Symbiotic microorganisms living in the digestive tracts of invertebrates can be crucial in host-symbiont interactions, as they play fundamental roles in important biological processes. Urbanization-related habitat alteration and disturbance, however, considerably affect the environment of host insects, from which their gut microbiota is derived. Still, relatively few studies, all on flying insects, have assessed the impact of urbanization on the gut microbiota of insects. Here, we compared the gut bacterial microbiota in rural and urban individuals of a flightless ground beetle, Carabus convexus, using next generation sequencing. Across the 48 gut samples we identified 1163 different bacterial operational taxonomic units (OTUs), forming significantly different gut bacterial communities in rural versus urban beetles. The taxonomic diversity of the gut bacterial microbiota expressed by the Hill numbers was significantly higher in rural than urban individuals, as well as in rural males vs. females. Smaller differences were found in functional diversity, assessed by the Rao’s quadratic entropy which was marginally significantly higher in urban than rural beetles.