Several find more recent studies reported that Wolbachia genes, in some cases even large chromosomal segments, have been horizontally transferred to host chromosomes. Such events have been described in a variety of insect and nematode hosts, including the adzuki bean beetle Callosobruchus chinensis, the fruit fly Drosophila ananassae, a parasitoid wasp of the genus Nasonia, the mosquito Aedes aegypti, the pea aphid Acyrthosiphon pisum, the longicorn beetle Monochamus alternatus and filarial nematodes of the genera Onchocerca,

Brugia and Dirofilaria [45–52]. Interestingly, some of these genes are highly transcribed suggesting that laterally transferred bacterial genes can be of functional importance [48–50]. In the present study, we report on the presence of Wolbachia infections in laboratory and natural populations of Glossina species. The characterization of these Wolbachia strains is based on the use of 16S rRNA, wsp and MLST gene markers. In addition, we report horizontal gene transfer events of Wolbachia genes to G. m. morsitans chromosomes. Methods Sample collection and DNA isolation Glossina specimens were collected in ten countries in Africa (Tanzania, South Africa, Zambia, Zimbabwe, Kenya, Senegal, Guinea, Ethiopia,

Uganda, and Democratic Republic of Congo – Zaire). Upon NVP-AUY922 concentration their arrival in the lab, all tsetse flies specimens have been immediately used for DNA extraction. DNA samples were stored at -20oC until their use. Laboratory strains from FAO/IAEA (Seibersdorf), Yale University (EPH), Slovak Academy of Sciences (SAS-Bratislava), Kenya (KARI-TRC), Burkina Faso (CIRDES) and Antwerp were also included in the analysis. DNA from adult flies was isolated according to Abd-Alla et al. 2007 [53], using the Qiagen DNeasy kit (Qiagen, Valencia, CA), following the manufacturers’ Methane monooxygenase instructions, except

for the samples from Antwerp and Bratislava, to which the CTAB (Cetyl trimethylammonium bromide) DNA isolation method was applied [54]. G. m. morsitans fertile females were maintained on blood meals supplemented with 10% (w/v) yeast extract (Becton Dickinson) and 20 ug/ml of tetracycline. Flies were fed every 48h for the duration of their life span. The resulting progeny are aposymbiotic (GmmApo) in that they lack their natural endosymbionts, Wigglesworthia and Wolbachia (Alam and Aksoy, personal communication). Aposymbiotic progeny were used for detection of nuclear Wolbachia DNA. PCR screen and MLST A total of 3750 specimens of nine Glossina species (G. m. morsitans, G. m. centralis, G. austeni, G. brevipalpis, G. pallidipes, G. p. palpalis, G. p. gambiensis, G. fuscipes fuscipes and G. tachinoides) were screened for the presence of Wolbachia strains.