1. Comparative performance of electronegative membrane filtration and automated concentrating pipette for detection of antibiotic resistance genes and microbial markers in river water samples.
- Author
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Siri Y, Sthapit N, Malla B, Raya S, and Haramoto E
- Subjects
- Escherichia coli genetics, Escherichia coli drug effects, RNA, Ribosomal, 16S analysis, Japan, Water Microbiology, Rivers microbiology, Environmental Monitoring methods, Filtration, Drug Resistance, Microbial genetics
- Abstract
The target viral and bacterial concentrations in river water are essential for environmental monitoring and public health studies. Filtration-based methods are commonly employed, yet challenges arise due to recoverability and filter pore size. This study aimed to compare the performance of electronegative membrane filtration (EMF) and automated Concentrating Pipette (CP) Select (InnovaPrep) methods for quantifying antibiotic resistance genes (ARGs), mobile genetic elements (MGEs), and bacterial and viral markers in river water samples. Fifty-four river water samples were collected from upstream and downstream locations in a river in Japan. The CP Select method was modified by adding MgCl
2 and using different tips. The recovery efficiencies for total coliforms and Escherichia coli were assessed, and class 1 integron-integrase gene (intI1), 16S rRNA, gene encoding sulfonamide resistance (sul1), cross-assembly phage (crAssphage), pepper mild mottle virus (PMMoV), and Escherichia coli gene (sfmD) were detected. CP Select showed recovery efficiencies of 45 %-63 % for total coliforms and 17 %-35 % for E. coli. The intI1, 16S rRNA, sul1, crAssphage, PMMoV, and sfmD concentrations using the modified CP Select method were 10.1 ± 0.5, 8.7 ± 0.2, 7.7 ± 0.2, 6.7 ± 0.2, 5.4 ± 0.2, and 3.5 ± 0.5 log10 copies/L, respectively. Higher intI1 and sul1 concentrations were observed downstream, with the highest contribution percentage (22 % and 21 %) using CP Select or EMF. The modified CP Select method with 0.05 μm tips yielded more quantifiable results for all target genes and greater PMMoV concentrations (p < 0.05). Positive correlations were found among bacterial, ARG/MGE, and viral markers (Spearman's ρ = 0.71 for 16S rRNA and sfmD, 0.88 for intI1 and sul1, and 0.64 for PMMoV and crAssphage). The modified CP Select method demonstrated effective recovery of bacteria and quantification of ARGs, MGEs, and microbial markers in river water. Further studies are required to validate these methods and confirm their applicability in diverse environmental contexts., Competing Interests: Declaration of competing interest No conflict of interest exists in the submission of this manuscript or personal relationships, and the manuscript is approved by all authors for publication. The work has not been published before and is not currently being considered for publication anywhere., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)- Published
- 2024
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