Project description:Antibiotic resistance associated with the expression of the clinically significant carbapenemases, IMP, KPC, and NDM and OXA-48 in Enterobacteriaceae is emerging as a worldwide calamity to health care. In Australia, IMP-producing Enterobacteriaceae is the most prevalent carbapenemase-producing Enterobacteriaceae (CPE). Genomic characteristics of such carbapenemase-producing Enterobacteriaceae (CPE) are well described, but the corresponding proteome is poorly characterised. We have thus developed a method to analyse dynamic changes in the proteome of CPE under antibiotic pressure. Specifically, we have investigated the effect of meropenem at sub-lethal concentrations to develop a better understanding of how antibiotic pressure leads to resistance. Escherichia coli, producing either NDM, IMP or KPC type carbapenemase were included in this study, and their proteomes were analysed in growth conditions with or without meropenem.
Project description:New and rapid antimicrobial susceptibility/resistance testing methods are required for bacteria from positive blood cultures. In the current study we developed and evaluated a targeted LC-MS/MS assay for the detection of beta-lactam, aminoglycoside and fluoroquinolone resistance mechanisms in blood cultures positive for E. coli or K. pneumoniae. Selected targets were the beta-lactamases SHV, TEM, OXA-1-like, CTX-M-1-like, CMY-2-like, chromosomal E. coli AmpC, OXA-48-like, NDM, VIM and KPC, the aminoglycoside modifying enzymes AAC(3)-Ia, AAC(3)-II, AAC(3)-IV, AAC(3)-VI, AAC(6’)-Ib, ANT(2”)-I and APH(3’)-VI, the 16S-RMTases ArmA, RmtB, RmtC and RmtF, the quinolone resistance mechanisms QnrA, QnrB, AAC(6’)-Ib-cr, the wildtype QRDR of GyrA, and for E. coli, the porins OmpC and OmpF. The developed assay was evaluated using 100 prospectively collected positive blood cultures, 100 negative blood cultures inoculated with isolates that were previously collected from blood cultures, and 48 isolates inoculated with isolates carrying genes of less prevalent resistance mechanisms.
