Transinfection of buffalo flies (Haematobia irritans exigua) with Wolbachia and effect on host biology

Buffalo flies (Haematobia irritans exigua) (BF) and closely related horn flies (Haematobia irritans irritans) (HF) are invasive haematophagous parasites with significant economic and welfare impacts on cattle production. Wolbachia are intracellular bacteria found widely in insects and currently of much interest for use in novel strategies for the area wide control of insect pests and insect-vectored diseases. In this paper, we report the transinfection of BF towards the development of area-wide controls. Three stages of BF; embryos, pupae and adult female flies, were injected with different Wolbachia strains (wAlbB, wMel and wMelPop). The success of transinfection and infection dynamics was compared by real-time PCR and FISH and fitness effects were assessed in transinfected flies. BF eggs were not easily injected because of their tough outer chorion and embryos were frequently damaged with less than 1% hatch rate of microinjected eggs. No Wolbachia infection was recorded in flies successfully reared from injected eggs. Adult and pupal injection resulted in higher survival rates and somatic and germinal tissue infections, with transmission to the succeeding generations on some occasions. Investigations of infection dynamics in flies from injected pupae confirmed that Wolbachia were actively multiplying in somatic tissues. Ovarian infections were confirmed with wMel and wMelPop in a number of instances, though not with wAlbB. Measurement of fitness traits indicated reduced longevity, decreased and delayed adult emergence, and reduced fecundity in Wolbachia-infected flies compared to mock-injected flies. Effects varied with the Wolbachia strain injected with most marked changes seen in the wMelPop-injected flies and least severe effects seen with wAlbB. Adult and pupal injection were the most suitable methods for transinfecting BF and all three strains of Wolbachia successfully replicated in somatic tissues. The Wolbachia-induced fitness effects seen in transinfected BF suggest potential for use of the wMel or wMelPop strains in Wolbachia-based biocontrol programmes for BF.