Whole Genome Sequencing of Extended-Spectrum Beta-Lactamase (ESBL)-Producing Escherichia coli Isolated From a Wastewater Treatment Plant in China

Abstract

Background and Objectives Wastewater treatment plants (WWTPs) are one of the major reservoirs for antimicrobial resistant bacteria (ARB) and antimicrobial resistance genes (ARGs) in the environment. Thus, the investigation on ARB and ARGs from WWTPs has attracted increasing attention in recent years. In order to uncover the resistome in a WWTP treating effluents from a pharmaceutical industry in China, the extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli strains were isolated and their whole genome sequences were obtained and analyzed. Moreover, metagenomic sequencing was applied to give a comprehensive view of antibiotic resistance in this WWTP. Methods 18 ESBL-producing E. coli strains were isolated from a WWTP located in Taizhou, China on April, 2017. All strains were sequenced using Illumina HiSeq 2000 sequencer. The whole genome sequences were assembled using SPAdes software and annotated with RAST server. Sequence types (STs), plasmids, ARGs and virulence genes were predicted from the genomes using MLST, Plasmid Finder, ResFinder and Virulence Finder, respectively. Metagenomic DNA of the same sample was extracted and sequenced using Illumina Hiseq X Ten platform. Metagenomic sequences were assembled using SOAPdenovo software. Results All 18 ESBL-producing E. coli strains were resistant to ampicillin, cefazolin, and ceftriaxone. Analysis of their genomes revealed that all strains carried beta-lactamase encoding genes and the most prevalent type was bla CTX–M . Various virulence genes and ARGs confronting resistance to other types of antimicrobial agents were also predicted. Further investigation on the metagenomics data indicated 11 ARGs with high amino acid identities to the known ARGs. Five of these ARGs, aadA1 , aac(6′)-lb-cr , flo(R) , sul2 and sul1 , were also present in the genomes of the ESBL-producing E. coli isolated from the same sample. Conclusion Our study revealed the resistome of a pharmaceutical WWTP by both culture-dependent and metegenomic methods. The existence of ESBL-producing E. coli strains, indicating that pharmaceutical WWTP can play a significant role in the emergence of ARB. The occurrence of ARGs annotated from the metagenomic data suggests that pharmaceutical WWTP can play a significant role in the emergence of ARGs. Our findings highlight the need for strengthening the active surveillance of ARB and ARGs from pharmaceutical industry.

Genomic Insights of Multidrug-Resistant Escherichia coli From Wastewater Sources and Their Association With Clinical Pathogens in South Africa

“…A huge diversity of virulence genes often associated with pathogenic E. coli was found in the genomes of isolates in this study ( Table 1). Similar virulence factors have been identified in environmental E. coli isolates obtained from surface water and WWTPs in previous studies (8,36). Most virulence genes were associated with the insertion sequences, suggesting that these are important in the mobilization of the bacterial virulome ( Table 4).…”

Whole Genome Sequencing and Characterization of Multidrug-Resistant (MDR) Bacterial Strains Isolated From a Norwegian University Campus Pond

“…The T3SS effector leucine-rich repeat protein EspR1 was present in strains NMBU_ W12E19 and NMBU_ W13E19, while only strain NMBU_W13E19 was positive for the EspR2 protein. Strain NMBU_W13E19 displayed the gene combination eilA, Air and lpfA (encoding for long polar fimbriae) that was recently reported from a bla CTX−M -producing E. coli strain isolated from a wastewater treatment plant in China (Jiang et al, 2019) and in eight food-producing animal isolates (seven turkey and one laying hen) in Poland (Zajac et al, 2019). This particular gene combination was also seen in the first clinical isolate of an E. coli harboring the mcr-1 gene in Mexico, isolated in 2017 (Merida-Vieyra et al, 2019), as well as in six ESBL-producing E. coli strains isolated from veterinary hospital staff and students in three United Kingdom veterinary hospitals (Royden et al, 2019).…”

<p>Co-Existence of <em>mcr-1</em> and <em>bla</em>_NDM-5 in an <em>Escherichia</em> <em>coli</em> Strain Isolated from the Pharmaceutical Industry, WWTP</p>

“…In our previous study, we obtained several ESBLproducing E. coli strains from a WWTP located in Taizhou, Zhejiang province, China. 9 This WWTP treats the wastewater from a pharmaceutical company which produces antimicrobial agents including beta-lactam antibiotics such as cefaclor, ceftizoxime, ceftibuten, cefuroxime sodium, cefprozil, cefdinir, and cefixime. Here, we reported the isolation of another strain, E. coli strain R15, from this WWTP.…”

“…The isolation and identification of R15 was performed as described previously. 9 Antimicrobial susceptibility testing by VITEK 2 Compact (bioMérieux, France) showed that R15 was resistant to various types of antimicrobial agents, including ampicillin (MIC ≥ 32 μg/mL), amoxicillin (MIC ≥32 μg/ mL), piperacillin (MIC ≥ 128 μg/mL), cefazolin (MIC ≥ 64 μg/mL), cefoxitin (MIC ≥ 64 μg/mL), ceftriaxone (MIC ≥ 64 μg/mL), cefepime (MI C≥ 64 μg/mL), ertapenem…”