Changes in the state of rivers resulting from the activity and expansion of urban areas are likely to affect aquatic populations by increasing stress and disease, with the microbiota playing a potentially important intermediary role. Unraveling the dynamics of microbial flora is therefore essential to better apprehend the impact of anthropogenic disturbances on the health of host populations and the ecological integrity of hydrosystems. In this context, the present study simultaneously examined changes in the microbial communities associated with mucosal skin and gut tissues of eight fish species along an urbanization gradient in the Orge River (France). 16S rRNA gene metabarcoding revealed that the structure and composition of the skin microbiota varied substantially along the disturbance gradient and to a lesser extent according to fish taxonomy. Sequences affiliated with the Gammaproteobacteria, in particular the genus Aeromonas, prevailed on fish caught in the most urbanized areas, whereas they were nearly absent upstream. This rise of opportunistic taxa was concomitant with a decline in phylogenetic diversity, suggesting more constraining environmental pressures. In comparison, fish gut microbiota varied much more moderately with the degree of urbanization, possibly because this niche might be less directly exposed to environmental stressors. Co-occurrence networks further identified pairs of associated bacterial taxa, co-existing more or less often than expected at random. Few correlations could be identified between skin and gut bacterial taxa, supporting the assumption that these two microbial niches are disconnected and do not suffer from the same vulnerability to anthropic pressures.