Application of nanopore sequencing to identify antimicrobial resistance genes, mobile genetic elements and virulence factors in clinical isolates

Abstract

AbstractThe global health challenge posed by the emergence of antibiotic resistance pathogen is further exacerbated in African countries by the indiscriminate use of antibiotics, poor surveillance and lack of stewardship programs. To address this issue, we employed the Oxford Nanopore Technologies (ONT) to sequence 17 clinical isolates from a referral hospital in Kenya. Our comprehensive bioinformatics approach facilitated the assembly, identification of sequence types and prediction of antimicrobial resistance genes, mobile genetic elements (plasmids and integrons) and virulence genes. Of the 17 isolates, five wereA. baumannii, fourE. coli, threeS. haemolyticus, three wereE. cloacae,whileS. aureusandE. faecaliswere single isolates. For the detection of AMR genes,A. baumanniiisolates harbored genes such asblaOXA-23which mediates resistance to carbapenems,E. coliandE. cloacaecarriedblaCTX-M-15which confers resistance to cephalosporins andS. haemolyticusharboredblaZ,responsible for resistance againstpenicillins.S. aureusco-habouredmecAandblaZ genes.In addition,, various other different AMR genes to chloramphenicol, macrolides, aminoglycosides, tetracycline were also observed. For plasmid replicons,E. colicarried the most number of plasmids and shared ColRNAI_1 and IncFIB(pB171)_1_pB171 withA. baumanniiand IncR_1 withE. cloacae.Many genes encoding various virulence factors includingfimA-Iand ompA, senBwere identified inE. coli, hlgA-Candhla/hly, hlb, hldinS. aureusandefaA,ebpA-CinE. faecalis.In conclusion, most isolates contained a combination of different AMR genes harbored in plasmids and integrons and virulence genes. This study provides significant information on genetic determinants of antibiotic resistant pathogens in clinical isolates and could assist in developing strategies that improve patient treatment.Author SummaryAntimicrobial resistance remains a major health challenge across the globe. The continued misuse and lack of proper monitoring has worsened the problem of antibiotic resistant infections. In this study, we sought to use nanopore sequencing to identify antibiotic resistance genes, mobile genetic elements and virulence factors from clinical isolates which showed resistance against commonly used antibiotics. We found the presence of resistance genes to multiple different antibiotics including beta-lactams, macrolides, tetracycline and aminoglycosides across multiple bacterial species. We also identified plasmid replicons and class I integrons which facilitate the spread of antimicrobial resistant genes. Furthermore, several virulence factors that help resistant bacteria to survive were identified. Overall, this study highlights the widespread issue of antibiotic resistance, factors contributing to its persistence in clinical isolates and utility of nanopore sequencing for monitoring genetic determinants of antimicrobial resistance.