96well-formatted CrfA assay for differentiation of Mycobacterium tuberculosis and Non-tuberculous mycobacteria

Failure in recognizing non-tuberculous mycobacteria (NTM) leads to misdiagnosis of multidrug-resistant Mycobacterium tuberculosis Complex (MTBC). There is an unmet need for diagnostic tools that can differentiate between NTMs and MTBC, and that are affordable for Low- and Middle-income Countries (LMIC). Earlier we developed a strip-based CrfA assay technology to detect the Carbapenem Resistance Factor A (CrfA) enzyme present only in MTBC. However, the strip-based CrfA assay had low turnaround time and lacked high-throughput capabilities. In this current research, we have developed a 96well-formatted CrfA assay for high-throughput detection of MTBC and differentiation with NTMs. This 96well-formatted CrfA assay displays a low turnaround time of 6-8 h with 100 % specificity and 93.75 % sensitivity on clinical samples. Based on these attributes, this 96well-formatted assay represents a valuable complementary tool to mitigate the misdiagnosis of chronic pulmonary tuberculosis with non-tuberculous mycobacteria in poorer nations.