Baseline Characterization of the Gut Microbiota of Field and Colony Populations of Phlebotomus tobbi and Preliminary Assessment of the Anti-Leishmanial Activity of Cultivable Bacteria

Sand fly midgut microbiota plays a critical role in shaping Leishmania development and vector competence, yet functional evidence from natural vector populations remains limited. In this study, sand flies were collected between 2020 and 2022 in Cukurova region, Türkiye to characterize the gut bacterial composition of Phlebotomus tobbi and evaluate the anti-leishmanial potential of cultivable isolates. A total of 1739 sand flies were captured (878 females, 861 males), of which Ph. tobbi was the predominant species (n = 1312). 16S rRNA amplicon sequencing (V4–V6) showed that the gut microbiota was dominated by Proteobacteria, with Erwinia aphidicola/persicina representing the most abundant species across all analyzed groups. Fourteen cultivable bacterial species were identified by MALDI-TOF MS, including Serratia liquefaciens, Pantoea agglomerans, and Micrococcus luteus. Functional XTT assays against Leishmania infantum promastigotes demonstrated variable inhibitory activity among isolates. The strongest leishmanicidal effects were observed with S. liquefaciens (32.3%) and M. luteus (28.8%). Morphological examination confirmed promastigote rounding and cell death in isolates showing >25% activity. These findings define the gut bacterial landscape of Ph. tobbi in an endemic region and identify bacterial taxa with in vitro anti-leishmanial activity, highlighting their potential for future microbiota-based or paratransgenic control strategies.