Abstract
There is limited information on the comparative genomic diversity of antibiotic-resistant Escherichia coli from wastewater. We sought to characterize environmental E. coli isolates belonging to various pathotypes obtained from a wastewater treatment plant (WWTP) and its receiving waters using whole-genome sequencing (WGS) and an array of bioinformatics tools to elucidate the resistomes, virulomes, mobilomes, clonality, and phylogenies. Twelve multidrug-resistant (MDR) diarrheagenic E. coli isolates were obtained from the final effluent of a WWTP, and the receiving river upstream and downstream of the WWTP were sequenced on an Illumina MiSeq machine. The multilocus sequence typing (MLST) analysis revealed that multiple sequence types (STs), the most common of which was ST69 (n = 4) and ST10 (n = 2), followed by singletons belonging to ST372, ST101, ST569, ST218, and ST200. One isolate was assigned to a novel ST ST11351. A total of 66.7% isolates were positive for β-lactamase genes with 58.3% harboring the blaTEM1B gene and a single isolate the blaCTX−M−14 and blaCTX−M−55 extended-spectrum β-lactamase (ESBL) genes. One isolate was positive for the mcr-9 mobilized colistin resistance gene. Most antibiotic resistance genes (ARGs) were associated with mobile genetic support: class 1 integrons (In22, In54, In191, and In369), insertion sequences (ISs), and/or transposons (Tn402 or Tn21). A total of 31 virulence genes were identified across the study isolates, including those responsible for adhesion (lpfA, iha, and aggR), immunity (air, gad, and iss), and toxins (senB, vat, astA, and sat). The virulence genes were mostly associated with IS (IS1, IS3, IS91, IS66, IS630, and IS481) or prophages. Co-resistance to heavy metal/biocide, antibiotics were evident in several isolates. The phylogenomic analysis with South African E. coli isolates from different sources (animals, birds, and humans) revealed that isolates from this study mostly clustered with clinical isolates. Phylogenetics linked with metadata revealed that isolates did not cluster according to source but according to ST. The occurrence of pathogenic and MDR isolates in the WWTP effluent and the associated river is a public health concern.
Highlights
The role of the environment in the spread of antibiotic resistance is an evolving issue [1]
The isolates belonged to the diarrheagenic group of E. coli; seven were enteroaggregative E. coli (EAEC), three were enteroinvasive E. coli (EIEC), with one enterohemorrhagic E. coli (EHEC), and one enteropathogenic E. coli (EPEC) (Supplementary Table 2)
Genomic insights reported in this study revealed the complexity and diversity of lineages, resistome, mobilome, and virulome of multi-drug resistant (MDR) E. coli found in wastewater and river water in Kwazulu Natal, South Africa, intimating that the aquatic environment contains a fluid and dynamic milieu of antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs)
Summary
The role of the environment in the spread of antibiotic resistance is an evolving issue [1]. With inadequately maintained sanitation infrastructure, lowand middle-income countries (LMICs) and emerging economies like South Africa face challenges with the release of untreated or poorly treated effluent into the environment, which may be a driver for the dissemination of antibiotic resistance in these settings [2]. Constant monitoring of WWTPs for the release of multi-drug resistant (MDR) bacteria into receiving waters via their effluents is important as it indicates what is disseminated to the environment. The WWTP investigated in this study is the largest in Pietermaritzburg, the provincial capital of KwaZulu-Natal in South Africa. Runoff from this WWTP is released into the Msunduzi River, a tributary that discharges into the Umgeni River [3]. The river water has previously been considered to be polluted with fecal matter and unsuitable for anthropogenic activities [4]
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