Antibiotic resistance patterns & sequencing of class I integron in uropathogenic escherichia coli in Lebanon. (c2009)

Abstract

Improper monitoring of antibiotic usage has lead to the development of antimicrobial resistance among uropathogenic Escherichia coli (UPEC). The rapid dissemination of antibiotic-resistance is mediated by gene transfer mechanisms involving mobile genetic elements including the recendy characterized gene cassettes in integrons. Integrons are conserved DNA sequences associated with multi-drug resistance. The aim of this study was to generate data on the prevalence and molecular basis of antimicrobial resistance in UPEC in Lebanon. The phenotypic resistance profiles of 9 currendy used antimicrobial agents were determined using the agar diffusion test on a total 100 UPEC isolates. Results obtained revealed that 60% of the isolates were resistant to ampicillin, 55% to trimethoprim/ sulfamethoxazole, 53% to streptomycin and only 5% to netilmicin. PCR assay was used to detect the presence of the class 1 integron variable region (VR) containing the gene cassettes. All VR negative isolates showed more susceptibility towards the used antimicrobial agents compared to the VR positive isolates. The VR positive isolates showed highest resistance to aminoglycosides and trimethoprim/sulfamethoxazole (96.7%). VR negative islolates were mainly resistant to ampicillin (48.6%), aminoglycosides (41%), and tetracycline (40%). The VR amplicons were then characterized by direct partial sequencing and restriction digestion with AluI, and accordingly, 30% of the isolates were found to be positive for the Class 1 integron VR, with a size ranging from 0.7 to 2.2 Kbp. VR positive isolates carried the genes 4frA7, 4frAI7-aadA5, 4frAl-aadA1, 4frAI2-orf5-aadA2 and blaoXA _ 30-aadAl. The predominant resistance genes were 4frA 17 and aadA5 (47% of the isolates) for trimethoprim/ sulfamethoxazole and streptomycin, respectively. The presence of a gene cassette was correlated with high resistance to the corresponding antibiotic, with 85.7% of the isolates harboring the aadA gene being resistant to streptomycin. Five different restriction patterns were detected; all isolates with the same class 1 integron VR amplicon size had the same restriction pattern. Characterization of class I integrons from UPEC isolates by direct sequencing revealed that those isolates exhibit a wide repertoire of genetic elements to sustain antimicrobial pressure. This study provided basal information for future pursuit and comparison especially with respect to epidemiologic distribution, antimicrobial resistance and evolution of these important pathogens.