This paper describes the first RT–PCR for discrimination of the nine African horse sickness virus (AHSV) serotypes. Nine pairs of primers were designed, each being specific for one AHSV serotype. The RT–PCR was sensitive and specific, providing serotyping within 24 h. Perfect agreement was recorded between the RT–PCR and virus neutralization for a coded panel of 56 AHSV reference strains and field isolates. Serotyping was achieved successfully with live and formalin-inactivated AHSVs, with isolates of virus after low and high passage through either tissue culture or suckling mouse brain, with viruses isolated from widely separated geographical areas and with viruses isolated up to 37 years apart. Overall, this RT–PCR provides a rapid and reliable method for the identification and differentiation of the nine AHSV serotypes, which is vital at the start of an outbreak to enable the early selection of a vaccine to control the spread of disease.


Article metrics loading...

Loading full text...

Full text loading...



  1. Borden, E. C., Shope, R. E. & Murphy, F. A. (1971). Physicochemical and morphological relationships of some arthropod-borne viruses to bluetongue virus – a new taxonomic group. Physicochemical and serological studies. Journal of General Virology 13, 261-271.[CrossRef] [Google Scholar]
  2. Bremer, C. W. (1976). A gel electrophoretic study of the protein and nucleic acid components of African horsesickness virus. Onderstepoort Journal of Veterinary Research 43, 193-199. [Google Scholar]
  3. Bremer, C. W., Huismans, H. & Van Dijk, A. A. (1990). Characterization and cloning of the African horsesickness virus genome. Journal of General Virology 71, 793-799.[CrossRef] [Google Scholar]
  4. Chomczynski, P. & Sacchi, N. (1987). Single-step method of RNA isolation by acid guanidinium thiocyanate–phenol–chloroform extraction. Analytical Biochemistry 162, 156-159. [Google Scholar]
  5. Du Toit, R. M. (1944). The transmission of bluetongue and horse sickness by Culicoides. Onderstepoort Journal of Veterinary Science and Animal Industry 19, 7-16. [Google Scholar]
  6. Grubman, M. J. & Lewis, S. A. (1992). Identification and characterization of the structural and nonstructural proteins of African horsesickness virus and determination of the genome coding assignments. Virology 186, 444-451.[CrossRef] [Google Scholar]
  7. Hamblin, C., Mertens, P. P. C., Mellor, P. S., Burroughs, N. J. & Crowther, J. R. (1991). A serogroup specific enzyme-linked immunosorbent assay for the detection and identification of African horse sickness viruses. Journal of Virological Methods 31, 285-292.[CrossRef] [Google Scholar]
  8. Huismans, H. & Els, H. J. (1979). Characterization of the tubules associated with the replication of three different orbiviruses. Virology 92, 397-406.[CrossRef] [Google Scholar]
  9. Iwata, H., Yamagawa, M. & Roy, P. (1992). Evolutionary relationships among the gnat-transmitted orbiviruses that cause African horse sickness, bluetongue, and epizootic hemorrhagic disease as evidenced by their capsid protein sequences. Virology 191, 251-261.[CrossRef] [Google Scholar]
  10. McIntosh, B. M. (1958). Immunological types of horse sickness virus and their significance in immunization.Onderstepoort Journal of Veterinary Research 27, 465-538. [Google Scholar]
  11. Mellor, P. S. (1994). Epizootiology and vectors of African horse sickness virus. Comparative Immunology, Microbiology and Infectious Diseases 17, 287-296.[CrossRef] [Google Scholar]
  12. Mizukoshi, N., Sakamoto, K., Iwata, A., Tsuchiya, T., Ueda, S., Watanabe, T., Kamada, M. & Fukusho, A. (1992). The complete sequence of African horsesickness virus serotype 4 (vaccine strain) RNA segment 5 and its predicted polypeptide compared with NS1 of bluetongue virus. Journal of General Virology 73, 2425-2428.[CrossRef] [Google Scholar]
  13. Moulay, S., Zientara, S., Sailleau, C. & Crucière, C. (1995). Detection of African horse sickness viruses by dot-blot hybridization using a digoxigenin-labelled probe. Molecular and Cellular Probes 9, 233-237.[CrossRef] [Google Scholar]
  14. Oellermann, R. A., Els, H. J. & Erasmus, B. J. (1970). Characterization of African horsesickness virus. Archiv für die gesamte Virusforschung 29, 163-174.[CrossRef] [Google Scholar]
  15. Reed, L. J. & Muench, H. (1938). A simple method for estimating fifty percent endpoints. American Journal of Hygiene 27, 493-497. [Google Scholar]
  16. Rodriguez, M., Hooghuis, H. & Castano, M. (1993). Current status of the diagnosis and control of African horse sickness. Veterinary Research 24, 189-197. [Google Scholar]
  17. Sailleau, C., Seignot, J., Hamblin, C. & Zientara, S. (1999). African horse sickness in Senegal: serotype identification and nucleotide sequence determination of segment S10 by RT–PCR. Veterinary Record (in press).
  18. Sakamoto, K., Mizukoshi, N., Apiwatnakorn, B., Iwata, A., Tsuchiya, T., Ueda, S., Imagawa, H., Sugiura, T., Kamada, M. & Fukusho, A. (1994). The complete sequences of African horsesickness virus serotype 4 (vaccine strain) RNA segment 2 and 6 which encode outer capsid protein. Journal of Veterinary Medical Science 56, 321-327.[CrossRef] [Google Scholar]
  19. Stone-Marschat, M., Carville, A., Skowronek, A. & Laegreid, W. W. (1994). Detection of African horse sickness virus by reverse transcription–PCR. Journal of Clinical Microbiology 32, 697-700. [Google Scholar]
  20. van Staden, V. & Huismans, H. (1991). A comparison of the genes which encode non-structural protein NS3 of different orbiviruses. Journal of General Virology 72, 1073-1079.[CrossRef] [Google Scholar]
  21. Verwoerd, D. W., Huismans, H. & Erasmus, B. J. (1979). Orbiviruses. In Comprehensive Virology, pp. 285-345. Edited by H. Fraenkel-Conrat & R. R. Wagner. New York: Plenum Press.
  22. Vreede, F. T. & Huismans, H. (1994). Cloning, characterization and expression of the gene that encodes the major neutralization-specific antigen of African horsesickness virus serotype 3. Journal of General Virology 75, 3629-3633.[CrossRef] [Google Scholar]
  23. Zientara, S., Sailleau, C., Moulay, S. & Crucière, C. (1994). Diagnosis of the African horse sickness virus serotype 4 by a one-tube, one manipulation RT–PCR reaction from infected organs. Journal of Virological Methods 46, 179-188.[CrossRef] [Google Scholar]
  24. Zientara, S., Sailleau, C., Moulay, S., Wade-Evans, A. & Crucière, C. (1995). Application of the polymerase chain reaction to the detection of African horse sickness viruses. Journal of Virological Methods 53, 47-54.[CrossRef] [Google Scholar]

Data & Media loading...

Most cited this month Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error