Wastewater pollution differently affects the antibiotic resistance gene pool and biofilm bacterial communities across streambed compartments

Wastewater discharges introduce antibiotic residues and antibiotic-resistant bacteria (ARB) into surface waters. Both inputs directly affect the streambed resistome, either by exerting a selective pressure that favour the proliferation of resistant phenotypes or by enriching the resident communities with wastewater-associated ARB. Here, we investigated the impact of raw and treated urban wastewater discharges on epilithic (growing on rocks) and epipsammic (growing on sandy substrata) streambed biofilms. The effects were assessed by comparing control and impact sites (i) on the composition of bacterial communities; (ii) on the abundance of twelve antibiotic resistance genes (ARGs) encoding resistance to beta-lactams, fluoroquinolones, sulphonamides, tetracyclines, macrolides and vancomycin, as well as the class 1 integron-integrase gene (intl1); (iii) on the occurrence of wastewater-associated bacteria, including putative pathogens, and their potential linkage to target ARGs. We measured more pronounced effects of raw sewage than treated wastewater at the three studied levels. This effect was especially noticeable in epilithic biofilms, which showed a higher contribution of wastewater-associated bacteria and ARB than in epipsammic biofilms. Comparison of correlation coefficients obtained between the relative abundance of both target ARGs and operational taxonomic units classified as either potential pathogens or nonpathogens yielded significant higher correlations between the former category and genes intl1, sul1, sul2 and ermB. Altogether, these results indicate that wastewater-associated micro-organisms, including potential pathogens, contribute to maintain the streambed resistome and that epilithic biofilms appear as sensitive biosensors of the effect of wastewater pollution in surface waters.