Establishment and evaluation of rapid detection of pyrazinamide resistance of Mycobacterium tuberculosis by probe enzyme digestion using Surveyor

doi:10.3969/j.issn.1000-6621.2021.08.011

Abstract

Abstract:

Objective To establish a probe enzyme digestion method using Surveyor for rapid detection of pyrazinamide (PZA) resistance detection of Mycobacterium tuberculosis (MTB) and evaluate the clinical value. Methods A total of 91 multidrug-resistant (MDR) MTB strains from Wuhan Pulmonary Hospital were collected. All strains were amplified using the above primers∶the downstream primer with the ratio of 2∶5, and the standard strain H37Rv of MTB was amplified using the upstream primer: the downstream primer with the ratio of 5∶2. Then, the PCR products were mixed in a ratio of 1∶1, and the electrophoresis was carried out after being digested by the endonuclease Surveyor enzyme. The gene mutation of pncA gene and its promoter sequence were determined based that the hybridization bands could be cleaved by the endonuclease Surveyor. The application value of endonuclease Surveyor digestion method was evaluated compared to BACTEC MGIT 960 method used to detect PZA resistance and pncA gene and its promoter gene sequencing method. Results For the detection of PZA resistance of MTB, compared to the BACTEC MGIT 960 method, the sensitivity, specificity and accuracy of the probe enzyme digestion method were 100.0% (29/29), 87.1% (54/62) and 91.2% (83/91), respectively. Compared to the sequencing results of pncA gene and its promoter region, the sensitivity, specificity and accuracy of the probe enzyme digestion method were 100.0% (37/37),100.0% (54/54) and 100.0% (91/91), respectively. Conclusion The probe enzyme digestion method demonstrated very high concordance with pncA gene and its promoter region, it might provide a new way for the detection of PZA resistance of MTB in areas without sequencing conditions.

Key words: Surveyor nuclease, Mycobacterium tuberculosis, Drug resistance, Pyrazinamide

HU Yan-jie, GONG Shi-wei, REN Yi. Establishment and evaluation of rapid detection of pyrazinamide resistance of Mycobacterium tuberculosis by probe enzyme digestion using Surveyor[J]. Chinese Journal of Antituberculosis, 2021, 43(8): 813-816. doi: 10.3969/j.issn.1000-6621.2021.08.011

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