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Antibiotic Resistance in Wastewater Treatment Plants

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A special issue of Antibiotics (ISSN 2079-6382). This special issue belongs to the section "Mechanism and Evolution of Antibiotic Resistance".

Deadline for manuscript submissions: 31 July 2026 | Viewed by 13145

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Dr. Qingbin Yuan

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Guest Editor

State Key Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing University, Nanjing 210023, China
Interests: detection of biological contaminants, geochemical behavior, and risk prevention and control; antibiotic resistance in the environment; applications of machine learning in the environment

Special Issue Information

Dear Colleagues,

The improper use of antibiotics worldwide has led to the development of antibiotic resistance, emerging as one of the greatest concerns of this century. Numerous strategies have been implemented to combat antibiotic resistance. While clinical strategies are crucial, managing antibiotics and antibiotic resistance in the environment is equally important, as antibiotic resistance can transfer across humans, animals and the environment, a concept known as the One Health concern. Wastewater treatment plants are major environmental reservoirs that not only receive wastewater containing antibiotics and antibiotic resistance genes (ARGs), but also release treated wastewater containing ARGs into various environments. Therefore, research on antibiotic resistance in wastewater treatment plants is essential for understanding the development of antibiotic resistance and formulating appropriate strategies.

In wastewater treatment plants, ARGs undergo complex processes, including reproduction, mutation and decay. They can also be horizontally transferred to new hosts, leading to the further development of antibiotic resistance. Conventional wastewater treatment processes, such as biological and disinfection processes, may not effectively eliminate ARGs, potentially releasing them into rivers, lakes and soils, depending on the wastewater disposal methods used.

Despite a growing number of scientific publications on the occurrence, distribution and removal of ARGs in wastewater treatment plants, several concerns remain to be addressed:

Methodologies for identifying antibiotic resistance genes and their hosts in wastewater;
Sources of antibiotic resistance genes in wastewater treatment plants;
The occurrence of antibiotic resistance genes in different physical states, especially extracellular ARGs, in wastewater treatment plants;
Horizontal gene transfer of ARGs within wastewater treatment plants;
Factors influencing the development of antibiotic resistance during wastewater treatment;
The release and fate of antibiotic resistance genes in the receiving environment;
Advanced techniques for the elimination of ARGs in wastewater.

This Special Issue addresses these issues in line with antibiotic management across various settings.

Dr. Qingbin Yuan
Guest Editor

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Keywords

antibiotic resistance genes
wastewater treatment
one health
distribution and removal of ARGs
horizontal gene transfer

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Published Papers (7 papers)

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Research

25 pages, 2146 KB

Open AccessArticle

Characterizing Aeromonas spp. as a Potential Sentinel Organism for Antimicrobial Resistance Dissemination in Wastewater and Drinking Water Treatment Systems: A Case Study in the Barcelona Metropolitan Area, Spain

by Laura Mondéjar, Victoria Ballén, Yaiza Gabasa, Laura Castellsagués, Anna Pinar-Méndez, Carles Vilaró, Belén Galofré, Aida González-Díaz, Sara Martí, Sergi Sanz and Sara M. Soto

Antibiotics 2026, 15(3), 301; https://doi.org/10.3390/antibiotics15030301 - 17 Mar 2026

Viewed by 530

Abstract

Background: Wastewater treatment plants (WWTPs) are hotspots of antimicrobial resistance (AMR) due to inputs from diverse anthropogenic sources. Aeromonas spp., ubiquitous in aquatic environments, often carry clinically relevant antibiotic resistance genes (ARGs) and can persist beyond fecal contamination indicators, making them promising sentinel organisms for AMR dissemination. The aim of this study was to assess the suitability of Aeromonas spp. in this role by characterizing resistance profiles, associated virulence factor genes (VFGs), genetic mobility, and persistence across wastewater and drinking water treatment processes in the Barcelona metropolitan area, Spain. Methods: Isolates were phenotypically characterized and screened for ARGs, VFGs, integrons, and heavy metal tolerance genes, followed by whole-genome sequencing (WGS). Biofilm formation was assessed in vitro. Conjugation assays with Escherichia coli evaluated horizontal gene transfer (HGT) potential. Results: A total of 428 antibiotic-resistant Aeromonas spp., the most abundant antibiotic-resistant bacteria isolated during the 2023 sampling campaigns from two WWTPs and one drinking water treatment plant (DWTP), were characterized. Trimethoprim/sulfamethoxazole (SXT) non-susceptibility was most frequent (72%), followed by cefoxitin resistance (65.4%). The sul1 (57.5%) and bla_MOX (78.6%) genes predominated among SXT- and β-lactam-resistant isolates. The merA gene was detected in 23.6%; 97.9% harbored at least one VFG (aerA, act, fla, alt, or hlyA), and 70.3% carried intI1. Half formed biofilm. Conjugation confirmed bi-directional HGT, and WGS revealed persistent ST3458 clones across treatment stages. Conclusions: WWTPs and DWTPs act as reservoirs of antibiotic-resistant Aeromonas spp., demonstrating persistence and HGT potential. Findings support their use as sentinel organisms for AMR surveillance in aquatic environments and for assessing treatment efficacy, highlighting variability across treatment types and locations, and reinforcing their relevance for urban water reclamation monitoring. Full article

