The tsetse fly (Glossina sp.) gut is colonized by maternally transmitted and environmentally acquired bacteria. Maternally transmitted symbionts are well characterized, but little is known about the origin and function of environmentally acquired bacteria inhabiting the tsetse midgut. To address this shortcoming, culture dependent and independent methods were used to characterize and quantify bacterial communities that reside within the midgut of tsetse flies collected at five geographically isolated locations in Kenya and Uganda. Bacteria were isolated from 83.33% of flies using culture dependent methods. In total, 38 strains were isolated. To increase the depth of this analysis, Illumina-based deep-sequencing of the V4 hypervariable region of the bacterial 16S ribosomal RNA (rRNA) gene was used to determine the relationship between the type and number of bacteria present in Glossina pallidipes midguts and geographic location. The midgut was dominated by the obligate endosymbiont, Wigglesworthia, but a diversity of other microbiota in individuals that were unique to each location was also observed. This was consistent between trypanosome infected and uninfected samples. The results of this study will increase the understanding of the breadth and depth of tsetse's midgut bacterial communities as they relate to the environmental acquisition of the fly's microbiome and its correlation to trypanosome vectorial capacity. Future studies can then be performed to determine the specific molecular mechanisms that underlie bacteria-mediated trypanocidal immunity in tsetse's gut.
