Project description:Acinetobacter baumannii is often highly resistant to multiple antimicrobials, posing a risk of treatment failure. Colistin is often chosen as a “last resort” for treatment of the bacterial infection, but resistance is easily developed when the bacteria is exposed to the drug. Thus a comprehensive analysis of colistin-mediated changes in colistin-susceptible and colistin-resistant A. baumannii is needed. In this study, we used a colistin-susceptible A. baumannii clinical isolate and a colistin-resistant isogenic mutant. Whole genome sequencing revealed 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, we observed that the colistin-resistant mutant had a shorter cell length than the parental isolate, and filamented cells were observed when both isolates were exposed to 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. This results helped a better understanding for the morphological difference between the colistin-susceptible and –resistant isolates and differed colistin-mediated responses in A. baumannii isolates by 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.
Project description:In past, resistance mechanisms have been identified by analysis of resistant isolates or defined mutants. Recently, high-throughput transposon mutagenesis coupled with sequencing (TraDIS-Xpress) is another approach proving useful for elucidating the roles of genes involved in the overall cellular response to a particular stress. In this study, we used TraDIS-Xpress to determine the role played by genes following exposure to colistin stress. Approximately 10^7 cells from the mutant library were inoculated into LB broth at a range of doubling concentrations of colistin ( 0.25 x MIC, 0.5 x MIC, 1 x MIC, 2 X MIC). Experiments were performed with no induction, or with induction using 0.2 or 1 mM of Isopropyl β-D-1-thiogalactopyranoside (IPTG). All experiments were performed in duplicate.
Project description:We report the transcriptional expression from wild type, ΔphoPQ, and ΔpmrAB to understand their contribution to colistin resistance.
Project description:This study aims to determine the epidemiology of Enterobacteriaceae resistant to antibiotics of last resort in pregnant women in labour at a tertiary hospital, Pretoria, South Africa. Rectal swabs shall be used to screen for colonisation with CRE and colistin-resistant Enterobacteriales in pregnant women during labour. Carbapenem and colistin-resistant Enterobacterales can cause the following infections: bacteraemia; nosocomial pneumonia; urinary tract infections, and intra-abdominal infections. Due to limited treatment options, infections caused by these multidrug-resistant organisms are associated with a mortality rate of 40-50%. Screening for colonisation of carbapenem-resistant Enterobacteriaceae (CRE) and colistin-resistant Enterobacteriaceae will help implement infection and prevention measures to limit the spread of these multidrug-resistant organisms.