Wolbachia successfully replicate in a newly established horn fly, Haematobia irritansirritans (L.) (Diptera: Muscidae) cell line

Abstract

BACKGROUND: Haematobia spp., horn flies (HF) and buffalo flies (BF), are economically important ectoparasites of dairy and beef cattle. Control of these flies relies mainly on treating cattle with chemical insecticides. However, the development of resistance to commonly used compounds is compromising the effectiveness of these treatments and alternative methods of control are required. Wolbachia are maternally transmitted endosymbiotic bacteria of arthropods that cause various reproductive distortions and fitness effects, making them a potential candidate for use in the biological control of pests. The first step towards this is the establishment and adaptation of xenobiotic infections of Wolbachia in target host cell lines.RESULTS: Here, we report the successful establishment of a continuous HF cell line (HIE-18) from embryonic cells and its stable transinfection with Wolbachia strains wAlbB native to mosquitoes, and wMel and wMelPop native to Drosophila melanogaster. HIE-18 cells were typically round and diploid with ten chromosomes (2n = 10) or tetraploid with 20 chromosomes (4n = 20), with a doubling time of 67.2 h. Wolbachia density decreased significantly in HIE-18 cells in the first 48 h of infection, possibly due to overexpression of antimicrobial peptides through the Imd immune signalling pathway. However, density recovered after this time and HIE-18 cell lines stably infected with the three strains of Wolbachia have now each been subcultured more than 50 times as persistently infected lines.CONCLUSION: The amenability of HF cells to infection with different strains of Wolbachia and the establishment of stable sustaining infections suggest the potential for use of Wolbachia in novel approaches for the control of Haematobia spp. Further, the availability of the HIE-18 cell line will provide an important resource for the study of genetics, host-parasite interactions and chemical resistance in Haematobia populations.

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

“…A similar result was obtained when wMelPop and wAlbB were injected into Anopheles gambiae adult mosquitoes [59]. The initial host immune response was anticipated as the densities of wAlbB, wMel and wMel-Pop Wolbachia in Haematobia cells were also observed to initially decrease, possibly due to an innate immune response mediated by the Imd pathway [48]. Real-time PCR analysis of dissected tissues nine days after injection showed Wolbachia to be present in all the vital somatic tissues (head, thoracic muscle, midgut and fat body), except for the ovarial tissues, suggesting that Wolbachia might need extra time to infect the ovaries.…”

“…As oviposition by BF may begin as early as three days after eclosion from the pupae and continue until death, knowledge of Wolbachia distribution and dynamics in injected females was critical for us to identify the optimal timing for collecting infected eggs for the establishment of an infected colony (11-15 days). Wolbachia density significantly decreased to day five possibly due to host immune response [48] but recovered by day eleven after injection. A similar result was obtained when wMelPop and wAlbB were injected into Anopheles gambiae adult mosquitoes [59].…”

“…Infected JW-18 cell lines were used for isolation of Wolbachia for the first three adult injection and embryonic microinjection batches. The Haematobia embryonic cell line (HIE-18) maintained in our laboratory without the use of antibiotics were transinfected with wAlbB (wAlbB-HIE-18), wMel (wMel-HIE-18) and wMelPop (wMelPop-HIE-18) as above [48]. The infected HIE-18 lines were cultured in 75 cm 2 flasks containing 12 ml of Schneider's medium supplemented with 10% FBS at 28 °C and subcultured every 5-6 days by splitting at a ratio of 1:2 into new flasks (Sigma Aldrich, NSW, Australia).…”

“…Hence, we established the HIE-18 cell line from HF to adapt wAlbB obtained from mosquito, wMel and wMelPop from Drosophila into the Haematobia spp. context prior to commencing BF microinjection [48].…”

Gut bacterial communities across 12 Ensifera (Orthoptera) at different feeding habits and its prediction for the insect with contrasting feeding habits

“…Surprisingly, Wolbachia was the only dominant genus in A.cinerea and accounted for 49.46%, but was unobserved in all Ensifera samples. Although there were significant interactions between Wolbachia and host from parasitic to mutualistic, the most frequently noted parasitic effects were reproductive manipulation by male-killing, feminisation, sperm-egg incompatibility, cytoplasmic incompatibility and parthenogenesis [54,55]. Therefore, we hypothesised that Wolbachia could be used as a biological control for Ensifera pests [56].…”