American Journal of Infectious Diseases and Microbiology
ISSN (Print): 2328-4056 ISSN (Online): 2328-4064 Website: Editor-in-chief: Maysaa El Sayed Zaki
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American Journal of Infectious Diseases and Microbiology. 2017, 5(4), 126-131
DOI: 10.12691/ajidm-5-4-1
Open AccessArticle

Diagnostic Performance of Polymerase Chain Reaction Targeting Insertion Sequence (IS6110) for the Detection of Extra Pulmonary Tuberculosis

A. Chitra1, B. Usharani2, S. Smita1, C. K. Vidya Raj1, S. Anbazhagi3 and M. Muthuraj1,

1State TB Training and Demonstration Centre, Intermediate Reference Laboratory, Government Hospital for Chest Diseases, Puducherry, India

2Department of Biomedical Genetics, Institute of Basic Medical Sciences, University of Madras, Tamil Nadu, India

3Department of Environment Science, Central University, Kerala, India

Pub. Date: August 10, 2017

Cite this paper:
A. Chitra, B. Usharani, S. Smita, C. K. Vidya Raj, S. Anbazhagi and M. Muthuraj. Diagnostic Performance of Polymerase Chain Reaction Targeting Insertion Sequence (IS6110) for the Detection of Extra Pulmonary Tuberculosis. American Journal of Infectious Diseases and Microbiology. 2017; 5(4):126-131. doi: 10.12691/ajidm-5-4-1


Background: The diagnostic challenges in extra-pulmonary tuberculosis remain to be addressed even though remarkable progress has been made in the diagnostics of pulmonary tuberculosis during the last decade. Methods: This study was conducted to evaluate the use of polymerase chain reaction (PCR) in diagnosis of definitive extra pulmonary tuberculosis patients, and to assess the performance of insertion sequence (IS)6110 based PCR assay as compared to conventional liquid culture by Microbial growth Indicator Tube (MGIT) 960 system. Results: A total of 792 clinical specimens were collected from clinically suspected extra pulmonary tuberculosis patients. The specimens included 22 ascitic fluids, 69 pleural fluids, 240 Cerebrospinal fluids (CSF), 386 endometrial tissues, 47 lymph nodes, 22 pus, one synovial fluid, one fallopian tube, two brain abscess and two ovarian cyst samples. All these clinical samples were subjected to Auromine O staining (FM) for acid fast bacilli (AFB) and culture on MGIT 960 tubes containing Modified Middlebrooks 7H9 broth medium. PCR was performed by targeting 123bp fragment of insertion sequence IS6110 of Mycobacterium tuberculosis. Our study of 792 specimens, revealed their 87.5% sensitivity to endometrial samples, 92.31% sensitivity for cerebrospinal fluid, and 66.66% sensitivity in Pleural fluid and 60% sensitivity in Lymph node samples. The combined sensitivity and specificity of the PCR IS6110 was calculated to be 85.71% and 82.91%, respectively. Conclusion: PCR using IS6110 primer was able to pick up more positivity in extra pulmonary samples as compared to conventional culture method for the detection of M. tuberculosis.

polymerase chain reaction mycobacterium tuberculosis extra pulmonary tuberculosis insertion sequence cerebrospinal fluid

