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Journal of Clinical Microbiology, Nov 1997, 2762-2766, Vol 35, No. 11
Copyright © 1997 by the American Society for Microbiology. All rights reserved.

Heminested PCR assay for detection of six genotypes of rabies and rabies-related viruses [In Process Citation]

PR Heaton, P Johnstone, LM McElhinney, R Cowley, E O'Sullivan and JE Whitby
Department of Virology, Veterinary Laboratories Agency, Addlestone, Surrey, United Kingdom. pheaton@hgmp.mrc.ac.uk

A heminested reverse transcriptase PCR (hnRT-PCR) protocol which is rapid and sensitive for the detection of rabies virus and rabies- related viruses is described. Sixty isolates from six of the seven genotypes of rabies and rabies-related viruses were screened successfully by hnRT-PCR and Southern blot hybridization. Of the 60 isolates, 93% (56 of 60) were positive by external PCR, while all isolates were detected by heminested PCR and Southern blot hybridization. We also report on a comparison of the sensitivity of the standard fluorescent-antibody test (FAT) for rabies antigen and that of hnRT-PCR for rabies viral RNA with degraded tissue infected with a genotype 1 virus. Results indicated that FAT failed to detect viral antigen in brain tissue that was incubated at 37 degrees C for greater than 72 h, while hnRT-PCR detected viral RNA in brain tissue that was incubated at 37 degrees C for 360 h.


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