Project description:Polymyxins are increasingly used as the critical last-resort therapeutic options for multidrug-resistant gram-negative bacteria. Unfortunately, polymyxin resistance has increased gradually for the last few years. Although studies on mechanisms of polymyxin are expanding, system-wide analyses of the underlying mechanism for polymyxin resistance and stress response are still lacking. To understand how Klebsiella pneumoniae adapt to colistin (polymyxin E) pressure, we carried out proteomic analysis of Klebsiella pneumoniae strain cultured with different concentrations of colistin. Our results showed that the proteomic responses to colistin treatment in Klebsiella pneumoniae involving several pathways, including (i) gluconeogenesis and TCA cycle; (ii) arginine biosynthesis; (iii) porphyrin and chlorophyll metabolism; and (iv) enterobactin biosynthesis. Interestingly, decreased abundance of class A β-lactamases including TEM, SHV-11, SHV-4 were observed in cells treated with colistin. Moreover, we also present comprehensive proteome atlases of paired polymyxin-susceptible and -resistant Klebsiella pneumoniae strains. The polymyxin-resistant strain Ci, a mutant of Klebsiella pneumoniae ATCC BAA 2146, showed missense mutation in crrB. The crrB mutant Ci, which displayed lipid A modification with 4-amino-4-deoxy-L-arabinose (L-Ara4N) and palmitoylation, showed striking increases of CrrAB, PmrAB, PhoPQ, ArnBCADT and PagP. We hypothesize that crrB mutations induce elevated expression of the arnBCADTEF operon and pagP via PmrAB and PhoPQ. Moreover, multidrug efflux pump KexD, which was induced by crrB mutation, also contributed to colistin resistance. Overall, our results demonstrated proteomic responses to colistin treatment and the mechanism of CrrB-mediate colistin resistance, which may further offer valuable information to manage polymyxin resistance.
Project description:The transcriptional, epigenomic, and genomic profiles of K. pneumoniae isolates were characterised to identify novel colistin and carbapenem resistance mechanisms. The genomic DNA and total RNA of the isolates were isolated and sequenced on PacBio.
Project description:To explore how multiple drug-resistant A. baumannii response to colistin resistance, we compared the genomic, transcriptional and proteomic profile of A. baumannii MDR-ZJ06 to that of induced colistin resistant strain ZJ06-200P5-1.
Project description:Acinetobacter baumannii is often highly resistant to multiple antimicrobials, posing a risk of treatment failure, and colistin is a "last resort" for treatment of the bacterial infection. However, colistin resistance is easily developed when the bacteria are exposed to the drug, and a comprehensive analysis of colistin-mediated changes in colistin-susceptible and -resistant A. baumannii is needed. In this study, using an isogenic pair of colistin-susceptible and -resistant A. baumannii isolates, alterations in morphologic and transcriptomic characteristics associated with colistin resistance were revealed. Whole-genome sequencing showed that the resistant isolate harbored a PmrBL208F mutation conferring colistin resistance, and all other single-nucleotide alterations were located in intergenic regions. Using scanning electron microscopy, it was determined that the colistin-resistant mutant had a shorter cell length than the parental isolate, and filamented cells were found when both isolates were exposed to the inhibitory concentration of colistin. When the isolates were treated with inhibitory concentrations of colistin, more than 80% of the genes were upregulated, including genes associated with antioxidative stress response pathways. The results elucidate the morphological difference between the colistin-susceptible and -resistant isolates and different colistin-mediated responses in A. baumannii isolates depending on their susceptibility to this drug.
Project description:Colistin is a crucial last-line drug used for the treatment of life-threatening infections caused by multi-drug resistant strains of the Gram-negative bacteria, Acinetobacter baumannii. However, colistin resistant A. baumannii isolates can be isolated following failed colistin therapy. Resistance is most often mediated by the addition of phosphoethanolamine (pEtN) to lipid A by PmrC, following missense mutations in the pmrCAB operon encoding PmrC and the two-component signal transduction system PmrA/PmrB. We recovered an isogenic pair of A. baumannii isolates from a single patient before (6009-1) and after (6009-2) failed colistin treatment that displayed low/intermediate and high levels of colistin resistance, respectively. To understand how increased colistin-resistance arose, we genome sequenced each isolate which revealed that 6009-2 had an extra copy of the insertion sequence element ISAba125 within a gene encoding an H-NS-family transcriptional regulator. Consequently, transcriptomic analysis of the clinical isolates identified was performed and more than 150 genes as differentially expressed in the colistin-resistant, hns mutant, 6009-2. Importantly, the expression of eptA, encoding a second lipid A-specific pEtN transferase, but not pmrC, was significantly increased in the hns mutant. This is the first time an H-NS-family transcriptional regulator has been associated with a pEtN transferase and colistin resistance.
Project description:Here we investigated colistin effects on A. baumannii, employing a Mass spectrometry approach and comparing a colistin-susceptible multidrug-resistantclinical isolate to its colistin-dependent subpopulation obtained by subsequent passages in moderate colistin concentrations. Colistin dependence might represent a stepping-stone to resistance, but the mechanisms of colistin dependence are far from clear.