|Kemp, William - Bill|
Submitted to: Archives of Insect Biochemistry and Physiology
Publication Type: Peer reviewed journal
Publication Acceptance Date: 5/2/2004
Publication Date: 9/1/2004
Citation: 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. Interpretive Summary: There is a need to increase the number of bee species used for agricultural pollination and a need to improve existing bee management systems. The alfalfa leafcutting bee, Megachile rotundata, is the bee species of choice for alfalfa seed production throughout western North America. Emphasis has been placed on the development of wintering procedures for the prepupal stage of development and improved incubation schedules to synchronize the emergence of M. rotundata adults with alfalfa bloom. The energy required for winter survival, post-diapause development, and the emergence of adult insects comes from intermediary metabolites that include stored internal lipids. Lipid extraction studies revealed that overwintering M. rotundata prepupae possessed copious quantities of internal lipids (20% of the fresh weight). High performance liquid chromatography-mass spectrometry showed that 88% of the internal lipids were a mixture of over 40 triacylglycerol components. Most of the triacylglycerol fatty acids contained one or more double bonds (76%) and the major fatty acid constituent (30%) was the monounsaturated fatty acid, palmitoleic acid (hexadec-9-enoic acid). The findings suggest that the large quantities of intracellular-stored triacyglycerols contribute to the overwintering physiology of M. rotundata by reducing intracellular water and thus increasing cyroprotectant concentrations, and likely serve an important bioenergetic role for pupal and emerged adult development.
Technical Abstract: Alfalfa leafcutting bees, Megachile rotundata (F.), overwinter as prepupae. The internal lipids were extracted from prepupae that had been wintered at 4 deg. C for seven months. Megachile rotundata prepupae possessed copious quantities of internal lipids (20% of the fresh weight) that were extracted with CHCl3/methanol (2:1). Transmission electron microscopy revealed that lipids were stored within very large intracellular vacuoles. Separation by silica chromatography revealed that 88% of the internal lipids were triacylglycerols. Ester derivatives of fatty acids from triacylglycerol components were analyzed by gas chromatography-mass spectrometry and 15 fatty acid constituents were identified. The majority (76%) of the triacylglycerol fatty acids were unsaturated fatty acids. The major triacylglycerol fatty acid constituent (30%) was the C16 monounsaturated fatty acid, palmitoleic acid (16:1, hexadec-9-enoic acid), with substantial amounts of linolenic acid (18:3, octadec-9,12,15-trienoic acid, 15%), palmitic acid (16:0, hexadecanoic acid, 14%) and oleic acid (18:1, octadec-9-enoic acid, 13%). Palmitoleic acid as the major fatty acid of an insect is an unusual occurrence as well as the presence of the 16-carbon polyunsaturated fatty acids, 16:2 and 16:3. The major intact triacylglycerol components were separated and identified by high performance liquid chromatography-mass spectrometry. A complex mixture of approximately 40 triacylglycerol components were identified and major components included palmitoyl palmitoleoyl oleoyl glycerol, palmitoyl palmitoleoyl palmitoleoyl glycerol, myristoyl palmitoleoyl palmitoleoyl glycerol, myristoleoyl palmitoyl palmitoleoyl glycerol and palmitoyl palmitoleoyl linolenoyl glycerol. The function of these internal lipids and their relevance to winter survival and post-wintering development of M. rotundata is discussed.