Detection of pyrazinamide-resistant Mycobacterium tuberculosis by PCR- SSCP of p nc A gene

Sun Shim Tae Sun Shim, Youngwhan Kim, Yong Chin Jae Yong Chin, C. M. Lim, Do Lee Sang Do Lee, Y. Koh, Sung Kim Woo Sung Kim, Soon Kim Dong Soon Kim, Dong Kim Won Dong Kim

Research output: Contribution to journalArticle

1 Citation (Scopus)

Abstract

Background: Recently the incidence of tuberculosis is increasing in many countries and control of the disease is further threatened by the emergence of multi-drug resistant tuberculosis. So rapid detection of drug. resistance is very important. Pyrazinamide (PZA) is a first-line chemotherapeutic agent for tuberculosis. Now in Korea, we perform PZase activity test instead of actual pyrazinamide susceptibility test for the detection of PZA resistant M. tuberculosis. Recently the pncA gene, encoding the PZase of M. tuberculosis, was completely sequenced. And it was reported that the mutation of pncA gene would be associated with PZA resistance of M. tuberculosis. Therefore we performed this study to evaluate the possibility for the rapid detection of PZA resistant M. tuberculosis using PCR-SSCP of pncA gene. Method: 44 cultured clinical isolates of M. tuberculosis, BCG Tokyo strain, BCG French strain, and one M bovis isolate were studied. We used H37Rv as the reference strain. The PZase activity test was done at the reference laboratory of Korean Tuberculosis Institute. DNA was extracted by bead-beater method and 561 bp fragment including pncA gene was amplified by PCR. The PCR product were digested by BstB I enzyme. SSCP was done using MDE gel. Of the 44 strains of M. tuberculosis, 22 strains were PZase-positive and other 22 strains were PZase negative. Results: Of the 22 PZase positive strains, 18 strains(82%) showed the same mobility compared with that of H37Rv and 4 (18%) showed different mobility. Of the 22 PZase-negative strains, 19 (86%) strains showed the same mobility pattern compared with that of H37Rv and 3(14%) showed different mobility. Naturally PZA-resistant BCG-French strain, BCG- Tokyo strain, and one M. bovis isolate showed the same band pattern each other, but their mobility were different from that of H37Rv. The results of PZase activity test and PCR-SSCP of pncA of M. tuberculosis were statistically significantly correlated each other (p<0.01). Conclusion: The PCR-SSCP after BstB I restriction of pncA gene of M. tuberculosis may be a useful method for the rapid detection of PZA-resistant M. tuberculosis.

Original languageEnglish
Pages (from-to)1178-1187
Number of pages10
JournalTuberculosis and Respiratory Diseases
Volume45
Issue number6
StatePublished - 1 Dec 1998

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Pyrazinamide
Single-Stranded Conformational Polymorphism
Mycobacterium tuberculosis
Polymerase Chain Reaction
Mycobacterium bovis
Genes
Tuberculosis
Tokyo
Multidrug-Resistant Tuberculosis
Korea
Gels
Mutation

