Abstract
Background: Diagnosis of extra-pulmonary tuberculosis (EPTB) in smear-negative patients can be difficult. We assessed prospectively the performance of Real Time PCR in diagnosing smear-negative tuberculosis (TB), which represents the most common form of TB in a low incidence setting.
Methods: The present study was undertaken to find the effectiveness of Polymerase Chain Reaction targetting IS6110, MPB64 and 16SrRNA genes and culture on L–J media in Ziehl-Neelsen stain negative samples of body fluids for rapid diagnosis of suspected cases of Extra pulmonary tuberculosis.
Results: A total of 62 samples were taken of which 2 samples were not adequate in amount and hence were excluded. All 6 (10%) culture positive samples were PCR positive, in addition PCR could detect 5 out of 60 (8.3%) samples to be positive which were negative by culture. Of the 11 positive samples only MTB was reported by PCR and No NTM was detected.
Conclusions: PCR is a sensitive method for rapid diagnosis of TB compared to the conventional ZN staining. PCR can serve as a sensitive diagnostic method for microbiological diagnosis of smear-negative TB in countries with a low TB prevalence.
Keywords: extra-pulmoary, smear negative tuberculosis, sputum smear negative, tuberculosis
References
- Negi SS, Khan SF, Gupta S, Pasha S T, Khare S, Lal S. Comparison of the conventional diagnostic modalities, bactec culture and polymerase chain reaction test for diagnosis of tuberculosis. Indian J Med Microbiol 2005;23:29-33.
- Index TB Guidelines. Guidelines on extra-pulmonary tuberculosis for India. World Health Organization; 2016. Available from: http://www.icmr.nic.in/guidelines/TB/IndexTB%20Guidelines%20-%20green%20colour%202594164. Pdf accessed on January 3, 2018.
- World Health Organization. Global tuberculosis control. WHO report 2009; WHO/HTM/TB/2009.
- Rattan A. PCR for diagnosis of tuberculosis: Where are we now? Indian J Tuberc2000;47:79-82.
- Montenegro SH, Gilman RH, Sheen P, Cama R, Caviedes L, Hopper T, et al. Improved detection of Mycobacterium tuberculosis in Peruvian children by use of a heminested IS6110 polymerase chain reaction assay. Clin Infect Dis 2003; 36:16-2.
- Kulkarni S, Singh P, Memon A, Nataraj G, Kanade S, Kelkar R, Rajan M. An in-house multiplex PCR test for the detection of Mycobacterium tuberculosis, its validation & comparison with a single target TB-PCR kit, Indian J Med Res 2012;135:788-94.
- Parandaman, V., S. Narayanan, and P. R. Narayanan. Utility of polymerase chain reaction using two probes for rapid diagnosis of tubercular pleuritis in comparison to conventional methods. Indian J. Med. Res.2000;112:47-51.
- Chakravorty S, Sen MK, Tyagi JS. Diagnosis of Extrapulmonary Tuberculosis by Smear, Culture, and PCR Using Universal Sample Processing Technology. Journal of Clinical Microbiology. 2005;43(9):4357-62.
- Luca Norbis, Riccardo Alagna, Enrico Tortoli, Luigi Ruffo Codecasa, Giovanni Battista Migliori et al. Challenges and perspectives in the diagnosis of extrapulmonary tuberculosis,Expert Review of Anti-infective Therapy 2014;12:633-47.
- Sunnetcioglu A, Sunnetcioglu M, Binici I, Baran AI, Karahocagil MK, Saydan MR. Comparative analysis of pulmonary and extrapulmonary tuberculosis of 411 cases. Annals of Clinical Microbiology and Antimicrobials. 2015;14:34.
- Shrivastava G, Bajpai T, Bhatambare GS, Patel KB. Genital tuberculosis: Comparative study of the diagnostic modalities. J Hum ReprodSci 2014;7:30-3
- Eisenach, K. D., M. D. Cave, J. H. Bates, and J. T. Crawford. Polymerase chain reaction amplification of repetitive DNA sequence specific for Mycobacterium tuberculosis. J. Infect. Dis. 1990;161: 977-981.
- Chitra, B. Usharani, S. Smita, C. K. Vidya Raj, S. Anbazhagi, 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), 126-31.
- Amer S, Hefnawy AE, Wahab NA, Okasha H, Baz A.Evaluation of different laboratory methods for rapid diagnosis of tuberculous pleurisy.Int J Mycobacteriol.2016 ;5:437- 45.
- Siddiqui M, Anuradha PR, Nagamanik Vishnup H.Comparison of conventional diagnostic modalities,BACTEC culture with polymerase chain reaction for diagnosis of extra-pulmonary tuberculosis.J Med Allied Sci 2013;3:53-8.
