Share This Article:

Comparison of the Sensitivities of Different PCR Assays for the Detection of Mycobacterium tuberculosis Complex Isolates from Five Regions of Cameroon

Abstract Full-Text HTML XML Download Download as PDF (Size:545KB) PP. 34-42
DOI: 10.4236/jtr.2015.32005    2,434 Downloads   3,153 Views   Citations

ABSTRACT

The accurate diagnosis of tuberculosis caused by members of the Mycobacterium tuberculosis complex (MTBC) has remained a major challenge in clinical laboratories world-wide. Several studies have evaluated the use of highly specific in-house PCR assays targeting the IS6110, hupB, rpoB, oxyR, and IS1081 genes in the detection of MTBC species with reports on variable sensitivities depending on the geographical sourcing of isolates. In the present investigations, we evaluated the sensitivities of these PCR assays on 125 MTBC cultured isolates from five (West, Centre, Littoral, North West and South West) of the ten regions of Cameroon. Of this number, 124 (99.2%), 117 (93.6%), 123 (99.1%), 119 (95.2%) and 118 (94.4%) were positive by the IS6110, hupB, rpoB, oxyR, and IS1081-based PCR assays respectively. A total of 110 (88%) of the cultured isolates were also identified as MTBC by standard biochemical tests. Of this number, 109 (99.1%), 104 (94.5%), 109 (99.1%), 106 (96.4%) and 104 (94.5%) were positive in the IS6110, hupB, rpoB, oxyR, and IS1081-based PCR assays respectively. Concordant PCR results were obtained for 108 of the 125 samples. The 15 isolates that were negative biochemically scored sensitivities ranging from 100% (for the IS6110 assay) to 86.7% (for the hupB and oxyR assay). The combination of the IS6110 assay, which turned out to be the most sensitive, and each of the other assays gave 100% sensitivity. We conclude that the combined targeting of the IS6110 and rpoB genes is likely to yield the most sensitive PCR procedure for the diagnosis of MTBC infection in the five regions of Cameroon.

Cite this paper

Cho-Ngwa, F. , Anyangwe, I. , Berinyuy, E. , Meriki, H. , Assam-Assam, J. and Titanji, V. (2015) Comparison of the Sensitivities of Different PCR Assays for the Detection of Mycobacterium tuberculosis Complex Isolates from Five Regions of Cameroon. Journal of Tuberculosis Research, 3, 34-42. doi: 10.4236/jtr.2015.32005.

Conflicts of Interest

The authors declare no conflicts of interest.

