Abstract
The emergence and spread of antimicrobial resistance (AMR) in clinically important bacterial pathogens has severely compromised the effectiveness of commonly used antibiotics in healthcare. Acquisition and transmission of AMR genes (ARGs) are often facilitated by sublethal concentrations of antibiotics in microbially dense environments. In this study, we use sewage samples (n = 381) collected from six Indian states between June and December 2023 to assess the concentration of eleven antibiotics, microbial diversity, and ARG richness. We find antibiotics from seven drug classes and detect over 2000 bacterial amplicon sequence variants (ASVs). Metagenomic (n = 220) and isolated genome sequences (n = 305) of aerobic and anaerobic bacterial species identify 82 ARGs associated with 80 mobile genetic elements (MGEs). These MGEs are predominantly present in multidrug-resistant (MDR) bacterial pathogens. Comparative core genome analysis of MDR bacterial isolates (n = 7166) shows strong genetic similarity between sewage-derived strains and clinical pathogens. Our results highlight sewage as a significant reservoir for ARGs, where genetic exchanges occur and facilitate the evolution and spread of AMR pathogens in both community and healthcare settings. Additionally, the dipstick-based assay developed for ARGs detection can be used for sewage surveillance in low-resource settings for better understanding of resistance prevalence.
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Data availability
All genomic datasets, including 16S metagenomic, shotgun metagenomic, and whole-genome sequencing (WGS) data generated in this study have been deposited in the National Center for Biotechnology Information (https://www.ncbi.nlm.nih.gov/), under BioProject ID PRJEB81436. All previously published genome sequence data used in the study are available in the NCBI database, with accession details provided in Supplementary Dataset 9b-j. The study also utilized publicly available databases, including CARD, KEGG, and NCBI, along with bioinformatics tools such as DADA2, Kraken2, and ABRicate among others.
Code availability
All codes used for data analysis and data generation have been deposited in a publicly accessible GitHub repository (https://github.com/daizee-gwh/urban-sewage-AMR-metagenomics).
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Acknowledgements
We extend our sincere gratitude to Prof. Ganesan Karthikeyan and Prof. Pramod K Garg for their generous support to complete this study. Authors also acknowledge Prof. Sudhansu Vrati, Prof. Arindam Maitra, and Dr. Onkar Nath Tiwari for their support to the execution of this research. This study was funded by the Department of Biotechnology, Government of India, under the surveillance project titled “Genomic Surveillance for SARS-CoV-2 in India: Indian SARS-CoV-2 Genomics Consortium (INSACOG)- Phase II: Sewage Surveillance” (Ref. RAD/22017/19/2022-KGD-DBT, dated 29/12/2022); Expansion of INSACOG- Wastewater Surveillance of SARS-CoV-2, Emerging Pathogens and AMR through Genomics-Based Methods (Ref No: GCI-13012/2/2025-GCl), as well as the Translational Research Program (TRP) (No. BT/PR30159/MED/15/188/2018).
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B.D. conceptualized and designed the research. D.P., D.T., R.S.K., V.G., L.N., P.J., P.K., J.S., S.K., C.B., K.K., S.B., S.L., S.T., R.K., P.B., M.B., and Y.K. performed experiments. S.M., D.P., and S.K.B. contributed to the investigation and sample collection. B.D., D.P., and A.M. contributed to the development of dipstick-based detection of ARGs. D.P., D.T., K.R.S., N.W., and B.D. contributed to formal analysis, validation and writing the original draft. All the authors approved the final version of the manuscript.
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Paul, D., Talukdar, D., Kapuganti, R.S. et al. Antibiotic contamination and antimicrobial resistance dynamics in the urban sewage microbiome in India. Nat Commun (2025). https://doi.org/10.1038/s41467-025-68034-3
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DOI: https://doi.org/10.1038/s41467-025-68034-3
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