work has been done to develop the tools necessary to study the abundance and diversity of native bees. We developed a protocol for using DNA methods (based on microsatellite markers) to determine the population genetics for 32 species of native bumble bees. These protocols can be used to reveal the genetic diversity and number of bumble bee colonies in an area based on a sample of collected bees. Additionally, these genetic markers have proven useful as taxonomic characters to identify difficult species (male bumble bees are particularly difficult to properly identify) , improving our ability to correctly identify species. Impact: These markers allow us to evaluate whether our native bumble bee populations are declining, and the status of a population can be evaluated with one, or only a few, years data rather than requiring several years of extensive sampling. This research was conducted under National Program 305 Crop Production, and directly contributes to research Component 6.0 Bees and Pollination. Research activities are further related to ARS Strategic Plan Goal 5 Protection and Enhance the Nation�s Natural Resource Base and Environment, Objective 5.1 Provide Science Based Knowledge and Education to Improve the Management of Forest, Rangelands, and Pastures. Enhanced Bee Identification. The extent of native bee declines has drawn national concern. However, a major roadblock exists to conducting an evaluation of our nation's pollinators, and that is, no means to readily and accurately identify all the bees collected from large surveys. Currently, only a handful of bee taxonomists are competent to provide identifications. To reduce this bottleneck, we developed, in collaboration with Patuxent Wildlife Refuge, a web-accessible guide to bees of the Eastern U.S. that allows non-specialists to accurately identify bees. We also created a guide to the genera of Megachilidae, a family of bees that includes the alfalfa leafcutting bee and blue orchard bee, and that shows the most promise for additional crop pollinators. Impact: These web products will expanded the capability of a great number of people to conduct pollination research. This research was conducted under National Program 305 Crop Production, and directly contributes to research Component 6.0 Bees and Pollination. Research activities are further related to ARS Strategic Plan Goal 5 Protection and Enhance the Nation�s Natural Resource Base and Environment, Objective 5.1 Provide Science Based Knowledge and Education to Improve the Management of Forest, Rangelands, and Pastures. Technology Transfer Number of New CRADAS and MTAS: 2 Number of Active CRADAS and MTAS: 2 Number of Patent Applications filed: 1 Number of Web Sites managed: 1 Number of Non-Peer Reviewed Presentations and Proceedings: 11 Number of Newspaper Articles,Presentations for NonScience Audiences: 15

Impacts
(N/A)

Publications

Winfree, R., Griswold, T.L., Kremen, C. 2007. Effect of human disturbance on bee communities in a forested ecosystem. Conservation Biology. 21(1): p. 213-223.
Yocum, G.D., Kemp, W.P., Bosch, J., Knoblett, J.N. 2006. Thermal history influences diapause development in the solitary bee Megachile rotundata. Journal of Insect Physiology. 52(11-12):1113-1120.
Frankie, G.W., Rizzardi, M., Vinson, S., Griswold, T.L., Ronchi, P. 2005. Changing bee composition and frequency on a flowering legume, Andira inermis (wright) kunth ex dc. during El Ni�o and La Ni�a years (1997-1999) in northwestern Costa Rica. Journal of Kansas Entomological Society. 78 (2) :100-117.
James, R.R. 2007. Chalkbrood. In: Shimanuki, H., Flottum, K., Harman, A., editors. ABC & XYZ of Bee Culture. 41st edition. Medina, OH: A.I. Root Company. p. 201.
James, R.R., Hayes, G. 2007. Microbial control of varroa: misadventures in the field. Journal of Anhui Agricultural University. 34(2) :162-166
Cane, J.H. 2006. An evaluation of pollination mechanisms for purple prairie-clover, Dalea purpurea (Fabaceae: Amorpheae). American Midland Naturalist. 156:193-197.
Cane, J.H., Sipes, S. 2006. Floral specialization by bees: analytical methods and a revised lexicon for oligolecty. In NM Waser and J. Ollerton, editors. Plant-pollinator interactions: from specialization to generalization. Univ. Chicago, Chicago, IL, USA. p.99-122.
Klein, A.M., Vaissiere, B.E., Cane, J.H., Steffan-Dewenter, I., Cunningham, S.A., Kremen, C., Tscharntke, T. 2007. Importance of pollinators in changing landscapes for world crops. Proceedings of the Royal Society of London B. 274 (1608):303-313
Tepedino, V., Bowlin, W., Griswold, T.L. 2006. Pollination biology of the endangered Blowout Penstemon (Penstemon haydenii S. Wats.: Scrophulariaceae) in Nebraska. The Journal of the Torrey Botanical Society. 133(4): 548-559
Pitts Singer, T. 2007. Olfactory response of megachilid bees, Osmia lignaria, Megachile rotundata and M. pugnata, to individual cues from old nest cavities. Environmental Entomology. 36: 402-408
Pitts Singer, T., James, R.R. 2007. Leafcutting bees. In: Shimanuki, H., Flottum, K., Harman, A., editors. ABC & XYZ of Bee Culture. 41st edition. Medina, OH: A.I. Root Company. p 492.
Clement, S.L., Griswold, T.L., Rust, R.W., Hellier, B.C., Stout, D.M. 2006. Bee associates of flowering Astragalus and Onobrychis genebank accessions at a Snake River site in Eastern Washington. Journal of Kansas Entomological Society. 79(3):254-260.
Cane, J.H. 2007. Alkali Bee. In: Shimanuki, H., Flottum, K., Harman, A., editors. ABC & XYZ of Bee Culture. 41st edition. Medina, OH. A.I. Root Company. p.11-12.
Cane, J.H. 2007. Bee, Definition of. In: Shimanuki, H., Flottum, K., Harman, A., editors. ABC & XYZ of Bee Culture. 41st edition. Medina, OH. A. I. Root Company. p.78-80.
James, R.R., Hayes, G., Leland, J.E. 2006. Field trials on the microbial control of varroa with the fungus Metarhizium anisopliae. American Bee Journal. November. p.968-972

