Sewer biofilm microbiome and antibiotic resistance genes as function of pipe material, source of microbes, and disinfection: field and laboratory studies.

Wastewater systems are recognized pathways for the spread of antibiotic resistant bacteria, but relatively little is known about the microbial ecology of the sewer environment. Sewer biofilm colonization by antibiotic resistance gene (ARG) carrying bacteria may impact interpretations of sewage epidemiology data, water quality during sewer overflows, and hazard to utility workers. The objectives of this research were to evaluate the (1) microbiome of real and simulated sewer biofilms and their potential to accumulate ARGs and (2) susceptibility of simulated sewer biofilms to bleach disinfection. First, biofilm samples were collected from sewer municipal systems. Next, an annular biofilm reactor was used to simulate the sewer environment while controlling the pipe material (concrete vs. PVC). The reactor was operated either as fed semi-batch with sewer sediment and synthetic wastewater (Sed-SB) or fed with a continuous flow of raw sewage (WW-CF). The abundance of ARGs, human fecal marker HF183, and 16S rRNA gene copies in these biofilm samples was measured with qPCR. Amplicon sequencing was performed to compare the prokaryotic diversity between samples. Finally, the susceptibility of reactor biofilm to a 4.6% bleach disnfection protocol was evaluated using viability qPCR and amplicon sequencing. Field and WW-CF biofilms contained the most ARG copies and the microbial community compositions varied between the different biofilm samples (field, Sed-SB, and WW-CF). Pipe material did not affect the abundance of ARGs in the reactor samples. However, log removal following bleach treatment suggested that the biofilm grown on PVC surface was primarily dislodged from the surface by the bleach treatment whereas more bacteria were lysed within the biofilm that remained on the concrete surface. Viable bacteria carrying ARGs were observed following 10 minutes of treatment. This study showed that sewer biofilms can accumulate bacteria carrying ARGs and that while bleach can reduce sewer biofilm density, the protocol tested here will not completely remove the biofilms.