- Lakshmi KR, Kumari VS, Vasundhara N, Suresh K. Detection of Extrapulmonary Tuberculosis from Various Samples in Sputum Smear Negative Patients. Int J Sci Stud 2016;3(10):63-6.
- Negi SS, Anand R, Pasha ST, Gupta S, Basir SF, Khare S, et al. Diagnostic potential of IS6110, 38kDa, 65kDa and 85B sequence-based polymerase chain reaction in the diagnosis of Mycobacterium tuberculosis in clinical samples. Indian J Med Microbiol. 2007;25:43–49.
- Kumar MV, Madhavan R, Narayanan S. Polymerase chain reaction targeting insertion sequence for the diagnosis of extrapulmonary tuberculosis. Indian J Med Res 2014;139:161-6
- Iqbal S, Ahmed R, Adhami SU, Mumtaz A. Importance of polymerase chain reaction in diagnosis of pulmonary and extra- pulmonary tuberculosis. Journal of Ayub Medical College, Abbottabad: JAMC. 2011;23(1):73–76.
- Sharma K, Appannanavar SB, Modi M, Singh M, Sharma A, Varma S. Role of multiplex polymerase chain reaction using IS6110 and protein b for the diagnosis of extra-pulmonary tuberculosis: North India. Indian J PatholMicrobiol 2015;58:27-30.
- KhaledGhaleb,MagdyAfifi, and Mohamad El-Gohary Assessment of Diagnostic Techniques of Urinary Tuberculosis Mediterr J Hematol Infect Dis. 2013; 5:1.
- Kaur J, Singh J, Mishra P. Comparative Evaluation of CFX96TM Real Time PCR with Conventional PCR for Rapid Diagnosis of Mycobacterium tuberculosis Complex in Clinical Isolates. Arch Clin Microbiol. 2016, 7:4.
- Ganavalli S. Ajantha, Praveen C. Shetty, Raghavendra D. Kulkarni, UmeshBiradar. PCR as a diagnostic tool for extra pulmonary tuberculosis.J Clin Diagn Res. 2013; 7: 1012–5.
- Tiwari V, Jain A, Verma RK. Applification of enzyme amplified mycobacterial DNA detection in the diagnosis of pulmonary and extrapulmonary tuberculosis. Indian J Med Res. 2003;118:224–8.
- Hemal AK, Gupta NP, Rajeev TP, Kumar R, Dar L, Seth P. Polymerase chain reaction in clinically suspected genito-urinary tuberculosis: Comparison with intravenous urography, bladder biopsy, and urine acid fast bacilli culture. Urology. 2000;56:570–74.
- Chakravorty S, Sen MK, Tyagi JS. Diagnosis of extrapulmonary tuberculosis by smear, culture and PCR using universal sample processing technology.J Clin Microbiol. 2005; 43: 4357-62.
- Narayanan S, Parandaman V, Narayanan PR, Venkatesan P, Girish C, Mahadevan S, Rajajee S. Evaluation of PCR Using TRC4 and IS6110 Primers in Detection of Tuberculous Meningitis. Journal of Clinical Microbiology. 2001;39:2006-8.
- Amin I, Idress M, Awan Z, Shahid M, Afzal S, Hussain A. PCR could be a method of choice for identification of both pulmonary and extrapulmonary tuberculosis. BMC Res Notes 2011; 4 : 332
- Iqbal S, Ahmed R, Adhami SU, Mumtaz A. Importance of polymerase chain reaction in diagnosis of pulmonary and extra-pulmonary tuberculosis. Journal of Ayub Medical College, Abbottabad: JAMC. 2011;23:73-6.
- Nagpal S, Chopra GS, Oberoi A, Singh N, Varghese SR. Conventional versus molecular methods for diagnosis of tuberculosis in a tertiary care center: A study from Punjab. CHRISMED J Health Res 2016;3:258-62.
- Hajia M, Rahbar M, Amini R. Is PCR assay reliable for diagnosis of extrapulmonary tuberculosis; Afr J Microbiol Res. 2009; 3(12):877-881.
- Beige J, Lokies J, Schaberg T, Finckh M, Fischer M. Mauch H, et al. Clinical evaluation of a Mycobacterium tuberculosis PCR assay. J ClinMicrobiol 1995; 33 : 90-5.
Corresponding Author
Dr Varun Goel
Department of Microbiology, Santosh Medical College & Hospital, Ghaziabad, UP., India
DOI: https://dx.doi.org/10.18535/jmscr/v6i5.60