Progress 10/01/05 to 09/30/06

Outputs
Progress Report 1. What major problem or issue is being resolved and how are you resolving it (summarize project aims and objectives)? How serious is the problem? Why does it matter? Pollinators are a critical component of Crop Production (ARS National Program 305), for without effective pollination, crops as diverse as alfalfa seed, almonds, tomatoes, sunflowers, tree fruits, berries, squash, and melons are failures. Non-Apis pollinators, such as the alfalfa leafcutting bee, alkali bee, and blue orchard bee are intensively managed for pollination of certain crops, and wild native bees are estimated to provide free pollination services worth $3 billion per year in the U.S. In addition, wild bees maintain plant diversity in our national parks and wild lands. The mission of Pollinating-Insect Biology, Management, & Systematics Research Unit (PIRU) is to improve both the quality and yield of insect- pollinated crops in the U.S. by increasing the availability of agriculturally important bee pollinators. This means improving solitary- bee management in alfalfa seed production, improving bumble bee production for greenhouse pollination, delivering the blue orchard bee as an orchard pollinator, and developing programs to maintain or supplement the bee populations necessary to meet the needs of U.S. crops and native plants. The major beneficiaries of the research outlined in this Project Plan are the U.S. alfalfa seed producers (NW Alfalfa Seed Growers Association, and The Alfalfa Alliance), who rely heavily on the superior performance of the alfalfa leafcutting bee and the alkali bee for pollination each year. This group enjoys a close relationship with the PIRU, and their members participate directly in the planning and execution of ongoing research. Additionally, PIRU research results are in demand by commercial seed and fruit producers, particularly almond growers, and organic fruit and nut producers, bee suppliers, as well as, federal, state, and private land managers, amateur gardeners, and non-profit non-governmental organizations. 2. List by year the currently approved milestones (indicators of research progress) Year 1 (2004) 1. Deliver improved pollination management systems for alfalfa seed production. a. Complete alkali bee brood provision analyses and draft provision sugar methods manuscript. Conduct alfalfa leafcutting bee density enclosure studies and analyze data. Collect samples and information for regional alfalfa leafcutting bee health survey. b. Initiate behavioral bioassays for nest attraction using alfalfa leafcutting bees. Perform chemical analyses of important attractants. c. Conduct temperature studies on summer development of phenologically distinct blue orchard bee populations and initiate pre-wintering studies. d. Complete life cycle respirometry, supercooling point, amino acid, lipid, and carbohydrate profiles for natural alfalfa leafcutting bee populations. Identify genes that signal change from diapause (hibernation- like state in insects) to post-diapause development and compare their expression levels during the winter through spring time development period. e. Demonstrate the blue orchard bee and new nesting block design in selected early-spring orchard crops. Complete the 6th, and final, year of cherry pollination demonstration in UT. Complete year 2 of organic almond pollination demonstration in CA. Complete year 1 of cherry pollination demonstration in MT. Complete initial field testing of current nest block prototype in UT pear-apple orchards. f. Complete first year experiments on wintering temperature, parasitism and body size. Select progeny of dispersing vs. non-dispersing females for future field trials. 2. Evaluate pollination efficacies of bees selected for potential to pollinate small fruit crops and native plants used in re-vegetation programs. a. Refine nest substrate options for Osmia aglaia in raspberries. Confirm basic breeding biologies of focal wildflower seed species. Begin sampling pollinator guild of wildflowers. Grow Osmia ribifloris and address dispersal problems. Publish cranberry pollinator efficacy study. 3. Develop novel disease and parasite control methods for bees. a. Refine molecular marker methods for chalkbrood detection. b. Apply fungicides to loose cells of the alfalfa leafcutting bee and test effect on chalkbrood in alfalfa leafcutting bee progeny. c. Measure chalkbrood spore concentration levels on bee nesting cells in commercial operations. Conduct biology studies on chalkbrood (growth and spore germination requirements, spore survival, etc.) d. Develop fermentation method for two species of fungi with potential for biological control of varroa mites in honey bees. Conduct field trials for microbial control of varroa in honey bees. 4. Enhance knowledge of native pollinators by conducting biological surveys. a. Develop prototype interactive key to the bees of the Mid-Atlantic States. Complete field (bee) surveys in Grand Staircase-Escalante National Monument. Draft revision of North American Stelis (group of parasitic bees of worldwide significance) for publication. Year 2 (2005) 1. Deliver improved pollination management systems for alfalfa seed production. a. Repeat the alfalfa leafcutting bee density study. b. Continue alfalfa leafcutting bee health survey and report to producers. c. Conduct temperature studies on summer development of phenologically distinct blue orchard bee populations and initiate pre-wintering studies. d. Continue behavioral bioassays for nest attraction using alfalfa leafcutting bees, begin tests with blue orchard bees. Perform chemical analyses of important chemical marker components. e. Complete summer development studies with the blue orchard bee. f. Complete analyses and draft manuscript on life cycle respirometry, intermediary metabolism in the alfalfa leafcutting bee under natural conditions. Monitor putative clones for life cycle expression patterns in the alfalfa leafcutting bee. Draft manuscript identifying apparent diapause gateways in the alfalfa leafcutting bee. g. Complete analysis of data and draft manuscript on UT cherry pollination. Complete year 3 of organic almond pollination demonstration in CA. Complete analysis of prototype performance, and develop modifications to be made to second generation prototype. h. Complete second year experiments on the effects of wintering temperature, parasitism and body size on winter diapause and survival for blue orchard bees and alfalfa leafcutting bees. 2. Evaluate pollination efficacies of bees selected for potential to pollinate small fruit crops and native plants used in re-vegetation programs. a. Trap-nest for Osmia atriventris for cranberry pollination. Trap-nest promising pollinators for wildflowers. Evaluate specific pollination needs of wildflowers. Evaluate trial plots. Sample and identify wildflower seed and flower pests. Publish cranberry pollen tube attrition study. 3. Develop novel disease and parasite control methods for bees. a. Refine molecular marker methods for Ascosphaera detection. Develop photo guide to disease in the alfalfa leafcutting bee. Test applications of systemic fungicides on chalkbrood levels in the alfalfa leafcutting bee and report to producers. b. Measure diurnal and distance-from-a-shelter effects on air loads of Ascosphaera spores. Measure levels of chalkbrood in feral populations of the alfalfa leafcutting bee. Continue basic biology studies on Ascosphaera aggregata. c. Conduct field trials of microbial control of varroa in honey bees. 4. Enhance knowledge of native pollinators by conducting biological surveys. a. Create a database of collection records of Osmia lignaria, O. ribifloris, and O. aglaia. Deliver web-based interactive key to the bees of the Mid-Atlantic states. Analyze faunal data as requested by Grand Staircase-Escalante Natl. Mon. Draft revision of N. Am. Dufourea for publication. Year 3 (2006) 1. Deliver improved pollination management systems for alfalfa seed production. a. Publish multi-year nest density study with the alkali bee. Estimate male pollination value, review for all bees. Implement large field trials with more optimal bee densities defined from enclosure study. Continue bee health survey and report to producers. b. Test ability to apply attractants to nesting boards. c. Complete life cycle respirometry, supercooling point, amino acid, lipid, and carbohydrate profiles for alfalfa leafcutting bee and blue orchard bee populations under natural and artificial conditions. Create cDNA library for blue orchard bee heat shock proteins via real-time-PCR for diapause and non-diapause condition. Sequence putative clones and generate DIG-labeled probes for expression assays via Northern blot. Publish data. d. Establish a more extensive contacts with almond and apple producers. Develop methods to propagate large numbers of blue orchard bees, such as on annual crops like canola. e. Determine whether blue orchard and alfalfa leafcutting bees have the ability to learn nesting cues that affect philopatry, conduct experiments with blue orchard bees to determine if mechanical stress (e.g. handling) increases dispersal rates. 2. Evaluate pollination efficacies of bees selected for potential to pollinate small fruit crops and native plants used in re-vegetation programs. a. Test trap-nested bee species for pollination efficacies. Cage pollination trial with O. atriventris on cranberry. Continue to evaluate O. aglaia pollinator efficacy on small fruits such as raspberries and blackberries. Evaluate overwintering management needs. Begin publishing wildflower breeding biologies. 3. Develop novel disease and parasite control methods for bees. a. Begin search for mycoviruses that can infect Ascosphaera. Begin testing the utility of ozone as a disinfectant for nesting materials for both honeybees and alfalfa leafcutting bees. Continue field testing the use of fungicides to control chalkbrood. b. Measure diurnal and distance-from-a-shelter effects on air loads of Ascosphaera spores. Estimate population influx of wild bees into commercial nesting blocks. Report to producers. c. Continue field tests of the fungus Metarhizium anisopliae as a microbial control of varroa testing application timing and dose. Work in collaboration with Earth Biosciences and commercial bee keepers. 4. Enhance knowledge of native pollinators by conducting biological surveys. a. Analyze faunal data as requested by Grand Staircase-Escalante Natl. Monument. Draft preliminary report on multi-year results. Initiate faunal studies at other sites. Year 4 (2007) 1. Deliver improved pollination management systems for alfalfa seed production. a. Publish carrying capacity and competition paper on the alkali bee. Continue large field trials with optimal bee densities. Analyze and evaluate the bee health survey and report to producers. b. Continue using the results of the behavioral bioassays for nest attraction compounds, apply compounds to nesting boards to test for attractants, and report to producers. c. Complete statistical treatment of data collected, draft manuscripts, and report to producers. d. Conduct field releases of bees in commercial orchards. Continue to develop mass propagation methods for the blue orchard bee. e. Conduct experiments to determine how and when learning takes place in emerging females, continue with philopatric studies. 2. Evaluate pollination efficacies of bees selected for potential to pollinate small fruit crops and native plants used in re-vegetation programs. a. Scale-up trap-nesting and production of promising pollinators. Evaluate bee performance in sundry nest substrates. Begin publishing pollinator efficiencies and conduct field trials of wildflower pollinators. Publish nesting biology of raspberry pollinator and report to producers. 3. Develop novel disease and parasite control methods for bees. a. Complete PCR processing of previous years' chalkbrood samples. b. Continue field testing the use of fungicides. Develop methods for culturing and formulating mycoviruses with activity against Ascosphaera. c. Correlate Ascosphaera spore loads on adults with chalkbrood levels in progeny. Develop quantitative PCR methods for detecting Ascosphaera spores and report to producers. d. Metarhizium anisopliae should be commercially available to beekeepers during this year. Begin tests on the applicability of using the fungus for other hive pests. 4. Enhance knowledge of native pollinators by conducting biological surveys. a. Develop prototype interactive key to the bee genera of the U.S. Analyze faunal data as requested by Grand Staircase-Escalante Natl. Mon. Draft final report. Conduct phylogenetic analysis of Megachilidae and draft manuscript. Year 5 (2008) 1. Deliver improved pollination management systems for alfalfa seed production. a. Analyze and publish carrying capacity of alfalfa and wildflower crops for the alkali bee. Continue large field trials with optimal bee densities. Analyze and evaluate large field trials with optimal bee densities. b. Analyze and evaluate bee health survey data and report to producers. c. Continue testing nesting board attractants. Analyze and evaluate application for commercial growers. Analyze chemical components of possible nest attractants. Submit final evaluation and recommendations. d. Evaluate potential for an almond pollination management program that uses the blue orchard bee. Transfer technology to CA organic orchard producers. e. Continue with bee learning experiments from FY2007, complete data analysis and write manuscript on female dispersal as impacted or not by mechanical stress. 2. Evaluate pollination efficacies of bees selected for potential to pollinate small fruit crops and native plants used in re-vegetation programs. a. Begin supplying growers with guidance and suitable pollinators for raspberries and for wildflowers. 3. Develop novel disease and parasite control methods for bees. a. Prepare a photo guide to disease in the alfalfa leafcutting bee. b. Continue field testing the use of fungicides. Continue work on mycoviruses for biological control of chalkbrood. c. Refine quantitative PCR methods. Test for Ascosphaera in environmental samples from commercial fields (pollen, soil, plants). d. Initiate bioassays testing microbial controls on parasites and predators in the alfalfa leafcutting bee. 4. Enhance knowledge of native pollinators by conducting biological surveys. a. Complete data capture of U.S. Apidae in collection. Deliver web-based interactive key to the bee genera of the U.S. Publish manuscript on multi- year faunal investigations with Grand Staircase-Escalante Natl. Mon. and other wildland sites. Conduct phylogenetic analysis of Osmiini and draft manuscript. 4a List the single most significant research accomplishment during FY 2006. This research was conducted under National Program 305 Crop Production, and directly contributes to research Component 6.0 Bees and Pollination, ARS Strategic Plan Goal 5 (Protection and Enhance the Nations Natural Resource Base and Environment). This year, we concluded a survey of the bees of Grand Staircase-Escalante National Monument, resulting in one of the largest, most complete pollinator surveys ever done. We developed the data set from continued sampling in the same plots over a period of four years. The survey results were used to address questions regarding both with-in year and between-year trends in bee population levels and phenologies. Additionally, our dataset includes a record of which flowering plant each bee was collected from, allowing for the analysis of patterns of specialization and generalization among bee species and pollinator guilds. Six hundred and forty three species were discovered within the monument, making it the richest bee landscape studied to date. This data contributes significantly to our understanding of the abundance and diversity of native pollinators within the U.S. during a time when conservation biologists and agriculturalists, alike, are concerned about our declining pollinator populations. 4b List other significant research accomplishment(s), if any. The research reported was conducted under National Program 305 Crop Production, and directly contributes to research Component 6.0 Bees and Pollination, and ARS Strategic Plan Goal 1 Enhance Economic Opportunities for Agricultural Producers, Objective 1.2 Contribute to the Efficiency of Agricultural Production Systems. Alfalfa seed producers can spend a significant portion of their production costs on pollination each year, yet the optimum bee stocking densities for this crop are unknown. This year we completed a three-year experiment where different numbers of bees were released in eight large, screen cages in an alfalfa field. Data were collected on pollination, bee success, and seed production. Preliminary analyses revealed that high bee densities are very detrimental to bee production, and that medium to low bee densities allow bees to reproduce. Knowing the appropriate number of bees to use in commercial alfalfa seed production could be of great economic value to seed producers. If the current recommendation of 40,000-60,000 bees per acre is wasteful, then growers could save money by using fewer bees, yet still obtain profitable seed yield. The research reported was conducted under National Program 305 Crop Production, and directly contributes to research Component 6.0 Bees and Pollination, and ARS Strategic Plan Goal 1 Enhance Economic Opportunities for Agricultural Producers, Objective 1.2 Contribute to the Efficiency of Agricultural Production Systems. We completed the collection of four years data on alfalfa leafcutting bee health and management factors from Idaho, Montana, Nevada, Oregon, Utah, Washington, Wyoming, and Canada, and published three reports in scientific journals. This database revealed information on bee health issues, including (1) that the health condition called "pollen balls" by growers occurs at the end of the nesting season, and is caused by several different mortality factors in very young larvae; (2) that the fungal disease "chalkbrood" is spread by emerging adult female bees, and can be controlled in part by treating the pre-emergence nest cells; and (3) that Canadian bees have more males per female (females do most of the pollination), but also have significantly higher numbers of live bees per pound, and higher adult survival during incubation and emergence. These results have helped improve our management recommendations to growers and have brought into focus what our future research needs are for improving bee health and pollination. 5. Describe the major accomplishments to date and their predicted or actual impact. Pollinator diversity and native plants. The research reported was conducted under National Program 305 Crop Production, and directly contributes to research Component 6.0 Bees and Pollination and ARS Strategic Plan Goal 5 Protection and Enhance the Nations Natural Resource Base and Environment. Little is known about the abundance and diversity of wild native bees, bees that not only contribute to crop pollination, but also maintain the diversity of our native plant species. Based on comprehensive biological surveys of six different ecosystems, including the Mojave Desert in Nevada, the Sierra Nevada Mts. and the Coast Range in California, Grand Staircase-Escalante National Monument and the Great Basin in Utah, and the Appalachia Mts. PIRU scientists have conducted one of the most complete pollinator surveys on record and assembled the largest systematic data set on local and regional bee species in the world. Six hundred and forty three bee species were discovered within the Grand Staircase-Escalante National Monument alone. PIRU scientists developed the data set from continued sampling in the same plots over a period of several years. These survey results are used to evaluate bee population levels, and times of activity. Additionally, the dataset includes a record of flowers the bees were collected from, revealing the patterns of specialization and generalization among bee species and pollination activity. This data set is also used to assess the effects of climate, landscape, and plant diversity on bee populations through time. Impact: The resulting information will aid land managers in preserving pollinator diversity, thus preserving vital pollination services and conserving rare bee species. Data from previously collected bee specimen labels have also been added to the data base, resulting in a data set of over 600,000 records. This data set will aid in the identification of alternative pollinators that could be domesticated for crop pollination. Development of a New Orchard Pollinator, the Blue Orchard Bee. The research reported was conducted under National Program 305, Crop Production, Component 6.0 Bees and Pollination, and ARS Strategic Plan Goal 1, Enhance Economic Opportunities for Agricultural Producers; Objective 1.2, Contribute to the Efficiency of Agricultural Production Systems. Considerable progress was made in developing the blue orchard bee as a fruit and nut tree pollinator. This includes: a) The publication of an 88-page manual, "How to manage the blue orchard bee as an orchard pollinator", b) The patent of a new, affordable and manageable polycarbonate-based nesting block for the blue orchard bee and the development of a related prototype emergence box, c) Publication of a six- year project demonstrating highly-significant yield increases in a commercial cherry orchard pollinated with blue orchard bees, d) The publication of research on successful methods to advance blue orchard bee emergence for almond pollination, and e) The publication or preparation of several research articles and two CRADA's on the developmental biology and rearing methods (wintering requirements, development and incubation temperature regimes, respirometry) of the blue orchard bee. Recent research has emphasized diapause induction and development, respirometry, and diapause-associated gene expression, as well as blue orchard bee toxicology related to fungicide sprays during bloom, and the use of visual and olfactory cues involved in nest location. Impact: Almonds is a lucrative, growing crop in S. California, but during the last three bloom cycles, producers experienced difficulty obtaining sufficient numbers of honey bees for pollination. In addition, the acreage of almond plantings is rapidly expanding, and the blue orchard bee could provide effective pollination for this crop. Epidemiology and control of disease in alfalfa leafcutting bees. Chalkbrood, is a disease caused by fungi in the genus Ascosphaera that attacks honey bees and non-Apis bees. Although not a serious problem in honey bees, this can be detrimental to alfalfa leafcutting bees. Alfalfa seed is an important crop in western North America and the alfalfa leafcutting bee Megachile rotunda is specifically managed for alfalfa pollination. Chalkbrood disease is such a problem in alfalfa leafcutting bees that U. S. alfalfa seed producers and growers must purchase and import about 50% of their leafcutting bees annually from Canada, where the disease is thought to be less prevalent. ARS scientists collected bee samples from alfalfa seed producers and commercial pollination providers from different growing regions, and during different times of year, in an effort to understand the epidemiology of chalkbrood disease, and other causes of bee mortality. Results indicated that non-chalkbrood mortality of immature bees incubated by producers was disproportionately higher than laboratory-incubated bees, which led to a review of existing management practices. ARS scientists compared the different on-farm methods for disinfecting bee nesting boards and other equipment used to rear bees and concluded that these management practices were inadequate to control chalkbrood. Scientists subsequently screened a series of fungicides for efficacy in controlling chalkbrood and found iprodione to be both effective and of low toxicity to the bee larvae. ARS research results were transmitted in a timely fashion through trade bulletins and meetings of the alfalfa seed grower groups. Impact: A better understanding of the epidemiology of chalkbrood disease has led to improved bee management techniques and increased production of alfalfa leafcutting bees. This has a positive impact for growers and producers, as pollination currently accounts for 20% of alfalfa seed production costs, so any reductions can significantly improve the overall economics of production. Restoration of the Nations Rangelands. The research reported here was conducted under National Program 305 Crop Production, and directly contributes to research Component 6.0 Bees and Pollination. Research activities are further related to ARS Strategic Plan Goal 2 Support Increased Economic Opportunities and Improved Quality of Life in Rural America, and Goal 5 Protection and Enhance the Nations Natural Resource Base and Environment, Objective 5.1 Provide Science Based Knowledge and Education to Improve the Management of Forest, Rangelands, and Pastures. Federal, state and private land managers in the western U.S. are critically in need of affordable native plant seed for revegetation of rangeland, and other wildlands, due to the destruction of native plant communities from invasive weeds (e.g. fireweed), natural disasters (e.g. wildfires), and destructive human activities. The U.S. government annually seeks tons of seed from dominant species of native wildflowers and herbaceous plants, but the exorbitant price of hand-collected wild seed stymies that effort. Farm production methods for wildflowers could provide an affordable seed source, and in the quantities necessary for revegetation efforts. ARS scientists, with funds from BLM and the Forest Service, found that bees are required for pollination of nearly all of the 15 target wildflower species desired for restoration and rehabilitation of Intermountain plant communities. ARS scientists have focused on two plant species, basalt milkvetch and royal penstemon, and found that they attract potentially manageable pollinators (e.g. mason bees and bumble bees), and several seed growers now have plants in trial production. Impact: This research has been critical to making wild-flower seed production amenable to current farming practices, and has contributed a valuable new crop to seed producers. 6. What science and/or technologies have been transferred and to whom? When is the science and/or technology likely to become available to the end- user (industry, farmer, other scientists)? What are the constraints, if known, to the adoption and durability of the technology products? Alfalfa leafcutting bee health improvement recommendations were delivered at local and regional alfalfa seed growers meetings, giving both oral and poster presentations, and a farmer-assisted demonstration trial on chalkbrood control was initiated this year in the Columbia Basin area of Washington. The Western Branch of the American Organization of Seed Certification Analysts met in Reno, Nevada, April, 2006 to discuss their concerns, strategies and policies regarding gene flow in alfalfa seed, especially with regard to transgenic varieties. PIRU provided advice on known and potential bee-mediated gene flow concerns from transgenic alfalfa, greatly assisting the organization in developing practical, yet effective, policies for growers. Public concerns regarding possible declines in pollinator populations has lead the National Academy of Sciences (NAS) to evaluate the status of pollinators through the National Research Council Workshop Committee on Status of Pollinators. This year, PIRU scientists were asked to present data and information on bee diversity and use in the western United States. The NAS will present a report on the current state of knowledge of pollination services and identify knowledge gaps. The opening of the Grand Staircase-Escalante National Monument Visitors' Center was held in June, 2006, and PIRU presented bee diversity survey results at the concurrent Science Symposium. In addition, the monument's new visitors center includes a large display on pollinators and their role in ecosystem function, a display based largely on information provided by PIRU. Additionally, land managers in the monument now have a complete and annotated list of bee species within their boundaries, and a database including information on the location, date of capture, floral host, and habitat type for each of the 90,000 specimens collected. PIRU research is often cited in the popular press, for example, PIRU research on alkali bees was extensively cited in an article about bee conservation for agriculture in a publication of the Natural Resources Defense Council, "The Vanishing", OnEarth Vol 28 (2). 2006; several newspaper articles have covered bee decline issues and PIRU microbial control of varroa mite research. Our methods for blue orchard bee management have also been used by the State of Utah to help in the pollination and re-establishment of a rare Phacelia plant. 7. List your most important publications in the popular press and presentations to organizations and articles written about your work. (NOTE: List your peer reviewed publications below). Cane, J.H. 2006. Honeybeesfriend or foe? Refuge Update (USFWS). 3(2): 16-17. Eardley, C., D. Roth, J. Clarke, S. Buchmann, & B. Gemmill, editors. 2006. Pollinators and pollination: A resource book for policy and practice. African Pollinator Initiative, Pretoria, South Africa, 77 pp. Lipper, Don. 2005. Is there some green in blue mason bees? A varroa free alternative for the almond industry? Pacific Nut Producer

Impacts
(N/A)

Publications

Pitts Singer, T., James, R.R. 2005. Emergence success and sex ratio of commercial alfalfa leafcutting bees, from the United States and Canada. Journal of Economic Entomology. 98(6):1785-1790.
James, R.R., Skinner, J.S. 2005. PCR diagnostic methods for Ascosphaera infections in bees. Journal of Invertebrate Pathology. 90:(2):98-103.
Cane, J.H. 2006. The Logan Beemail Shelter: a practical, portable unit for managing cavity-nesting agricultural pollinators. American Bee Journal. 146(7):611-613.
Cane, J.H., Minckley, R., Kervin, L., Roulston, T., Williams, N.M. 2006. Complex responses within a desert bee guild (Hymenoptera: Apiformes) to urban habitat fragmentation. Ecological Applications. 16(2):632-644.
Cane, J.H. 2005. Pollination potential of the bee Osmia aglaia for cultivated raspberries and blackberries (Rubus: Rosaceae). Hortscience. 40:1705-1708.
Sipes, S.D., Tepedino, V. 2005. Pollen-host specificity and evolutionary patterns of host switching in a clade of specialist bees (Apoidea: Diadasia). Biological Journal of the Linnean Society, London. 86:487-505.
Toler, T.R., Evans, E.W., Tepedino, V.J. 2005. Pan-trapping for bees (Hymenoptera: Apiformes) in Utah's west desert: the importance of color diversity. Pan Pacific Entomology. 81(3/4):103-113.
Bartholomew, C.S., Prowell, D., Griswold, T.L. 2006. An annotated checklist of bees (hymenoptera: apoidea) in longleaf pine savannas on the western edge of the east gulf coastal plain. Journal of Kansas Entomological Society. 79:184-198
Bosch, J., Kemp, W.P., Trostle, G.E. 2006. Bee population returns and cherry yields in an orchard pollinated with Osmia lignaria (Hymenoptera: Megachilidae). Journal of Economic Entomology. 99(2):408-413.
Guedot, C., Pitts Singer, T.L., Buckner, J.S., Bosch, J., Kemp, W.P. 2006. Olfactory cues and nest recognition in the solitary bee Osmia lignaria. Physiological Entomology. 31:110-119.
Guedot, C., Bosch, J., Kemp, W.P., James, R.R. 2006. Effects of three- dimensional and color patterns on nest location and progeny mortality in alfalfa leafcutting bee (Hymenoptera: Megachilidae). Journal of Environmental Entomology. 99(3):626-633.
Guedot, C., Bosch, J., Kemp, W.P. 2005. The relative importance of vertical and horizontal visual cues in nest location by Megachile rotundata. Journal of Apicultural Research. 44(3):109-115.
Kemp, W.P., Bosch, J. 2005. Effect of temperature on Osmia lignaria (hymenoptera:megachilidae) prepupa-adult development, survival, and emergence. Journal of Economic Entomology. 98(6):1917-1923.
Meikle, W.G., Holst, N., Mercadier, G., Derouane, F., James, R.R. 2006. Evaluation of Beauveria bassiana (Balsamo) Vuillemin (Deuteromycota: Hyphomycetes) strains isolated from varroa mites in southern Franc. Journal of Apicultural Research.

