PCR-Based Replicon Typing of Plasmids in E.Coli Producing Esbls in Patients with UTI in Iraq

Abstract

Objectives:The aim of this work was to study the distribution of antibiotics resistance genes of ESBLs in clinical E. coli in patients with UTI as well as identification and classification of bacterial plasmids associated with virulence and antimicrobial resistance by using PCR-Based Replicon Typing (PBRT).

Materials and methods: ESBLs- producing E. coli were preliminary screened by cephalosporin confirmation tests. Antimicrobial susceptibility test was performed of all ESBLs- producing E. coli. For genotypic detection, CTX-M, TEM and SHV ESBL genes were determined by PCR. PBRT was performed by PBRT 2.0 KIT (Diatheva, Italy, 2020), as well as ESBLs resistance transfer assay was performed by conjugation method.Results: The prevalence of ESBL in this study is high. Overall, bla CTX-M was the commonest genotype (93.6%) in E. coli by PCR. 26 replicons were detected in CTX-M producing E. coli strains:" HI- 1,  HI- 2, I1- α, I- 2, X- 1, X- 2,X- 3,X -4, L, I1y, N, FIA, FIB, FIC, FII, W, Y, P1,I1 A/C, T, k, U, R, B/O, HIB-M and FIB-M "; 25 replicons  were detected of TEM and SHV by PBRT. After conjugation was performed; transconjugants could be obtained at ratio 100% in E. coli producing CTX-M; and 88% in TEM and SHV.

Conclusions: The PBRT scheme exhibited anexcellent specificity and sensitivity, detecting new epidemiologically relevant replicons: P1, N2 and X4 in ESBLs-producing E. coli. The PBRT method for the(plasmids analysis) by the(PBRT 2.0) kitretrenches the detection time and provided that simplifies the steps for identification and classification ofE. coli plasmids.