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Archives of Virology

, Volume 160, Issue 5, pp 1259–1266 | Cite as

Comparative evaluation of nucleic acid-based assays for detection of Japanese encephalitis virus in swine blood samples

  • H. Dhanze
  • K. N. Bhilegaonkar
  • G. V. P. P. S. Ravi Kumar
  • P. Thomas
  • H. B. Chethan Kumar
  • M. Suman Kumar
  • S. Rawat
  • P. Kerketta
  • D. B. Rawool
  • A. Kumar
Original Article

Abstract

Japanese encephalitis is an emerging mosquito-borne flaviviral zoonotic disease. The present study was undertaken with the objective of developing rapid and sensitive nucleic-acid-based assays for detection of Japanese encephalitis virus (JEV) in swine blood samples. Three nucleic-acid-based assays, viz., reverse transcription polymerase chain reaction (RT-PCR), reverse transcription loop-mediated isothermal amplification (RT-LAMP), and real-time RT-PCR, were developed and compared in terms of their diagnostic efficacy. All three assays were found to be 100 per cent specific. The minimum detection limit of RT-LAMP and real-time RT-PCR was 12 copies/µl, while RT-PCR could detect 1.2 × 105 copies/µl. On comparison, RT-LAMP and real-time RT-PCR were 4-log more sensitive than RT-PCR. The applicability of the assays was evaluated by screening 135 field swine blood samples, of which 24 (17.77 %) were positive by RT-LAMP and real-time RT-PCR and only six (4.44 %) were positive by RT-PCR. The viral load in swine blood samples ranged between 2 × 106 and 4.8 × 109 copies per ml of blood by real-time RT-PCR. The comparative diagnostic sensitivity and specificity of RT-LAMP vis-à-vis real-time RT-PCR was found to be 100 %, while the sensitivity and specificity of RT-PCR vis-à-vis real-time RT-PCR was found to be 25 % and 100 %, respectively. Thus, the use of RT-PCR may cause the incidence of JEV in the swine population to be underestimated, while the real-time RT-PCR reported here is the test of choice for reference laboratories, and the newly developed one-step RT-LAMP assay will be suitable for field-level testing.

Keywords

Reverse Transcription Polymerase Chain Reaction West Nile Virus Japanese Encephalitis Virus Japanese Encephalitis Classical Swine Fever Virus 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Conflict of interest

The authors declare no conflict of interest.

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Copyright information

© Springer-Verlag Wien 2015

Authors and Affiliations

  • H. Dhanze
    • 1
  • K. N. Bhilegaonkar
    • 2
  • G. V. P. P. S. Ravi Kumar
    • 2
  • P. Thomas
    • 2
  • H. B. Chethan Kumar
    • 2
  • M. Suman Kumar
    • 3
  • S. Rawat
    • 4
  • P. Kerketta
    • 2
  • D. B. Rawool
    • 2
  • A. Kumar
    • 2
  1. 1.Division of Veterinary Public HealthIndian Veterinary Research InstituteBareillyIndia
  2. 2.Indian Veterinary Research InstituteBareillyIndia
  3. 3.Department of VPH and EpidemiologyCollege of Veterinary Science and Animal HusbandryJunagadhIndia
  4. 4.Department of VPH and EpidemiologyCollege of Veterinary ScienceMeerutIndia

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