Progress 10/01/04 to 09/30/05

Outputs
1. What major problem or issue is being resolved and how are you resolving it (summarize project aims and objectives)? How serious is the problem? What does it matter? Pollinators are critical elements of ARS National Program 305, Crop Production Bees and Pollination. Without effective pollination, insect- pollinated crops such as alfalfa seed, almonds, sweet cherries, berries, apples, melons, and tomatoes are failures. The annual combined value of U. S. crops requiring cross-pollination is over $10 billion; the bulk of this is accomplished using the honey bee. However, some crops are not adequately serviced by honeybees, such as alfalfa seed and blueberries, and others have suffered recently because the number of honey bee colonies in the U.S. has declined drastically due to high overwintering mortality associated with parasites such as Varroa and tracheal mites. U. S. beekeeping associations estimated that 50% of commercial hives were lost during the winter of 2004-05, and the price of hive rentals for pollination nearly doubled the following spring due to an ensuing shortage. The Pollinating Insect-Biology, Management, & Systematics Research Unit (PIRU) Project Plan was initiated on 1 October 2003, with the overriding goal to improve the profitability of U.S. production of insect-pollinated crops by diversifying our agricultural pollinator portfolio to improve the yield and quality of product. To achieve this goal, priority will be given to: 1) Delivering improved pollination management systems for alfalfa seed production using the alfalfa leafcutting bee and the alkali bee, and delivering a new pollination management system for orchards using the blue orchard bee; 2) Evaluating pollination efficacies of confined and free-flying populations of select new bee pollinators for small fruit crops and forbs in demand for re-vegetation, and developing practical management protocols to encourage adoption of the more promising species as custom pollinators; 3) Developing effective control options and delivery systems for the management of chalkbrood, other diseases, parasites, and predators in commercial-scale pollinator populations; and, 4) Conducting biological surveys, establishing monitoring programs, and expanding current knowledge of bee systematics in natural and agricultural systems. Anticipated products of the research include pollination and bee management handbooks, patented nesting and shelter designs, and peer-reviewed publications. The major customers of the research outlined in this Project Plan are the U.S. alfalfa seed producers (NW Alfalfa Seed Growers Association, and The Alfalfa Alliance), who rely heavily on the superior performance of the alfalfa leafcutting bee and the alkali bee for pollination each year. This group enjoys a close relationship with the PIRU, and their members participate directly in the planning and execution of ongoing research. Additionally, PIRU research results are in demand by commercial seed and fruit producers, particularly organic tree fruit and nut producers in the case of the blue orchard bee, as well as custom pollination providers, federal, state, and private land managers, amateur gardeners, and non- profit non-governmental organizations. Potential benefits expected from attaining the objectives outlined in the current Project Plan include increased productivity and profitability through the identification and management of superior, affordable pollinators, and improved sustainability through the diversification of a pollinator portfolio for U.S. Agriculture. 2. List the milestones (indicators of progress) from your Project Plan. Objective 1 Deliver improved pollination management systems for alfalfa seed production using the alfalfa leafcutting bee and the alkali bee, and a new pollination management system for orchards using the blue orchard bee. Sub-Objective 1.1: For the two primary alfalfa pollinators, the alkali bee and the alfalfa leafcutting bee, we will determine cost-effective, optimal stocking densities for pollinators that maximize seed production while minimally compromising bee reproduction. FY2004 Milestones: Complete alkali bee brood provision analyses and draft provision sugar methods manuscript. Conduct alfalfa leafcutting bee density enclosure studies and analyze data. Collect samples and information for regional bee health survey. FY2005 Milestones: Analyze data and publish provision mass pollen/sugar paper. Repeat bee density study with different densities and analyze data. Continue bee health survey and report to producers. FY2006 Milestones: Publish multi-year nest density study with the alkali bee. Estimate male pollination value, review for bees. Implement large field trials with more optimal bee densities defined from enclosure study. Continue bee health survey and report to producers. FY2007 Milestones: Publish carrying capacity and competition paper on the alkali bee. Continue large field trials with optimal bee densities. Continue bee health survey and report to producers. FY2008 Milestones: Analyze and publish carrying capacity of alfalfa and wildflower crops for the alkali bee. Analyze and evaluate large field trials with optimal bee densities. Analyze and evaluate bee health survey data and report to producers. Sub-Objective 1.2: To better enhance nesting of the alfalfa leafcutting bee in clean, artificial nesting boards and to better ensure field fidelity of bees released for pollination, we will investigate the origins of cues that attract bees to previously used nesting boards. FY2004 Milestone: Initiate behavioral bioassays for nest attraction using alfalfa leafcutting bees. Perform chemical analyses of important attractants. FY2005 Milestones: Continue tests with alfalfa leafcutting bees; begin tests with blue orchard bees. Perform chemical analyses of important components. FY2006 Milestone: Test ability to apply attractants to nesting boards. FY2007 Milestones: Field tests of nesting board attractants in all species where applicable and report to producers. FY2008 Milestones: Continue testing nesting board attractants. Analyze and evaluate application for commercial growers. Submit final evaluation and recommendations. Sub-Objective 1.3: Improve summer and pre-wintering temperature regimes for the commercial (and artificial) management of blue orchard bee populations. FY2004 Milestone: Conduct temperature studies on summer development of phenologically distinct blue orchard bee populations and initiate pre-wintering studies. FY2005 Milestone: Complete summer development studies with the blue orchard bee. FY2006 Milestone: Complete pre-wintering studies with the blue orchard bee. FY2007 Milestone: Complete statistical treatment of data collected, draft manuscripts, and report to producers. FY2008 Milestone: Submit final evaluation and recommendation of optimal summer and pre-wintering temperature regimes for the blue orchard bee. Sub-Objective 1.4: Support improvements in pre-wintering and wintering temperature regimes for the commercial (and artificial) management of the alfalfa leafcutting bee and the blue orchard bee through improving our understanding of intermediary metabolic processes and diapause gateways. FY2004 Milestone: Complete life cycle respirometry, supercooling point, amino acid, lipid, and carbohydrate profiles for natural alfalfa leafcutting bee populations. Identify genes to signal change from diapause (hibernation-like state in insects) to post-diapause development and compare their expression levels during the winter through springtime development period. FY2005 Milestone: Complete analyses and draft manuscript on life cycle respirometry, intermediary metabolism in the alfalfa leafcutting bee under natural conditions. Monitor putative clones for life cycle expression patterns in the alfalfa leafcutting bee. Draft manuscript identifying apparent diapause gateways in the alfalfa leafcutting bee. FY2006 Milestone: Complete life cycle respirometry, supercooling point, amino acid, lipid, and carbohydrate profiles for alfalfa leafcutting bee and blue orchard bee populations under natural and artificial conditions. Create cDNA library for blue orchard bee HSPs via RT-PCR for diapause and non-diapause condition. Sequence putative clones and generate DIG-labeled probes for expression assays via Northern blot. FY2007 Milestone: Complete analyses and draft manuscripts on life cycle respirometry, intermediary metabolism for alfalfa leafcutting bee and blue orchard bees under natural and artificial conditions. Monitor putative clones for life cycle expression patterns in the blue orchard bee. Draft manuscript identifying diapause gateways in the blue orchard bee. FY2008 Milestone: Submit final comparison and evaluation of the physiological basis of optimal summer, pre-wintering, and wintering temperature regimes for the alfalfa leafcutting bee and the blue orchard bee. Sub-Objective 1.5: Demonstrate the blue orchard bee and new nesting block design in selected early-spring orchard crops. FY2004 Milestone: Complete year 6 (of 6) in cherry pollination demonstration in UT. Complete year 2 of organic almond pollination demonstration in CA. Complete year 1 of cherry pollination demonstration in MT. Complete initial field testing of current nest block prototype in UT pear-apple orchards. FY2005 Milestone: Complete analysis of data and draft manuscript on UT cherry pollination. Complete year 3 of organic almond pollination demonstration in CA. Complete analysis of prototype performance, and develop modifications to be made to second generation prototype. FY2006 Milestone: Submit final evaluation and recommendation of optimal cherry pollination management program using the blue orchard bee. Complete year 4 of organic almond pollination demonstration in CA. Complete testing of second generation prototype in UT pear-apple orchards. FY2007 Milestone: Complete year 5 of organic almond pollination demonstration in CA. Complete analysis of data and draft manuscript on organic almond pollination. Prepare final evaluation and recommendation for transfer to mass market. FY2008 Milestone: Submit final evaluation and recommendation of optimal almond pollination management program using the blue orchard bee. Transfer technology to CA organic orchard producers. Sub-Objective 1.6: Improve the establishment of artificially managed blue orchard bee populations used for commercial pollination by minimizing the dispersal of pre-nesting females. FY2004 Milestone: Complete first year experiments on wintering temperature, parasitism and body size. Select progeny of dispersing vs. non-dispersing females for field trials in FY2005 and beyond. FY2005 Milestone: Complete second year experiments on wintering temperature, parasitism and body size. Complete first year experiment with philopatric females. FY2006 Milestone: Complete second year experiment with philopatric females. FY2007 Milestone: Complete third year experiment with philopatric females.FY2008 Milestone: Complete data analysis and write manuscript on female dispersal. Submit final evaluation and recommendation of optimal blue orchard bee population management. Objective 2 Evaluate pollination efficacies of confined and free- flying populations of select new bee pollinators for small fruit crops and regional forbs in demand for re-vegetation, and develop practical management protocols to encourage adoption of the more promising species as custom pollinators. FY2004 Milestone: Refine nest substrate options for Osmia aglaia in raspberries. Confirm basic breeding biologies of focal wildflower seed species. Begin sampling pollinator guild of wildflowers. Grow Osmia ribifloris and address dispersal problems. Publish cranberry pollinator efficacy study. FY2005 Milestone: Trap-nest for O. atriventris for cranberry pollination. Trap-nest promising pollinators for wildflowers. Evaluate specific pollination needs of wildflowers. Evaluate trial plots. Sample, ID wildflower seed and flower pests. Publish cranberry pollen tube attrition study. FY2006 Milestone: Test trap-nested bee species for pollination efficacies. Cage pollination trial with O. atriventris on cranberry. Evaluate pollinator efficacy on highbush blueberries. Evaluate overwintering management and needs. Begin publishing wildflower breeding biologies. Try O. ribifloris free-flight in blueberries. FY2007 Milestone: Scale-up trap-nesting and production of promising pollinators. Evaluate bee performance in sundry nest substrates. Begin publishing pollinator efficiencies and conduct field trials of wildflower pollinators. Publish nesting biology of raspberry pollinator and report to producers. FY2008 Milestone: Begin supplying growers with guidance and suitable pollinators for raspberries and for wildflowers. Objective 3 Develop novel control options and delivery systems for the management of chalkbrood disease, other diseases, parasites, and predators in commercial-scale pollinator populations. Sub-Objective 3.1: Determine the abundance and diversity of chalkbrood in commercial fields, and conduct a survey of bee larvae to determine the frequency of infection for each pathogen species present. FY2004 Milestone: Sample alfalfa leafcutting bee larvae from commercial fields, and assess levels of chalkbrood infection. Refine molecular marker methods for chalkbrood detection. FY2005 Milestone: Sample alfalfa leafcutting bee larvae from commercial fields, and assess levels of chalkbrood infection. Refine molecular marker methods for Ascosphaera detection. Develop photo guide to disease in the alfalfa leafcutting bee. FY2006 Milestone: Conduct survey of chalkbrood prevalence in the alfalfa leafcutting bee from commercial alfalfa fields and report to producers. FY2007 Milestone: Conduct survey of chalkbrood prevalence in the alfalfa leafcutting bee from commercial alfalfa fields and report to producers. FY2008 Milestone: Conduct survey of chalkbrood prevalence in the alfalfa leafcutting bee from commercial alfalfa fields. Draft manuscript on multi- year study and report to producers. Sub-Objective 3.2: Develop new control methods for chalkbrood, focusing on chemical fungicides, sanitation methods, and microbial control strategies. FY2004 Milestone: Apply fungicides to loose cells of the alfalfa leafcutting bee and test effect on chalkbrood in alfalfa leafcutting bee progeny. FY2005 Milestone: In commercial fields, test applications of systemic fungicides on chalkbrood levels in the alfalfa leafcutting bee and report to producers. FY2006 Milestone: Begin search for mycoviruses that can infect Ascosphaera. FY2007 Milestone: Develop method for culturing and formulating mycoviruses with activity against Ascosphaera. FY2008 Milestone: Conduct small scale field tests of mycoviruses for biological control of chalkbrood and report to producers. Sub-Objective 3.3: Determine the epizootiology of chalkbrood, and in particular, determine the major means by which the disease is spread and maintained in a bee population. FY2004 Milestone: Measure chalkbrood spore concentration levels on bee nesting cells in commercial operations. Conduct biology studies on chalkbrood (growth and spore germination requirements, spore persistence, etc.). FY2005 Milestone: Measure diurnal and distance-from-a-shelter effects on air loads of Ascosphaera spores. Measure levels of chalkbrood in feral populations of the alfalfa leafcutting bee. Continue basic biology studies on Ascosphaera aggregata. FY2006 Milestone: Measure diurnal and distance-from-a-shelter effects on air loads of Ascosphaera spores. Estimate population influx of wild bees into commercial nesting blocks. Report to producers. FY2007 Milestone: Correlate Ascosphaera spore loads on adults with chalkbrood levels in progeny. Develop quantitative PCR methods for detecting Ascosphaera spores and report to producers. FY2008 Milestone: Refine quantitative PCR methods. Test for Ascosphaera in environmental samples from commercial fields (pollen, soil, plants). Sub-Objective 3.4: Develop microbial control strategies for controlling predators and parasites in bees. FY2004 Milestone: Develop fermentation method for two species of fungi with potential for biological control of varroa mites in honey bees. Conduct field trials for microbial control of varroa in honey bees. FY2005 Milestone: Conduct field trials of microbial control of varroa in honey bees. Develop a laboratory population of the checkered flower beetle. FY2006 Milestone: Develop bioassays for the checkered flower beetle, test fungi for pathogenicity. Conduct non-target tests of the candidate fungi on bees. FY2007 Milestone: Field test microbial control strategy for predatory beetles in commercial fields and report to producers. FY2008 Milestone: Field test microbial control strategy for predatory beetles in commercial fields and report to producers. Objective 4 Enhance knowledge of native pollinators by conducting biological surveys, establishing monitoring programs, and expanding knowledge of bee systematics in agricultural and natural systems. Sub-Objective 4.1: Data capture of specimen records of Megachilidae will be collected in support of Objectives 1 & 2. FY2004 Milestone: Develop prototype interactive key to the bees of the Mid-Atlantic States. FY2005 Milestone: Capture collection records of Osmia lignaria, O. ribifloris, and O. aglaia. Deliver web-based interactive key to the bees of the Mid- Atlantic states. FY2006 Milestone: Complete data capture of U.S. Megachilidae in BBSL collection. FY2007 Milestone: Develop prototype interactive key to the bee genera of the U.S. FY2008 Milestone: Complete data capture of U.S. Apidae in collection. Deliver web-based interactive key to the bee genera of the U.S. Sub-Objective 4.2: A multi-year (bee) faunal survey of Grand Staircase- Escalante National Monument initiated in 2000 will continue, with plans for a final field season in FY2004. FY2004 Milestone: Complete field (bee) surveys in Grand Staircase- Escalante National Monument. FY2005 Milestone: Analyze faunal data as requested by Grand Staircase-Escalante Natl. Mon. FY2006 Milestone: Analyze faunal data as requested by Grand Staircase-Escalante Natl. Mon. Draft preliminary report on multi-year results. FY2007 Milestone: Analyze faunal data as requested by Grand Staircase-Escalante Natl. Mon. Draft final report. FY2008 Milestone: Publish manuscript on multi-year faunal investigations with Grand Staircase-Escalante Natl. Mon. collaborators. Sub-Objective 4.3: Systematic studies in support of Objectives 1 & 2 will concentrate on candidate pollinator-rich Megachilidae. FY2004 Milestone: Draft revision of North American Stelis (group of parasitic bees of worldwide significance) for publication. FY2005 Milestone: Draft revision of N. Am. Dufourea for publication. FY2006 Milestone: Draft revision of bright metallic Osmia for publication. FY2007 Milestone: Conduct phylogenetic analysis of Megachilidae and draft manuscript. FY2008 Milestone: Conduct phylogenetic analysis of Osmiini and draft manuscript. 3a List the milestones that were scheduled to be addressed in FY 2005. For each milestone, indicate the status: fully met, substantially met, or not met. If not met, why. 1. Sub-Objective 1.1: For the two primary alfalfa pollinators, the alkali bee and the alfalfa leafcutting bee, we will determine cost-effective, optimal stocking densities for pollinators that maximize seed production while minimally compromising bee reproduction. FY2005 Milestones: Analyze data and publish provision mass pollen/sugar paper. Repeat bee density study with different densities and analyze data. Continue bee health survey and report to producers. FY2005 Progress: A quantitative evaluation of sugar quality in bee brood provisions continues. Completed a publication on the interaction between bee population dynamics, flowering, and pollination rates for alfalfa and the alfalfa leafcutting bee. The bee health survey has been expanded to include 13 alfalfa leafcutting bee managers in seven northwestern states, up from 7 the year before, and we will complete the third and final year this year. We have been soliciting these producers for bee samples four times a year, and collect information on their bee and alfalfa management methods. We have published three papers from data collected so far, and are in the process of conducting further data analysis. New questions for experimentation are emerging from such analyses. Milestone Fully Met 2. Sub-Objective 1.2: To better enhance nesting of the alfalfa leafcutting bee in clean, artificial nesting boards and to better ensure field fidelity of bees released for pollination, we will investigate the origins of cues that attract bees to previously used nesting boards. FY2005 Milestones: Continue tests with alfalfa leafcutting bees; begin tests with blue orchard bees. Perform chemical analyses of important components. FY2005 Progress: Behavioral bioassays using a Y-tube apparatus was used to test bee attraction to nest materials. These studies were completed for alfalfa leafcutting bees and blue orchard bees, and a manuscript has been prepared. Bioassays and field trials were initiated using touch responses. Observation hives and chemical analyses were conducted to detect chemical markers used by nesting females, and a manuscript has been prepared. Milestone Fully Met 3. Sub-Objective 1.3: Improve summer and pre-wintering temperature regimes for the commercial (and artificial) management of blue orchard bee populations. FY2005 Milestone: Complete summer development studies with the blue orchard bee. FY2005 Progress: Studies to determine the degree- day requirements for the blue orchard bee were completed. Bee respiratory activity, ovarian development, and survival to adulthood were compared for three different prewintering conditions. In addition, bees from a warm climate (S. California) were compared with bees from a cold climate (N. Utah)to determine the effect of phenotype on summer development. This research is part of a PhD dissertation in collaboration with the University of Bologna, Italy. Milestone Fully Met 4. Sub-Objective 1.4: Support improvements in pre-wintering and wintering temperature regimes for the commercial (and artificial) management of the alfalfa leafcutting bee and the blue orchard bee through improving our understanding of intermediary metabolic processes and diapause gateways. FY2005 Milestone: Complete analyses and draft manuscript on life cycle respirometry, intermediary metabolism in the alfalfa leafcutting bee under natural conditions. Monitor putative clones for life cycle expression patterns in the alfalfa leafcutting bee. Draft manuscript identifying apparent diapause gateways in the alfalfa leafcutting bee. FY2005 Progress: Completed publications on (1) the effect of pre- wintering and wintering temperature regimes on weight loss, survival, and emergence time in the mason bee, (2) overwintering lipid content in the alfalfa leafcutting bee, and (3) temporal variation in gene expression in the alfalfa leafcutting bee. Milestone Fully Met 5. Sub-Objective 1.5: Demonstrate the blue orchard bee and new nesting block design in selected early-spring orchard crops. FY2005 Milestone: Complete analysis of data and draft manuscript on UT cherry pollination. Complete year 3 of organic almond pollination demonstration in CA. Complete analysis of prototype performance, and develop modifications to be made to a second generation prototype. FY2005 Progress: Prepared a manuscript describing the results of UT cherry pollination by the blue orchard bee. Completed a CRADA with Dadant on the propagation of blue orchard bees in cherry and apple orchards in MT, demonstrating that nesting blocks can be moved during the nesting period. Completed another year of field testing the release of blue orchard bees for pollination in a California organic almond orchard. Discovered that the CA phenotype is better adapted to long summers than is the UT phenotype. Milestone Fully Met 6. Sub-Objective 1.6: Improve the establishment of artificially managed blue orchard bee populations used for commercial pollination by minimizing the dispersal of pre-nesting females. FY2005 Milestone: Complete second year experiments on wintering temperature, parasitism and body size. FY2005 Progress: In the first year, body size was determined not to be related to dispersal. A comparison was made between dispersal of progeny from bees that dispersed the first year and those that did not, thus completing the second year of the study. Again, no correlation was found. We will now test if handling (mechanical stress and removing the nest) affects bee tendency to disperse. Milestone Fully Met 7. Objective 2 Evaluate pollination efficacies of confined and free-flying populations of select new bee pollinators for small fruit crops and regional forbs in demand for re-vegetation, and develop practical management protocols to encourage adoption of the more promising species as custom pollinators. FY2005 Milestone: Trap-nest for Osmia atriventris for cranberry pollination. Trap-nest promising pollinators for wildflowers. Evaluate specific pollination needs of wildflowers. Evaluate trial plots. Sample Idaho wildflower seed and flower pests. Publish cranberry pollen tube attrition study. FY2005 Progress: The native bee Osmia aglaia was found to be an effective raspberry pollinator, and a manuscript was prepared and is in press. Osmia aglaia set fruits equivalent in size and shape to fruit set by honeybee pollination, even when Osmia aglaia populations were stocked at a much lower density. Osmia aglaia populations increased 2-3 times in one season in commercial fields when provided with nesting b1ocks slightly modified from those manufactured for alfalfa leafcutting bees. Field plots were used to assess pollination needs of select wildland flowers, and these were found to require pollinators for good seed production. Bees responsible for pollination of these plants were identified. A collaborator is propagating a new line of cranberries, so cranberry research has been delayed until next year. Milestone Substantially Met 8. Sub-Objective 3.1: Determine the abundance and diversity of chalkbrood in commercial fields, and conduct a survey of bee larvae to determine the frequency of infection for each pathogen species present. FY2005 Milestone: Sample alfalfa leafcutting bee larvae from commercial fields and assess levels of chalkbrood infection. Refine molecular marker methods for Ascosphaera detection. Develop photo guide to disease in the alfalfa leafcutting bee. FY2005 Progress: A third year of the bee health survey was conducted, and the fourth year started, allowing for a good assessment of chalk brood infection levels in alfalfa seed production areas. The PCR methods for Ascosphaera detection have been fully developed, as have species-specific markers, and the methods are in press. The species identifications for Ascosphaera have been completed for samples from ten farms. A photo guide was not completed because not all the diseases have been identified. Milestone Substantially Met 9. Sub-Objective 3.2: Develop new control methods for chalkbrood, focusing on chemical fungicides, sanitation methods, and microbial control strategies. FY2005 Milestone: In commercial fields, test applications of systemic fungicides on chalkbrood levels in the alfalfa leafcutting bee and report to producers. FY2005 Progress: A large scale, replicated, field trial involving four growers was completed, and fungicide applications were found to decrease chalk brood levels. A second field trial, using a range of doses, has been initiated to determine if better control can be achieved. Also, a new, biologically based fungicide was identified as active against chalkboard but safe for the bees. Milestone Fully Met 10. Sub-Objective 3.3: Determine the epizootiology of chalkbrood, and in particular, determine the major means by which the disease is spread and maintained in a bee population. FY2005 Milestone: Air samples for Ascosphaera spores were made using a personal air sampler and all-glass impingers. Samples were taken over 12 hours from two different shelters. A sufficient number of spores were collected to be able to visuall quantify them. However, we have determined that the most prevalent means of disease transmission has been the loose nest cell material, and not the air around the shelters. Milestone Fully Met 11. Sub-Objective 3.4: Develop microbial control strategies for controlling predators and parasites in bees. FY2005 Milestone: Conduct field trials of microbial