The Global Regulatory Cyclic AMP Receptor Protein (CRP) Controls Multifactorial Fluoroquinolone Susceptibility in Salmonella enterica Serovar Typhimurium

Abstract

Fluoroquinolone antibiotics are prescribed for the treatment of infections, but resistance to this family of antibiotics is growing. Here we report that loss of the global regulatory protein cyclic AMP (cAMP) receptor protein (CRP) or its allosteric effector, cAMP, reduces susceptibility to fluoroquinolones. A Δ mutation was synergistic with the primary fluoroquinolone resistance allele , thus able to contribute to clinically relevant resistance. Decreased susceptibility to fluoroquinolones could be partly explained by decreased expression of the outer membrane porin genes and with a concomitant increase in the expression of the ciprofloxacin resistance efflux pump gene in Δ cells. Expression of , which encode the DNA supercoiling enzyme GyrAB, which is blocked by fluoroquinolones, and expression of, which encodes the dominant supercoiling-relaxing enzyme topoisomerase I, were unchanged in Δ cells. Yet Δ cells maintained a more relaxed state of DNA supercoiling, correlating with an observed increase in topoisomerase IV () expression. Surprisingly, the Δ mutation had the unanticipated effect of enhancing fitness in the presence of fluoroquinolone antibiotics, which can be explained by the observation that exposure of Δ cells to ciprofloxacin had the counterintuitive effect of restoring wild-type levels of DNA supercoiling. Consistent with this, Δ cells did not become elongated or induce the SOS response when challenged with ciprofloxacin. These findings implicate the combined action of multiple drug resistance mechanisms in Δ cells: reduced permeability and elevated efflux of fluoroquinolones coupled with a relaxed DNA supercoiling state that buffers cells against GyrAB inhibition by fluoroquinolones.

Horizontal transfer of antibiotic resistance genes in clinical environments

“…Fluoroquinolones target DNA topoisomerases, and resistance requires amino acid changes to reduce antibiotic affinity for the topoisomerase called DNA gyrase. In most bacteria, including Enterobacteriaceae, clinical resistance to fluoroquinolones can only be achieved through the additive contributions of multiple genetic loci (Hooper 2001;Corkill et al 2005;Kary et al 2017;Osei Sekyere and Amoako 2017). Fluoroquinolones have a demonstrated ability to induce natural competence in Streptococcus (Prudhomme et al 2006); thus, antibiotic exposure increases the potential for HGT and consolidation of chromosomal resistance alleles.…”

Analysis of multidrug resistance in Streptococcus suis ATCC 700794 under tylosin stress

“…Some studies have provided evidence that the "site" mutations of EF-G result in large changes in gene expression such as cyaA (adenylate cyclase) and topA (topoisomerase I) [24]. The topA gene acts jointly with topoisomerase IV genes (parC and parE) and relaxes DNA supercoils in opposition to GyrAB activity, while the GyrAB protein is the most important site to form the resistance of fluoroquinolone [21]. Previous studies have reported that the tetracycline (TC) antibiotics inhibit bacterial protein synthesis by preventing the attachment of aminoacyl tRNA to the ribosomal acceptor A site [50].…”

“…It is well known that elongation factors play important roles in forming bonds between the tRNA attached peptidyltransferase and the next amino acid, ribosomal translation, causing premature dissociation of the peptidyl-tRNA from the ribosome and protein synthesis [16][17][18][19]. Previous studies have shown that elongation factors can not only inhibit bacterial protein synthesis using the antibiotics tetracycline, tylosin and fluoroquinolone [20,21], but also take part in imparting drug resistance in Escherichia coli. Moreover, elongation factors have been reported to be differentially expressed in tylosin-resistant Mycoplasma gallisepticum [22][23][24].…”

Modulation of the cAMP levels with a conserved actinobacteria phosphodiesterase enzyme reduces antimicrobial tolerance in mycobacteria

“…Cyclic AMP signalling, which is one of the best studied signalling pathways in biology, has been implicated in AMR 4,5 . Several earlier studies have linked cAMP signalling to antibiotic resistance in gram-negative bacteria 6,7 . For example, Alper et al .…”

“…Moreover, Kary et al . reported that Salmonella typhimurium strains lacking Crp (a cAMP-activated global transcriptional regulator) or with altered levels of cAMP exhibit reduced susceptibility to fluoroquinolones as a result of decreased permeability and increased efflux of this class of antibiotics 7 .…”