Abstract:

Objective Evaluation of multicolor melting curve analysis (MMCA) in the detection of mycobacterium tuberculosis resistance in sputum Samples.Methods 235 morning sputum specimen, which were concentrated acid-fast bacterium-positive and/or M.tuberculosis molecular test positive, were collected. Then, the MMCA drug-resistant detection, mycobacterium fluid culture and proportion drug-susceptibility testing (DST) were conducted simultaneously. The results of the MMCA and DST were compared using Kappa test. Sanger sequencing was conducted for those samples that had discordant results between MMCA and DST.Results Concordance rates between MMCA and DST for rifampicin (RFP), isoniazid (INH), ethambutol (EMB), levofloxacin (Lfx), moxifloxacin (Mfx), and multidrug-resistant tuberculosis (MDR-TB) were 94.04%(221/235), 90.63% (213/235), 80.00% (188/235), 94.89% (223/235), 77.87% (183/235), and 89.36% (210/235), respectively. The Kappa value between these two methods for RFP, INH, Lfx and MDR-TB were 0.88, 0.81, 0.88, and 0.77, respectively, indicating an excellent consistency. The discordant samples were verified by Sanger sequencing. The total coincidence rates of sequencing results and MMCA for sputum and culture isolates were 89.80%(132/147) and 98.64%(145/147), respectively.Conclusion Direct and rapid detection of drug resistance of mycobacterium tuberculosis in sputum samples by MMCA has good concordance rates with DST or Sanger Gene sequencing. MMCA is easy to operate, rapid and accurate, and it will play an important role in the diagnosis of tuberculosis and rapid screening for drug-resistant tuberculosis.

Key words: Tuberculosis, multidrug-resistant, Fluoroimmunoassay, Molecular probe techniques, drug-susceptibility testing, Comparative study