control of varroa in honey bees. Develop a laboratory population of the checkered flower beetle. FY2005 Progress: Field trials testing the efficacy of a fungal biological control agent, Metarhizium anisopliae, on varroa mites continued. A commercial supplier of the fungus is now collaborating with this research, and we found their strain of the fungus to infect and kill varroa mites, but not the bees. A field trial in bee hives is currently underway. Due to the success of the varroa control project, and the time required for these experiments, checkered flower beetle research has been suspended. Milestone Substantially Met 12. Sub-Objective 4.1: Data capture of specimen records of Megachilidae will be collected in support of Objectives 1 & 2. FY2005 Milestone: Capture collection records of Osmia lignaria, O. ribifloris, and O. aglaia. Deliver web-based interactive key to the bees of the Mid-Atlantic states. FY2005 Progress: An interactive key to six genera of Mid-Atlantic bees has been completed and is located on the web at www.discoverlife.org. Interactive keys for additional bee genera are under development. In addition, interactive keys to the genera of the bee family Megachilidae have been completed for North America and East Africa. Milestone Fully Met 13. Sub-Objective 4.2: A multi-year bee faunal survey of Grand Staircase- Escalante National Monument initiated in 2000 will continue, with plans for a final field season in FY2004. FY2005 Milestone: Analyze faunal data as requested by Grand Staircase-Escalante Natl. Mon. FY2005 Progress: Precipitation was unusually high during the winter of 2004-2005, so bee populations were sampled an additional year to determine the impact of that rainfall. Fifty-six genera and 638 species of bees have been described from the first field surveys, which represents bee diversity greater than anywhere else described in North America. Bees showed a bimodal distribution during the season, with peaks in late spring and again in late summer. This project has resulted in the initiation of three new surveys: Dugway Proving Grounds, UT; Clarke, Co, NV; and Yosemite National Park, CA; and sampling was conducted this year. Milestone Fully Met 14. Sub-Objective 4.3: Systematic studies in support of Objectives 1 & 2 will concentrate on candidate pollinator-rich Megachilidae. FY2005 Milestone: Draft revision of N. Am. Dufourea for publication. FY2005 Progress: A draft revision of Dufourea has been initiated. Milestone Fully Met 3b List the milestones that you expect to address over the next 3 years (FY 2006, 2007, and 2008). What do you expect to accomplish, year by year, over the next 3 years under each milestone? Objective 1 Deliver improved pollination management systems for alfalfa seed production using the alfalfa leafcutting bee and the alkali bee, and a new pollination management system for orchards using the blue orchard bee. Sub-Objective 1.1: For the two primary alfalfa pollinators, the alkali bee and the alfalfa leafcutting bee, we will determine cost-effective, optimal stocking densities for pollinators that maximize seed production while minimally compromising bee reproduction. FY2006 Milestones: Publish multi-year nest density study with the alkali bee. Estimate male pollination value, review for bees. Implement large field trials to better define the optimal bee densities derived from enclosure study. Conclude the bee health survey and report to producers. FY2007 Milestones: Publish carrying capacity and competition paper on the alkali bee. Continue large field trials with optimal bee densities, conduct an economic analysis. Analyze and evaluate bee health survey data and report to producers. FY2008 Milestones: Analyze and publish carrying capacity of alfalfa and wildflower crops for the alkali bee. Analyze and evaluate large field trials with optimal bee densities, develop an economic model to assess optimal densities based on current seed and bee prices. Sub-Objective 1.2: To better enhance nesting of the alfalfa leafcutting bee in clean, artificial nesting boards and to better ensure field fidelity of bees released for pollination, we will investigate the origins of cues that attract bees to previously used nesting boards. FY2006 Milestone: Test ability to apply attractants to nesting boards. FY2007 Milestone: Conduct field tests of nesting board attractants in all species where applicable and report to producers. FY2008 Milestones: Continue testing nesting board attractants. Analyze and evaluate application for commercial growers. Submit final evaluation and recommendations. Sub-Objective 1.3: Due to (1) the transfer of the principal investigator, (2) the fact that most of the goals of this objective have already been met, and (3) a new shortage of honeybees for almond pollination, this objective will terminate after 2006, and more emphasis will be placed Sub- Objective 1.5 and on technology transfer for the commercial (and artificial) management of blue orchard bee populations. FY2006 Milestone: Complete statistical treatment of data collected, and draft manuscripts. Sub-Objective 1.4: As with Sub-Objective 1.3, this objective will terminate after 2006, and more emphasis will be placed Sub-Objective 1.5 and on technology transfer for the commercial (and artificial) management of blue orchard bee populations, due to (1) the transfer of the principal investigator, (2) the fact that most of the goals of this objective have already been met, and (3) a new shortage of honeybees for almond pollination. FY2006 Milestone: Complete analysis of life cycle respirometry, supercooling point, amino acid, lipid, and carbohydrate profiles for alfalfa leafcutting bee and blue orchard bee populations under natural and artificial conditions, and report the analysis. Sub-Objective 1.5: Demonstrate the effectiveness of the blue orchard bee for early-spring orchard crops such as almonds or apples. FY2006 Milestone: Establish a more extensive contact with almond or apple producers. Develop methods to propagate large numbers of blue orchard bees, such as on annual crops like canola. FY2007 Milestone: Conduct field releases of bees in commercial orchards. Continue to develop mass propagation methods for the blue orchard bee. FY2008 Milestone: Submit final evaluation and recommendation of optimal almond pollination management program using the blue orchard bee. Report to producers and bee managers. Sub-Objective 1.6: Improve the establishment of artificially managed blue orchard bee populations used for commercial pollination by minimizing the dispersal of pre-nesting females. FY2006 Milestone: Determine whether blue orchard and alfalfa leafcutting bees have the ability to learn nesting cues that affect philopatry, conduct experiments with blue orchard bees to determine if mechanical stress (e.g. handling) increases dispersal rates. FY2007 Milestone: Conduct experiments to determine how and when learning takes place in emerging females, continue with philopatry experiments testing effects of mechanical stress. FY2008 Milestone: Continue with experiments from FY2007, complete data analysis and write manuscript on female dispersal. Objective 2 Evaluate pollination efficacies of confined and free- flying populations of select new bee pollinators for small fruit crops and regional forbs in demand for re-vegetation, and develop practical management protocols to encourage adoption of the more promising species as custom pollinators. FY2006 Milestone: Test trap-nested bee species for pollination efficacies. Cage pollination trial with O. atriventris on cranberry. Continue to evaluate O. aglaia pollinator efficacy on small fruits such as raspberries and blackberries. Evaluate overwintering management needs. Begin publishing wildflower breeding biologies. FY2007 Milestone: Scale-up trap-nesting and production of promising pollinators. Evaluate bee performance in sundry nest substrates. Begin publishing pollinator efficiencies and conduct field trials of wildflower pollinators. Publish nesting the biology of raspberry pollinator and report to producers. FY2008 Milestone: Begin supplying growers with guidance and suitable pollinators for raspberries and for wildflowers. Objective 3 Develop novel control options and delivery systems for the management of chalkbrood, other diseases, parasites, and predators in commercial-scale pollinator populations. Sub-Objective 3.1: Determine the abundance and diversity of chalkbrood in commercial fields, and conduct a survey of bee larvae to determine the frequency of infection for each pathogen species present. FY2006 Milestone: Continue survey of chalkbrood prevalence in the alfalfa leafcutting bee from commercial alfalfa fields, continue PCR processing of samples. FY2007 Milestone: Complete PCR processing of previous years chalkbrood samples. Draft manuscript on multi-year study and report to producers. FY2008 Milestone: Prepare a photo guide to disease in the alfalfa leaf cutting bee. Sub-Objective 3.2: Develop new control methods for chalkbrood, focusing on chemical fungicides, sanitation methods, and microbial control strategies. FY2006 Milestone: Begin search for mycoviruses that can infect Ascosphaera. Begin testing the utility of ozone as a disinfectant for nesting materials for both honeybees and alfalfa leafcutting bees. Continue field testing the use of fungicides for control of chalkbrood in alfalfa leafcutting bees. FY2007 Milestone: Continue field testing the use of fungicides. Develop method for culturing and formulating mycoviruses with activity against Ascosphaera. FY2008 Milestone: Continue field testing the use of fungicides. Continue work with mycoviruses with activity against Ascosphaera. Report to producers. Sub-Objective 3.3: Determine the epizootiology of chalkbrood, and in particular, determine the major means by which the disease is spread and maintained in a bee population. FY2006 Milestone: Measure diurnal and distance-from-a-shelter effects on air loads of Ascosphaera spores. Develop real-time PCR methods for Ascosphaera to assist in quantifying spore loads. Estimate population influx of wild bees into commercial nesting blocks and report to producers. FY2007 Milestone: Correlate Ascosphaera spore loads on adults with chalkbrood levels in progeny. FY2008 Milestone: Test for Ascosphaera in environmental samples from commercial fields (pollen, soil, plants). Prepare a report on the mechanisms for disease spread of chalkbrood in the alfalfa leafcutting bee. Sub-Objective 3.4: Develop microbial control strategies for controlling predators and parasites in bees. FY2006 Milestone: Continue field tests of the fungus Metarhizium anisopliae as a microbial control of varroa testing application timing and dose. Work in collaboration with Earth Biosciences and commercial bee keepers. FY2007 Milestone: Metarhizium anisopliae should be commercially available to beekeepers during this year. Begin tests on the applicability of using the fungus for other hive pests. FY2008 Milestone: Initiate bioassays testing microbial controls on parasites and predators in the alfalfa leafcutting bee. Objective 4 Enhance knowledge of native pollinators by conducting biological surveys, establishing monitoring programs, and expanding knowledge of bee systematics in agricultural and natural systems. Sub-Objective 4.1: Data capture of specimen records of Megachilidae will be collected in support of Objectives 1 & 2. FY2006 Milestone: Complete data capture of U.S. Megachilidae in PIRU collection. FY2007 Milestone: Develop prototype interactive key to the bee genera of the U.S. FY2008 Milestone: Complete data capture of U.S. Apidae in collection. Deliver web-based interactive key to the bee genera of the U.S. Sub-Objective 4.2: A multi-year faunal survey of Grand Staircase- Escalante National Monument initiated in 2000 will continue, with plans for a final field season in 2003. FY2006 Milestone: Analyze faunal data as requested by Grand Staircase-Escalante Natl. Mon. Draft preliminary report on multi-year results. Initiate faunal studies at other sites. FY2007 Milestone: Analyze faunal data as requested by Grand Staircase- Escalante Natl. Mon. and draft final report. FY2008 Milestone: Publish manuscripts on multi-year faunal investigations with Grand Staircase- Escalante Natl. Monument, and other wildland sites. Sub-Objective 4.3: Systematic studies in support of Objectives 1 & 2 will concentrate on candidate pollinator-rich Megachilidae. FY2006 Milestone: Draft revision of bright metallic Osmia for publication. FY2007 Milestone: Conduct phylogenetic analysis of Megachilidae and draft manuscript. FY2008 Milestone: Conduct phylogenetic analysis of Osmiini and draft manuscript. 4a What was the single most significant accomplishment this past year? We experimentally demonstrated an easy to use method for reducing chalkbrood infections in the alfalfa leafcutting bee. Chalkbrood was reduced in growers fields by treating the overwintering bee cells with fungicide just prior to incubation. Chalkbrood is a disease of bees caused by a fungus that infects bee larvae. This disease is a serious mortality factor in alfalfa leafcutting bees managed in U.S. alfalfa seed fields. An epidemiological approach was used to solving the problem. Previous strategies of sterilizing the nesting boards and other equipment have proven inadequate, as has removing the overwintering nests from the nesting boards, a method designed to eliminate the contamination of emerging bees that have to chew their way through dead, infective siblings. We found that adult bees still become contaminated with spores when they emerge from "loose cells" (cells that have been removed from the nesting blocks), and presumably transfer these spores to their offspring. However, this contamination can be reduced by treating loose cells with the fungicide Rovral. Other fungicides have been screened for activity against chalkbrood and safety to bees, and two new biological chemicals were identified this year that show good potential. Now that an application strategy is available, new compounds have a possible utility for alfalfa seed growers. 4b List other significant accomplishments, if any. Using a Native Species of Mason Bee for Berry Pollination. We demonstrated that Osmia aglaia, a bee native to Oregon and California, and is an effective pollinator of red raspberries, and amenable to being managed. This research was conducted in collaboration with commercial cane berry producers, in their fields. O. aglaia pollination activities yielded red raspberry fruits equivalent in size and number to those set by honey bees, even when the O. aglaia are stocked at much lower densities than the honey bees. Furthermore, O. aglaia populations increased two to three fold in a single season in commercial raspberry fields when using our newly designed, costeffective nesting shelters and substrates. The nesting blocks required only a minor modification to nesting blocks already commercially available for alfalfa leafcutting bees. A new collaboration with Xerces Society and Oregon State University is expected to help develop this bee further for use by producers of organic cane berries. The Alfalfa Bee Health Survey. This annual survey was begun in 2002, and continued this year. Sampling "kits" are currently sent to 20 different alfalfa seed producers and pollination providers, representing 25 commercial operations throughout western North America, at regular intervals throughout the year. This year, fall and winter samples of alfalfa leafcutting bee cells were collected and analyzed, and individual reports were sent to collaborators concerning their fields. Data from 2003 and 2004 were also analyzed concerning the period from incubation to emergence of commercial bees and bee health from different growing regions were compared. Results show that bees raised in Canada are healthier than bees raised in the U.S., and that the sex ratio of emerged bees is more male biased in the Canadian populations than the U.S. populations. Furthermore, during this incubation period, more bees died as prepupae than as pupae or adults, especially if reared on the farm by growers or bee keepers, as compared to in the laboratory. These findings have led to new hypothesis regarding which management practices that differ between Canada and the U.S. might explain the poorer health of bees raised in the U.S. Old Nest Components as Attractants to New Female Bees. It has long been recognized by alfalfa seed growers and bee managers that alfalfa leafcutting bee and blue orchard bee females prefer to nest in previously used nesting materials. We found that volatile compounds emanating from leaf pieces used in lining nest cells, and feces left attached to cast-off cocoons were attractive to leafcutting bees, but other nesting materials were not. For the orchard bee, volatile compounds from caste-off cocoons and the paper straws used in the nest cavities were the attractive materials. Understanding that the bees are attracted to chemical components of old nest parts is a first step in identifying important nest attractants that could be used to enhance commercial nesting and discourage dispersal of economically important bees. Wild Bee Biogeography and Diversity. We developed an electronic data base for U. S. National Pollinating Insects Collection in Logan, UT. Approximately 4,000 potential pollinators (bee species) reside in the U.S. What is largely lacking in order to utilize and conserve these bees for agricultural pollination needs is knowledge on their biology: What plants do they pollinate? Where do they live? When are they active? Where do they nest? We have increased our knowledge of these specifications by linking collection specimens with the data on their records. Extensive multi-year field surveys of bee fauna from four sites: Clark Co. NV, Yosemite Natl. Park, Dugway Proving Grounds, Grand Staircase-Escalante Natl. Monument has fed the database. Over 600,000 records are now in the database, including all identified U.S. specimens. Records of flowers visited by each bee species are an especially valuable component of this database, one which we are providing to the Natural Resources Conservation Service at their request. 5. Describe the major accomplishments over the life of the project, including their predicted or actual impact. The 60-month Project Plan for the PIRU was initiated 1 October 2003. The research reported was conducted under National Program 305 Crop Production, and directly contributes to research Component 6.0 Bees and Pollination. Research activities are further related to ARS Strategic Plan Goal 1 Enhance Economic Opportunities for Agricultural Producers, Objective 1.2 Contribute to the Efficiency of Agricultural Production Systems. Considerable progress was made in developing the blue orchard bee as a fruit tree pollinator. This includes: a) The publication of an 88-page manual, "How to manage the blue orchard bee as an orchard pollinator", b) The patent of a new, affordable and manageable polycarbonate-based nesting block for the blue orchard bee and the development of a related prototype emergence box, c) Publication of a six- year project demonstrating highly-significant yield increases in a commercial cherry orchard pollinated with blue orchard bees, d) The publication of research on successful methods to advance blue orchard bee emergence for almond pollination, and e) The publication or preparation of several research articles and two CRADAs on the developmental biology and rearing methods (wintering requirements, development and incubation temperature regimes, respirometry) of the blue orchard bee. Recent research has emphasized diapause induction and development, respirometry, and diapause-associated gene expression, as well as blue orchard bee toxicology related to fungicide sprays during bloom, and the use of visual and olfactory cues involved in nest location. Almonds is a lucrative, growing crop in S. California, but during the last three bloom cycles, producers experienced difficulty obtaining sufficient numbers of honey bees for pollination. In addition, the acreage of almond plantings is rapidly expanding, and the first new plantings are expected to come into production around 2010. The blue orchard bee could provide effective pollination for this new crop. The research reported was conducted under National Program 305 Crop Production, and directly contributes to research Component 6.0 Bees and Pollination. Research activities are further related to ARS Strategic Plan Goal 2 Support Increased Economic Opportunities and Improved Quality of Life in Rural America, and Goal 5 Protection and Enhance the Nations Natural Resource Base and Environment, Objective 5.1 Provide Science Based Knowledge and Education to Improve the Management of Forest, Rangelands, and Pastures. An ongoing interdisciplinary research program to establish successful agricultural production and marketing of eight or more broadly adapted wildflower species native to the Great Basin and adjacent ecological regions. The Bureau of Land Management and the U.S. Forest Service annually broadcast 3-10 million pounds of grass, shrub and forb seed across public lands in the Great Basin to promote re-vegetation on degraded or burned rangeland and forest. Current seed mixes contain <1% forb seed, far short of the annual need for up to 2 million pounds of select native wildflower seed. We determined each forb species breeding biology and pollination needs, evaluated its bee fauna for potentially manageable species, and developed protocols and starting populations of effective pollinators that can be managed in an on-farm setting for profitable forb seed production. To date, we found that the forb species preferred for revegitation differ widely in the diversity of native bee species which they attract, and all but a few require bee visitation to set seed. Manageable captive populations of two Osmia species are in hand for pollinating the first of the desired forb species, and alfalfa leafcutting bees and honey bees are being tested as alternative pollinators. The research reported was conducted under National Program 305, Crop Production, and directly contributes to research Component 6.0 Bees and Pollination. Research activities are further related to ARS Strategic Plan Goal 3 Enhance Protection and Safety of the Nations Agriculture and Food Supply, Objective 3.2, Develop and Deliver Science-Based Information Technologies to Reduce the Number and Severity of Agricultural Pest, Insect, Weed, and Disease Outbreaks. Alfalfa leaf cutting bees are managed for pollination of alfalfa seed crops, and an industry has developed around the need to supply bees. Currently, seed growers purchase 50% or more of their bees from Canada every year due to a high mortality of the bees that they return out of the field. We quantified the causes of the high mortality of larvae from U.S. fields, and found it to be due primarily to disease (chalkbrood), "pollen ball" (undefined, early stage mortality), and parasitic wasps. Some mortality also occurs during overwintering. We found that chalkbrood is spread primarily by female bees who become contaminated with spores when emerging, and found that this can be reduced with the use of fungicides, and overwintering mortality can be reduced by more careful control of incubation conditions. We hypothesize that late summer and fall storage conditions may also affect the ability of bees to survive the winter, especially if the bees are maintained under conditions that are too warm, forcing them to burn fatbody reserves. The bee health survey has emphasized that control measures are seriously needed for parasitic wasps, and the main causes for "pollen balls" need to be identified so that it too can be controlled. Results from these studies have been presented at seed grower meetings in the U.S. and Canada. Chalkbrood control methods are expected to be available to growers within the next year. 6. What science and/or technologies have been transferred and to whom? When is the science and/or technology likely to become available to the end- user (industry, farmer, other scientists)? What are the constraints, if known, to the adoption and durability of the technology products? Varroa destructor is a mite that is a serious parasite of honey bees that can cause hive fatality. It was estimated that 50% of all commercial hives in the U.S. were lost during the winter of 2004-2005, and the cause was primarily due to pesticide resistance in the mite leading to failure of control. We have been working on developing a microbial control for varroa mites using a fungus, and has shown that it can reduce mite loads and increase winter survival of hives. We have found an industry partner interested in producing the fungus for varroa control. The company has already registered this fungus with the EPA for pest control. Field trials are currently underway to identify an application method that is easy, yet effective, and to better define the best dosage and timing of application. This product is expected to be commercialized in the near future, and would be utilized by both commercial and hobbyist beekeepers. The major issue holding up commercialization is the limited availability of fungus production facilities. The National Park Service has identified the inventory of native pollinators as a high priority for management planning. In response to this need we are providing information on native pollinators to land managers, and recently completed the fifth year of a survey of the bee pollinators of Grand Staircase-Escalante National Monument. This work has highlighted the importance of long-term studies for monitoring the health of pollinator services. Similar projects are currently underway in the John Muir National Historic Site and Great Smoky Mountains National Park. The expanding demand for this information among Federal land management agencies has led to the development of joint efforts on native pollinators and their impact on rare plants, fire and grazing management in Zion National Park, Yosemite National Park and with the Bureau of Land Management Clark County, NV. We also lead the development of a world checklist of bees at the request of the U.S. Geological Survey. A prototype using one subfamily of bees was initiated and completed during 2005. (Project Plan Objective 4.2, Milestones 1 and 2; NP 305, Action Plan 6 Bees and Pollination; Performance Measure 1.2.4 7. List your most important publications in the popular press and presentations to organizations and articles written about your work. (NOTE: List your peer reviewed publications below). Cane, J. 2005. Invited presentation. "How much pollen and nectar constitute larval provisions of the alfalfa leaf-cutting bee?" Annual Meeting of the Northwest Alfalfa Seed Growers Association. Jan. 30-Feb. 1, 2005. Boise, Idaho. p.49-50. Cane, J. 200. Invited presentation. "Bees and seed production for native plant restoration in wildlands". In a symposium "The Well Bee-ing of Pollinators and their Impact on US Agriculture and Natural Ecosystems," at the Annual Meeting of the Entomological Society of America, in Salt Lake City, Utah, Nov. 14-17, 2004. Griswold, T. 2004. Invited presentation. Patterns of bee biodiversity in North America. In a symposium "New insights into bee phylogeny" Annual Meeting of the Entomological Society of America, Salt Lake City, Utah, Nov. 14-17, 2004. James, R.R. 2005. Invited presentation. Chalkbrood, a U.S. Perspective. Annual Meeting of the Canadian Forage Seed Producers, in Winnipeg, Canada. Jan. 22-25, 2005. James, R.R. 2005. Invited presentation. Varroa mite control with fungal pathogens: will this little piggy get to market? In a symposium "Use of Pathogens Against Incursion Pests," Annual Meeting of the Society for Invertebrate Pathology, Anchorage, Alaska. Aug. 7-11, 2005. Messinger, O. and Griswold, T. 2005. Invited presentation. The bees of GSENM: Four years, six hundred species and counting. GSENM Science Forum, Escalante, Utah, 10 June 2005. Pitts-Singer, T.L. 2005. Invited presentation. Attraction of leafcutting bees to old nest material and individuals cues. Annual Meeting of the Northwest Alfalfa Seed Growers Association. Jan. 30-Feb. 1, 2005. Boise, Idaho. p.51. Pitts-Singer, T.L. 2004. Invited presentation. Management of alfalfa leafcutting bees over four decades. In a symposium "The Well Bee-ing of Pollinators and their Impact on US Agriculture and Natural Ecosystems," at the Annual Meeting of the Entomological Society of America, in Salt Lake City, Utah, Nov. 14-17, 2004.

