Phenotypic Detection of AAC(6 ′)-Ib-Cr Expression in Enterobacteriacae Isolates

Abstract

Background: Quinolone resistance in Enterobacteriaceae usually results from mutations in genes carried by chromosomally encoded type II topoisomerases, efflux pumps, or porin-related proteins. Since 1998, plasmid-mediated quinolone resistances (PMQR) have been described. Recently, a new mechanism of transferable quinolone resistance was reported: enzymatic inactivation of certain quinolones. AAC(6′)-Ib-cr is a variant of AAC(6′)-Ib which encodes an aminoglycoside acetyltransferase that confers reduced susceptibility to ciprofloxacin or norfloxacin.. PMQR determinants are often combined with extended-spectrum beta-lactamases (ESBLs) leading to organisms possessing multi drug resistance. Aims & Objectives: To study the incidence of AAC(6′)-Ib-cr expression in Enterobacteriacae isolates by phenotypic method and to find any correlation between the genetic expression of AAC(6′)-Ib-cr along with ESBL production. Materials and methods: The 430 clinical Enterobacteriaceae isolates exhibiting ciprofloxacin and norfloxacin resistance were included in this study. They were grown in Luria-Bertani Broth ( LB broth) containing norfloxacin (8 µg/ml), with intermittent shaking for 18 h at 35°C. Ten microliters of each culture medium was applied on the blank disk set on a Mueller-Hinton agar plate inoculated with E. coli ATCC 25922 and incubated for 18 h at 35°C. A significant decrease of a growth-inhibitory zone by ≥ 10 mm was considered as a positive test for the production of aac(6′)-Ib-cr, while aac(6′)-Ib-cr production was considered not to be produced in strains which did not show any reduction in the zone of inhibition. Results: The overall expression of the AAC(6′)-Ib-cr did not show much variation with almost half of the isolates from all the different samples showing AAC(6′)-Ib-cr production with blood isolates showing a production as high as 66.77%.). A significant association was noted with ESBL production and the simultaneous expression of AAC(6′)-Ib-cr in both E.coli (P value <0.0001) and K.pneumoniae isolates (P value = 0.0137 )which were the two most predominant isolates in the study. A significant association (P value 0.0442) was observed between aminoglycoside resistance and AAC(6’)-Ib-cr expression in E.coli isolates only, while no significant association was observed (P value= 0.7763) in K. pneumonia isolates. Conclusion: AAC(6′)-Ib-cr, though being a recently described gene has high degree of expression in Enterobacteriaceae isolates, which not only confers resistance to fluroquinolones and aminoglycosides but also is coexpressed with ESBL genes, thus producing a scenario of multi drug resistant organisms, further increasing the burden on our limited armamentarium against these bacteria.