Modulation of trypanosome establishment in Glossina palpalis palpalis by its microbiome in the Campo sleeping sickness focus, Cameroon.

The purpose of this study was to investigate factors involved in vector competence by analyzing whether the diversity and relative abundance of the different bacterial genera inhabiting the fly's gut could be associated with its trypanosome infection status. This was investigated on 160 randomly selected G. p. palpalis flies - 80 trypanosome-infected, 80 uninfected - collected in 5 villages of the Campo trypanosomiasis focus in South Cameroon. Trypanosome species were identified using specific primers, and the V4 region of the 16S rRNA gene of bacteria was targeted for metabarcoding analysis in order to identify the bacteria and determine microbiome composition. A total of 261 bacterial genera were identified of which only 114 crossed two barriers: a threshold of 0.01% relative abundance and the presence at least in 5 flies. The secondary symbiont Sodalis glossinidius was identified in 50% of the flies but it was not considered since its relative abundance was much lower than the 0.01% relative abundance threshold. The primary symbiont Wigglesworthia displayed 87% relative abundance, the remaining 13% were prominently constituted by the genera Spiroplasma, Tediphilus, Acinetobacter and Pseudomonas. Despite a large diversity in bacterial genera and in their abundance observed in micobiome composition, the statistical analyzes of the 160 tsetse flies showed an association with flies' infection status and the sampling sites. Furthermore, tsetse flies harboring Trypanosoma congolense Savanah type displayed a different composition of bacterial flora compared to uninfected flies. In addition, our study revealed that 36 bacterial genera were present only in uninfected flies, which could therefore suggest a possible involvement in flies' refractoriness; with the exception of Cupriavidus, they were however of low relative abundance. Some genera, including Acinetobacter, Cutibacterium, Pseudomonas and Tepidiphilus, although present both in infected and uninfected flies, were found to be associated with uninfected status of tsetse flies. Hence their effective role deserves to be further evaluated in order to determine whether some of them could become targets for tsetse control of fly vector competence and consequently for the control of the disease. Finally, when comparing the bacterial genera identified in tsetse flies collected during 4 epidemiological surveys, 39 genera were found to be common to flies from at least 2 sampling campaigns.

The composition and abundance of bacterial communities residing in the gut of Glossina palpalis palpalis captured in two sites of southern Cameroon.

BACKGROUND: A number of reports have demonstrated the role of insect bacterial flora on their host's physiology and metabolism. The tsetse host and vector of trypanosomes responsible for human sleeping sickness (human African trypanosomiasis, HAT) and nagana in animals (African animal trypanosomiasis, AAT) carry bacteria that influence its diet and immune processes. However, the mechanisms involved in these processes remain poorly documented. This underscores the need for increased research into the bacterial flora composition and structure of tsetse flies. The aim of this study was to identify the diversity and relative abundance of bacterial genera in Glossina palpalis palpalis flies collected in two trypanosomiasis foci in Cameroon. METHODS: Samples of G. p. palpalis which were either negative or naturally trypanosome-positive were collected in two foci located in southern Cameroon (Campo and Bipindi). Using the V3V4 and V4 variable regions of the small subunit of the 16S ribosomal RNA gene, we analyzed the respective bacteriome of the flies' midguts. RESULTS: We identified ten bacterial genera. In addition, we observed that the relative abundance of the obligate endosymbiont Wigglesworthia was highly prominent (around 99%), regardless of the analyzed region. The remaining genera represented approximately 1% of the bacterial flora, and were composed of Salmonella, Spiroplasma, Sphingomonas, Methylobacterium, Acidibacter, Tsukamurella, Serratia, Kluyvera and an unidentified bacterium. The genus Sodalis was present but with a very low abundance. Globally, no statistically significant difference was found between the bacterial compositions of flies from the two foci, and between positive and trypanosome-negative flies. However, Salmonella and Serratia were only described in trypanosome-negative flies, suggesting a potential role for these two bacteria in fly refractoriness to trypanosome infection. In addition, our study showed the V4 region of the small subunit of the 16S ribosomal RNA gene was more efficient than the V3V4 region at describing the totality of the bacterial diversity. CONCLUSIONS: A very large diversity of bacteria was identified with the discovering of species reported to secrete anti-parasitic compounds or to modulate vector competence in other insects. For future studies, the analyses should be enlarged with larger sampling including foci from several countries.