Phlebotomine sand flies are dipterans of relevance due to their role as vectors of several pathogens worldwide. Bacteria in the gut of sand flies possibly affect their vectorial capacity and competence to transmit parasites. A retrospective study was performed in sand fly specimens that had previously been collected in four localities of the state of Chiapas during the period 2009–2011 to detectWolbachiaandBartonellaand their possible coinfection withLeishmania. For the molecular detection of bacteria, we used primers and conditions that had previously been reported. A total of 531 sand fly specimens of 10 species were analyzed. FourWolbachiastrains were detected in five sand fly species, showing a prevalence of 8.6%. All theWolbachiastrains had previously been reported in other taxa. In one sand fly species, we also detected a new lineage ofBartonellaevidenced by a phylogenetic analysis. No sand fly specimens showed coinfections of these bacteria andLeishmania. The bacteria found in the phlebotomine sand flies are possibly transmitted by plant-mediated horizontal transmission and during blood meal feeding.