Isolation in Natural Host Cell Lines of Wolbachia Strains wPip from the Mosquito Culex pipiens and wPap from the Sand Fly Phlebotomus papatasi

Endosymbiotic intracellular bacteria of the genus Wolbachia are harboured by many species of invertebrates. They display a wide range of developmental, metabolic and nutritional interactions with their hosts and may impact the transmission of arboviruses and protozoan parasites. Wolbachia have occasionally been isolated during insect cell line generation. Here, we report the isolation of two strains of Wolbachia, wPip and wPap, during cell line generation from their respective hosts, the mosquito Culex pipiens and the sand fly Phlebotomus papatasi. wPip was pathogenic for both new C. pipiens cell lines, CPE/LULS50 and CLP/LULS56, requiring tetracycline treatment to rescue the lines. In contrast, wPap was tolerated by the P. papatasi cell line PPL/LULS49, although tetracycline treatment was applied to generate a Wolbachia-free subline. Both Wolbachia strains were infective for a panel of heterologous insect and tick cell lines, including two novel lines generated from the sand fly Lutzomyia longipalpis, LLE/LULS45 and LLL/LULS52. In all cases, wPip was more pathogenic for the host cells than wPap. These newly isolated Wolbachia strains, and the novel mosquito and sand fly cell lines reported here, will add to the resources available for research on host–endosymbiont relationships, as well as on C. pipiens, P. papatasi, L. longipalpis and the pathogens that they transmi