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[1]  Dye, C., Global epidemiology of tuberculosis. Lancet, 938-940. 2006.
[2]  S.P. Agarwal, L.S. Chauhan, Tuberculosis control in India. Directorate General of Health Services/Ministry of Health and Family Welfare, New Delhi, India. 2005.
[3]  Ellner, J. J, The emergence of extensively drug-resistant tuberculosis: a global health crisis requiring new interventions. Part II. Scientific advances that may provide solutions. Clin Transl Sci, 2. 80-84.2009.
[4]  Manju Purohit., Tehmina Mustafa, Laboratory Diagnosis of Extra-pulmonary Tuberculosis (EPTB) in Resource-constrained Setting: State of the Art, Challenges and the Need. J Clin Diag Res, 9. 1-6. 2015.
[5]  Makeshkumar, V., Radha Madhavan., Sujatha Narayanan, Polymerase chain reaction targeting insertion sequence for the diagnosis of extra pulmonary tuberculosis. Indian J Med Res, 139. (6)-166.2014.
[6]  Soni, H., Musser, J.M, Molecular diagnosis of mycobacteria. Clin Chem, 47. 809-14. 2001.
[7]  Hale, Y.M., Pfyffer, G.E., Salfinger, M,Laboratory diagnosis of mycobacterial infections: new tools and lessons learned. Clin Infect Dis, 33. 834-46. 2001.
[8]  Montenegro, S.H., Gilman, R.H., Sheen, P., Cama, R., Caviedes, L., Hopper, T, Improved detection of M. tuberculosis in Peruvian Children by use of hemi nested IS6110 PCR assay. Clin Infect Dis, 36 16-23. 2006.
[9]  Telenti, A., Honore, N., Brenasconi, C., March, J.O., Takiff, H.E., Cole, S.T. Genotyping assessment of isoniazid and rifampicin resistance in Mycobacterium tuberculosis: a blind study at reference laboratory level. J Clin Microbiol, 35.719-723.1997.
[10]  Usharani Brammacharry,Muthaiah Muthuraj,Diagnostic value of Polymerase Chain Reaction for the rapid diagnosis of clinically suspected tuberculous meningitis, Int J Pharm Bio Sci, 5. 70-77. 2014.
[11]  Muthuraj, M., Jagadeesan, S., Ayalusamy, N. Molecular epidemiological study of pyrazinamide resistance in clinical isolates of Mycobacterium tuberculosis from South India. Int J Mol Sci, 11. 2670-2680. 2011.
[12]  Fletcher, H.A., Donoghue, H.D., Taylor, G.M., Adri, G.M., Van der Zanden, A.G.M.,Spigelman, M.Molecular analysis of Mycobacterium tuberculosis DNA from a family of 18th century Hungarians. Microbiology, 149. 143-151.2005.
[13]  Tiwari, V., Jain, A., Verma, R.K.Amplification of enzyme amplified mycobacterial DNA detection in the diagnosis of pulmonary and extra pulmonary tuberculosis. Indian J Med Res, 118. 224-248. 2003.
[14]  Kaneko, K., Onodera, O., Miyatake, T., Tsuji, S.Rapid diagnosis of tuberculous meningitis by polymerase chain reaction (PCR). Neurology, 40. 1617-1618. 1990.
[15]  Eisenach, K.D., Cave, M.D., Bates, J.H., Crawford, J.T. Polymerase chain reaction amplification of a repetitive DNA sequence specific for Mycobacterium tuberculosis. J Infect Dis, 161. 977-981. 1990.
[16]  Negi, S.S., Anand, R., Pasha, S.T., Gupta, S., Blasir, S.F., Khare, S.Diagnostic potential of IS6110, 38 kda, 65 kda and 85B sequence based polymerase chain reaction in the diagnosis of Mycobacterium tuberculosis in clinical specimens. Indian J Med Microbio, 25. 43-9. 2007.
[17]  Ogusuk, M.M., Salem, J. I. Analysis of different primers used in the PCR method: diagnosis of tuberculosis in the state of Amazonas. Brazil J Bras Pnemo, 30. 343-9. 2004.
[18]  Narayanan, S., Parandaman, V., Narayanan, P.R., Venkatesan, P., Girish, C., Mahadevan, S. Evaluation of PCR using TRC (4) and IS6110 primers in detection of tuberculous meningitis. J Clin Microbiol, 39. 2006-2008. 2001.
[19]  Van Embden, J.D.A., Cave, M.D., Crawford, J.T., Dale, J.W., Eisenach, K.D., Gicquel, B.Strains identification of Mycobacterium tuberculosis by DNA fingerprinting: recommendations for standardized methodology. J Clin Microbiol, 31. 406-409. 1993.
[20]  Cole, S.T., Brosch, R., Parkhill, J., Garnier, T., Churcher, C., Harris, D. Deciphering the biology of Mycobacterium tuberculosis from the complete genome sequence. Nature, 393.537-544.1998.
[21]  Small, P.M., Hopewell, P.C., Singh, S.P., Paz, A., Parsonnet, J., Ruston, D.C. The epidemiology of tuberculosis in san Francisco. A population-based study using conventional and molecular methods. N Engl J Med, 330.1703-1709.1994.