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This protocol describes standard rearing of N. vitripennis strains on Sarcophaga bullata hosts. By using incubators at different temperatures, Nasonia’s development rate can be adjusted to conform to the investigator’s schedule. Nasonia will produce diapause larvae when reared at 18C with a short-day light cycle (Saunders 1965); these can be archived at 4C for over a year. Diapausing Nasonia may lose their Wolbachia infections (Perrot-Minnot et al. 1996). N. longicornis, N. giraulti and N. oneida can be reared under similar conditions, with the caveat that N. giraulti and N. oneida lay all-diapause broods more frequently and are therefore best reared at 25C.
Problem: low sex ratio (mostly males)
Solution: Crowding by multiple males may prevent some females from mating, who then produce all male broods and repeat the cycle. Remove as many females as possible using paintbrush, or, if necessary, knock out with CO2 and sort. Separate up to 20 females into a new vial, add 2–4 males, paint a dot of honey water on the wall of the tube, and allow mating to occur for 4h or overnight. All-male broods means no female mated. Recover strains from a culture backed up at 4C.
JHW and DL acknowledge support from the NIH 1 R24 GM084917-01 and assistance from Rachel Edwards, Jon Giebel, Michael Clark and Rhitoban Raychoudhury.