Resistome characterisation from catchment to effluent in a treatment plant without hospital inputs: implications for AMR monitoring

ABSTRACT

Graphical abstract illustrating the study design and key findings using icons and arrows. On the left, three rounded rectangles each labelled Catchment 1, Catchment 2, and Catchment 3 contain house icons representing domestic households. A vertical dark teal bar connects all three catchments with the label No hospital input, indicating the absence of hospital wastewater inputs. A large horizontal arrow points right from the catchments into a central box labelled Domestic Wastewater Treatment. Inside this box, three dark grey squares represent the sequential treatment stages: primary sedimentation, trickling filter, and sludge tank. Below the treatment box, four icons depicting racks of test tubes are labelled IN (influent), PS (primary sedimentation), EF (effluent), and SL (sludge), connected by downward arrows showing the sampling points. Beneath the sampling point icons, a row of bacteria and DNA double-helix icons illustrates the progressive reduction in microbial and genetic material through treatment: large bacteria and large red DNA helices at IN reduce in size at PS, become very small at EF, while SL shows bacteria and DNA helices again at moderate size. On the right, two dark teal boxes display the key findings in white text: the upper box reads ARGs persisting despite treatment: aadA7, blaOXY, qepA; the lower box reads Candidate biomarkers: aadA7, sul1_2, blaOXA48.

Wastewater surveillance is increasingly used to monitor antimicrobial resistance (AMR) at community scales. This study aimed to (1) assess AMR removal across treatment stages, (2) identify persistent versus responsive antimicrobial resistance genes (ARGs), and (3) characterise resistome profile of a wastewater treatment plant (WWTP) in Northeast England with no hospital inputs. Analyses included physicochemical profiling, microbial culturing, and high-throughput quantitative polymerase chain reaction targeting ARGs. Presumptive extended spectrum β-lactamase (ESBL)-producing Escherichia coli were effectively reduced by over two logs in the WWTP, although viable ESBL E. coli was still present in effluent. Comparative analyses revealed significant reductions along treatment stages for most ARGs. However, some ARGs, such as aadA7, blaOXY, and qepA, showed no significant reduction, indicating some ARGs persist through treatment, an important consideration for future monitoring programmes. The resistome profiles from three sub-catchments were similar, with only one ARG (lnuC) showing significant variation. This suggests that domestic-only catchments without hospital inputs may exhibit relatively uniform ARG profiles, although these results are specific to the WWTP studied and requires further validation. Finally, three ARGs, aadA7, sul1_2, and blaOXA48 were identified as potential biomarkers for AMR monitoring in WWTP, based on their abundance, strong correlation patterns, and clinical relevance.