|O'Regan, Edel -|
|Quinn, Teresa -|
|Pages, Jean-Marie -|
|Porwollik, Steffen -|
|Mcclelland, Michael -|
|Fanning, Seamus -|
Submitted to: Antimicrobial Agents and Chemotherapy
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: November 6, 2009
Publication Date: January 5, 2010
Citation: O'Regan, E., Quinn, T., Frye, J.G., Pages, J., Porwollik, S., Cray, P.J., Mcclelland, M., Fanning, S. 2010. Fitness Costs and Stability of a High-Level Ciprofloxacin Resistance Phenotype in Salmonella enterica Serotype Enteritidis: Reduced Infectivity Associated with Decreased Expression of Salmonella Pathogenicity Island 1 Genes. Antimicrobial Agents and Chemotherapy. 54(1):367-374. Interpretive Summary: Antimicrobial resistance in bacteria has been shown to often affect bacterial fitness which is related to survival. To investigate how resistant bacteria may adapt to maintaining resistance, fluoroquinolone (ciprofloxacin) resistant Salmonella mutant strains were compared to their sensitive wild-type parental strains. Two resistant mutant strains displayed altered morphology on agar and by electron microscopy, reduced growth rates, motility, invasiveness into host cells and increased sensitivity to environmental stresses when compared to their parental strains. Microarray data revealed decreased expression of virulence and motility genes in both mutants. Upon passage on antibiotic-free agar, strains regained all abilities with the exception of motility. Potential mechanisms associated with reversal of the resistance phenotype and fitness costs were attributed to several compensatory mutations in regulation and gene expression. In conclusion, ciprofloxacin resistance in Salmonella Enteritidis is associated with fitness costs. In the absence of antibiotic selection pressure isolates may acquire mutations enabling reversion to a lower-level ciprofloxacin resistance phenotype associated with lower fitness costs.
Technical Abstract: The fitness costs associated with high-level fluoroquinolone resistance were examined in phenotypically and genotypically characterized ciprofloxacin-resistant Salmonella Enteritidis mutants (104-cip and 5408-cip, MIC > 32 µg/ml). The stability of the fluoroquinolone resistance phenotype in both mutants was investigated to assess whether clones with better fitness could emerge in the absence of antibiotic selective pressure. 104-cip and 5408-cip displayed altered morphology on agar and by electron microscopy, reduced growth rates, motility and invasiveness in Caco-2 cells and increased sensitivity to environmental stresses. Microarray data revealed decreased expression of virulence and motility genes in both mutants. Two clones (104-revert and 1A-revertC2) with ciprofloxacin MICs of 3 and 2 µg/ml were recovered from separate lineages of 104-cip after 20 and 70 passages on antibiotic-free agar, respectively. All fitness costs, except motility, were reversed in 104-revert. Potential mechanisms associated with reversal of the resistance phenotype were examined. Compared to 104-cip, both 104-revert and 1A-revertC2 showed decreased expression of acrB and soxS but still over-expressed marA. Both acquired additional mutations in soxR and parC and 1A-revertC2 harboured two mutations in marA. The altered porin and LPS profiles observed in 104-cip were partially reversed. In contrast, 5408-cip showed no reversal in fitness costs and maintained its high-level ciprofloxacin resistance for 200 passages on antibiotic-free agar. In conclusion, high-level ciprofloxacin resistance in Salmonella Enteritidis is associated with fitness costs. In the absence of antibiotic selection pressure isolates may acquire mutations enabling reversion to an intermediate-level ciprofloxacin resistance phenotype associated with less significant fitness costs.