Genome-Resolved Metagenomics and Antibiotic Resistance Genes Analysis in Reclaimed Water Distribution Systems

Changzhi Wang,Pei-Ying Hong

doi:10.3390/w12123477

Abstract

Water reuse is increasingly pursued to alleviate global water scarcity. However, the wastewater treatment process does not achieve full removal of biological contaminants from wastewater, hence microorganisms and their genetic elements can be disseminated into the reclaimed water distribution systems (RWDS). In this study, reclaimed water samples are investigated via metagenomics to assess their bacterial diversity, metagenome-assembled genomes (MAGs) and antibiotic resistance genes (ARGs) at both point of entry (POE) and point of use (POU) in 3 RWDS. The number of shared bacterial orders identified by metagenome was higher at the POE than POU among the three sites, indicating that specific conditions in RWDS can cause further differentiation in the microbial communities at the end of the distribution system. Two bacterial orders, namely Rhizobiales and Sphingomonadales, had high replication rates in two of the examined RWDS (i.e., site A and B), and were present in higher relative abundance in POU than at POE. In addition, MAG and ARG relative abundance exhibited a strong correlation (R2 = 0.58) in POU, indicating that bacteria present in POU may have a high incidence of ARG. Specifically, resistance genes associated with efflux pump mechanisms (e.g., adeF and qacH) increased in its relative abundance from POU to POE at two of the RWDS (i.e., site A and B). When correlated with the water quality data that suggests a significantly lower dissolved organic carbon (DOC) concentration at site D than the other two RWDS, the metagenomic data suggest that low DOC is needed to maintain the biological stability of reclaimed water along the distribution network.

Highlights

Water scarcity is predicted to affect at least 45 countries worldwide by the year 2050 [1]
Short reads of metagenome-assembled genomes (MAGs) from each water sample were extracted by Samtools [23] and they were mapped to nucleotide sequences of antibiotic resistance genes (ARGs)
This study demonstrates the use of metagenomics and relevant bioinformatics to assess reclaimed water quality along the distribution networks

Summary

Introduction

Water scarcity is predicted to affect at least 45 countries worldwide by the year 2050 [1]. Alternative water resources like treated wastewater are increasingly being used for both indirect potable (e.g., drinking water in Singapore and California) and non-potable (e.g., irrigating agricultural crops in California) purposes. Wastewater is conventionally treated through multiple barriers comprising of various technologies, for example clarifiers, activated sludge tanks, sand filters, and/or disinfection. Depending on the reuse purpose, local guidelines allow permissible levels of biochemical oxygen demand (BOD5 , an indicator of organic carbon content), total suspended solids (TSS) and pH in the reuse water. Agency (US-EPA) suggests ≤ 10 mg/L BOD5 and 2 NTU (for turbidity) in treated wastewater to be used for surface or spray irrigation of any food crops [2].

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Conclusion