Characterization of Wigglesworthia glossinidia amino acid profiles relative to Glossina morsitans pregnancy

Tsetse flies (Glossina morsitans), the obligate vectors of Old-World trypanosomes (Trypanosoma spp.) that cause African trypanosomiasis, have a bacteriome composed of bacteriocytes that house the symbiont Wigglesworthia glossinidia. The Gammaproteobacteria Wigglesworthia function to supplement necessary B vitamins lacking in the hematophagous diet of the tsetse host. In exchange, Wigglesworthia, auxotrophic for 15 of the 20 proteogenic amino acids, receive most of their amino acids from either the protein-rich blood diet of tsetse or directly from tsetse hosts. We hypothesize that the provisioning of tsetse-supplied amino acids into the bacteriome is greater when Wigglesworthia-supplied B-vitamins are in higher demand, specifically during tsetse pregnancy. To characterize the amino acid profiles of bacteriomes during tsetse pregnancy, bacteriomes from age-matched aposymbiotic, virgin and mated females are dissected and their bacteriocytes are lysed to release Wigglesworthia. Wigglesworthia are then purified from tsetse cells and their amino acid profiles are characterized with liquid chromatography-tandem mass spectrometry (LC-MS/MS). Amino acids will be identified using Freestyle (Thermofisher Scientific) and Qual Browser (Xcalibur, Thermofisher Scientific) and quantified by comparison to internal standards. Wigglesworthia-free bacteriomes will provide baseline amino acid estimates. Insight into the mutualistic nature of this host-symbiont relationship provides opportunities for development of novel disease management strategies. Additionally, elucidation of the coordination of metabolic processes, likely mediated through partner fidelity feedback between tsetse and Wigglesworthia, offers insight into factors regulating heterospecific stability not only for this symbiosis but also for other animal-associated microbiota.