Impacts
(N/A)

Publications

Bess, J.A., O'Neill, K.M., Kemp, W.P. 2004. Leafhopper assemblages on native and reseeded grasslands in southwestern Montana. Western North American Naturalist. 64(4):518-531.
Bosch, J., Kemp, W.P. 2005. Alfalfa leafcutting bee population dynamics, flower availability, and pollination rates in two Oregon alfalfa fields. Journal of Economic Entomology. 98(4):1077-1086.
Bosch, J., Kemp, W.P. 2004. Effect of pre-wintering and wintering temperature regimes on weight loss, survival, and emergence time in the mason bee Osmia cornuta (hymenoptera: megachilidae). Apidologie. 35:469- 479.
Buckner, J.S., Kemp, W.P., Bosch, J. 2004. Characterization of triacylglycerols from overwintering prepupae of the alfalfa pollinator Megachile rotundata (Hymenoptera: Megachilidae). Archives of Insect Biochemistry and Physiology. 57(1):1-14.
Yocum, G.D., Kemp, W.P., Bosch, J., Knoblett, J.N. 2005. Temporal variation in overwintering gene expression and respiration in the solitary bee Megachile rotundata. Journal of Insect Physiology. 51(6):621-629.
Ladurner, E., Bosch, J., Kemp, W.P., Maini, S. 2005. Assessing delayed and acute toxicity of five formulated fungicides to Osmia lignaria Say and Apis mellifera. Apidologie. 36:449-460.
Ladurner, E., Bosch, J., Kemp, W.P., Maini, S. 2005. Evaluation of a standard artificial flower design to feed individual bees known amounts of pesticides. Apidologie. 36:379-387.
James, R.R., Pitts Singer, T. 2005. Ascosphaera aggregata contamination on alfalfa leafcutting bees in a loose cell incubation system. Journal of Invertebrate Pathology. 89:176-178.
James, R.R. 2005. Impact of disinfecting nesting boards on chalkbrood control in the alfalfa leafcutting bee. Journal of Economic Entomology. 98(4):1094-1100.
James, R.R. 2005. Temperature and chalkbrood development in the alfalfa leafcutting bee. Apidologie. p. 15-23.
James, R.R., Buckner, J.S. 2004. Lipids stimulate spore germination in the entomopathogenic ascomycete Ascosphaera aggregata. Mycopathologia. 158:293- 302
Cane, J.H. 2005. Pollination needs of arrowleaf balsamroot, Balsamorhiza sagittata (Heliantheae: Asteraceae). Western North American Naturalist. 65(3):359-364.
Cane, J.H., Minckley, R.L., Kervin, L., Roulston, T. 2005. Temporally persistent patterns of incidence and abundance in a pollinator guild at annual and decadal scales: the bees of Larrea tridentata. Biological Journal of the Linnean Society, London. 85:319-329.
Sampson, B.J., Stringer, S.J., Cane, J.H., Spiers, J.M. 2004. Screen house evaluations of a blue orchard bee osmia ribifloris (hymenoptera: apidae) as a supplemental pollinator for southeastern blueberries. Blueberry Research Extension North American Workers Conference Proceedings. vol.3(15) pp.381-392.
Pitts Singer, T. 2004. Examination of "Pollen Balls" in the nests of the alfalfa leafcutting bee, Megachile rotundata. Journal of Apicultural Research. 43(2):40-46.
Tepedino, V.J., Sipes, S.D., Griswold, T.L. 2004. Reproduction and demography of townsendia aprica (asteraceae), a rare endemic of the southern Utah plateau. Western North American Naturalist. 64(4):465-470