(This article belongs to the Special Issue Antibiotic Resistance in Wastewater Treatment Plants)

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12 pages, 1058 KB

Open AccessArticle

Environmental Dissemination of Antimicrobial Resistance: A Resistome-Based Comparison of Hospital and Community Wastewater Sources

by Taito Kitano, Nobuaki Matsunaga, Takayuki Akiyama, Takashi Azuma, Naoki Fujii, Ai Tsukada, Hiromi Hibino, Makoto Kuroda and Norio Ohmagari

Antibiotics 2026, 15(1), 99; https://doi.org/10.3390/antibiotics15010099 - 19 Jan 2026

Viewed by 876

Abstract

Background/Objectives: Comparative analysis of antimicrobial resistomes in hospital and community wastewater can provide valuable insights into the diversity and distribution of antimicrobial resistance genes (ARGs), contributing to the advancement of the One Health approach. This study aimed to characterize and compare the resistome profiles of wastewater sources from a hospital and community. Methods: Longitudinal metagenomic analysis was conducted on wastewater samples collected from the National Center for Global Health and Medicine (hospital) and a shopping mall (community) in Tokyo, Japan, between December 2019 and September 2023. ARG abundance was quantified using reads per kilobase per million mapped reads (RPKM) values, and comparative analyses were performed to identify the significantly enriched ARGs in the two sources. Results: A total of 46 monthly wastewater samples from the hospital yielded 825 unique ARGs, with a mean RPKM of 2.5 across all detected genes. In contrast, 333 ARGs were identified in the three shopping mall wastewater samples, with a mean RPKM of 2.1. Among the ARGs significantly enriched in the hospital samples, 23, including genes conferring resistance to aminoglycosides (nine groups) and β-lactam antibiotics (eight groups), exhibited significantly high RPKM values. No ARGs were found to be significantly enriched in the community wastewater samples. Conclusions: This study highlights the higher diversity and abundance of ARGs, particularly those conferring resistance to aminoglycosides and β-lactam antibiotics including carbapenems, in hospital wastewater than in community wastewater. These findings underscore the importance of continuous resistome monitoring of hospital wastewater as part of the integrated One Health surveillance strategy. Full article

(This article belongs to the Special Issue Antibiotic Resistance in Wastewater Treatment Plants)

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19 pages, 1684 KB

Open AccessArticle

Effectiveness of Implementing Hospital Wastewater Treatment Systems as a Measure to Mitigate the Microbial and Antimicrobial Burden on the Environment

by Takashi Azuma, Miwa Katagiri, Takatoshi Yamamoto, Makoto Kuroda and Manabu Watanabe

Antibiotics 2025, 14(8), 807; https://doi.org/10.3390/antibiotics14080807 - 7 Aug 2025

Cited by 1 | Viewed by 3362

Abstract

Background: The emergence and spread of antimicrobial-resistant bacteria (ARB) has become an urgent global concern as a silent pandemic. When taking measures to reduce the impact of antimicrobial resistance (AMR) on the environment, it is important to consider appropriate treatment of wastewater from medical facilities. Methods: In this study, a continuous-flow wastewater treatment system using ozone and ultraviolet light, which has excellent inactivation effects, was implemented in a hospital in an urban area of Japan. Results: The results showed that 99% (2 log₁₀) of Gram-negative rods and more than 99.99% (>99.99%) of ARB comprising ESBL-producing Enterobacterales were reduced by ozone treatment from the first day after treatment, and ultraviolet light-emitting diode (UV-LED) irradiation after ozone treatment; UV-LED irradiation after ozonation further inactivated the bacteria to below the detection limit. Inactivation effects were maintained throughout the treatment period in this study. Metagenomic analysis showed that the removal of these microorganisms at the DNA level tended to be gradual in ozone treatment; however, the treated water after ozone/UV-LED treatment showed a 2 log₁₀ (>99%) removal rate at the end of the treatment. The residual antimicrobials in the effluent were benzylpenicillin, cefpodoxime, ciprofloxacin, levofloxacin, azithromycin, clarithromycin, doxycycline, minocycline, and vancomycin, which were removed by ozone treatment on day 1. In contrast, the removal of ampicillin and cefdinir ranged from 19% to 64% even when combined with UV-LED treatment. Conclusions: Our findings will help to reduce the discharge of ARB and antimicrobials into rivers and maintain the safety of aquatic environments. Full article

