Indian Journal Of Animal Research

Standardization and application of immunofluorescence and immuno- peroxidase tests for detection of bluetongue virus antigen
 

DOI: 10.18805/ijar.B-3626    | Article Id: B-3626 | Page : 1074-1079
Citation :- Standardization and application of immunofluorescence and immuno- peroxidase tests for detection of bluetongue virus antigen.Indian Journal Of Animal Research.2019.(53):1074-1079
Yashitha Priya, Kalyani Putty, Sunil. R. Patil, Y. Narsimha Reddy, Y. Vishnuvardhan Reddy, J. Shiva Jyothi, B. Susmitha and P.P. Rao kalyaniputty@gmail.com
Address : College of Veterinary Science, P.V.N.R. Telangana Veterinary University, Rajendrangar, Hyderabad-500 030, Telangana, India.
Submitted Date : 4-05-2018
Accepted Date : 24-08-2018
First Online:

Abstract

India is enzootic for bluetongue (BT), a predominant disease of small ruminants. The most important task in the control of disease is rapid and sensitive detection of virus. The present study was undertaken to standardize immunofluorescence (IFT) and immunoperoxidase tests (IPT) employing BTV serogroup specific VP7 monoclonal antibodies (MAbs), polyclonal homologous, and polyclonal heterologous antisera against specific serotypes of BTV for detection of BTV antigen. Serial tenfold dilutions of BTV-9 were tested for limit of detection (LoD) of IFT, IPT, and molecular assays by using MAbs against VP7, homologous anti-BTV-9 serum, and heterologous anti-BTV-16 serum. LoD of IFT was found to be 101 TCID50/mL using MAbs against VP7, anti BTV-9 serum, and anti BTV-16 serum. LoD of IPT was found to be 101 TCID50/mL, 102 TCID50/mL, and 102 TCID50/mL using MAbs against VP7, anti BTV-9 serum and anti BTV-16 serum, respectively. LoD of RT-PCR was 101 TCID50/mL and that of real time PCR was 100 TCID50/mL. This standardized assay was then applied for BTV detection in BTV suspected field samples collected from BT outbreaks followed by confirmation with virus isolation and NS3 group specific PCR. The current study shows that IFT and IPT are specific tests for group specific BTV identification. For IFT, monoclonal and polyclonal (homologous and heterologous) source of antibodies had similar sensitivity in the ability of BTV detection; whereas the most sensitive mode of detection by IPT was with MAbs. 

Keywords

Bluetongue Immunofluorescence test Immunoperoxidase test LoD VP7 monoclonal antibody.

