3b). The Protein Tyrosine Kinase inhibitor Wolbachia-free G. m. morsitans line contained only the

smaller 453 bp version of the fbpA gene, suggesting again that this gene fragment is the result of a horizontal gene transfer event to the host chromosome. Figure 3 Overview of deleted fragments in two Wolbachia genes A) PCR amplified products from G. m. morsitans (GmmY and Gtet) of the 16S rRNA and fbpA genes were resolved on 2.5% agarose gels stained with ethidium bromide. A 100-bp ladder was used as size standard. The input of the negative (neg) control was water. B) 16S rRNA and fbpA fragments from tsetse flies Wolbachia strains aligned with the corresponding regions of strain wMel. Red dashes represent the deletion region, the numbers show the positions before and after the deletions in respect to the wMel genome. The blue arrows

represent the corresponding wMel genes. Deleted fragments were detected in G. m. morsitans samples (Gmormor: GmmY, 12.3A, 24.4A, 30.9D, 32.3D and Gtet). The right-left red arrows below the number indicate the size of deletion in base pairs. Tissue AG-014699 nmr specific detection of cytoplasmic and nuclear Wolbachia markers The tissue specific distribution of the Wolbachia markers in G. m. morsitans were tested in ovary, salivary gland, midgut and selleck screening library carcass in normal and tetracycline-treated (Wolbachia-cured) flies. Two 16S rRNA PCR products (438 and 296 bp as described in Figure 3, corresponding to cytoplasmic and nuclear Wolbachia markers) could be amplified from ovary and testes tissues of uncured flies, while only the truncated 296 bp product that corresponds to the nuclear Wolbachia marker was amplified from all of the tissues (Figure 4). In contrast, the fragment that corresponds to the cytoplasmic 16S rRNA marker could not be amplified from any of the

tissues of Wolbachia cured tetracycline-treated flies, including the reproductive organs (ovary and testes) (Fig. 4). The amplification of the larger product that from corresponds to the cytoplasmic Wolbachia only from testes and ovary tissues of adults suggests that Wolbachia is restricted to the gonadal tissues in this species. Unlike for the 16S rRNA, a single wsp PCR product was observed in all tissues of Wolbachia infected and cured adults (Fig. 4). While it was not possible to differentiate between amplifications of cytoplasmic and nuclear Wolbachia, amplification from tetracycline treated adults suggests a horizontal transfer event also for the wsp gene. The size heterogeneity was also observed for fbpA. The larger 509 bp amplification which corresponds to the cytoplasmic marker was restricted to the reproductive tissues of the tsetse flies while the smaller derived 453 bp product corresponding to the nuclear marker was present in all tissues of infected and cured adults, suggesting horizontal transfer of fbpA to the G. m. morsitans genome (Fig. 4). Figure 4 Tissue tropism of Wolbachia infections in G. m. morsitans. G. m.