(This article belongs to the Special Issue Antibiotic Resistance in Wastewater Treatment Plants)

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14 pages, 1052 KB

Open AccessArticle

Phenotypic Detection of Extended-Spectrum β-Lactamase and Carbapenemase-Producing Enterobacteriaceae from Wastewater Treatment Plants in Ouagadougou, Burkina Faso

by Inayatou Hamma Yacouba, Yacouba Konaté, Bapio Valérie Elvira Jean Télesphore Bazie and Boukary Sawadogo

Antibiotics 2025, 14(7), 641; https://doi.org/10.3390/antibiotics14070641 - 25 Jun 2025

Viewed by 1340

Abstract

Background/Objectives: The overuse of antibiotics has led to the emergence of antibiotic-resistant bacteria, posing a global public health challenge. Among these, extended-spectrum β-lactamase and carbapenemase-producing Enterobacteriaceae (ESBL-E and CPE) are of particular concern due to their potential to spread resistance in various environments. Understanding the prevalence and spread of these bacteria in the influent and effluent of wastewater treatment plants is essential. Methods: This study examined ESBL-E and CPE in wastewater from three WWTPs in Ouagadougou, Burkina Faso, and was conducted between February and August 2024. Phenotypic detection of ESBL-E was performed on the isolates using the double-disk synergy test and the combination disk test, whereas the CPE detection employed the combination disk test and the modified Hodge test. Results: A total of 250 Enterobacteriaceae isolates were found, with Escherichia coli, Klebsiella spp., Enterobacter spp., and Buttiauxella spp. the most predominant taxa. Phenotypic analysis revealed a high prevalence of ESBL-E, particularly in influent samples, with rates ranging from 55 to 98% across the WWTPs. CPE detection showed varying prevalence, with higher proportions identified in effluent samples, ranging from 37 to 68%, depending on the plant. These findings highlight the critical role of WWTPs in the persistence and potential spread of antibiotic-resistant bacteria. Conclusions: This study underscores the urgent need for improved wastewater treatment technologies and comprehensive monitoring systems to reduce the dissemination of ESBL-E and CPE in the environment. Addressing these challenges is crucial for mitigating the public health risks associated with antimicrobial resistance. Full article

(This article belongs to the Special Issue Antibiotic Resistance in Wastewater Treatment Plants)

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11 pages, 1872 KB

Open AccessArticle

Simulated Discharge of Ballast Water Reveals Potential Contribution to Spread of Antibiotic Resistance Genes in Geographically Isolated Receiving Waters

by Jianhong Shi, Chengyuan Ji, Rui Wang, Chaoli Sun and Baoyi Lv

Antibiotics 2025, 14(4), 340; https://doi.org/10.3390/antibiotics14040340 - 26 Mar 2025

Cited by 2 | Viewed by 1145

Abstract

Background/Objectives: The propagation of antibiotic resistance genes (ARGs) poses a huge threat to environmental and human health. The ballast water from ships has been recognized as an important vector of ARGs. However, little is known about how ballast water from geographically isolated water affects ARGs in receiving waters. Methods: Herein, we investigated the changes in ARGs in receiving water by microcosm experiments simulating the discharge of ballast water. Results: The simulated discharge of ballast water increased the abundances of target ARGs, which were 1.3–5.6-fold higher in the mixture of ballast water and receiving water (microcosm M) than in receiving water at the end of the experiment. The enrichment of target ARGs was significantly associated with MGEs. Moreover, the discharge of ballast water changed the microbial communities in receiving water. Further network analysis identified potential ARG hosts, such as Pseudohongiellaa and Amphritea, with the abundance in microcosm M (0.23% and 0.036%) being higher than in receiving water (0.09% and 0.006%), the changes of which might be responsible for ARG variations. Conclusions: Overall, our findings suggest the discharge of ballast water might promote the spread of ARGs in different geographical waters and the corresponding ecological risks should not be ignored. Full article

(This article belongs to the Special Issue Antibiotic Resistance in Wastewater Treatment Plants)

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16 pages, 2846 KB

Open AccessArticle

Isolation and Characterization of Lytic Phages Infecting Clinical Klebsiella pneumoniae from Tunisia

by Donia Mourali, Rahma Kazdaghli, Marwa Gara-Ali, Houda Ben-Miled, Lucas Mora-Quilis, Pilar Domingo-Calap and Kamel Ben-Mahrez

