Metagenomic analysis of antibiotic resistance across the wastewater process

Bacterial resistance to antimicrobials is a global health threat. Within the One Health context, water from regions with high antibiotic usage, such as clinical and urban areas, collects at wastewater treatment plants (WWTPs). In the WWTP, the activated sludge becomes a complex environment where var...

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Main Authors: Stephanie Pillay, Ramin Shirali Hossein Zade, Paul van Lent, David Calderón-Franco, Thomas Abeel
Format: Article
Language:English
Published: Elsevier 2025-03-01
Series:Heliyon
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Online Access:http://www.sciencedirect.com/science/article/pii/S2405844025013003
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author Stephanie Pillay
Ramin Shirali Hossein Zade
Paul van Lent
David Calderón-Franco
Thomas Abeel
author_facet Stephanie Pillay
Ramin Shirali Hossein Zade
Paul van Lent
David Calderón-Franco
Thomas Abeel
author_sort Stephanie Pillay
collection DOAJ
description Bacterial resistance to antimicrobials is a global health threat. Within the One Health context, water from regions with high antibiotic usage, such as clinical and urban areas, collects at wastewater treatment plants (WWTPs). In the WWTP, the activated sludge becomes a complex environment where various antimicrobials and microorganisms converge. While significant research has focused on the influent, activated sludge, and effluent, upstream and downstream sectors around the WWTP are often neglected. We conducted a systematic analysis using five publicly available metagenomic datasets (n=164) from different WWTP sectors and adjacent freshwater systems: upstream (n=14), influent (n=14), activated sludge (n=109), effluent (n=14), and downstream (n=13) to identify and characterise the microbiome, resistome, and mobilome. Opportunistic pathogenic bacteria, such as Pseudomonas, Aeromonas, and Acidovorax, were found in all WWTP sectors, with abundances exceeding 9% in the influent. ESKAPE pathogens, including Klebsiella pneumoniae and Enterobacter species, were identified in the effluent with abundances over 1%. We detected 230 antibiotic resistance genes (ARGs) throughout the WWTP. FTU and CKO β-lactamase gene families dominated the upstream, effluent, and downstream sectors, while the OXA β-lactamase gene family was highly abundant in the influent and activated sludge. ARGs, such as the OXA β-lactamase gene family, were linked to plasmids. Class-1 integrons, associated with the sul1 gene, a marker for anthropogenic pollution, were prevalent in the effluent and downstream sectors. Integrative elements (ICEclc, Tn4371, and PGI2), linked to ARGs, were identified in all sectors, increasing AMR dissemination. These integrative elements conferred resistance to antibiotics, including sulfonamides, tetracyclines and carbapenems. Our findings highlight the presence of ARGs and mobile genetic elements in WWTPs and nearby freshwater systems, raising concerns about AMR transmission to humans, animals, and the environment. This study emphasises the need for effective AMR monitoring and strategies in wastewater treatment to protect public and environmental health.
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spelling doaj-art-907c05ebba3a44d0a097aef3e8e01e6b2025-08-20T03:05:55ZengElsevierHeliyon2405-84402025-03-01115e4291910.1016/j.heliyon.2025.e42919Metagenomic analysis of antibiotic resistance across the wastewater processStephanie Pillay0Ramin Shirali Hossein Zade1Paul van Lent2David Calderón-Franco3Thomas Abeel4Delft University of Technology, Delft, the NetherlandsDelft University of Technology, Delft, the Netherlands; Department of Biomedical Data Sciences, Leiden University Medical Center, Leiden, the Netherlands; Leiden Center for Computational Oncology, Leiden, the NetherlandsDelft University of Technology, Delft, the NetherlandsHologenomix B.V, Delft, the NetherlandsDelft University of Technology, Delft, the Netherlands; Infectious Disease and Microbiome Program, Broad Institute of MIT and Harvard, Cambridge, USA; Corresponding author.Bacterial resistance to antimicrobials is a global health threat. Within the One Health context, water from regions with high antibiotic usage, such as clinical and urban areas, collects at wastewater treatment plants (WWTPs). In the WWTP, the activated sludge becomes a complex environment where various antimicrobials and microorganisms converge. While significant research has focused on the influent, activated sludge, and effluent, upstream and downstream sectors around the WWTP are often neglected. We conducted a systematic analysis using five publicly available metagenomic datasets (n=164) from different WWTP sectors and adjacent freshwater systems: upstream (n=14), influent (n=14), activated sludge (n=109), effluent (n=14), and downstream (n=13) to identify and characterise the microbiome, resistome, and mobilome. Opportunistic pathogenic bacteria, such as Pseudomonas, Aeromonas, and Acidovorax, were found in all WWTP sectors, with abundances exceeding 9% in the influent. ESKAPE pathogens, including Klebsiella pneumoniae and Enterobacter species, were identified in the effluent with abundances over 1%. We detected 230 antibiotic resistance genes (ARGs) throughout the WWTP. FTU and CKO β-lactamase gene families dominated the upstream, effluent, and downstream sectors, while the OXA β-lactamase gene family was highly abundant in the influent and activated sludge. ARGs, such as the OXA β-lactamase gene family, were linked to plasmids. Class-1 integrons, associated with the sul1 gene, a marker for anthropogenic pollution, were prevalent in the effluent and downstream sectors. Integrative elements (ICEclc, Tn4371, and PGI2), linked to ARGs, were identified in all sectors, increasing AMR dissemination. These integrative elements conferred resistance to antibiotics, including sulfonamides, tetracyclines and carbapenems. Our findings highlight the presence of ARGs and mobile genetic elements in WWTPs and nearby freshwater systems, raising concerns about AMR transmission to humans, animals, and the environment. This study emphasises the need for effective AMR monitoring and strategies in wastewater treatment to protect public and environmental health.http://www.sciencedirect.com/science/article/pii/S2405844025013003MetagenomicsAntibiotic resistant bacteriaAntibiotic resistance genesMobile genetic elementsWastewater treatment plantMeta-analysis
spellingShingle Stephanie Pillay
Ramin Shirali Hossein Zade
Paul van Lent
David Calderón-Franco
Thomas Abeel
Metagenomic analysis of antibiotic resistance across the wastewater process
Heliyon
Metagenomics
Antibiotic resistant bacteria
Antibiotic resistance genes
Mobile genetic elements
Wastewater treatment plant
Meta-analysis
title Metagenomic analysis of antibiotic resistance across the wastewater process
title_full Metagenomic analysis of antibiotic resistance across the wastewater process
title_fullStr Metagenomic analysis of antibiotic resistance across the wastewater process
title_full_unstemmed Metagenomic analysis of antibiotic resistance across the wastewater process
title_short Metagenomic analysis of antibiotic resistance across the wastewater process
title_sort metagenomic analysis of antibiotic resistance across the wastewater process
topic Metagenomics
Antibiotic resistant bacteria
Antibiotic resistance genes
Mobile genetic elements
Wastewater treatment plant
Meta-analysis
url http://www.sciencedirect.com/science/article/pii/S2405844025013003
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AT davidcalderonfranco metagenomicanalysisofantibioticresistanceacrossthewastewaterprocess
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