References

  1. Anderson G A, Phillips D L, Waldvogel A S, and Osburn B I (1989) Detection of bluetongue virus in bovine fetuses using the avidin-    biotin complex immunoperoxidase method. Journal of VeterinaryDiagnostic Investigation 1:45-49.
  2. Bowne JG, Luedke AJ, Jochim MM, and Metcalf HE (1968) Bluetongue disease in cattle. Journal of American Veterinary and Medical Association 153:662-668.
  3. Cherrington, J.M., Ghalib, H.W., Dvis, W.C. and Osbum, B.I. (1985) Monoclonal antibodies raised against bluetongue virus detect viral antigen in infected tissues using an indirect immunoperoxidase method. In: T.L Barber, M.M. Jochim and B.I.Osbom (Eds), Bluetongue and Related Orbiviruses. Alan Liss, New York, pp. 505-510.
  4. Fernandes MV (1973) Isolation and propagation of bluetongue virus in tissue culture. AmericanJournal of Veterinary Research 20:398-408.
  5. Foster N M, Luedke A J and Metcalf HE (1972) Bluetongue in sheep and cattle efficacy of embryonating chicken eggs in viral isolations. American Journal of Veterinary Research 33:77-81.
  6. Gould A R, Hyatt A D, Eaton B T, White J R, Hooper P T, Blacksell S D and Le Blancsmith P M (1989) Current techniques in rapid bluetongue virus diagnosis. Austrian Veterinary Journal, 66 (12):450-454.
  7. Lakshmi.I.K, Putty K, Raut S S, Patil S R, Rao P P, Bhagyalakshmi B, Jyothi Y K, Susmitha B, Reddy Y V, Kasulanati S, Jyothi J S and Reddy Y N (2018) Standardisation and application of real time PCR for rapid detection of bluetongue virus ;Veterinary World, 11(4): 452-458 
  8. Li T,Yang Y, Xu Q, Sun E, Feng Y, Lv S, Zhang Q, Wang H and Wu D (2015) DNA Vaccine prime and recombinant FPV vaccine boost : an important candidate immunization strategy to control bluetongue virus type I. Applied Microbiology and Bitechnology, 99 (20): 8643-52.
  9. Maan S, Maan NS, Samuel AR, Rao S, Attoui H, Mertens PP (2007) Analysis and phylogenetic comparisons of full-length VP2 genes of the 24 bluetongue virus serotypes. Journal of General Virology. Feb;88(Pt 2):621-30.
  10. Maan S, Maan NS, Belaganahalli MN,Potgieter AC, Kumar V, Batra K, et al. (2016) Development and evaluation of real time RT-PCR assays for detection and typing of Bluetongue virus. PLoS ONE 11(9): e0163014 doi: 10.1371/journal.pone.0163014.
  11. MacLachlan N J, Jagels G, Rossitto P V, Moore P F and Heidner H W (1990) The pathogenesis of experimental bluetongue virus infection of calves. Veterinary Pathology 27, 223-229.
  12. MacLachlan N J, Drew C P, Darpel K E and Worwa G (2009). The pathology and pathogenesis of Bluetongue. Journal of Comparative Pathology 141, 1-16.
  13. MacLachlan N J and Gard G (2009) Clinical signs and pathology. In: Bluetongue. [Mellor P, Baylis M, Mertens PPC] Academic Press, London. 285–293.
  14. MacLachlan N J (2004) Bluetongue: pathogenesis and duration of viraemia. Veterinaria Italiana 40, 462-467.
  15. McPhee D A, Parsonson I M and Della-Porta A J (1982) Comparative studies on the growth of Australian bluetongue virus serotypes in continuous cell lines and embryonated chicken eggs. Veterinary Microbiology 7:401-410.
  16. O’Brien CA, Hobson-Peters J, and Yam AWY, Colmant AMG, McLean BJ, Prow NA, et al. (2015) Viral RNA intermediates as targets for detection and discovery of novel and emerging Mosquito-Borne viruses. PLoS Negl Trop Dis 9 (3): e0003629.
  17. Parsonson I M, Della-Porta A J, McPhee D A (1981) Isolation of bluetongue virus serotype 20 from the semen of an experimentally-    infected bull. Australian Veterinary Journal 57:252-253.
  18. Pini A (1976) Study on the pathogenesis of bluetongue: replication of the virus in the organs of infected sheep. Onderstepoort Journal of Veterinary Research 43: 159-164.
  19. Ruckerbauer G M, Gray D P, Girard A, Bannister G L and Boulanger P (1967) Studies on bluetongue, detection of the virus in infected materials by immunofluorescence. Canadian Journal of Comparitive Medicine and Veterinary Science. 31: 175-181.
  20. Spreull J (1905) Malarial catarrhal fever (bluetongue) of sheep in South Africa. Journal of Comparative Pathology and Therapeutics 18: 321–337.
  21. Wechsler S J and McHolland L E (1988) Susceptibility of 14 cell lines to bluetongue virus infection. Journal of Clinical Microbiology 26: 2324–2327.
  22. Wechsler S J, Austin K J, William and Wilson C (1990) Limits of detection of bluetongue virus with different assay systems. Journal of Veterinary Diagnostic Investigation 2: 103-106.
  23. Whetter L E, Maclachlan N J, Gebhard D H Heidner H W and Moore P F (1989) Bluetongue Virus-Infection of Bovine Monocytes. Journal of General Virology 70: 1663-1676.
  24. Wright I M (2013) Serological and genetic characterisation of putative new serotypes of bluetongue virus and epizootic haemorrhagic disease virus isolated from an Alpaca, South Africa: North-West University, Potchefstroom Campus; 2013. 

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