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Journal of Clinical Microbiology, August 2007, p. 2398-2403, Vol. 45, No. 8
0095-1137/07/$08.00+0     doi:10.1128/JCM.00292-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Development and Evaluation of a Test for Tuberculosis in Live European Badgers (Meles meles) Based on Measurement of Gamma Interferon mRNA by Real-Time PCR{triangledown}

J. Sawyer,1 D. Mealing,1 D. Dalley,2 D. Davé,2 S. Lesellier,2 S. Palmer,2 J. Bowen-Davies,3,{dagger} T. R. Crawshaw,4 and M. A. Chambers2*

Technology Transfer Unit, Biotechnology Department,1 TB Research Group, Department of Statutory and Exotic Bacteria, Veterinary Laboratories Agency Weybridge, New Haw, Addlestone, Surrey KT15 3NB, United Kingdom,2 TBD Wildlife Unit, Aston Down, Stroud, Gloucestershire GL6 8GA, United Kingdom,3 Veterinary Laboratories Agency Starcross, Staplake Mount, Starcross, Exeter EX6 8PE, United Kingdom4

Received 6 February 2007/ Returned for modification 14 March 2007/ Accepted 21 May 2007

A real-time PCR assay for the measurement of gamma interferon (IFN-{gamma}) mRNA in European badger (Meles meles) blood cultures was developed. The levels of IFN-{gamma} mRNA in blood cultures stimulated with either bovine or avian tuberculin or specific mycobacterial antigens were compared with those in a nonstimulated control blood culture as the basis for determining the tuberculosis (TB) status of live badgers. The assay was validated by testing 247 animals for which there were matching data from postmortem examination and culture of tissues. Relative changes in the levels of IFN-{gamma} mRNA in response to bovine tuberculin and specific antigens were found to be greater among badgers with tissues positive for TB on culture. The test was at its most accurate (87% of test results were correct) by using blood cultures containing bovine tuberculin as the antigen and when the response to avian tuberculin was taken into account by subtracting the avian tuberculin response from the bovine tuberculin response. At a specificity of 90.7%, the test was 70.6% sensitive. At the same specificity, the current serological enzyme-linked immunosorbent assay for TB in badgers was only 53% sensitive. This work demonstrates that measurement of IFN-{gamma} mRNA by real-time PCR is a valid method for the detection of TB in live badgers and may provide an alternative to the current serological methods of diagnosis, the Brock test. The testing procedure can be completed within 5 h of receipt of the blood culture samples. In addition, the use of a molecular biology-based test offers the potential to fully automate the testing procedure through the use of robotics.

* Corresponding author. Mailing address: TB Research Group, Department of Statutory and Exotic Bacteria, Veterinary Laboratories Agency Weybridge, New Haw, Addlestone, Surrey KT15 3NB, United Kingdom. Phone: 44 (0) 1932 357494. Fax: 44 (0) 1932 357260. E-mail: m.a.chambers{at}vla.defra.gsi.gov.uk

{triangledown} Published ahead of print on 30 May 2007.

{dagger} Present address: Home Office, P.O. Box 1138, Swindon SN1 2RZ, United Kingdom.

Journal of Clinical Microbiology, August 2007, p. 2398-2403, Vol. 45, No. 8
0095-1137/07/$08.00+0     doi:10.1128/JCM.00292-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Chambers, M. A., Crawshaw, T., Waterhouse, S., Delahay, R., Hewinson, R. G., Lyashchenko, K. P. (2008). Validation of the BrockTB Stat-Pak Assay for Detection of Tuberculosis in Eurasian Badgers (Meles meles) and Influence of Disease Severity on Diagnostic Accuracy. J. Clin. Microbiol. 46: 1498-1500 [Abstract] [Full Text]  
  • Harrington, N. P., Surujballi, O. P., Waters, W. R., Prescott, J. F. (2007). Development and Evaluation of a Real-Time Reverse Transcription-PCR Assay for Quantification of Gamma Interferon mRNA To Diagnose Tuberculosis in Multiple Animal Species. CVI 14: 1563-1571 [Abstract] [Full Text]  


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