Diagnostic Performance of Multiplex Allele-specific Polymerase Chain Reaction for Isoniazid Resistance in Mycobacterium tuberculosis : Comparison with Phenotypic Mycobacteria Growth Indicator Tube Drug-susceptibility Testing.

BACKGROUND

Rapid detection of isoniazid (INH) resistance remains important for tuberculosis management, although targeted molecular assays may not capture the full genetic heterogeneity underlying phenotypic resistance. This study evaluated multiplex allele-specific polymerase chain reaction (MAS-PCR) for detecting INH resistance-associated molecular patterns in Mycobacterium tuberculosis .

METHODS

This cross-sectional diagnostic accuracy study included 100 sputum specimens, comprising 70 acid-fast bacilli smear-positive and 30 smear-negative specimens. Phenotypic INH susceptibility was determined using Mycobacteria Growth Indicator Tube drug-susceptibility testing (DST-MGIT). MAS-PCR interrogated resistance-associated regions, with particular emphasis on katG315 and ahpC. Diagnostic performance and agreement were assessed in the primary evaluable population of 64 smear-positive specimens with paired MAS-PCR and DST-MGIT results.

RESULTS

Among the 64 evaluable specimens, MAS-PCR yielded 5 true-positive, 3 false-negative, 0 false-positive, and 56 true-negative results. Sensitivity was 62.5% (95% confidence interval [CI]: 24.5%-91.5%), specificity of 100.0% (95% CI: 93.6%-100.0%), positive predictive value of 100.0% (95% CI: 47.8%-100.0%), negative predictive value of 94.9% (95% CI: 85.9%-98.9%), and accuracy of 95.3% (95% CI: 86.9%-99.0%). Overall agreement was 95.3%, with substantial agreement beyond chance (Cohen's κ≈0.745). The exact McNemar's test showed no evidence of systematic directional discordance ( P = 0.250). The targeted katG315 mutation-associated pattern occurred exclusively among phenotypically resistant specimens, whereas no targeted ahpC pattern was detected.

CONCLUSION

MAS-PCR demonstrated high specificity and substantial agreement with DST-MGIT among evaluable smear-positive specimens but incomplete sensitivity for phenotypic INH resistance. These findings support its use as a complementary targeted molecular approach rather than a replacement for phenotypic DST.