engleski

Rapid determination of antibiotic resistance mechanisms of pathogenic bacteria by CAF-/CAF+ technique

Over the past several decades, the medical community has become increasingly concerned over the ability of bacteria to develop resistance to antibacterial drugs. Many bacteria have become resistant to multiple classes of antibiotics, especially in hospitals. Determination of antibiotic resistance mechanisms of pathogenic bacteria can provide information that will help in infection control and surveillance of bacterial resistance to antibiotics. In this study we demonstrated novel, rapid and accurate method for determination of antibiotic resistance mechanisms of pathogenic bacteria. The method is based on proteome analysis exclusively driven by peptide/protein de novo sequencing. The method is based on chemically activated fragmentation in negative and positive ion mode (CAF-/CAF+), a newly developed technique that enables tandem mass spectrometry acquisition (MS/MS) from both positive and negative ion modes. Peptides identified as overlapped amino acid sequences read from both ion modes could unambiguously assign peptide after de novo sequencing and protein database identification. Using this method, the key proteins responsible for the antibiotic resistance of pathogenic bacteria such as clinical Pseudomonas aeruginosa aeruginosa, uropathogenic Escherichia coli and Proteus mirabilis mirabilis were identified. Induction of AmpC enzymes was found in three P. aeruginosa strains resistant to meropenem. Sensitivity of CAF-/CAF+ method enabled even determination of particular mutations between the AmpC enzymes in tested strains. The interaction of various proteins such as β-lactamase and outer membrane protein A was found as mechanism of penicillin G resistance in E. coli MFBF10519. Aforementioned method enabled the determination of differentially expressed proteins (antibiotic susceptibility vs. antibiotic resistance) responsible for specific types of antimicrobial resistivity and is presented as a promising de novo sequencing tool.

antibiotics resistance; bacteria; CAF-/CAF+ technique

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