References

[1] Katoch, V.M. (2004) Infections Due to Non-Tuberculous Mycobacteria (NTM). Indian Journal of Medical Research, 120, 290-304.
[2] World Health Organisation (2010) Tuberculosis, WHO Fact Sheet No. 104, November 2010.
http://www.Who.Int/Mediacentre/Factsheets/Fs104/En/
[3] World Health Organisation (2008) Global Tuberculosis Control: Surveillance, Planning, Financing: WHO Report 393.
[4] Kent, P.T. and Kubica, G.P. (1985) Public Health Mycobacteriology: A Guide to the Level III Laboratory. USDHHS. Centres for Disease Control, Atlanta, 159-184.
[5] Katoch, V.M. (2003) Advances in Molecular Diagnosis of Tuberculosis. Medical Journal Armed Forces of India, 59, 182-186. http://dx.doi.org/10.1016/S0377-1237(03)80001-1
[6] Katoch, V.M. (2004) Newer Diagnostic Techniques for Tuberculosis. Indian Journal of Medical Research, 120, 418- 428.
[7] Balasingham, S.V., Davidsen, T., Szpinda, I., Frye, S.A. and Tønjum, T. (2009) Molecular Diagnostics in Tuberculosis. Molecular Diagnosis and Therapy, 13, 137-151. http://dx.doi.org/10.1007/BF03256322
[8] Eisenach, K.D., Cave, M.D., Bates, J.H. and Crawford, J.T. (1990) Polymerase Chain Reaction Amplification of a Repetitive DNA Sequence Specific for Mycobacterium tuberculosis. Journal of Infectious Diseases, 161, 977-981. http://dx.doi.org/10.1093/infdis/161.5.977
[9] Niyaz, A., Ashok, K.M., Utpal, M., Virender, K.B. and Sunita, G. (1998) PCR-Based Rapid Detection of Mycobacterium tuberculosis in Blood from Immuno-Competent Patients with Pulmonary Tuberculosis. Journal of Clinical Microbiology, 36, 3094-3095.
[10] Mokaddas, E. and Ahmad, S. (2007) Development and Evaluation of a Multiplex PCR for Rapid Detection and Differentiation of Mycobacterium tuberculosis Complex Members from Non-Tuberculous Mycobacteria. Japanese Journal of Infectious Disease, 60, 140-144.
[11] Prabhakar, S., Mishra, A., Singhal, A., Katoch, V.M., Thakral, S.S., Tyagi, J.S. and Prasad, H.K. (2004) Use of the Hupb Gene Encoding a Histone-Like Protein of Mycobacterium tuberculosis as a Target for Detection and Differentiation of M. tuberculosis and M. Bovis. Journal Clinical Microbiology, 42, 2724-2732.
http://dx.doi.org/10.1128/JCM.42.6.2724-2732.2004
[12] International Union against Tuberculosis and Lung Disease (1998) The Public Health Service National Tuberculosis Reference Laboratory and the National Laboratory Network: Minimum Requirements, Role and Operation in a Low- Income Country. Paris, 62-71.
[13] Wilson, K. (1990) Preparation of Genomic DNA from Bacteria. In: Ausbel, F.M., Brent, R., Kingston, R.E., Moore, D.D., Seidman, J.G., Smith, J.A., et al., Eds., Current Protocols in Molecular Biology, Volume 1, Green and Wiley- Interscience, New York, 241-242.
[14] Ogusku, M.M. and Salem, J.I. (2004) Analysis of Different Primers Used in the PCR Method: Diagnosis of Tuberculosis in the State of Amazonas. Jornal Brasileiro De Pneumologia, 30, 343-349.
[15] Van Soolingen, D., De Haas, P.W., Hermans, P.W., Groenen, P.M.A. and Van Embden, J.D.A. (1993) Comparison of Various Repetitive DNA Elements as Genetic Markers for Strain Differentiation and Epidemiology of Mycobacterium tuberculosis. Journal of Clinical Microbiology, 31, 1987-1995.
[16] Kim, B.J., Lee, S.H., Lyu, M.A., Kim, S.J., Bai, G.H., et al. (1999) Identification of Mycobacterial Species by Comparative Sequence Analysis of the RNA Polymerase Gene (Rpob). Journal of Clinical Microbiology, 37, 1714-1720.
[17] Nassar, A.F., Miyashiro, S., Oliveira, C.G., Pacheco, W.A. and Ogata, R.A. (2007) Isolation and Identification of Bovine Tuberculosis in a Brazilian Herd (São Paulo). Memórias do Instituto Oswaldo Cruz, 102, 639-642. http://dx.doi.org/10.1590/S0074-02762007005000073
[18] Kato-Maeda, M., Bifani, P.J., Krieswirth, B.N. and Small, P.M. (2001) The Nature and Consequence of Genetic Variability in Mycobacterium tuberculosis. Journal of Clinical Investigation, 107, 533-537.
http://dx.doi.org/10.1172/JCI11426
[19] Bemer-Melchior, P. and Drugeon, H. (1999) Inactivation of Mycobacterium tuberculosis for DNA Typing Analysis. Journal of Clinical Microbiology, 37, 2350-2351.

  
comments powered by Disqus

Copyright © 2020 by authors and Scientific Research Publishing Inc.

Creative Commons License

This work and the related PDF file are licensed under a Creative Commons Attribution 4.0 International License.