Biomedical Science Letters (대한의생명과학회지)

The Korean Society for Biomedical Laboratory Sciences (대한의생명과학회)
권호 표지

Performance of Quantitative Real-Time PCR for Detection of Tuberculosis in Granulomatous Lymphadenitis Using Formalin-Fixed Paraffin-Embedded Tissue

  • Munkhdelger, Jijgee (Department of Pathology, Wonju College of Medicine, Yonsei University) ;
  • Mia-Jan, Khalilullah (Department of Pathology, Wonju College of Medicine, Yonsei University) ;
  • Lee, Dongsup (Department of Clinical Laboratory Science, Hyejeon College) ;
  • Park, Sangjung (Department of Biomedical Laboratory Science, College of Health Sciences, Yonsei University) ;
  • Kim, Sunghyun (Department of Biomedical Laboratory Science, College of Health Sciences, Yonsei University) ;
  • Choi, Yeonim (Department of Biomedical Laboratory Science, College of Health Sciences, Yonsei University) ;
  • Wang, Hye-Young (M&D, Inc., Wonju Eco Environmental Technology Center) ;
  • Jeon, Bo-Young (Department of Biomedical Laboratory Science, College of Health Sciences, Yonsei University) ;
  • Lee, Hyeyoung (Department of Biomedical Laboratory Science, College of Health Sciences, Yonsei University) ;
  • Park, Kwang Hwa (Department of Pathology, Wonju College of Medicine, Yonsei University)
  • Received : 2013.03.13
  • Accepted : 2013.04.25
  • Published : 2013.06.29
Download PDF
⟨ Previous Next ⟩

Abstract

Although culture is the gold standard method to identify mycobacteria, its use in tuberculous lymphadenitis (TBL) is limited due to formalin fixation of the submitted specimens. We evaluated the performance of quantitative real-time PCR (q-PCR) for Mycobacterium Tuberculosis (MTB) in granulomatous lymphadenitis using formalin-fixed paraffin-embedded (FFPE) tissues. From 2000 to 2010, a total number of 117 cases of lymph node samples with granulomatous inflammation which were surgically removed and fixed in formalin were studied. Hematoxylin & Eosin (H&E) and Ziehl-Neelsen-stained (ZN) slides were reviewed. qPCR using Real TB-Taq$^{(R)}$ was performed for all cases to identify Mycobacterium tuberculosis. Thirteen non-tuberculous lymphadenopathy cases were used as negative control. Cervical lymph nodes were more frequently affected (60%, 70/117) than other sites. ZN stain for acid fast bacilli was positive in 19 (16.24%) cases. qPCR for tuberculosis was positive in 92 (78.63%) cases. Caseous necrosis was found in 103 (88.03%) cases. While the ZN stain and qPCR were both negative in all control cases, the qPCR showed a significantly higher positive rate (78.63% vs. 16.24%) compared to ZN stain in histologically diagnosed TBL. Quantitative real-time PCR proves to be more sensitive than ZN stain for diagnosis of tuberculous lymphadenitis.