Keywords

  • Drug resistance
  • PCR
  • Pyrazinamide
  • SSCP
  • Tuberculosis

Cite this

Tae Sun Shim, S. S., Kim, Y., Jae Yong Chin, Y. C., Lim, C. M., Sang Do Lee, D. L., Koh, Y., ... Won Dong Kim, D. K. (1998). Detection of pyrazinamide-resistant Mycobacterium tuberculosis by PCR- SSCP of p nc A gene. Tuberculosis and Respiratory Diseases, 45(6), 1178-1187.
Tae Sun Shim, Sun Shim ; Kim, Youngwhan ; Jae Yong Chin, Yong Chin ; Lim, C. M. ; Sang Do Lee, Do Lee ; Koh, Y. ; Woo Sung Kim, Sung Kim ; Dong Soon Kim, Soon Kim ; Won Dong Kim, Dong Kim. / Detection of pyrazinamide-resistant Mycobacterium tuberculosis by PCR- SSCP of p nc A gene. In: Tuberculosis and Respiratory Diseases. 1998 ; Vol. 45, No. 6. pp. 1178-1187.
@article{4812f529ee7d467d918dc8ecebdad116,
title = "Detection of pyrazinamide-resistant Mycobacterium tuberculosis by PCR- SSCP of p nc A gene",
abstract = "Background: Recently the incidence of tuberculosis is increasing in many countries and control of the disease is further threatened by the emergence of multi-drug resistant tuberculosis. So rapid detection of drug. resistance is very important. Pyrazinamide (PZA) is a first-line chemotherapeutic agent for tuberculosis. Now in Korea, we perform PZase activity test instead of actual pyrazinamide susceptibility test for the detection of PZA resistant M. tuberculosis. Recently the pncA gene, encoding the PZase of M. tuberculosis, was completely sequenced. And it was reported that the mutation of pncA gene would be associated with PZA resistance of M. tuberculosis. Therefore we performed this study to evaluate the possibility for the rapid detection of PZA resistant M. tuberculosis using PCR-SSCP of pncA gene. Method: 44 cultured clinical isolates of M. tuberculosis, BCG Tokyo strain, BCG French strain, and one M bovis isolate were studied. We used H37Rv as the reference strain. The PZase activity test was done at the reference laboratory of Korean Tuberculosis Institute. DNA was extracted by bead-beater method and 561 bp fragment including pncA gene was amplified by PCR. The PCR product were digested by BstB I enzyme. SSCP was done using MDE gel. Of the 44 strains of M. tuberculosis, 22 strains were PZase-positive and other 22 strains were PZase negative. Results: Of the 22 PZase positive strains, 18 strains(82{\%}) showed the same mobility compared with that of H37Rv and 4 (18{\%}) showed different mobility. Of the 22 PZase-negative strains, 19 (86{\%}) strains showed the same mobility pattern compared with that of H37Rv and 3(14{\%}) showed different mobility. Naturally PZA-resistant BCG-French strain, BCG- Tokyo strain, and one M. bovis isolate showed the same band pattern each other, but their mobility were different from that of H37Rv. The results of PZase activity test and PCR-SSCP of pncA of M. tuberculosis were statistically significantly correlated each other (p<0.01). Conclusion: The PCR-SSCP after BstB I restriction of pncA gene of M. tuberculosis may be a useful method for the rapid detection of PZA-resistant M. tuberculosis.",
keywords = "Drug resistance, PCR, Pyrazinamide, SSCP, Tuberculosis",
author = "{Tae Sun Shim}, {Sun Shim} and Youngwhan Kim and {Jae Yong Chin}, {Yong Chin} and Lim, {C. M.} and {Sang Do Lee}, {Do Lee} and Y. Koh and {Woo Sung Kim}, {Sung Kim} and {Dong Soon Kim}, {Soon Kim} and {Won Dong Kim}, {Dong Kim}",
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Tae Sun Shim, SS, Kim, Y, Jae Yong Chin, YC, Lim, CM, Sang Do Lee, DL, Koh, Y, Woo Sung Kim, SK, Dong Soon Kim, SK & Won Dong Kim, DK 1998, 'Detection of pyrazinamide-resistant Mycobacterium tuberculosis by PCR- SSCP of p nc A gene', Tuberculosis and Respiratory Diseases, vol. 45, no. 6, pp. 1178-1187.

Detection of pyrazinamide-resistant Mycobacterium tuberculosis by PCR- SSCP of p nc A gene. / Tae Sun Shim, Sun Shim; Kim, Youngwhan; Jae Yong Chin, Yong Chin; Lim, C. M.; Sang Do Lee, Do Lee; Koh, Y.; Woo Sung Kim, Sung Kim; Dong Soon Kim, Soon Kim; Won Dong Kim, Dong Kim.

In: Tuberculosis and Respiratory Diseases, Vol. 45, No. 6, 01.12.1998, p. 1178-1187.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Detection of pyrazinamide-resistant Mycobacterium tuberculosis by PCR- SSCP of p nc A gene

AU - Tae Sun Shim, Sun Shim

AU - Kim, Youngwhan

AU - Jae Yong Chin, Yong Chin

AU - Lim, C. M.

AU - Sang Do Lee, Do Lee

AU - Koh, Y.

