Antibiotic Resistance and Sewage-Associated Marker Genes in Untreated Sewage and a River Characterized During Baseflow and Stormflow.

Warish Ahmed,Erin M Symonds,Stuart L Simpson,Erica Donner,Sayalee Joshi,David Aster,Kerry A Hamilton,Pradip Gyawali

doi:10.3389/fmicb.2021.632850

Abstract

Since sewage is a hotspot for antibiotic resistance genes (ARGs), the identification of ARGs in environmental waters impacted by sewage, and their correlation to fecal indicators, is necessary to implement management strategies. In this study, sewage treatment plant (STP) influent samples were collected and analyzed using quantitative polymerase chain reaction (qPCR) to investigate the abundance and correlations between sewage-associated markers (i.e., Bacteroides HF183, Lachnospiraceae Lachno3, crAssphage) and ARGs indicating resistance to nine antibiotics (belonging to aminoglycosides, beta-lactams, sulfonamides, macrolides, and tetracyclines). All ARGs, except blaVIM, and sewage-associated marker genes were always detected in untreated sewage, and ermF and sul1 were detected in the greatest abundances. intl1 was also highly abundant in untreated sewage samples. Significant correlations were identified between sewage-associated marker genes, ARGs and the intl1 in untreated sewage (τ = 0.488, p = 0.0125). Of the three sewage-associated marker genes, the BIO-ENV procedure identified that HF183 alone best maximized correlations to ARGs and intl1 (τ = 0.590). Additionally, grab samples were collected from peri-urban and urban sites along the Brisbane River system during base and stormflow conditions, and analyzed for Escherichia coli, ARGs, the intl1, and sewage-associated marker genes using quantitative polymerase chain reaction (qPCR). Significant correlations were identified between E. coli, ARGs, and intl1 (τ = 0.0893, p = 0.0032), as well as with sewage-associated marker genes in water samples from the Brisbane River system (τ = 0.3229, p = 0.0001). Of the sewage-associated marker genes and E. coli, the BIO-ENV procedure identified that crAssphage alone maximized correlations with ARGs and intl1 in river samples (τ = 0.4148). Significant differences in E. coli, ARGs, intl1, and sewage-associated marker genes, and by flow condition (i.e., base vs. storm), and site types (peri-urban vs. urban) combined were identified (R = 0.3668, p = 0.0001), where percent dissimilarities between the multi-factorial groups ranged between 20.8 and 11.2%. Results from this study suggest increased levels of certain ARGs and sewage-associated marker genes in stormflow river water samples compared to base flow conditions. E. coli, HF183 and crAssphage may serve as potential indicators of sewage-derived ARGs under stormflow conditions, and this merits further investigation. Data presented in this study will be valuable to water quality managers to understand the links between sewage pollution and ARGs in urban environments.

Highlights

Sewage treatment plants (STPs) collect sewage from various sources, including households, hospitals, commercial and industrial sites followed by treatment processes to remove biological and chemical contaminants before the treated water is discharged back into the environment or recycled for commercial enterprises
All water samples collected during the baseflow and stormflow from peri-urban and urban sites were positive for E. coli
The antibiotic resistance genes (ARGs) blactx−m−32, sul2, and vanA were not detected in any water sample collected during baseflow from these peri-urban and urban sites. intl1 and ARG sul1 were most frequently detected in both baseflow and stormflow, the frequency of detection was greater in peri-urban sites compared to urban sites

Summary

Introduction

Sewage treatment plants (STPs) collect sewage from various sources, including households, hospitals, commercial and industrial sites followed by treatment processes to remove biological and chemical contaminants before the treated water is discharged back into the environment or recycled for commercial enterprises. Sewage contamination results in the dissemination of pathogens, nutrients, toxicants, endocrine disruptors, antibiotic resistant bacteria (ARB) and antibiotic resistance genes (ARGs) into the environment (Rodriguez-Manzano et al, 2010). Antibiotic resistant bacteria pose significant human health risks. At least 2.8 million people contract an antibiotic-resistant infection, and more than 35,000 deaths occur each year in the United States as per CDC’s 2019 AR Threats Report (CDC, 2019). In Australia, 290– 1,600 deaths/year are due to antibiotic resistance (Australian Commission on Safety and Quality in Health Care (ACSQHC), 2019). ARGs are considered emerging contaminants in the aquatic environments since they can be transferred to pathogenic bacteria in ecosystems via horizontal gene transfer (HGT) (Nguyen et al, 2019). The World Health Organization (WHO) identified water safety and improved sanitation as critical components in preventing the spread of ARB (WHO, 2014)

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