Host range of antibiotic resistance genes in wastewater treatment plant influent and effluent

Abstract

Wastewater treatment plants (WWTPs) collect wastewater from various sources for a multi-step treatment process. By mixing a large variety of bacteria and promoting their proximity, WWTPs constitute potential hotspots for the emergence of antibiotic resistant bacteria. Concerns have been expressed regarding the potential of WWTPs to spread antibiotic resistance genes (ARGs) from environmental reservoirs to human pathogens. We utilized epicPCR (Emulsion, Paired Isolation and Concatenation PCR) to detect the bacterial hosts of ARGs in two WWTPs. We identified the host distribution of four resistance-associated genes (tetM, int1, qacEΔ1and blaOXA-58) in influent and effluent. The bacterial hosts of these resistance genes varied between the WWTP influent and effluent, with a generally decreasing host range in the effluent. Through 16S rRNA gene sequencing, it was determined that the resistance gene carrying bacteria include both abundant and rare taxa. Our results suggest that the studied WWTPs mostly succeed in decreasing the host range of the resistance genes during the treatment process. Still, there were instances where effluent contained resistance genes in bacterial groups not carrying these genes in the influent. By permitting exhaustive profiling of resistance-associated gene hosts in WWTP bacterial communities, the application of epicPCR provides a new level of precision to our resistance gene risk estimates.

Figure 1.

(A) Order level classification of the 16S rRNA gene sequences. Microbial community composition was analyzed from both plants (VI = Viikinmäki, SU = Suomenoja) in the influent and effluent water on three subsequent days. (B) Principal coordinate analysis (PCoA) plot from the OTU relative abundance with the Bray–Curtis dissimilarity. The influent clusters from two plants clustered together, whereas the effluent samples were more distinct at the studied plants.

Figure 2.

Presence of ARGs in microbial families in the two wastewater purification plants. The phylogenetic tree was constructed from family-level OTUs based on 16S rRNA gene amplicon sequences. An ARG was considered to be present in a bacterial genus when found in 2/2 or 3/3 samples (depending on whether all three samples passed the QC step) from the influent or effluent water at the same plant. The inner circle (INF) denotes the influent and the outer circle (EFF) the effluent water. The results are presented at family level and bars in the INF and EFF rings display relative abundance.

Figure 3.

Bacterial families with resistance gene detected by epicPCR in influent and/or effluent of the two plants (Viikinmäki and Suomenoja). Relative abundance of the family in total 16S rRNA gene amplicon sequencing is shown with the size of the sphere. Open sphere: gene not detected with epicPCR, closed sphere: gene detected with epicPCR.