AU - Woo Sung Kim, Sung Kim

AU - Dong Soon Kim, Soon Kim

AU - Won Dong Kim, Dong Kim

PY - 1998/12/1

Y1 - 1998/12/1

N2 - Background: Recently the incidence of tuberculosis is increasing in many countries and control of the disease is further threatened by the emergence of multi-drug resistant tuberculosis. So rapid detection of drug. resistance is very important. Pyrazinamide (PZA) is a first-line chemotherapeutic agent for tuberculosis. Now in Korea, we perform PZase activity test instead of actual pyrazinamide susceptibility test for the detection of PZA resistant M. tuberculosis. Recently the pncA gene, encoding the PZase of M. tuberculosis, was completely sequenced. And it was reported that the mutation of pncA gene would be associated with PZA resistance of M. tuberculosis. Therefore we performed this study to evaluate the possibility for the rapid detection of PZA resistant M. tuberculosis using PCR-SSCP of pncA gene. Method: 44 cultured clinical isolates of M. tuberculosis, BCG Tokyo strain, BCG French strain, and one M bovis isolate were studied. We used H37Rv as the reference strain. The PZase activity test was done at the reference laboratory of Korean Tuberculosis Institute. DNA was extracted by bead-beater method and 561 bp fragment including pncA gene was amplified by PCR. The PCR product were digested by BstB I enzyme. SSCP was done using MDE gel. Of the 44 strains of M. tuberculosis, 22 strains were PZase-positive and other 22 strains were PZase negative. Results: Of the 22 PZase positive strains, 18 strains(82%) showed the same mobility compared with that of H37Rv and 4 (18%) showed different mobility. Of the 22 PZase-negative strains, 19 (86%) strains showed the same mobility pattern compared with that of H37Rv and 3(14%) showed different mobility. Naturally PZA-resistant BCG-French strain, BCG- Tokyo strain, and one M. bovis isolate showed the same band pattern each other, but their mobility were different from that of H37Rv. The results of PZase activity test and PCR-SSCP of pncA of M. tuberculosis were statistically significantly correlated each other (p<0.01). Conclusion: The PCR-SSCP after BstB I restriction of pncA gene of M. tuberculosis may be a useful method for the rapid detection of PZA-resistant M. tuberculosis.

AB - Background: Recently the incidence of tuberculosis is increasing in many countries and control of the disease is further threatened by the emergence of multi-drug resistant tuberculosis. So rapid detection of drug. resistance is very important. Pyrazinamide (PZA) is a first-line chemotherapeutic agent for tuberculosis. Now in Korea, we perform PZase activity test instead of actual pyrazinamide susceptibility test for the detection of PZA resistant M. tuberculosis. Recently the pncA gene, encoding the PZase of M. tuberculosis, was completely sequenced. And it was reported that the mutation of pncA gene would be associated with PZA resistance of M. tuberculosis. Therefore we performed this study to evaluate the possibility for the rapid detection of PZA resistant M. tuberculosis using PCR-SSCP of pncA gene. Method: 44 cultured clinical isolates of M. tuberculosis, BCG Tokyo strain, BCG French strain, and one M bovis isolate were studied. We used H37Rv as the reference strain. The PZase activity test was done at the reference laboratory of Korean Tuberculosis Institute. DNA was extracted by bead-beater method and 561 bp fragment including pncA gene was amplified by PCR. The PCR product were digested by BstB I enzyme. SSCP was done using MDE gel. Of the 44 strains of M. tuberculosis, 22 strains were PZase-positive and other 22 strains were PZase negative. Results: Of the 22 PZase positive strains, 18 strains(82%) showed the same mobility compared with that of H37Rv and 4 (18%) showed different mobility. Of the 22 PZase-negative strains, 19 (86%) strains showed the same mobility pattern compared with that of H37Rv and 3(14%) showed different mobility. Naturally PZA-resistant BCG-French strain, BCG- Tokyo strain, and one M. bovis isolate showed the same band pattern each other, but their mobility were different from that of H37Rv. The results of PZase activity test and PCR-SSCP of pncA of M. tuberculosis were statistically significantly correlated each other (p<0.01). Conclusion: The PCR-SSCP after BstB I restriction of pncA gene of M. tuberculosis may be a useful method for the rapid detection of PZA-resistant M. tuberculosis.

KW - Drug resistance

KW - PCR

KW - Pyrazinamide

KW - SSCP

KW - Tuberculosis

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VL - 45

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JO - Tuberculosis and Respiratory Diseases

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SN - 1738-3536

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Tae Sun Shim SS, Kim Y, Jae Yong Chin YC, Lim CM, Sang Do Lee DL, Koh Y et al. Detection of pyrazinamide-resistant Mycobacterium tuberculosis by PCR- SSCP of p nc A gene. Tuberculosis and Respiratory Diseases. 1998 Dec 1;45(6):1178-1187.