Asian Research Thesis Index

Abstract

Escherichia coli lineage ST131 predominates across various spectra of extra-intestinal infections. Distinctive resistance profile, diverse armamentarium of virulence factors and rapid global dissemination of ST131 E. coli makes it an intriguing pathogen of significant importance. However, little is known about the prevalence and various attributes of ST131 associated with urinary tract infections in Pakistan. Evolutionary dimensions of ST131 linage and corresponding molecular markers encoding drug resistance remained largely unexplored and prevalence data are extremely scarce in South Asian countries. We scrutinized prevalence and various genetic attributes of E. coli ST131 isolates involved in urinary tract infections by processing 148 randomly selected samples during August 2012 and August 2014. Phylogenetic grouping, fumC/fimH and O-typing was done by PCR based methods. Antibiotic susceptibility assays and phenotypic detection of ESBLs were carried out according to the CLSI guidelines of year 2013 and ESBL isolates were screened for the prevalence of blaCTX-M-15. Plasmid encoded qnrA, qnrB and qnrS genes were identified by DNA amplification and sequencing. Mutations in genes gyrA, gyrB, parC and parE were identified among fluoroquinolone resistant isolates. Minimum inhibitory concentrations (MICs) for ciprofloxacin and levofloxacin were determined by using broth dilution method. Phylogenetic analysis was performed by PCR using three gene classification systems (Clermont et al., 2000). Overall, 59% of the UPEC isolates belonged to phylogenetic group B2 which was followed by group D=28%, B1=8% and A=5%. Among 18 different ST-types ST131 was the most dominant lineage as 46% of the isolates belonged to ST 131 lineage. By, CH-typing 46% of the ST 131 isolates belonged to sub-group H30. Higher numbers of identified ST 131 isolates 74% were MDR and 44% showed ESBL phenotypes while 100% of the isolates carried blaCTX-M-15. Overall, resistant patterns of ST 131 isolates confirmed least resistance against tigecycline, meropenam and amikacin. Likewise, resistance against tazocine, nitrofurantoin, sulzone and minocycline remained below threshold level. For fosfomycin, (FF) and gentamicin (CN) resistance were 13% and 28% respectively. Higher resistances was noticed against all four generations of cephalosporins, (53%-78%). Higher percentages of ST 131 isolates were resistant to co-trimoxazole (85%), amoxicillin clavulanic acid 65% and fluoroquinolones 60%. Overall, minimum inhibitory concentrations of ciprofloxacin remained above 256μg/ml in ST 131 and non ST 131 isolates. In comparison to MICs of ciprofloxacin, MICs of levofloxacin in most of the isolates varied between 8μg/ml-64μg/ml. In this study, majority of the fluoroquinolone resistant UPEC ST 131 isolates carried at least two or more non-synonymous mutations in gyrA, parC and parE genes. These isolates showed higher MICs for both ciprofloxacin and levofloxacin. Conclusively, higher percentages of the ST 131 isolates showed antibiotic resistance against trimethoprimsulphamethoxazolen, cephalosporin, fluoroquinolone, and majority of these isolates were multi-drug resistant. In addition, fluoroquinolone resistant UPECs isolates shared lineage of a clonal sub-group. An allelic combination of gyrA/parC/parE genes of sub-clone O25b-ST131 H30-R was associated with higher MICs of ciprofloxacin and levofloxacin. Overall, it is the first report from Pakistan that provides details insights about susceptibility patterns and genetic attributes of pandemic MDR ST131 strains involved in UTIs.

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