Clinical Evaluation of an Oligonucleotide Array for Detection of First- and Second-Line Drug Resistance of Mycobacterium tuberculosis Complex in a Northern Taiwan Medical Center (I)

Tuberculosis (TB) is caused by Mycobacterium tuberculosis complex (MTBC). World Health Organization estimated that there were 900 million new TB cases and 480,000 new multidrug-resistant TB cases in 2013. TB is the second leading cause of death among infectious diseases. The turn-around time for identification and drug susceptibility testing of M. tuberculosis is at least four weeks. A rapid assay method is urgently needed for promopt and proper TB treatment and infectious control. Drug-resistant TB is associated with mutations in several genes, including the rpoB gene for rifampin (RIF), the katG gene and the inhA regulatory region for isoniazid (INH), the embB gene for ethambutol (EMB), the rpsL and rrs genes for streptomycin (SM), g^rA and gyrB for ofloxacin (OFX), rrs for kanamycin (KM) and amikacin (AN), the tylA and rrs genes for caperomycin (CM). A prototype array-based assay using digoxin labeled primers to PCR-amplified target genes and consequent hybridization with a nylon membrane was developed. Results of primerary evaluation of the prototype chip using 1,426 MGIT-positive culture showed that sensitivities were 91.7% (RIF), 88.9% (INH), 100% (EMB), 82.4% (SM), 100% (OFX), 100% (KM) and 100% (AN); specificity were: 98.9% (RIF), 99.4% (INH), 99.5% (EMB), 100% (SM), 100% (OFX), 100% (KM) and 99.5% (AN); PPV were: 91.7% (RIF), 97.0% (INH), 92.9% (EMB), 100% (SM), 100% (OFX), 100% (KM) and 66.7 % (AN); NPV were: 98.9% (RIF), 97.7% (INH), 100% (EMB), 98.4% (SM), 100% (OFX), 100% (KM) and 100% (AN), respectively. The objectives ot the study are to conduct a pre-clinical trial and to finalize the formate of an array for the clinical trail in the first year; to conduct a multicenter clinical trail using a commercial kit including array and reader in the second year. Subproject 1 will use the array to test a chieve of multidrug-resistant and extensively drug-resistant M. tuberculosis clinical isolates in Taiwan Centers for Disease Control. Subprojects 2 and 3 will use the array to detect clinical isolates and MGIT- positive culture. Finally, the overall performances of the array for detecting the first and second-line drug-resistance of M. tuberculosis will be evaluated. The goal to this project is to develop a new diagnostics for better control of drug-resistant TB.