Molecular analysis of multidrug resistance in clinical isolates of Shigella spp. from 2001–2010 in Kolkata, India: role of integrons, plasmids, and topoisomerase mutations

To understand the genetic basis of high drug resistance in Shigella, 95 clinical isolates of Shigella spp. (2001-2010) were obtained from the Infectious Diseases Hospital, Kolkata, India. Ninety-three isolates were resistant to three or more antibiotics. Resistance to nalidixic acid, trimethoprim, streptomycin, and co-trimoxazole was most common in this population. Dendrogram analysis showed that S. sonnei strains were more clonally related when compared to the other Shigella species. The role of mobile genetic elements and chromosome-borne resistance factors was analyzed in detail. Integron analysis indicated the preponderance of class 2 and atypical class 1 integrons in that population. Typical class 1 integron was present in only one S. sonnei isolate and harbored trimethoprim resistance-encoding gene dfrV, while atypical class 1 integrons harbored dfrA1-aadA or blaOXA-aadA gene cassettes responsible for resistance to trimethoprim, aminoglycosides, and β-lactams. Class 2 integrons harbored either dfrA1-sat-aadA or dfrA1-sat gene cassettes. Most importantly, a novel gene cassette array InsE-InsO-dfrA1-sat was found in class 2 integron of S. sonnei NK4846. Many of the resistance traits for antibiotics such as trimethoprim, co-trimoxazole, kanamycin, ampicillin, and tetracycline were transferred from parent Shigella isolates to recipient Escherichia coli during conjugation, establishing the role of plasmids in horizontal transfer of resistance genes. Multiple mutations such as S80→I, S83→L, and D87→G/N/Y in quinolone resistance determining regions of topoisomerases from the representative quinolone-resistant isolates could explain the spectrum of minimal inhibitory concentration values for various quinolones. To the best of our knowledge, this is the first comprehensive report that describes the contribution of mobile (plasmids, integrons, and quinolone resistance genes named qnr) and innate genetic elements (mutations in topoisomerases) in determining the resistance phenotype of all the four species of Shigella over a span of ten years.