Objective To compare the expression levels of pncA gene in pyrazinamide （PZA）-resistant and multidrug-resistant Mycobacterium tuberculosis before and after PZA drug stimulation， and to explore the relationship between PZA-resistant Mycobacterium tuberculosis and pncA gene expression. Methods PZA-resistant and multidrug-resistant Mycobacterium tuberculosis was cultured， and the minimal inhibitory concentration （MIC） of rifampicin and PZA was detected. The strains were cultured in medium containing 0 and 50 μg/mL PZA， and the expression level of pncA was detected by real-time fluorescence quantitative PCR. Results The MIC of PZA of 21 strains of Mycobacterium tuberculosis was ≥ 1 600 μg/mL，400-800 μg/mL for 12 strains， and 100-200 μg/mL in 5 strains，and their corresponding PZA resistance levels were defined as high，medium and low，respectively. The median MIC of rifampicin was 128， 128 and 64μg/mL， respectively. Chi-square test showed that there was no significant difference between any two groups （all P > 0.05）. According to the relative quantitative 2-ΔΔCt method， the expression levels of pncA in the high，medium and low levels of PZA drug-resistant Mycobacterium tuberculosis was 0.904（0.202， 2.653）， 1.157（0.658， 1.511） and 1.147（0.372， 1.347）， and 1.544（0.611， 3.191）， 0.846（0.421， 1.237） and 0.726 （0.225， 3.017） in the 50 μg/mL treatment group，respectively. There was no significant difference in pncA gene expression between the control and PZA drug-containing culture groups（P > 0.05）. Conclusion No significant changes are observed in the expression levels of pncA in multidrug-resistant and PZA-resistant Mycobacterium tuberculosis before and after PZA drug stimulation.