Comparison of PCR-RFLP and biochemical methods for detection of human pulmonary tuberculosis agents

Subject Areas : Molecular Microbiology

Behnam Rafiee ¹ (Department of Microbiology, Qom Branch, Islamic Azad University, Qom, Iran)
Ali asghar Farazi ² (Department of Infectious and Tropical Diseases, Arak University of Medical Sciences, Arak, Iran)
Davoud Sadeghi ³ (Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, Iran)
Sepideh Ghani ⁴ (Young Researcher’s Club, Arak Branch, Islamic Azad University, Arak, Iran)
Razieh Nazari ⁵ (Department of Microbiology, Qom Branch, Islamic Azad University, Qom, Iran)
Rohulah Keshavarz ⁶ (Razi Vaccine & Serum Research Institute, Karaj, Iran)
Keyvan Tadayon ⁷ (Razi Vaccine & Serum Research Institute, Karaj, Iran)
Nader Msavari ⁸ (Razi Vaccine & Serum Research Institute, Karaj, Iran)

Received: 2010-05-15 Accepted : 2010-08-16 Published : 2010-07-01

Keywords: PCR-RFLP, Mycobacterium tuberculosis complex, oxyR pseudogene,

Abstract :

Background and Objective: Tuberculosis is one of important cause of death in several countries. Mycobacterium tuberculosis complex is consisted of homogenous slow growing mycobacteria that their isolation and identification are difficult and time consuming. The objective of this study was to evaluate a molecular method for differentiation of Mycobacterium tuberculosis from Mycobacterium bovis, and at the same time, to compare the method with biochemical tests. Material and methods: In this study 68 sputum specimens were collected from the smear positive patients who hospitalized in health centers of Markazi province. The samples were cultivated in the specialized cultures. Also, 10 isolates of Mycobacterium bovis that were obtained from different area of Iran as well as 6 standard strains have been used for comparison. After performance of biochemical tests, oxyR pseudogene was amplified by PCR and the PCR products were digested by AluI endonuclease. Results:  The digestion on PCR products of Mycobacterium tuberculosis and Mycobacterium bovis showed one and three fragment panel, respectively, on the gele. Only one isolate of human Mycobacterium bovis strain has been identified. The results of PCR-RFLP were consistent with biochemical tests. Conclusion: Since PCR-RFLP is a rapid and accurate method for differentiation between Mycobacterium bovis and other Mycobacterium tuberculosis complex and the results are important in terms of diagnosis and treatment of tuberculosis.

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