Developing a Transcriptomic Assay for Rapid Antimicrobial Susceptibility Testing of Mycobacterium abscessus in Clinical Microbiology Laboratories

About This Grant

PROJECT SUMMARY AND ABSTRACT Nontuberculous mycobacteria (NTM) cause chronic pulmonary infections in the elderly and those with immunosuppression or pre-existing lung disease. NTMs are intrinsically resistant to most classes of antibiotics, and patients require treatment with two or more antibiotics for at least 12 months. While phenotypic and genotypic AST antimicrobial susceptibility testing (AST) performed in clinical microbiology laboratories is used by physicians to select antibiotics, its many limitations result in delays and uncertainty in the treatment of NTM infections. Improved AST methods for NTMs are essential for advancing outcomes from these infections, whose burden in the US now exceeds that of tuberculosis. Transcriptomic AST is an exciting new approach that shows great promise in solving gaps in NTM AST. This K08 award proposes to develop rapid transcriptomic AST for identifying susceptibility to clarithromycin and imipenem for the effective treatment of Mycobacterium abscessus. Both aims will use a combination of RNA-Seq and the NanoString platform to develop and validate a 10-gene transcriptomic AST assay that uses multiplexed RNA quantification directly from specimens without the need for RNA amplification of purification. Clarithromycin resistance is mediated by multiple resistance mechanisms and requires a combination of genotypic and phenotypic AST that can take weeks to return. Aim 1 will develop a rapid transcriptomic AST method that detects inducible and acquired clarithromycin resistance in a single assay. The breakdown of imipenem (half-life 0.6 days) during phenotypic AST (3-5 days) can lead to false-positive AST results showing imipenem resistance and result in physicians not prescribing an effective antibiotic. Aim 2 will determine the true imipenem susceptibility of clinical isolates using time-kill assays and then develop transcriptomic AST to accurately predict imipenem susceptibility without false imipenem resistance due to antibiotic degradation. By developing transcriptomic AST for clarithromycin and imipenem, this project will allow the rapid and reliable selection of antibiotics critical for the effective treatment of Mycobacterium abscessus. Dr. Poonawala is a physician-scientist trained in infectious diseases and clinical microbiology dedicated to improving the diagnosis of mycobacterial infections. Dr. Poonawala will be mentored by Dr. Bree Aldridge (rational design of tuberculosis (TB) treatment combinations using quantitative measurements of antibiotic activity) and Dr. Charles L. Daley (evaluation of novel diagnostics and treatments for TB and NTMs). Dr. Rebecca Batorsky (bioinformatics), Dr. Reeti Khare (clinical microbiology of mycobacteria), and Dr. Linden T. Hu (diagnostic development and career advice) will form his scientific advisory committee. In developing transcriptomic AST to address gaps in the AST of Mycobacterium abscessus, Dr. Poonawala will gain skills in molecular diagnostic development, bioinformatics, and advanced antimicrobial susceptibility testing. Upon completion of this K08, Dr. Poonawala will be ready for an independent career in developing mycobacterial diagnostics.