Role of the two component signal transduction system CpxAR in conferring cefepime and chloramphenicol resistance in Klebsiella pneumoniae NTUH-K2044

• Published in: PloS one 2012-04, Vol.7 (4), p.e33777
• Main Authors: Srinivasan, Vijaya Bharathi, Vaidyanathan, Vasanth, Mondal, Amitabha, Rajamohan, Govindan
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Amino acids; Anti-Bacterial Agents - toxicity; Antibiotics; Antiinfectives and antibacterials; Antimicrobial resistance; Bacillus subtilis; Bacterial Outer Membrane Proteins; Bacterial physiology; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Base Sequence; Beta lactamases; beta-Lactam Resistance - genetics; Bile; Biology; Blotting, Western; Cefepime; Cellular signal transduction; Cephalosporins - toxicity; Chloramphenicol; Chloramphenicol - toxicity; Chloromycetin; Chromosomes; Complementation; Computational Biology; Cross infection; DNA, Bacterial - genetics; Drug resistance; Efflux; Electrophoretic Mobility Shift Assay; Environmental changes; Environmental conditions; Escherichia coli; Gene expression; Health aspects; Kinases; Klebsiella; Klebsiella Infections - drug therapy; Klebsiella Infections - genetics; Klebsiella Infections - microbiology; Klebsiella pneumoniae; Klebsiella pneumoniae - drug effects; Klebsiella pneumoniae - growth & development; Klebsiella pneumoniae - pathogenicity; Lactams; Medicine; Membrane composition; Membrane proteins; Microbial drug resistance; Microbial Sensitivity Tests; Molecular Sequence Data; Multidrug resistance; Mycobacterium tuberculosis; Nosocomial infections; Operon - genetics; Osmotic Pressure; Oxidative Stress; Pathogens; Physiological aspects; Pneumonia; Protein folding; Protein Kinases - genetics; Protein Kinases - metabolism; Proteins; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Bacterial - genetics; RNA, Messenger - genetics; Salmonella enterica; Sequence Homology, Amino Acid; Signal transduction; Signaling; Stimulus-response coupling; Strain; Stresses
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0033777

Klebsiella pneumoniae is a gram-negative, non-motile, facultative anaerobe belonging to the Enterobacteriaceae family of the γ-Proteobacteria class in the phylum Proteobacteria. Multidrug resistant K. pneumoniae have caused major therapeutic problems worldwide due to emergence of extended-spectrum β-lactamase producing strains. Two-component systems serve as a basic stimulus-response coupling mechanism to allow organisms to sense and respond to changes in many different environmental conditions including antibiotic stress. In the present study, we investigated the role of an uncharacterized cpxAR operon in bacterial physiology and antimicrobial resistance by generating isogenic mutant (ΔcpxAR) deficient in the CpxA/CpxR component derived from the hyper mucoidal K1 strain K. pneumoniae NTUH-K2044. The behaviour of ΔcpxAR was determined under hostile conditions, reproducing stresses encountered in the gastrointestinal environment and deletion resulted in higher sensitivity to bile, osmotic and acid stresses. The ΔcpxAR was more susceptible to β-lactams and chloramphenicol than the wild-type strain, and complementation restored the altered phenotypes. The relative change in expression of acrB, acrD, eefB efflux genes were decreased in cpxAR mutant as evidenced by qRT-PCR. Comparison of outer membrane protein profiles indicated a conspicuous difference in the knock out background. Gel shift assays demonstrated direct binding of CpxR(KP) to promoter region of ompC(KP) in a concentration dependent manner. The Cpx envelope stress response system is known to be activated by alterations in pH, membrane composition and misfolded proteins, and this systematic investigation reveals its direct involvement in conferring antimicrobial resistance against clinically significant antibiotics for the very first time. Overall results displayed in this report reflect the pleiotropic role of the CpxAR signaling system and diversity of the antibiotic resistome in hyper virulent K1 serotype K. pneumoniae NTUH-K2044.