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Brazilian Journal of Microbiology

, Volume 50, Issue 1, pp 287–296 | Cite as

Quantitative RT-PCR assays for identification and typing of the Equine encephalosis virus

  • Sushila MaanEmail author
  • Manjunatha N. BelaganahalliEmail author
  • Narender Singh Maan
  • Abraham C. Potgieter
  • Peter P. C. Mertens
Veterinary Microbiology - Research Paper

Abstract

Equine encephalosis (EE) is an acute, arthropod-borne, noncontagious, febrile disease of equids. The clinical signs of EE are similar to milder forms of African horse sickness (AHS) and the two diseases can be easily confused. The Equine encephalosis virus (EEV) is a distinct virus species within the genus Orbivirus, family Reoviridae, with ten linear segments of dsRNA genome. Seven distinct serotypes of EEV have been recognised on the basis of sequence analyses of Seg-2. The need for differential diagnosis of similar forms of EE and AHS warranted the development of molecular diagnostic methods for specific detection and identification of EEV. We report the development of quantitative real-time RT-PCR assay for detection of any member of the EEV species targeting the highly conserved EEV Seg-9. Similar serotype-specific qRT-PCR assays were designed for each of the seven EEV serotypes targeting genome Seg-2, encoding the serotype determining VP2 protein. These assays were evaluated using different EEV serotypes and other closely related orbiviruses. They were shown to be EEV virus species–specific, or EEV type–specific capable of detecting 1 to 13 copies of viral RNA in clinical samples. The assays failed to detect RNA from closely related orbiviruses, including AHSV and Peruvian horse sickness virus (PHSV) isolates.

Keywords

Equine encephalosis Equine encephalosis virus EEV TaqMan assays Real-time RT-PCR Serotype-specific assays Serogroup-specific assays Orbivirus Reference Collection Culicoides 

Notes

Acknowledgments

This work was supported by the Biotechnology and Biological Sciences Research Council (BBSRC) (grant no. BB/L004690/1) and Department for Environment, Food and Rural Affairs (Defra) (SE2617) to PM. Department of Biotechnology, Government of India (DBT, GOI) (grant no. BT/IN/Indo-UK/FADH/46/SM/2013.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

42770_2018_34_MOESM1_ESM.docx (22 kb)
ESM 1 (DOCX 21 kb)

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Copyright information

© Sociedade Brasileira de Microbiologia 2018

Authors and Affiliations

  • Sushila Maan
    • 1
    • 2
    • 3
    Email author
  • Manjunatha N. Belaganahalli
    • 2
    • 4
    Email author
  • Narender Singh Maan
    • 1
    • 2
  • Abraham C. Potgieter
    • 5
  • Peter P. C. Mertens
    • 2
    • 6
  1. 1.College of Veterinary SciencesLLR University of Veterinary and Animal SciencesHisarIndia
  2. 2.The Pirbright InstitutePirbrightUK
  3. 3.Department of Animal Biotechnology, College of Veterinary SciencesLala Lajpat Rai University of Veterinary and Animal Sciences (LUVAS)HisarIndia
  4. 4.One Health Institute, School of Veterinary medicineUniversity of CaliforniaCaliforniaUSA
  5. 5.Deltamune (Pty) LtdLytteltonSouth Africa
  6. 6.Chair of Virology, School of Veterinary Medicine and ScienceUniversity of NottinghamSutton Bonington CampusUK

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