Abstract
Presently the gold standard diagnostic technique for rabies is the direct immunofluorescence assay (dFA) which is very expensive and requires a high level of expertise. There is a need for more economical and user friendly tests, particularly for use in developing countries. We have established one such test called the direct rapid immunohistochemical test (dRIT) for diagnosis of rabies using brain tissue. The test is based on capture of rabies nucleoprotein (N) antigen in brain smears using a cocktail of biotinylated monoclonal antibodies specific for the N protein and color development by streptavidin peroxidase-amino ethyl carbazole and counter staining with haematoxollin. The test was done in parallel with standard FAT dFA using 400 brain samples from different animals and humans. The rabies virus N protein appears under light microscope as reddish brown particles against a light blue background. There was 100 % correlation between the results obtained by the two tests. Also, interpretation of results by dRIT was easier and only required a light microscope. To conclude, this newly developed dRIT technique promises to be a simple, cost effective diagnostic tool for rabies and will have applicability in field conditions prevalent in developing countries.
Keywords: Rabies, Post-mortem diagnosis, Fluorescent antibody technique, Immunohistochemistry, Direct rapid immunohistochemical test (dRIT)
References
- 1.Dean D. J., Abelesth M. K., Atanasiu P. The Fluorescent antibody test. In: Meslin F. X., Koprowsky H., Kaplan M. M., editors. Laboratory techniques in rabies. 4th ed. Geneva: World Health Organization; 1996. pp. 88–95. [Google Scholar]
- 2.Durr S., Naissengar S., Mindekem R., et al. Rabies diagnosis for developing countries. PLoS Negl Trop Dis. 2008;2:e206. doi: 10.1371/journal.pntd.0000206. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 3.Lembo T., Niezgoda M., Velasco-Villa A., et al. Evaluation of a direct, rapid immunohistochemical test for rabies diagnosis. Emerg Infect Dis. 2006;12:310–313. doi: 10.3201/eid1202.050812. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 4.Menezes R. Rabies in India. CMAJ. 2008;178:564–566. doi: 10.1503/cmaj.071488. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 5.Nagarajan T., Mohanasubramanian B., Seshagiri E. V., et al. Molecular epidemiology of rabies virus isolates in India. J Clin Microbiol. 2006;44:3218–3224. doi: 10.1128/JCM.00801-06. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 6.Perrin P., Rollin P. E., Sureau P. A rapid rabies enzyme immuno-diagnosis (RREID): a useful and simple technique for the routine diagnosis of rabies. J Biol Stand. 1986;14:217–222. doi: 10.1016/0092-1157(86)90006-5. [DOI] [PubMed] [Google Scholar]
- 7.Reddy G. B., Singh R., Singh R. P., et al. Molecular characterization of Indian rabies virus isolates by partial sequencing of nucleoprotein (N) and phosphoprotein (P) genes. Virus Genes. 2011;43:13–17. doi: 10.1007/s11262-011-0601-0. [DOI] [PubMed] [Google Scholar]
- 8.Saturday G A, King R, Fuhrmann L. 2009. Validation and operational application of a rapid method for rabies antigen detection. US Army Med Dep J, Jan–March, 42–45. [PubMed]
- 9.Sudarshan M. K., Madhusudana S. N., Mahendra B. J., et al. Assessing the burden of human rabies in India: results of a national multi-center epidemiological survey. Int J Infect Dis. 2007;11:29–35. doi: 10.1016/j.ijid.2005.10.007. [DOI] [PubMed] [Google Scholar]
- 10.Tao X. Y., Niezgoda M., Du J. L., et al. The primary application of direct rapid immunohistochemical test to rabies diagnosis in China. Chin J Exp Clin Virol. 2008;22(3):168–170. [PubMed] [Google Scholar]
- 11.World Health Organization. WHO Tech Rep Ser 935. Geneva: WHO; 2005. Expert consultation on rabies, 1st report. [PubMed] [Google Scholar]