Identification of Clinically Important Microbes by Genomic and Proteomic Methods

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Abstract

Following a quantitative blood-based assay detecting M. tuberculosis-specific peptides in digested serum samples, we have identified target peptides for the detection of important nontuberculous mycobacterial species. In addition, a blood-based molecular assay was developed for detection of M. avium complex (MAC) using CRISPR methodology. We unveiled two mechanisms of resistance responsible for the progressive development of extremely high-level imipenem resistance in M. abscessus: i) A partial deletion in the porin gene cooccurred with a first fourfold increase in MIC. ii) An increase in transcription level and enzymatic activity of M. abscessus β-lactamase accounted for the subsequent eightfold increase in MIC leading to a high-resistance phenotype. We developed a quantitative droplet digital PCR (ddPCR) scheme for rapid detection of clarithromycin resistance discriminating between full-length or truncated (inactive) erm(41) inducible resistance in M. abscessus and a probe based ddPCR screening assay for assessment of 23S rRNA rrl mutational resistance in M. abscessus and MAC.