Experimental validation of a rearing protocol for laboratory assays utilizing Osmia lignaria Say (Hymenoptera: Megachilidae)

Authors: Williams, Mary-Kate; BOYLE, NATALIE - Pennsylvania State University; SCHAEFFER, ROBERT - Utah State University; Cox-Foster, Diana

Submitted to: Journal of Insect Science
Publication Type: Pre-print Publication
Publication Acceptance Date: 7/23/2025
Publication Date: 10/6/2025
Citation: Williams, M.F., Boyle, N.K., Schaeffer, R.N., Cox-Foster, D.L. 2025. Experimental validation of a rearing protocol for laboratory assays utilizing Osmia lignaria Say (Hymenoptera: Megachilidae). Journal of Insect Science. Volume 25, Issue 5. https://doi.org/10.1093/jisesa/ieaf081.
DOI: https://doi.org/10.1093/jisesa/ieaf081

Interpretive Summary: The blue orchard bee (Hymenoptera: Megachilidae: Osmia lignaria Say) is a solitary bee native to North America that is increasingly used, propagated, and sold for pollination of orchard crops. While methods exist to rear blue orchard bees in a laboratory setting, the current protocols are inconsistent, and most do not follow bee development to adult emergence. Variability in these methods makes comparisons challenging. Here, we present a protocol that was used to rear the blue orchard bee in a laboratory setting under environmental conditions for northern Utah. Our protocol used 3-D printed well plates with an well diameter tailored for propagating the blue orchard bees. Each well permits observation of individual bees without disturbing their development. To validate the protocol, we monitored the development across larval instars, prepupal, pupal, and adult stages using a dissection scope and X-ray imaging. Although we observed variation in bee's development and percent weight loss between study years, we can fairly reliably rear bees to adult emergence,. Our protocol can be altered to fit any experiment and adapted to investigate other above-ground cavity-nesting bee species in a laboratory setting. Additionally, our methods can be used to explore how multiple factors like pesticides and pathogens affect bee health.

Technical Abstract: The blue orchard bee (Hymenoptera: Megachilidae: Osmia lignaria Say) is a solitary bee native to North America that is increasingly used, propagated, and sold for pollination of rosaceous orchard crops. While methods exist to rear blue orchard bees in a laboratory setting, present protocols are inconsistent, vary in diet manipulation, and most do not follow bee development to adult emergence. Variability in published methods makes standardized comparisons within or across species challenging. Here, we present a replicable protocol that was employed over two years to rear O. lignaria in a laboratory setting while mirroring their ecology from northern Utah (United States). Our protocol used 3-D printed well plates with an appropriate well diameter size recommended for propagating O. lignaria. Each well permits the user to observe detailed life stages of individual O. lignaria bees without disturbing their development. To validate the protocol, we monitored O. lignaria development across three larval instars, prepupal, pupal, and adult stages using a dissection scope and X-ray imaging. Although we observed variation in O. lignaria’s development and percent weight loss between study years, we can fairly reliably rear bees to adult emergence, where approximately 49% successfully emerged. Our protocol can be altered to fit any experiment and adapted to investigate other above-ground cavity-nesting bee species in a laboratory setting. Additionally, our methods can be used to explore how multiple factors and stressors influence bee health, where present research appears limited.