Keywords

References

  1. Ahmed HG, Nassar AS, Ginawi I. Screening for tuberculosis and its histological pattern in patients with enlarged lymph node. Patholog Res Int. 2011. 2011: 417635.
  2. Bruijnesteijn Van Coppenraet ES, Lindeboom JA, Prins JM, Peeters MF, Claas EC, Kuijper EJ. Real-time PCR assay using fine-needle aspirates and tissue biopsy specimens for rapid diagnosis of mycobacterial lymphadenitis in children. J Clin Microbiol. 2004. 42: 2644-2650. https://doi.org/10.1128/JCM.42.6.2644-2650.2004
  3. Causse M, Ruiz P, Gutierrez-Aroca JB, Casal M. Comparison of two molecular methods for rapid diagnosis of extrapulmonary tuberculosis. J Clin Microbiol. 2011. 49: 3065-3067. https://doi.org/10.1128/JCM.00491-11
  4. Daley P, Thomas S, Pai M. Nucleic acid amplification tests for the diagnosis of tuberculous lymphadenitis: a systematic review. Int J Tuberc Lung Dis. 2007. 11: 1166-1176.
  5. Greco S, Girardi E, Navarra A, Saltini C. Current evidence on diagnostic accuracy of commercially based nucleic acid amplification tests for the diagnosis of pulmonary tuberculosis. Thorax. 2006. 61: 783-790. https://doi.org/10.1136/thx.2005.054908
  6. Huang CT, Tsai YJ, Shu CC, Lei YC, Wang JY, Yu CJ, Lee LN, Yang PC. Clinical significance of isolation of nontuberculous mycobacteria in pulmonary tuberculosis patients. Respir Med. 2009. 103: 1484-1491. https://doi.org/10.1016/j.rmed.2009.04.017
  7. Johansen IS, Thomsen VO, Forsgren A, Hansen BF, Lundgren B. Detection of Mycobacterium tuberculosis complex in formalinfixed, paraffin-embedded tissue specimens with necrotizing granulomatous inflammation by strand displacement amplification. J Mol Diagn. 2004. 6: 231-236. https://doi.org/10.1016/S1525-1578(10)60515-6
  8. Kerleguer A, Fabre M, Bernatas JJ, Gerome P, Nicand E, Herve V, Koeck JL. Clinical evaluation of the gen-probe amplified mycobacterium tuberculosis direct test for rapid diagnosis of tuberculosis lymphadenitis. J Clin Microbiol. 2004. 42: 5921-5922. https://doi.org/10.1128/JCM.42.12.5921-5922.2004
  9. Kwon KS, Oh CK, Jang HS, Lee CW, Jun ES. Detection of mycobacterial DNA in cervical granulomatous lymphadenopathy from formalin-fixed, paraffin-embedded tissue by PCR. J Dermatol. 2000. 27: 355-360. https://doi.org/10.1111/j.1346-8138.2000.tb02184.x
  10. Lee HS, Park KU, Park JO, Chang HE, Song J, Choe G. Rapid, sensitive, and specific detection of Mycobacterium tuberculosis complex by real-time PCR on paraffin-embedded human tissues. J Mol Diagn. 2011. 13: 390-394. https://doi.org/10.1016/j.jmoldx.2011.02.004
  11. Linasmita P, Srisangkaew S, Wongsuk T, Bhongmakapat T, Watcharananan SP. Evaluation of real-time polymerase chain reaction for detection of the 16S ribosomal RNA gene of Mycobacterium tuberculosis and the diagnosis of cervical tuberculous lymphadenitis in a country with a high tuberculosis incidence. Clin Infect Dis. 2012. 55: 313-321. https://doi.org/10.1093/cid/cis401
  12. Osores F, Nolasco O, Verdonck K, Arevalo J, Ferrufino JC, Agapito J, Huayanay L, Gotuzzo E, Maguina C. Clinical evaluation of a 16S ribosomal RNA polymerase chain reaction test for the diagnosis of lymph node tuberculosis. Clin Infect Dis. 2006. 43: 855-859. https://doi.org/10.1086/507536
  13. Park DY, Kim JY, Choi KU, Lee JS, Lee CH, Sol MY, Suh KS. Comparison of polymerase chain reaction with histopathologic features for diagnosis of tuberculosis in formalin-fixed, paraffin-embedded histologic specimens. Arch Pathol Lab Med. 2003. 127: 326-330.
  14. Polesky A, Grove W, Bhatia G. Peripheral tuberculous lymphadenitis: epidemiology, diagnosis, treatment, and outcome. Medicine (Baltimore). 2005. 84: 350-362. https://doi.org/10.1097/01.md.0000189090.52626.7a
  15. Rimek D, Tyagi S, Kappe R. Performance of an IS6110-based PCR assay and the Cobas Amplicor MTB PCR system for detection of Mycobacterium tuberculosis complex DNA in human lymph node samples. J Clin Microbiol. 2002. 40: 3089-3092. https://doi.org/10.1128/JCM.40.8.3089-3092.2002
  16. Schulz S, Cabras AD, Kremer M, Weirich G, Miethke T, Bosmuller HC, Hofler H, Werner M, Fend F. Species identification of mycobacteria in paraffin-embedded tissues: frequent detection of nontuberculous mycobacteria. Mod Pathol. 2005. 18: 274-282. https://doi.org/10.1038/modpathol.3800289
  17. Selva E, Hofman V, Berto F, Musso S, Castillo L, Santini J, Dellamonica P, Hofman P. The value of polymerase chain reaction detection of Mycobacterium tuberculosis in granulomas isolated by laser capture microdissection. Pathology. 2004. 36: 77-81. https://doi.org/10.1080/00313020310001644516
  18. Singh KK, Muralidhar M, Kumar A, Chattopadhyaya TK, Kapila K, Singh MK, Sharma SK, Jain NK, Tyagi JS. Comparison of in house polymerase chain reaction with conventional techniques for the detection of Mycobacterium tuberculosis DNA in granulomatous lymphadenopathy. J Clin Pathol. 2000. 53: 355-361. https://doi.org/10.1136/jcp.53.5.355
  19. Yang YC, Lu PL, Huang SC, Jenh YS, Jou R, Chang TC. Evaluation of the Cobas TaqMan MTB test for direct detection of Mycobacterium tuberculosis complex in respiratory specimens. J Clin Microbiol. 2011. 49: 797-801. https://doi.org/10.1128/JCM.01839-10