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Antibiotic resistance genes, antibiotic residues, and microplastics in influent and effluent wastewater from treatment plants in Norway, Iceland, and Finland
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Researchers used Oxford Nanopore metagenomic sequencing, qPCR, HPLC, and µFTIR spectroscopy to simultaneously track antibiotic resistance genes, antibiotic residues, and microplastics in wastewater treatment plant influents and effluents in Norway, Iceland, and Finland. Sequencing identified 193 unique ARGs, with patterns suggesting that treatment processes affect ARG abundance but do not eliminate them, and that MPs co-occur with resistance-promoting conditions.
ABSTRACT Monitoring antimicrobial resistance genes (ARGs) in wastewater influent (pre-treatment) and effluent (post-treatment) reveals their circulation in communities via wastewater surveillance (WS), possible amplification during treatment, and potential public health risks from gene releases into surface water. This study used Oxford Nanopore (ONP) metagenomic sequencing and qPCR to track ARGs in wastewater treatment plants (WWTPs) influents and effluents in Mekjarvik (Norway), Reykjavik (Iceland), and Mariehamn (Åland-Finland). High-Performance Liquid Chromatography (HPLC) monitored antibiotic residues and Micro-Fourier Transform Infrared Spectroscopy (µ-FTIR) used for microplastics (MPs) in Mekjarvik and Reykjavik. Metagenomic analysis identified 193 unique ARGs, with the highest average (±SD) in Reykjavik (66.3 ± 4.1), followed by Mekjarvik (61.3 ± 14.1) and Mariehamn (18.0 ± 2.2). While treatment generally reduced ARGs, ONP and qPCR detected amplification of some carbapenemase and ESBL genes. ONP sequencing linked many ARGs to plasmids, co-occurring with metal stress genes. The most prevalent plasmids— Col440I , IncQ2 , and ColRNAI —were found across all WWTPs. Mercury-related genes dominated metal stress genes (64.9%), followed by multimetal (23.7%) and copper (6.4%) stress genes. Among 45 antibiotics screened in Mekjarvik and Reykjavik, only sulfamethoxazole and sulfapyridine were consistently quantified, while azithromycin, ciprofloxacin, and ofloxacin were often below detection limits. MPs were highest in Reykjavik influent (8200 MPs/m³) and Mekjarvik influent (5900 MPs/m³). Treatment effectively reduced larger MPs but was less effective against smaller particles. Polyethylene (∼60%) was the most dominant MP type, except in Mekjarvik influent, where polypropylene (∼50%) prevailed. This study reveals distinct ARG and antibiotic residue patterns in wastewater. While treatment significantly reduced ARGs, antibiotic residues, and larger MPs, it did not eliminate them, posing risks for environmental pollution. ARGs related to carbapenemase and ESBL persisted, and fine MPs increased post-treatment. These findings underscore the need to monitor both influent and effluent to have information about removal efficiencies and needs to protect downstream water environments. The detection of diverse ARGs, plasmids, and genes of critical pathogens like Acinetobacter baumannii , Pseudomonas aeruginosa , Escherichia coli , and Salmonella spp. in wastewater effluent presents a significant environmental challenge and emerging pollutant for recipient waters. Highlights Influent and effluent wastewater samples from Norway, Iceland, and Finland were analyzed. Resistance genes were analyzed via high-throughput qPCR and Oxford Nanopore (ONP) metagenomics. Dominant ARG groups in Mekjarvik and Mariehamn were tetracycline and macrolide but Quinolone, and macrolide in Reykjavik. Only sulfamethoxazole and sulfapyridine were consistently detected in Mekjarvik and Reykjavik, out of 45 screened antibiotics. Treatment effectively removed larger MPs but was less effective against smaller ones.
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