Green fluorescent protein (GFP)-labeling of enterobacteria associated with fruit flies (Diptera: Tephritidae) and persistence in their natural host Rhagoletis completa Cresson

Fruit flies (Diptera: Tephritidae) are a highly successful, widespread group of insects that cause economic damage in agriculture. Data available so far on the composition of the bacterial community associated with their digestive tract indicate that members of Enterobacteriaceae are the species most often isolated. Bacteria naturally occurring in insect guts may be engineered and used to study the spatial and functional interactions of microbes within the insect system and offer one route to meet the demand for novel insect pest management strategies. With this aim we introduced by conjugation the gfp gene carried by the suicide plasmid pTn5gfpmut1 into Klebsiella oxytoca and Raoultella (formerly Klebsiella) spp. strains isolated from the oesophageal bulb of the fruit flies Ceratitis capitata (Wiedemann) and Rhagoletis completa Cresson, respectively. The GFP-encoding gene was stably maintained in two tested transgenic strains, both originally isolated from R. completa. In one case, GFP-labeled bacterial cells were used to feed larvae and adults of the original host. Genetically modified bacteria were able to colonize the gut of larvae and persisted through all larval instars to pupal stage.