Progress 10/01/03 to 09/30/04

Outputs
1. What major problem or issue is being resolved and how are you resolving it (summarize project aims and objectives)? How serious is the problem? What does it matter? Pollinators are critical elements of ARS National Program 305, Crop Production - Bees and Pollination. Without effective pollination, insect- pollinated crops such as almond, blueberry, sweet cherry, or cranberry, are failures. The annual combined value of U.S. crops requiring cross- pollination is over $10 billion; the bulk of this is accomplished using the honey bee. However, some crops have suffered recently because the number of honey bee colonies has actually declined due to depressed honey prices and increased management costs associated with the arrival of enemies such as Varroa and tracheal mites. For example, 3% of the California almond producers were unable to rent honey bees for pollination during the 2000-2002 pollination seasons. The overarching goal for the for the Pollinating Insect-Biology, Management, & Systematics Research Unit (PIRU) Project Plan, initiated 1 October 2003, is improved profitability, for U.S. producers of agricultural crops that require cross-pollination, through the diversification of our agricultural pollinator portfolio. In order to achieve this goal, priority will be given to: 1) Delivering improved pollination management systems for alfalfa seed production using the alfalfa leafcutting bee and the alkali bee, and a new pollination management system for orchards using the blue orchard bee; 2) Evaluating pollination efficacies of confined and free-flying populations of select new bee pollinators for small fruit crops and forbs in demand for re- vegetation, and developing practical management protocols to encourage adoption of the more promising species as custom pollinators; 3) Developing novel control options and delivery systems for the management of chalkbrood, other diseases, parasites, and predators in commercial- scale pollinator populations; and, 4) Conducting biological surveys, establishing monitoring programs, and expanding current knowledge of bee systematics in natural and agricultural systems. Anticipated products of the research include pollination and bee management handbooks, patented nesting and shelter designs, and peer-reviewed publications. The major customers of this research are the U.S. alfalfa seed producers (NW Alfalfa Seed Growers Association, and The Alfalfa Alliance), who rely heavily on the superior performance of the alfalfa leafcutting bee and the alkali bee for pollination each year. This group enjoys a close relationship with the PIRU, and their members participate directly in the planning and execution of ongoing research. Additionally, PIRU research results are in demand by commercial seed and fruit producers, particularly organic tree fruit and nut producers in the case of the blue orchard bee, as well as custom pollination providers, federal, state, and private land managers, amateur gardeners, and non-profit non-governmental organizations. Potential benefits expected from attaining the objectives outlined in the current Project Plan include increased productivity and profitability through the identification and management of superior, affordable pollinators, and improved sustainability through the diversification of a pollinator portfolio for U.S. Agriculture. 2. List the milestones (indicators of progress) from your Project Plan. Objective 1 - Deliver improved pollination management systems for alfalfa seed production using the alfalfa leafcutting bee and the alkali bee, and a new pollination management system for orchards using the blue orchard bee. Sub-Objective 1.1: For the two primary alfalfa pollinators, the alkali bee and the alfalfa leafcutting bee, we will determine cost-effective, optimal stocking densities for pollinators that maximize seed production while minimally compromising bee reproduction. FY2004 Milestones: Complete alkali bee brood provision analyses and draft provision sugar methods manuscript. Conduct alfalfa leafcutting bee density enclosure studies and analyze data. Collect samples and information for regional bee health survey. FY2005 Milestones: Analyze data and publish provision mass pollen/sugar paper. Repeat bee density study with different densities and analyze data. Continue bee health survey and report to producers. FY2006 Milestones: Publish multi-year nest density study with the alkali bee. Estimate male pollination value, review for bees. Implement large field trials with more "optimal" bee densities defined from enclosure study. Continue bee health survey and report to producers. FY2007 Milestones: Publish carrying capacity and competition paper on the alkali bee. Continue large field trials with optimal bee densities. Continue bee health survey and report to producers. FY2008 Milestones: Analyze and publish carrying capacity of alfalfa and wildflower crops for the alkali bee. Analyze and evaluate large field trials with optimal bee densities. Analyze and evaluate bee health survey data and report to producers. Sub-Objective 1.2: To better enhance nesting of the alfalfa leafcutting bee in clean, artificial nesting boards and to better ensure field fidelity of bees released for pollination, we will investigate the origins of cues that attract bees to previously used nesting boards. FY2004 Milestone: Initiate behavioral bioassays for nest attraction using alfalfa leafcutting bees. Perform chemical analyses of important attractants. FY2005 Milestones: Continue tests with alfalfa leafcutting bees; begin tests with blue orchard bees. Perform chemical analyses of important components. FY2006 Milestone: Test ability to apply attractants to nesting boards. FY2007 Milestones: Field tests of nesting board attractants in all species where applicable and report to producers. FY2008 Milestones: Continue testing nesting board attractants. Analyze and evaluate application for commercial growers. Submit final evaluation and recommendations. Sub-Objective 1.3: Improve summer and pre-wintering temperature regimes for the commercial (and artificial) management of blue orchard bee populations. FY2004 Milestone: Conduct temperature studies on summer development of phenologically distinct blue orchard bee populations and initiate pre-wintering studies. FY2005 Milestone: Complete summer development studies with the blue orchard bee. FY2006 Milestone: Complete pre-wintering studies with the blue orchard bee. FY2007 Milestone: Complete statistical treatment of data collected, draft manuscripts, and report to producers. FY2008 Milestone: Submit final evaluation and recommendation of optimal summer and pre-wintering temperature regimes for the blue orchard bee. Sub-Objective 1.4: Support improvements in pre-wintering and wintering temperature regimes for the commercial (and artificial) management of the alfalfa leafcutting bee and the blue orchard bee through improving our understanding of intermediary metabolic processes and diapause gateways. FY2004 Milestone: Complete life cycle respirometry, supercooling point, amino acid, lipid, and carbohydrate profiles for natural alfalfa leafcutting bee populations. Identify genes to signal change from diapause (hibernation-like state in insects) to post-diapause development and compare their expression levels during the winter through springtime development period. FY2005 Milestone: Complete analyses and draft manuscript on life cycle respirometry, intermediary metabolism in the alfalfa leafcutting bee under natural conditions. Monitor putative clones for life cycle expression patterns in the alfalfa leafcutting bee. Draft manuscript identifying apparent diapause gateways in the alfalfa leafcutting bee. FY2006 Milestone: Complete life cycle respirometry, supercooling point, amino acid, lipid, and carbohydrate profiles for alfalfa leafcutting bee and blue orchard bee populations under natural and artificial conditions. Create cDNA library for blue orchard bee HSP's via RT-PCR for diapause and non-diapause condition. Sequence putative clones and generate DIG-labeled probes for expression assays via Northern blot. FY2007 Milestone: Complete analyses and draft manuscripts on life cycle respirometry, intermediary metabolism for alfalfa leafcutting bee and blue orchard bees under natural and artificial conditions. Monitor putative clones for life cycle expression patterns in the blue orchard bee. Draft manuscript apparent identifying diapause gateways in the blue orchard bee. FY2008 Milestone: Submit final comparison and evaluation of the physiological basis of optimal summer, pre-wintering, and wintering temperature regimes for the alfalfa leafcutting bee and the blue orchard bee. Sub-Objective 1.5: Demonstrate the blue orchard bee and new nesting block design in selected early-spring orchard crops. FY2004 Milestone: Complete year 6 (of 6) in cherry pollination demonstration in UT. Complete year 2 of organic almond pollination demonstration in CA. Complete year 1 of cherry pollination demonstration in MT. Complete initial field testing of current nest block prototype in UT pear-apple orchards. FY2005 Milestone: Complete analysis of data and draft manuscript on UT cherry pollination. Complete year 3 of organic almond pollination demonstration in CA. Complete analysis of prototype performance, and develop modifications to be made to second generation prototype. FY2006 Milestone: Submit final evaluation and recommendation of optimal cherry pollination management program using the blue orchard bee. Complete year 4 of organic almond pollination demonstration in CA. Complete testing of second generation prototype in UT pear-apple orchards. FY2007 Milestone: Complete year 5 of organic almond pollination demonstration in CA. Complete analysis of data and draft manuscript on organic almond pollination. Prepare final evaluation and recommendation for transfer to mass market. FY2008 Milestone: Submit final evaluation and recommendation of optimal almond pollination management program using the blue orchard bee. Transfer technology to CA organic orchard producers. Sub-Objective 1.6: Improve the establishment of artificially managed blue orchard bee populations used for commercial pollination by minimizing the dispersal of pre-nesting females. FY2004 Milestone: Complete first year experiments on wintering temperature, parasitism and body size. Select progeny of dispersing vs. non-dispersing (philopatric) females for field trials in FY2005 and beyond. FY2005 Milestone: Complete second year experiments on wintering temperature, parasitism and body size. Complete first year experiment with philopatric females. FY2006 Milestone: Complete second year experiment with philopatric females. FY2007 Milestone: Complete third year experiment with philopatric females.FY2008 Milestone: Complete data analysis and write manuscript on female dispersal. Submit final evaluation and recommendation of optimal blue orchard bee population management. Objective 2 - Evaluate pollination efficacies of confined and free- flying populations of select new bee pollinators for small fruit crops and regional forbs in demand for re-vegetation, and develop practical management protocols to encourage adoption of the more promising species as custom pollinators. FY2004 Milestone: Refine nest substrate options for Osmia aglaia in raspberries. Confirm basic breeding biologies of focal wildflower seed species. Begin sampling pollinator guild of wildflowers. Grow Osmia ribifloris and address dispersal problems. Publish cranberry pollinator efficacy study. FY2005 Milestone: Trap-nest for O. atriventris for cranberry pollination. Trap-nest promising pollinators for wildflowers. Evaluate specific pollination needs of wildflowers. Evaluate trial plots. Sample, ID wildflower seed and flower pests. Publish cranberry pollen tube attrition study. FY2006 Milestone: Test trap-nested bee species for pollination efficacies. Cage pollination trial with O. atriventris on cranberry. Evaluate pollinator efficacy on highbush blueberries. Evaluate overwintering management and needs. Begin publishing wildflower breeding biologies. Try O. ribifloris free-flight in blueberries. FY2007 Milestone: Scale-up trap-nesting and production of promising pollinators. Evaluate bee performance in sundry nest substrates. Begin publishing pollinator efficiencies and conduct field trials of wildflower pollinators. Publish nesting biology of raspberry pollinator and report to producers. FY2008 Milestone: Begin supplying growers with guidance and suitable pollinators for raspberries and for wildflowers. Objective 3 - Develop novel control options and delivery systems for the management of chalkbrood disease, other diseases, parasites, and predators in commercial-scale pollinator populations. Sub-Objective 3.1: Determine the abundance and diversity of chalkbrood in commercial fields, and conduct a survey of bee larvae to determine the frequency of infection for each pathogen species present. FY2004 Milestone: Sample alfalfa leafcutting bee larvae from commercial fields, and assess levels of chalkbrood infection. Refine molecular marker methods for chalkbrood detection. FY2005 Milestone: Sample alfalfa leafcutting bee larvae from commercial fields, and assess levels of chalkbrood infection. Refine molecular marker methods for Ascosphaera detection. Develop photo guide to disease in the alfalfa leafcutting bee. FY2006 Milestone: Conduct survey of chalkbrood prevalence in the alfalfa leafcutting bee from commercial alfalfa fields and report to producers. FY2007 Milestone: Conduct survey of chalkbrood prevalence in the alfalfa leafcutting bee from commercial alfalfa fields and report to producers. FY2008 Milestone: Conduct survey of chalkbrood prevalence in the alfalfa leafcutting bee from commercial alfalfa fields. Draft manuscript on multi- year study and report to producers. Sub-Objective 3.2: Develop new control methods for chalkbrood, focusing on chemical fungicides, sanitation methods, and microbial control strategies. FY2004 Milestone: Apply fungicides to loose cells of the alfalfa leafcutting bee and test effect on chalkbrood in alfalfa leafcutting bee progeny. FY2005 Milestone: In commercial fields, test applications of systemic fungicides on chalkbrood levels in the alfalfa leafcutting bee and report to producers. FY2006 Milestone: Begin search for mycoviruses that can infect Ascosphaera. FY2007 Milestone: Develop method for culturing and formulating mycoviruses with activity against Ascosphaera. FY2008 Milestone: Conduct small scale field tests of mycoviruses for biological control of chalkbrood and report to producers. Sub-Objective 3.3: Determine the epizootiology of chalkbrood, and in particular, determine the major means by which the disease is spread and maintained in a bee population. FY2004 Milestone: Measure chalkbrood spore concentration levels on bee nesting cells in commercial operations. Conduct biology studies on chalkbrood (growth and spore germination requirements, spore persistence, etc.). FY2005 Milestone: Measure diurnal and distance-from-a-shelter effects on air loads of Ascosphaera spores. Measure levels of chalkbrood in feral populations of the alfalfa leafcutting bee. Continue basic biology studies on Ascosphaera aggregata. FY2006 Milestone: Measure diurnal and distance-from-a-shelter effects on air loads of Ascosphaera spores. Estimate population influx of "wild bees" into commercial nesting blocks. Report to producers. FY2007 Milestone: Correlate Ascosphaera spore loads on adults with chalkbrood levels in progeny. Develop quantitative PCR methods for detecting Ascosphaera spores and report to producers. FY2008 Milestone: Refine quantitative PCR methods. Test for Ascosphaera in environmental samples from commercial fields (pollen, soil, plants). Sub-Objective 3.4: Develop microbial control strategies for controlling predators and parasites in bees. FY2004 Milestone: Develop fermentation method for two species of fungi with potential for biological control of varroa mites in honey bees. Conduct field trials for microbial control of varroa in honey bees. FY2005 Milestone: Conduct field trials of microbial control of varroa in honey bees. Develop a laboratory population of the checkered flower beetle. FY2006 Milestone: Develop bioassays for the checkered flower beetle, test fungi for pathogenicity. Conduct non-target tests of the candidate fungi on bees. FY2007 Milestone: Field test microbial control strategy for predatory beetles in commercial fields and report to producers. FY2008 Milestone: Field test microbial control strategy for predatory beetles in commercial fields and report to producers. Objective 4 - Enhance knowledge of native pollinators by conducting biological surveys, establishing monitoring programs, and expanding knowledge of bee systematics in agricultural and natural systems. Sub-Objective 4.1: Data capture of specimen records of Megachilidae will be collected in support of Objectives 1 & 2. FY2004 Milestone: Develop prototype interactive key to the bees of the Mid-Atlantic States. FY2005 Milestone: Capture collection records of Osmia lignaria, O. ribifloris, and O. aglaia. Deliver web-based interactive key to the bees of the Mid- Atlantic states. FY2006 Milestone: Complete data capture of U.S. Megachilidae in BBSL collection. FY2007 Milestone: Develop prototype interactive key to the bee genera of the U.S. FY2008 Milestone: Complete data capture of U.S. Apidae in collection. Deliver web-based interactive key to the bee genera of the U.S. Sub-Objective 4.2: A multi-year (bee) faunal survey of Grand Staircase- Escalante National Monument initiated in 2000 will continue, with plans for a final field season in FY2004. FY2004 Milestone: Complete field (bee) surveys in Grand Staircase-Escalante National Monument. FY2005 Milestone: Analyze faunal data as requested by Grand Staircase-Escalante Natl. Mon. FY2006 Milestone: Analyze faunal data as requested by Grand Staircase-Escalante Natl. Mon. Draft preliminary report on multi-year results. FY2007 Milestone: Analyze faunal data as requested by Grand Staircase-Escalante Natl. Mon. Draft final report. FY2008 Milestone: Publish manuscript on multi-year faunal investigations with Grand Staircase-Escalante Natl. Mon. collaborators. Sub-Objective 4.3: Systematic studies in support of Objectives 1 & 2 will concentrate on candidate pollinator-rich Megachilidae. FY2004 Milestone: Draft revision of North American Stelis (group of parasitic bees of worldwide significance) for publication. FY2005 Milestone: Draft revision of N. Am. Dufourea for publication. FY2006 Milestone: Draft revision of bright metallic Osmia for publication. FY2007 Milestone: Conduct phylogenetic analysis of Megachilidae and draft manuscript. FY2008 Milestone: Conduct phylogenetic analysis of Osmiini and draft manuscript. 3. Milestones: A. List the milestones (from the list in Question #2) that were scheduled to be addressed in FY 2004. How many milestones did you fully or substantially meet in FY 2004 and indicate which ones were not fully or substantially met, briefly explain why not, and your plans to do so. The 60-month Project Plan for the PIRU was initiated 1 October 2003. Substantial progress has been made on each Year 1 Milestone listed under the four major Objectives. Objective 1 - Deliver improved pollination management systems for alfalfa seed production using the alfalfa leafcutting bee and the alkali bee, and a new pollination management system for orchards using the blue orchard bee. Sub-Objective 1.1: For the two primary alfalfa pollinators, the alkali bee and the alfalfa leafcutting bee, we will determine cost-effective, optimal stocking densities for pollinators that maximize seed production while minimally compromising bee reproduction. FY2004 Milestone: Complete alkali bee brood provision analyses and draft provision sugar methods manuscript. Conduct bee density enclosure studies and analyze data. Collect samples and information for bee health survey. FY2004 Progress: The draft manuscript on the analytical method to quantify (brood) provision sugars is in the final stages of preparation. The alfalfa leafcutting bee density study conducted in 2003 yielded interesting and suggestive, but not conclusive, results due to lack of good alfalfa bloom in our first year field; the study is being repeated in summer 2004 on a now-established alfalfa field. Samples and information from alfalfa leafcutting bee managers in seven northwestern states continue to be solicited for the bee health survey. Data accumulated in our lab, information from manager surveys, and temperature and relative humidity records from Hobo Dataloggers are being analyzed. New questions for experimentation are emerging from such analyses, while some results already are yielding publications (e.g., pollen ball/immature death analysis; adult emergence and sex ratio). Sub-Objective 1.2: To better enhance nesting of the alfalfa leafcutting bee in clean, artificial nesting boards and to better ensure field fidelity of bees released for pollination, we will investigate the origins of cues that attract bees to previously used nesting boards. FY2004 Milestone: Initiate behavioral bioassays for nest attraction using alfalfa leafcutting bees. Perform chemical analyses of important attractants. FY2004 Progress: Behavioral bioassays using a Y-tube apparatus to test attraction to nest materials as well as extracts of nest material were initiated in 2003 and will be completed at the end of Summer 2004. Chemical analyses will begin in Fall 2004, after all results are obtained from Y-tube tests, and will continue in 2005. Sub-Objective 1.3: Improve summer and pre-wintering temperature regimes for the commercial (and artificial) management of blue orchard bee populations. FY2004 Milestone: Conduct temperature studies on summer development of phenologically distinct blue orchard bee populations and initiate pre-wintering studies. FY2004 Progress: Pre-wintering temperature studies were completed as planned, and data are currently being analyzed. Sub-Objective 1.4: Support improvements in pre-wintering and wintering temperature regimes for the commercial (and artificial) management of the alfalfa leafcutting bee and the blue orchard bee through improving our understanding of intermediary metabolic processes and diapause gateways. FY2004 Milestone: Complete life cycle respirometry, supercooling point, amino acid, lipid, and carbohydrate profiles for natural alfalfa leafcutting bee populations. Identify genes to signal change from diapause (hibernation-like state in insects) to post-diapause development and compare their expression levels during the winter through springtime development period. FY2004 Progress: Alfalfa leafcutting bee life cycle respirometry, supercooling point, and amino acid profiles have been completed; lipid and carbohydrate profiles have been delayed by the need to develop improved analytical methods for quantification (currently underway). Genes that signal change from diapause (hibernation-like state in insects) to post-diapause development in the alfalfa leafcutting bee have been identified; a manuscript describing these results was drafted and subsequently accepted for publication. Sub-Objective 1.5: Demonstrate the blue orchard bee and new nesting block design in selected early-spring orchard crops. FY2004 Milestone: Complete year 6 (of 6) in cherry pollination demonstration in UT. Complete year 2 of organic almond pollination demonstration in CA. Complete year 1 of cherry pollination demonstration in MT. Complete initial field testing of current nest block prototype in UT pear-apple orchards. FY2004 Progress: Year six (and final) of the UT blue orchard bee cherry pollination demonstration was completed; a manuscript describing the results of this demonstration is nearly ready for journal submission. Pollination demonstrations (with the blue orchard bee) were completed in MT cherries and CA almonds. Field testing of our patented nest block prototype yielded very promising results in a UT pear and apple orchard. Sub-Objective 1.6: Improve the establishment of artificially managed blue orchard bee populations used for commercial pollination by minimizing the dispersal of pre-nesting females. FY2004 Milestone: Complete first year experiments on wintering temperature, parasitism and body size. Select progeny of dispersing vs. non-dispersing females for field trials in FY2005 and beyond. FY2004 Progress: Dispersing and non- dispersing females were collected in Spring 2004. Marked Osmia lignaria females were released from a common emergence area in a local apple orchard. Nests being provisioned by marked bees in shelters at nearby and distant locations were marked accordingly and subsequently collected from the field. Nests are currently being maintained in the laboratory and will overwinter in cold storage. Surviving progeny from nearby (non- dispersing) and distant (dispersing) nests will be used in investigations for continuing this project in 2005. Objective 2 - Evaluate pollination efficacies of confined and free- flying populations of select new bee pollinators for small fruit crops and regional forbs in demand for re-vegetation, and develop practical management protocols to encourage adoption of the more promising species as custom pollinators. FY2004 Milestone: Refine nest substrate options for Osmia aglaia in raspberries. Confirm basic breeding biologies of focal wildflower seed species. Begin sampling pollinator guild of wildflowers. Grow Osmia ribifloris and address dispersal problems. FY2004 Progress: The bee Osmia aglaia has been shown to be an effective raspberry pollinator. Pollination needs and breeding biology of one target wildflower, Balsamorhiza sagittata, have been summarized, a manageable pollinator identified, and a manuscript submitted. An inexpensive, practical prototype nesting shelter was designed and will be tested in two states in cool and hot seasons for acceptance and thermal performance. It is acceptable to the two bee species thus far tried. Objective 3 - Develop novel control options and delivery systems for the management of chalkbrood disease, other diseases, parasites, and predators in commercial-scale pollinator populations. Sub-Objective 3.1: Determine the abundance and diversity of chalkbrood in commercial fields, and conduct a survey of bee larvae to determine the frequency of infection for each pathogen species present. FY2004 Milestone: Sample alfalfa leafcutting bee larvae from commercial fields, and assess levels of chalkbrood infection. Refine molecular marker methods for chalkbrood detection. FY2004 Progress: A second year of the bee health survey was conducted, and the third year started. The number of growers participating in the survey has grown from three to twelve, so the 2003 field season really comprises the beginning of a new, more expanded survey done in collaboration with Theresa Pitts-Singer, who is looking at health issues other than chalkbrood. The PCR methods for Ascosphaera detection have been fully developed, and DNA extractions of field samples bees have been completed. Sub-Objective 3.2: Develop new control methods for chalkbrood, focusing on chemical fungicides, sanitation methods, and microbial control strategies. FY2004 Milestone: Apply fungicides to loose cells of the alfalfa leafcutting bee and test effect on chalkbrood in alfalfa leafcutting bee progeny. FY2004 Progress: A large scale, replicated, field trial involving four growers has been initiated. Loose cells were treated with Rovral to determine if this method of fungicide application will reduce chalkbrood incidence. Preliminary laboratory studies were used to determine which fungicide to use and at what rate. Prior to starting the grower-assisted trial, we tested the safety of this fungicide, in the laboratory, when loose cells are treated. Sub-Objective 3.3: Determine the epizootiology of chalkbrood, and in particular, determine the major means by which the disease is spread and maintained in a bee population. FY2004 Milestone: Measure chalkbrood spore concentration levels on bee nesting cells in commercial operations. Conduct biology studies on chalkbrood (growth and spore germination requirements, spore persistence, etc.). FY2004 Progress: The number of Ascosphaera spores that get on emerging adult bees was been determined for the loose cells system. Females pick-up significantly fewer spores in a loose cell system than when solid wood boards are used. However, the females are still being contaminated with a large number of spores, which probably serves as the main source of transmission for the disease. The thermal limits for Ascosphaera aggregata growth and spore germination were determined. Also, the effect of temperature on disease development was determined. The optimum temperature for in vitro growth and spore germination was higher than the optimum temperature for infection. Previously, we found that lipids stimulated spore germination in Ascosphaera aggregata. This year we determined that the response was not nutritional, but