Antibiotic resistance gene sequencing is necessary to reveal the complex dynamics of immigration from sewers to activated sludge

Claire Gibson; Susanne A Kraemer; Natalia Klimova; Bing Guo; Dominic Frigon

doi:10.3389/fmicb.2023.1155956

Antibiotic resistance gene sequencing is necessary to reveal the complex dynamics of immigration from sewers to activated sludge

Front Microbiol. 2023 Apr 26:14:1155956. doi: 10.3389/fmicb.2023.1155956. eCollection 2023.

Authors

Claire Gibson¹, Susanne A Kraemer^{1

2}, Natalia Klimova¹, Bing Guo³, Dominic Frigon¹

Affiliations

¹ Department of Civil Engineering and Applied Mechanics, McGill University, Montreal, QC, Canada.
² Aquatic Contaminants Research Division, Environment and Climate Change Canada, Montreal, QC, Canada.
³ Department of Civil and Environmental Engineering, Centre for Environmental Health and Engineering, University of Surrey, Surrey, United Kingdom.

Abstract

Microbial community composition has increasingly emerged as a key determinant of antibiotic resistance gene (ARG) content. However, in activated sludge wastewater treatment plants (AS-WWTPs), a comprehensive understanding of the microbial community assembly process and its impact on the persistence of antimicrobial resistance (AMR) remains elusive. An important part of this process is the immigration dynamics (or community coalescence) between the influent and activated sludge. While the influent wastewater contains a plethora of ARGs, the persistence of a given ARG depends initially on the immigration success of the carrying population, and the possible horizontal transfer to indigenously resident populations of the WWTP. The current study utilized controlled manipulative experiments that decoupled the influent wastewater composition from the influent microbial populations to reveal the fundamental mechanisms involved in ARG immigration between sewers and AS-WWTP. A novel multiplexed amplicon sequencing approach was used to track different ARG sequence variants across the immigration interface, and droplet digital PCR was used to quantify the impact of immigration on the abundance of the targeted ARGs. Immigration caused an increase in the abundance of over 70 % of the quantified ARGs. However, monitoring of ARG amplicon sequence variants (ARG-ASVs) at the immigration interface revealed various immigration patterns such as (i) suppression of the indigenous mixed liquor ARG-ASV by the immigrant, or conversely (ii) complete immigration failure of the influent ARG-ASV. These immigration profiles are reported for the first time here and highlight the crucial information that can be gained using our novel multiplex amplicon sequencing techniques. Future studies aiming to reduce AMR in WWTPs should consider the impact of influent immigration in process optimisation and design.

Keywords: activated sludge; antimicrobial resistance; community assembly; community coalescence; immigration; wastewater.

Grants and funding

This work was funded by NSERC through a Discovery grant (NSERC RGPIN-2016-06498) and a Strategic Program grant (NSERC STPGP 521349-18). NSERC had no role in the design of this study. CG and BG were partly funded by the McGill University Engineering Doctoral Award.