Molecular investigation of the chlorine and antibiotic resistance mechanisms of Escherichia coli isolated from natural water sources in the Western Cape

[摘要] Water is used for various purposes and contamination can have severe implications ifuntreated. One of the most common and cost effective water disinfectants, especially usedin developing countries, is chlorine. However, microorganisms have developed differentmechanisms in response to environmental stress conditions, such as the viable but nonculturable(VBNC) effects possibly displayed in this study, enabling them to survive.Chlorine may also exert several effects on microorganisms, such as the expression ofmulti-substrate efflux pumps, decreased membrane permeability and transport inhibitionthat may lead to chlorine tolerance and antimicrobial resistance. In a descriptive andcomparative study, the molecular characteristics of E. coli strains isolated fromenvironmental waters in the Western Cape and the possible relationship betweenchlorination and antimicrobial resistance were investigated.Water and biofilm samples were exposed to chlorine, as well as efflux pump inhibitor (EPI)concentrations, and surviving E. coli strains were tested for their phenotypic characteristicsincluding antimicrobial susceptibility profiles and morphological types. Candidate genespossibly involved in resistance to antimicrobials, disinfection and efflux pumps weredetected with polymerase chain reaction (PCR) and sequenced. Sequencing analysis andhomology searches were done and E. coli strains were typed as either EnteropathogenicE. coli strains (EPEC) or Enterotoxigenic E. coli strains (ETEC) on the presence ofvirulence genes.All water and biofilm sources examined were heavily polluted with E. coli, and a highenumeration level of this indicator organism of faecal contamination was recorded.Chlorine tolerance was found to be associated with antimicrobial resistance. Addition ofEPI with exposure to chlorine decreased enumeration levels of these organisms,suggesting that efflux pumps may play a role in tolerance to chlorine. Severalmorphological patterns were described amongst the E. coli strains and a change in thiswas recorded after exposure to chlorine. Highly resistant antibiograms displayed by theisolated strains included ampC β-lactamase producing E. coli strains and extendedspectrum β-lactamases (ESBLs). Amplification of the candidate genes selected for heatshock,oxidative stress genes and efflux pump were most frequently detected while thestructural genes involved in fluoroquinolones (FQs) resistance were detected lessfrequently in the selected strains. Sequencing of these amplified candidate genesdemonstrated various changes in amino acid sequences, including one commonmutational pathway taken by E. coli when exposed to stress conditions. Further homologysearches of the sequenced candidate genes illustrated similarities in 19 pathogenic and 14non-pathogenic E. coli as well as 3 Shigella strains. Detection of virulence genes foundthree EPEC strains (bfpA, eaeA), two EPEC (eaeA), ten EPEC (bfpA) and one ETECstrain (st) amongst the isolates.This study underlines the need for monitoring our water sources, which poses a publichealth risk due to incomplete chlorination, antimicrobial resistance and the spread ofclinically relevant pathogenic strains.

[发布机构] Stellenbosch University