physical. The spores appear to be bi-polar with one end being lipophilic and the other hydrophilic. An understanding of the significance of this in the initiation of infection is being sought. Sub-Objective 3.4: Develop microbial control strategies for controlling predators and parasites in bees. FY2004 Milestone: Develop fermentation method for two species of fungi with potential for biological control of varroa mites in honey bees. Conduct field trials for microbial control of varroa in honey bees. FY2004 Progress: Procedures for growing Metarhizium anisopliae on rice and Hirsutella thompsonii on bran, sufficient for producing enough spores in-house for field trials have been developed. The stability of M. anisopliae was found to be optimal when the spores were not dried, and adequate stability for H. thompsonii was never found. A large field trial using whole bee hives was conducted with H. thompsonii, but no significant effect was seen, resulted in the abandonment of this fungus as a potential biocontrol agent. On the other hand, M. anisopliae applications were found to significantly increase the overwintering survival of honey bee colonies. Objective 4 - Enhance knowledge of native pollinators by conducting biological surveys, establishing monitoring programs, and expanding knowledge of bee systematics in agricultural and natural systems. Sub-Objective 4.1: Data capture of specimen records of Megachilidae will be collected in support of Objectives 1 & 2. FY2004 Milestone: Develop prototype interactive key to the bees of the Mid-Atlantic States. FY2004 Progress: A prototype interactive key to six genera of Mid-Atlantic bees has been completed and is located on the web at www.discoverlife.org. Interactive keys for additional bee genera are under development. In addition, interactive keys to the genera of the bee family Megachilidae have been completed for North America and East Africa. Sub-Objective 4.2: A multi-year (bee) faunal survey of Grand Staircase- Escalante National Monument initiated in 2000 will continue, with plans for a final field season in FY2004. FY2004 Milestone: Complete field (bee) surveys in Grand Staircase-Escalante National Monument. FY2004 Progress: Field surveys of bees in Grand Staircase-Escalante National Monument were completed in 2004. The specimen data has been entered into a relational database and identifications are in progress. Sub-Objective 4.3: Systematic studies in support of Objectives 1 & 2 will concentrate on candidate pollinator-rich Megachilidae. FY2004 Milestone: Draft revision of North American Stelis (group of parasitic bees of worldwide significance) for publication. FY2004 Progress: A draft revision of Stelis has been completed for the 59 species in North American and progress has been made on revising the manuscript. B: List the milestones (from the list in Question #2) that you expect to address over the next 3 years (FY 2005, 2006, & 2007). What do you expect to accomplish, year by year, over the next 3 years under each milestone? Objective 1 - Deliver improved pollination management systems for alfalfa seed production using the alfalfa leafcutting bee and the alkali bee, and a new pollination management system for orchards using the blue orchard bee. Sub-Objective 1.1: For the two primary alfalfa pollinators, the alkali bee and the alfalfa leafcutting bee, we will determine cost-effective, optimal stocking densities for pollinators that maximize seed production while minimally compromising bee reproduction. FY2005 Milestones: Analyze data and publish provision mass pollen/sugar paper. Repeat bee density study with different densities and analyze data. Continue bee health survey and report to producers. FY2006 Milestones: Publish multi-year nest density study with the alkali bee. Estimate male pollination value, review for bees. Implement large field trials with more "optimal" bee densities defined from enclosure study. Continue bee health survey and report to producers. FY2007 Milestones: Publish carrying capacity and competition paper on the alkali bee. Continue large field trials with optimal bee densities. Continue bee health survey and report to producers. Sub-Objective 1.2: To better enhance nesting of the alfalfa leafcutting bee in clean, artificial nesting boards and to better ensure field fidelity of bees released for pollination, we will investigate the origins of cues that attract bees to previously used nesting boards. FY2005 Milestone: Continue tests with alfalfa leafcutting bees; begin tests with blue orchard bees. Perform chemical analyses of important components. FY2006 Milestone: Test ability to apply attractants to nesting boards. FY2007 Milestone: Conduct field tests of nesting board attractants in all species where applicable and report to producers. Sub-Objective 1.3: Improve summer and pre-wintering temperature regimes for the commercial (and artificial) management of blue orchard bee populations. FY2005 Milestone: Complete summer development studies with the blue orchard bee. FY2006 Milestone: Complete pre-wintering studies with the blue orchard bee. FY2007 Milestone: Complete statistical treatment of data collected, draft manuscripts, and report to producers. Sub-Objective 1.4: Support improvements in pre-wintering and wintering temperature regimes for the commercial (and artificial) management of the alfalfa leafcutting bee and the blue orchard bee through improving our understanding of intermediary metabolic processes and diapause gateways. FY2005 Milestone: Complete analyses and draft manuscript on life cycle respirometry, intermediary metabolism in the alfalfa leafcutting bee under natural conditions. Monitor putative clones for life cycle expression patterns in the alfalfa leafcutting bee. Draft manuscript identifying apparent diapause gateways in the alfalfa leafcutting bee. FY2006 Milestone: Complete life cycle respirometry, supercooling point, amino acid, lipid, and carbohydrate profiles for alfalfa leafcutting bee and blue orchard bee populations under natural and artificial conditions. Create cDNA library for blue orchard bee HSP's via RT-PCR for diapause and non-diapause condition. Sequence putative clones and generate DIG- labeled probes for expression assays via Northern blot. FY2007 Milestone: Complete analyses and draft manuscripts on life cycle respirometry, intermediary metabolism for alfalfa leafcutting bee and blue orchard bees under natural and artificial conditions. Monitor putative clones for life cycle expression patterns in the blue orchard bee. Draft manuscript apparent identifying diapause gateways in the blue orchard bee. Sub-Objective 1.5: Demonstrate the blue orchard bee and new nesting block design in selected early-spring orchard crops. FY2005 Milestone: Complete analysis of data and draft manuscript on UT cherry pollination. Complete year 3 of organic almond pollination demonstration in CA. Complete analysis of prototype performance, and develop modifications to be made to second generation prototype. FY2006 Milestone: Submit final evaluation and recommendation of optimal cherry pollination management program using the blue orchard bee. Complete year 4 of organic almond pollination demonstration in CA. Complete testing of second generation prototype in UT a pear-apple orchard. FY2007 Milestone: Complete year 5 of organic almond pollination demonstration in CA. Complete analysis of data and draft manuscript on organic almond pollination. Prepare final evaluation and recommendation for transfer to mass market. Sub-Objective 1.6: Improve the establishment of artificially managed blue orchard bee populations used for commercial pollination by minimizing the dispersal of pre-nesting females. FY2005 Milestone: Complete second year experiments on wintering temperature, parasitism and body size. Complete first year experiment with non-dispersing (philopatric) females. FY2006 Milestone: Complete second year experiment with philopatric females. FY2007 Milestone: Complete third year experiment with philopatric females. Objective 2 - Evaluate pollination efficacies of confined and free- flying populations of select new bee pollinators for small fruit crops and regional forbs in demand for re-vegetation, and develop practical management protocols to encourage adoption of the more promising species as custom pollinators. FY2005 Milestone: Trap-nest for O. atriventris for cranberry pollination. Trap-nest promising pollinators for wildflowers. Evaluate specific pollination needs of wildflowers. Evaluate trial plots. Sample, ID wildflower seed and flower pests. Publish cranberry pollen tube attrition study. FY2006 Milestone: Test trap-nested bee species for pollination efficacies. Cage pollination trial with O. atriventris on cranberry. Evaluate pollinator efficacy on highbush blueberries. Evaluate overwintering management and needs. Begin publishing wildflower breeding biologies. Try O. ribifloris free-flight in blueberries. FY2007 Milestone: Scale-up trap-nesting and production of promising pollinators. Evaluate bee performance in sundry nest substrates. Begin publishing pollinator efficiencies and conduct field trials of wildflower pollinators. Publish nesting the biology of raspberry pollinator and report to producers. Objective 3 - Develop novel control options and delivery systems for the management of chalkbrood, other diseases, parasites, and predators in commercial-scale pollinator populations. Sub-Objective 3.1: Determine the abundance and diversity of chalkbrood in commercial fields, and conduct a survey of bee larvae to determine the frequency of infection for each pathogen species present. FY2005 Milestone: Sample alfalfa leafcutting bee larvae from commercial fields and assess levels of chalkbrood infection. Refine molecular marker methods for Ascosphaera detection and develop a photo guide to disease in the alfalfa leafcutting bee. FY2006 Milestone: Conduct survey of chalkbrood prevalence in the alfalfa leafcutting bee from commercial alfalfa fields and report to producers. FY2007 Milestone: Conduct survey of chalkbrood prevalence in the alfalfa leafcutting bee from commercial alfalfa fields and report to producers. Sub-Objective 3.2: Develop new control methods for chalkbrood, focusing on chemical fungicides, sanitation methods, and microbial control strategies. FY2005 Milestone: In commercial fields, test applications of systemic fungicides on chalkbrood levels in the alfalfa leafcutting bee and report to producers. FY2006 Milestone: Begin search for mycoviruses that can infect Ascosphaera. FY2007 Milestone: Develop method for culturing and formulating mycoviruses with activity against Ascosphaera. Sub-Objective 3.3: Determine the epizootiology of chalkbrood, and in particular, determine the major means by which the disease is spread and maintained in a bee population. FY2005 Milestone: Measure diurnal and distance-from-a-shelter effects on air loads of Ascosphaera spores. Measure levels of chalkbrood in feral populations of the alfalfa leafcutting bee. Continue basic biology studies on Ascosphaera aggregata. FY2006 Milestone: Measure diurnal and distance-from-a-shelter effects on air loads of Ascosphaera spores. Estimate population influx of "wild bees" into commercial nesting blocks and report to producers. FY2007 Milestone: Correlate Ascosphaera spore loads on adults with chalkbrood levels in progeny. Develop quantitative PCR methods for detecting Ascosphaera spores and report to producers. Sub-Objective 3.4: Develop microbial control strategies for controlling predators and parasites in bees. FY2005 Milestone: Conduct field trials of microbial control of varroa in honey bees. Develop a laboratory population of the checkered flower beetle. FY2006 Milestone: Develop bioassays for the checkered flower beetle, test fungi for pathogenicity. Conduct non-target tests of the candidate fungi on bees. FY2007 Milestone: Field test microbial control strategy for predatory beetles in commercial fields and report to producers. Objective 4 - Enhance knowledge of native pollinators by conducting biological surveys, establishing monitoring programs, and expanding knowledge of bee systematics in agricultural and natural systems. Sub-Objective 4.1: Data capture of specimen records of Megachilidae will be collected in support of Objectives 1 & 2. FY2005 Milestone: Capture collection records of Osmia lignaria, O. ribifloris, and O. aglaia. Deliver web-based interactive key to the bees of the Mid-Atlantic states. FY2006 Milestone: Complete data capture of U.S. Megachilidae in PIRU collection. FY2007 Milestone: Develop prototype interactive key to the bee genera of the U.S. Sub-Objective 4.2: A multi-year faunal survey of Grand Staircase- Escalante National Monument initiated in 2000 will continue, with plans for a final field season in 2003. FY2005 Milestone: Analyze faunal data as requested by Grand Staircase-Escalante Natl. Mon. FY2006 Milestone: Analyze faunal data as requested by Grand Staircase-Escalante Natl. Mon. Draft preliminary report on multi-year results. FY2007 Milestone: Analyze faunal data as requested by Grand Staircase-Escalante Natl. Mon. and draft final report. Sub-Objective 4.3: Systematic studies in support of Objectives 1 & 2 will concentrate on candidate pollinator-rich Megachilidae. FY2005 Milestone: Draft revision of North American Dufourea for publication. FY2006 Milestone: Draft revision of bright metallic Osmia for publication. FY2007 Milestone: Conduct phylogenetic analysis of Megachilidae and draft manuscript. 4. What were the most significant accomplishments this past year? A. Single most significant accomplishment during FY 2004 (one per Research (OOD) Project): R. R. James determined the thermal limits for Ascosphaera aggregata growth and spore germination, and was surprised to find that spores can germinate at temperatures as high as 45DGC; spores germinated at this temperature soon die, however, and the maximum temperature for sustained (and maximum) growth was determined to be 35DGC. Ascosphaera aggregata is a fungal disease of the alfalfa leafcutting bee that is widely used in the production of alfalfa seed in North America. The effect of temperature on disease development was also determined, and the optimum temperature for in vitro growth was the condition that led to the least number of infected hosts, indicating that some kind of interaction between the host and the pathogen occurred at high temperatures which inhibited the ability of the fungus to cause disease. Temperatures above 25DGC can be stressful to the host, and so James hypothesizes that one or more temperature stress responses may have an added benefit of acting as a defense against pathogen invasion, such as the induction of prophenol oxidases that are associated with the immune response in insects. These results have direct impact on the development of disease management protocols in commercial scale alfalfa leafcutting bee populations throughout the western United States. B. Other significant accomplishment(s): T. L. Pitts-Singer and R.R. James revised and improved the regional Alfalfa Leafcutting Bee Health Survey during 2004. This project began in 2002, was expanded during 2003 and 2004 at the request of the Northwest Alfalfa Seed Growers Association, and will continue for at least one more year. Sampling "kits" are currently sent to 20 different alfalfa seed producers and pollination providers, representing 25 commercial operations throughout western North America, at regular intervals throughout the year. Survey results have already contributed to the publication of an article by Pitts-Singer describing various categories of immature bee death and emphasizing the importance of recognizing the categories in order to determine the causes of such mortality. With addition of 2004 survey results, Pitts-Singer will report to seed producers on adult bee survival and emergence after incubation, comparing emergence in the laboratory to emergence under conditions provided by bee managers participating in the survey. Survey results have direct impact on the improvement of management systems for commercial scale alfalfa leafcutting bee populations throughout the western United States. J. H. Cane expanded investigations on the pollination efficacy of the alkali bee (Nomia melanderi) a regionally important pollinator of alfalfa production systems. Survey results indicate that few if any growers rely solely on the alkali bee, however; growers typically use the alkali bee in combination with the alfalfa leafcutting bee, Megachile rotundata. Both bees effectively pollinate seed alfalfa, at least when measured for single floral visits by individual bees. Cane used a measure of pollination ability of individual alkali bees to extrapolate to large populations (1.4 million nesting females) pollinating an 80 acre commercial field of seed alfalfa in WA. On average, 43% of flowers pollinated by alkali bees produced a seed pod. This proportion held up across the entire field, with no reduction of curl set with distance. This compares well with earlier greenhouse pollination studies, wherein 48% of tripped flowers set pods. The grower reports harvesting 1000 lbs/acre of clean seed off of this 2-year-old field, despite imperfect irrigation later in the season. Thus, it appears that pod production resulting from pollination by alkali bees was near alfalfa's upper limit in this field, even 12 mile distant from the source aggregation of its pollinator, the alkali bee. At these stocking densities, purchased leaf- cutting bees are superfluous and may in fact deter population growth by the alkali bee. These results have direct impact on the improvement of management systems for commercial scale alfalfa pollinator populations throughout Great Basin and nearby regions of the western United States. The Bureau of Land Management and the U.S. Forest Service annually broadcast 3-10 million pounds of grass, shrub and forb seed across public lands in the Great Basin to promote re-vegetation on degraded or burned rangeland and forest. Current seed mixes contain <1% forb seed, far short of the annual need for up to 2 million pounds of select native wildflower seed. In order to develop the technology to obtain such quantities in the future, J.H. Cane is participating in an ongoing interdisciplinary research program to establish successful agricultural production and marketing of eight or more broadly adapted wildflower species native to the Great Basin and adjacent ecological regions. In 2004, far-flung wild populations of a highly desired species now added by BLM were located and their bee faunas systematically sampled, in addition to those of 3 other species not previously sampled. Native bees continue to be the dominant visitors for all but one of the species (for which a wasp and now 2 native bees have been found primary). Bees of the genus Osmia dominate faunas at each location and for all but one species, which primarily hosts a specialist species of Andrena. Manual pollination experiments with a third wildflower species (Cleome lutea) shows the necessity of bee visitation for fruit and seed set; it is fully self-compatible. Forty nesting blocks at 8 locations were recovered across a 3000' elevation gradient, their contents overwintered and identified. Fresh nesting blocks are being returned to the productive sites. Three of these trap- nested bee species, all candidate pollinators, prospered when confined to northern sweetvetch. Cane has successfully propagated 1- and 2-yr-old plants of 4 wildflower species now, which have been transplanted to field cage sites. Growth has been rapid, bloom sporadic, so they will be usable for pollination trials in 2005. During 2004, T. L. Griswold established three new biological surveys of native bees: Dugway Proving Grounds (UT), eastern Mojave Desert (NV), and Yosemite National Park (CA). All were solicited by the agencies represented and largely funded by them. The surveys will assist land managers in their efforts to conserve bees and the pollination services that native bees provide, as well as establish baseline information to assess current and future impacts of human activities on lands under Federal management. W.P. Kemp and T. L. Pitts-Singer completed the initial (field) performance testing of the blue orchard bee "Solitary Bee Emergence Box" in CA and UT and submitted an Invention Disclosure (#0051.04), which has received provisional approval for the preparation of a patent submission by the Northern Plains, Southern Plains and Midwest Areas Patent Committee. The emergence box allows for the improved control of pollinator emergence during the bloom period of early spring tree fruit crops such as almonds and cherries, which is frequently punctuated by periods of inclement weather not conducive to bee emergence, and is designed to be used together with the previously patented Solitary Bee Nesting Block. The emergence box is the second in an anticipated series of three patents which will define a standardized system for managing the blue orchard as a reliable pollinator in U.S. tree fruit crops. C. Significant activities that support special target populations. None D. Progress Report opportunity to submit additional programmatic information to your Area Office and NPS (optional for all in-house ("D") projects and the projects listed in Appendix A; mandatory for all other subordinate projects). None 5. Describe the major accomplishments over the life of the project, including their predicted or actual impact. The 60-month Project Plan for the PIRU was initiated 1 October 2003. J.H. Cane is participating in an ongoing interdisciplinary research program to establish successful agricultural production and marketing of eight or more broadly adapted wildflower species native to the Great Basin and adjacent ecological regions. The Bureau of Land Management and the U.S. Forest Service annually broadcast 3-10 million pounds of grass, shrub and forb seed across public lands in the Great Basin to promote re- vegetation on degraded or burned rangeland and forest. Current seed mixes contain <1% forb seed, far short of the annual need for up to 2 million pounds of select native wildflower seed. Dr. Cane's responsibility is the determination of each forb species' breeding biology and pollination needs, the evaluation of its bee fauna for potentially manageable species, and development of protocols and starting populations of effective pollinators that can be managed in an on-farm setting for profitable forb seed production. To date, Dr. Cane has found that the preferred forb species differ widely in the diversity of native bee species that they attract and all but a few require bee visitation to set seed. Manageable captive populations of 2 Osmia species are in hand for pollinating the first of the desired forb species. The National Park Service has identified the inventory of native pollinators as a high priority for management planning. In response to this need T.L. Griswold is providing information on native pollinators to land managers, and recently completed the fourth year of a survey of the bee pollinators of Grand Staircase-Escalante National Monument. This work has highlighted the importance of long-term studies for monitoring the health of pollinator services. Similar projects are currently underway in the John Muir National Historic Site and Great Smoky Mountains National Park. The expanding demand for this information among Federal land management agencies has led to the development of joint efforts on native pollinators and their impact on rare plants, fire and grazing management in Zion National Park, Yosemite National Park and with the Bureau of Land Management Clark County, NV. Griswold also leads the development of a world checklist of bees at the request of the U.S. Geological Survey. A prototype using one subfamily of bees was initiated and completed during 2004. 6. What science and/or technologies have been transferred and to whom? When is the science and/or technology likely to become available to the end- user (industry, farmer, other scientists)? What are the constraints, if known, to the adoption and durability of the technology products? J.H. Cane in collaboration with an alfalfa seed producer has demonstrated a new sub-irrigation technology for an alkali bee nesting bed. This technology has been found to be effective in delivering adequate and unimpeded subsurface moisture. Working with an advanced soils class at Utah State University, its performance was modeled against current technology, showing that the new system uses 90% less water than the current system, has better lateral dispersion, and is much less likely to contaminate ground-water with salts leached from the surface. Cost is comparable with current systems. Stable or in a few cases increasing nesting populations of alkali bees have resulted from informing alfalfa seed growers of minimum blooming alfalfa acreages calculated to fulfill bees' foraging needs of their respective (censused) nesting aggregations in southeastern WA. This has protected an estimated 13 million nesting individuals (yielding 5 million pounds of seed) despite acreage withdrawals owing to seed surpluses as well as cheap alfalfa leaf-cutting bees. The alkali bee has been shown to effectively pollinate commercial yellow Spanish onion for hybrid seed production. Adoption awaits industry interest. The research team of W.P. Kemp has taken important steps to transfer blue orchard bee technology to the U.S. fruit sector, including: a) The publication of an 88-page manual ("How to manage the blue orchard bee as an orchard pollinator"), b) The patent of a new, affordable and manageable polycarbonate-based nesting block for the blue orchard bee and the development of a related prototype emergence box, c) The completion of a six-year project demonstrating highly-significant yield increases in a commercial cherry orchard pollinated with blue orchard bees, d) The publication of research on successful methods to advance blue orchard bee emergence for almond pollination, and e) The publication or preparation of several research articles and two CRADAs on the developmental biology and rearing methods (wintering requirements, development and incubation temperature regimes, respirometry) of the blue orchard bee. Current research is emphasizing diapause induction and development, respirometry, and diapause-associated gene expression, as well as blue orchard bee toxicology as related to fungicide sprays during bloom, and the use of visual and olfactory cues involved in nest location. The sustainability of this technology is high and the adoption is expanding. The research team of W. P. Kemp has transferred the results from a series of experiments that revealed that much can be done to improve nest location in commercial populations of the alfalfa leafcutting bee in seed production systems. These experiments showed that the use of the third dimension in nesting board design can significantly reduce the percentage of wrong-hole visitations by alfalfa leafcutting bees. Separating the alfalfa leafcutting bee nesting boards when setting them up nesting shelters is a simple way to provide additional orientation cues and decrease wrong hole visitations by actively nesting females. Other ways to improve nest location include setting boards at different depths within the shelter (some advanced and some recessed), placing them in irregular patterns (some vertical and some horizontal), or developing the manufacturing capability to produce nesting boards with a 3-dimensional front surface. Reducing wrong hole visitations will decrease nest location times, which in turn will positively influence pollination efficiency, brood production, and, ultimately, seed yields. A large number of seed producers are now placing nesting boards in configurations that will provide more orientation cues for nesting females, and at least two commercial nesting board producers have expressed interest in the development of nesting boards with a 3-dimensional front face. 7. List your most important publications in the popular press and presentations to organizations and articles written about your work. 1. Kemp, W.P. 2003. Invited presentation. 11th National Small Farm Trade Show and Conference. Columbia, MO. November 6-8, 2003. 2. Kemp, W.P. 2004. Invited presentation California Rare Fruit Growers. Pamona, CA. June 18-19, 2004. 3. Kemp, W.P. 2004. Invited presentation. Heartland Apiculture Society Third Conference. Lebanon, TN. July 8-10, 2004. 4. Cane, J.H. 2004. Invited presentation/poster. National Alfalfa Seed Growers. Reno, NV. January 19-21, 2004. 5. James, R.R. 2004. Invited presentation/poster. National Alfalfa Seed Growers. Reno, NV. January 19-21, 2004. 6. Pitts-Singer, T.L. 2004. Invited presentation/poster. National Alfalfa Seed Growers. Reno, NV. January 19-21, 2004. 7. Pitts-Singer, T.L. 2004. Invited presentation. Montana Alfalfa Seed Growers Annual meeting. Billings, MT. February 19, 2004. 8. Pitts-Singer, T.L. 2004. Invited presentation. Wyoming Alfalfa Seed Growers Annual meeting. Cody, WY. February 18, 2004.

