Comparison of Phenotypic and Genotypic methods for Pyrazinamide susceptibility testing.
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Date
2009-04
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Abstract
Objectives: To evaluate Pyrazinamide (PZA) susceptibility results obtained by phenotypic MGIT 960 TB system
against enzymatic Pyrazinamidase assay and genotypic pncA gene sequencing. To find the prevalence of infections
caused by M. bovis in PZA resistant M. tuberculosis complex isolates.
Methods: 33 consecutive PZA resistant and 30 consecutive PZA susceptible isolates reported for PZA susceptibility
testing by MGIT 960 TB system were included in this study. Presence of active pyrazinamidase enzyme was sought
by using the Wayne assay. The pncA gene was amplified by PCR and then sequenced to screen mutations. All the PZA
resistant isolates were further spoligotyped to identify M. bovis, if present.
Results: Of 33 PZA resistant strains by MGIT 960, 31 were Wayne assay negative and two were positive. Of the 30
susceptible PZA strains six were Wayne assay negative reporting false resistance. PncA gene sequencing revealed that 32 of
the 33 MGIT PZA resistant isolates had diverse nucleotide changes scattered throughout the pncA gene (one isolate did not
show any mutation). Of the 30 phenotypically susceptible isolates, 21 were wild types whilst nine isolates showed the
presence of a silent mutation C-T at codon 195. Fifteen mutations found in this study has not been described earlier. Not
a single isolate of M. bovis was detected among PZA resistant M. tuberculosis complex isolates.
Conclusion: MGIT 960 showed better concordance with sequencing results in comparison with Wayne assay. In
present study, a high proportion (85%) of MDR-TB isolates from patients receiving anti-TB treatment were found to be resistant to PZA.
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Keywords
Pyrazinamide susceptibility, M. tuberculosis, pncA gene mutations
Citation
Shenai Shubhada, Rodrigues Camilla, Sadani Meeta, Sukhadia Nesline, Mehta Ajita. Comparison of Phenotypic and Genotypic methods for Pyrazinamide susceptibility testing. Indian Journal of Tuberculosis. 2009 Apr; 56(2): 82-90.