Wastewater-based surveillance (WBS) has been widely adopted as a cost-effective and population-representative infectious disease monitoring tool and is increasingly being applied to bacterial, fungal, and antimicrobial resistance gene (ARG) targets. However, some of these targets may persist in pipe biofilms and detach into wastewater, complicating accurate interpretation for WBS. To investigate contributions from biofilms to wastewater pathogen and ARG signals, paired sink drain biofilm, branch drain plumbing biofilm (“sewer biofilm”), and wastewater samples were collected from five hospital wards over a four-month period and analyzed using both 16S rRNA gene amplicon sequencing and probe-capture-enrichment metagenomics. Overall, sewer biofilm bacterial communities were as diverse as wastewater but with fewer pathogens detected. Across wards, on average, an estimated 9% of wastewater bacterial communities could be attributed to sewer biofilm communities. Many pathogens were consistently detected both in sewer biofilm and wastewater, including environmentally persistent and/or biofilm-associated taxa (e.g., Pseudomonas aeruginosa, Stenotrophomonas maltophilia). While many ARGs overlapped between wastewater and biofilms (e.g., tetA, sul1, blaCTX-M, mcr-5.1, and vanA), others were significantly enriched in sewer biofilms (e.g., qacL, van-operon genes, and OXA genes). Together, these findings confirm that wastewater pathogen and resistome profiles integrate inputs from both human shedding and pipe-resident microbial reservoirs and therefore need to be considered when selecting WBS targets and interpreting signal.