Abstract
OBJECTIVES: Antibiotic resistance genes (ARGs), one of the key drivers of the growing antibiotic resistance, present a formidable public health concern. Wastewater may act as a key repository for both antibiotic-resistant bacteria and ARGs, facilitating their transfer across microbial communities. Given the limited knowledge of clinically relevant ARGs in community wastewater of Bangladesh, this study explored the genetic diversity and occurrence of ARGs from community wastewater discharges using a culture-independent approach. METHODS: Wastewater samples (n = 70) were collected from different sites in Dhaka and Cox's Bazar between April and May 2023. These samples underwent Nanotrap®-based culture-independent microbial concentration, followed by nucleic acid extraction and purification. Nine clinically important ARGs (blaTEM, blaCTX-M, blaSHV, blaKPC, blaNDM, blaOXA, mcr-1, tolC, and acrA) were detected using uniplex polymerase chain reaction (PCR), followed by Sanger sequencing. Bioinformatic analyses further explored the genetic diversity of the ARGs, as well as mutational analysis and molecular docking of Klebsiella pneumoniae carbapenemase (KPC). RESULTS: blaTEM, blaCTX-M, blaOXA, tolC, and acrA were the most frequently detected ARGs (90 % each), followed by blaNDM (85.7 %), blaKPC (84.3 %), blaSHV (77.1 %), and mcr-1 (52.9 %). The frequency of all ARGs was higher in Cox's Bazar compared to Dhaka, except for mcr-1. Co-occurrences of all nine ARGs were observed in 47.1 % of samples. Mutational analysis of KPC revealed unique missense mutations, and further docking demonstrated altered binding affinities and interactions between KPC-2 mutants and β-lactam antibiotics. CONCLUSION: Our study provides the baseline data of clinically significant ARGs in local community wastewater in Bangladesh, reflecting the need for further surveillance to identify hotspots and guide antibiotic stewardship and wastewater management policies.