Impacts
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Publications

Bosch, J., Kemp, W.P. 2003. Effect of wintering duration and temperature on survival and emergence time in the orchard pollinator Osmia lignaria (Hymenoptera: Megachilidae). Environmental Entomology. 32(4):711-716.
Ladurner, E., Bosch, J., Maini, S., Kemp, W.P. 2003. A method to feed bees (Hymenoptera: Apiformes) known amounts of pesticides. Apidologie. 34:597- 602.
Tepedino, V.J., Messinger, S.M. 2004. Cymopterus beckii, a rare protogynous umbellifer of capital reef national park, central Utah. Madrono. 51(3:271-274.
Cane, J.H., Schiffhauer, D. 2003. Dose-response relationships between pollination and fruiting refine pollinator comparisions for cranberry (Vaccinium macrocarpon ait.). American Journal of Botany. 90:1425-1432
Arneson, L.C., Tepedino, V.J., Smith, S.L. 2004. Reproductive success of a rare fire-follower (Iliamna bakeri: Malvaceae), and its association with a native specialist bee (Diadasia nitidifrons: Apidae), in northeastern California. Northwest Science.78(2):141-149.
James, R.R. 2004. Chalkbrood in the alfalfa leafcutting bee: dna methods for detecting infections. National Entomological Society of America Annual Meeting, October 26-29, 2003, Cincinnati, OH.
Nguyen, K.B., Shapiro Ilan, D.I., Stuart, R.J., James, R.R., Adams, B.J. 2004. Heterorhabditis mexicana n. sp. (heterorhabditidae: rhabditida) from tamaulipas, mexico, and morphological studies of the bursa of heterorhabditis spp. Nematology. 6:231-244.
Stuart, R.J., Shapiro Ilan, D.I., James, R.R., Nguyen, K.B., Mccoy, C.W. 2004. Virulence of new strains of the entomopathogenic nematode steinernema riobrave to larvae of the citrus root weevil diaprepes abbreviatus. Biological Control. v.30. p.439-445.
James, R.R., Buckner, J.S., Freeman, T.P. 2003. Cuticular lipids and silverleaf whitefly stage affect conidial germination of Beauveria bassiana and Paecilomyces fumosoroseus. Journal of Invertebrate Pathology. 84:67-74.
Kemp, W.P., Bosch, J., Dennis, B. 2004. Oxygen consumption during the life cycle of the prepupa-wintering bee Megachile rotundata (f.) and the adult- wintering bee Osmia lignaria say (Hymenoptera: Megachilidae. Annals of the Entomological Society of America.97(1):161-170.
Kanga, L.H., James, R.R., Jones, W.A. 2003. Field trials using the fungal pathogen, Metarhizium anisopliae (Deutermycetes: Hyphomycete) to control the ectoparasitic mite, Varroa destructor (Acari: Varroidae) in honey bee colonies. Journal of Economic Entomology. 96(4):1091-1099.
Cane, J.H. 2003. Exotic non-social bees (hymenoptera: apoidea) in north America: ecological implications. In:Strickler, K., Cane, J.H.,editors. For nonnative crops, whence pollinator for the future? Lanham, MD. Thomas Say Publications in Entomology. p.113-126
Strickler, K., Cane, J.H. 2003. Introduction. In: Strickler, K., Cane, J.H. ,editors. For non-native crops, whence pollinators of the future?. Lanham, MD. Thomas Say Publications in Entomology. p.1-9.
Strickler, K., Cane, J.H. 2004. For nonnative crops, whence pollinators of the future?. Lanham, MD. Thomas Say Publications in Entomology. 204.
Pitts, J.P., Parker, F.D., Pitts Singer, T. 2004. A review of the sphaeropthalma uro species-group (hymenoptera: mutillidae), with taxonomic changes. Journal of Kansas Entomological Society.77(3):222-234.
Mcleay, D., Genthner, F., James, R.R., Lazarovits, G., Percy, D. 2004. Guidance document for testing the pathogenicity and toxicity of new microbial substances to aquatic and terrestrial organisms. Laboratory Publication.Report number EPS 1/RM/44. Environment Canada, Ottawa, Ontario, Canada. 171