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Fluoroquinolone resistance in Enterobacteriales is developed by chromosomal and plasmid-mediated mechanisms. Plasmids play an important role in dissemination of resistant genes and they carry genes that protect bacteria in different stress-induced situations. In this study, we studied Escherichia coli strains, each carried one plasmid-mediated quinolone resistance determinant namely, qnrA1, qnrB1, qnrC1 , and qnrD1. We exposed 0.5 McFarland density of each strain to 0.5 mg/L ciprofloxacin from the period of 30, 60, 90, and 120 min over 24 h. All treated strains were further exposed to a constantly increasing 1, 2, 4, and 8 mg/L ciprofloxacin solution through 24, 48, and 120 h. In given timepoints, RNA was extracted from all treated strains. Expression of qnrA1, qnrB1, qnrC1 , and qnrD1 was investigated by quantitative PCR. Mutations in gyrA and parC genes were analyzed by PCR and nucleic acid sequencing. In this study, during 0.5 mg/L ciprofloxacin exposition, the following expression levels were detected: 1.2 for qnrA1 , 1.47 for qnrD1 , 12.44 for qnrC1 , and 80.63 for qnrB1 . In case of long-term study, we selected a resistant strain in qnrB1 -positive E. coli , and its expression increased from 105.91 to 212.31. On the contrary, plasmid copy number increased in time from 1 to 4.13. No mutations in gyrA or in parC chromosomal genes of treated strains were detected. Our results show tha qnrB1 -positive E. coli strain was able to develop fluoroquinolone resistance by upregulated qnrB1 expression that was linked to a minor increase in plasmid copy number but no mutations occurred in gyrA or parC.

Plasmid copy number and qnr gene expression in selection of fluoroquinolone-resistant Escherichia coli