Antibiotics 2024, 13(12), 1154; https://doi.org/10.3390/antibiotics13121154 - 2 Dec 2024

Cited by 4 | Viewed by 2606

Abstract

Background: Klebsiella pneumoniae is an opportunistic pathogen that causes a wide range of infections worldwide. The emergence and spread of multidrug-resistant clones requires the implementation of novel therapeutics, and phages are a promising approach. Results: In this study, two Klebsiella phages, KpTDp1 and KpTDp2, were isolated from wastewater samples in Tunisia. These phages had a narrow host range and specifically targeted the hypervirulent K2 and K28 capsular types of K. pneumoniae. Both phages have double-stranded linear DNA genomes of 49,311 and 49,084 bp, respectively. Comparative genomic and phylogenetic analyses placed phage KpTDp2 in the genus Webervirus, while phage KpTDp1 showed some homology with members of the genus Jedunavirus, although its placement in a new undescribed genus may be reconsidered. The replication efficiency and lytic ability of these phages, combined with their high stability at temperatures up to 70 °C and pH values ranging from 3.5 to 8.2, highlight the potential of these phages as good candidates for the control of hypervirulent multidrug-resistant K. pneumoniae. Methods: Phage isolation, titration and multiplicity of infection were performed. The stability of KpTDp1 and KpTDp2 was tested at different pH and temperatures. Genomic characterization was done by genome sequencing, annotation and phylogenetic analysis. Conclusions: The ability of KpTDp1 and KpTDp2 to lyse one of the most virulent serotypes of K. pneumoniae, as well as the stability of their lytic activities to pH and temperature variations, make these phages promising candidates for antibacterial control. Full article

(This article belongs to the Special Issue Antibiotic Resistance in Wastewater Treatment Plants)

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14 pages, 1405 KB

Open AccessArticle

Contrasting Dynamics of Intracellular and Extracellular Antibiotic Resistance Genes in Response to Nutrient Variations in Aquatic Environments

by Lele Liu, Xinyi Zou, Yuan Cheng, Huihui Li, Xueying Zhang and Qingbin Yuan

Antibiotics 2024, 13(9), 817; https://doi.org/10.3390/antibiotics13090817 - 28 Aug 2024

Cited by 4 | Viewed by 2204

Abstract

The propagation of antibiotic resistance in environments, particularly aquatic environments that serve as primary pathways for antibiotic resistance genes (ARGs), poses significant health risks. The impact of nutrients, as key determinants of bacterial growth and metabolism, on the propagation of ARGs, particularly extracellular ARGs (eARGs), remains poorly understood. In this study, we collected microorganisms from the Yangtze River and established a series of microcosms to investigate how variations in nutrient levels and delivery frequency affect the relative abundance of intracellular ARGs (iARGs) and eARGs in bacterial communities. Our results show that the relative abundance of 7 out of 11 representative eARGs in water exceeds that of iARGs, while 8 iARGs dominate in biofilms. Notably, iARGs and eARGs consistently exhibited opposite responses to nutrient variation. When nutrient levels increased, iARGs in the water also increased, with the polluted group (COD = 333.3 mg/L, COD:N:P = 100:3:0.6, m/m) and the eutrophic group (COD = 100 mg/L, COD:N:P = 100:25:5, m/m) showing 1.2 and 3.2 times higher levels than the normal group (COD = 100 mg/L, COD:N:P = 100:10:2, m/m), respectively. In contrast, eARGs decreased by 6.7% and 8.4% in these groups. On the other hand, in biofilms, higher nutrient levels led to an increase in eARGs by 1.5 and 1.7 times, while iARGs decreased by 17.5% and 50.1% in the polluted and eutrophic groups compared to the normal group. Moreover, while increasing the frequency of nutrient delivery (from 1 time/10 d to 20 times/10 d) generally did not favor iARGs in either water or biofilm, it selectively enhanced eARGs in both. To further understand these dynamics, we developed an ARGs-nutrient model by integrating the Lotka–Volterra and Monod equations. The results highlight the complex interplay of bacterial growth, nutrient availability, and mechanisms such as horizontal gene transfer and secretion influencing ARGs’ propagation, driving the opposite trend between these two forms of ARGs. This contrasting response between iARGs and eARGs contributes to a dynamic balance that stabilizes bacterial resistance levels amid nutrient fluctuations. This study offers helpful implications regarding the persistence of bacterial resistance in the environment. Full article

(This article belongs to the Special Issue Antibiotic Resistance in Wastewater Treatment Plants)

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