Hostname: page-component-cd9895bd7-dk4vv Total loading time: 0 Render date: 2024-12-23T20:11:09.178Z Has data issue: false hasContentIssue false
  • English
  • Français

Establishment and evaluation of real-time PCR for West Nile virus detection

Published online by Cambridge University Press:  24 April 2009

Shi Li-Jun ,
Lu Mao-Min ,
Li Gang ,
Li Cheng-Yao  and
Zhang Jin-Gang
Shi Li-Jun
Affiliation:
Institute of Transfusion Medicine, Academy of Military Medical Sciences, Beijing 100850, China Institute of Animal and Veterinary Medicine, Chinese Academy of Agricultural Science, Beijing 100193, China
Lu Mao-Min
Affiliation:
Institute of Transfusion Medicine, Academy of Military Medical Sciences, Beijing 100850, China
Li Gang
Affiliation:
Institute of Animal and Veterinary Medicine, Chinese Academy of Agricultural Science, Beijing 100193, China
Li Cheng-Yao
Affiliation:
College of Bio-technology, Southern Medical University, Guangzhou 510515, China
Zhang Jin-Gang*
Affiliation:
Institute of Transfusion Medicine, Academy of Military Medical Sciences, Beijing 100850, China
*
*Corresponding author. E-mail: [email protected]
Get access Rights & Permissions [Opens in a new window]

Abstract

A rapid real-time polymerase chain reaction (RT-PCR) for detecting West Nile virus (WNV) was established. Primers were designed according to the sequence of the capsid protein gene of WNV by Primer Premier 5.0. In this way, an inexpensive assay using the intercalating dye SYBR Green I was developed and validated. The amplifying curve showed that this method could successfully amplify 102 copies/μl of the WNV gene, while reference to Japanese encephalitis virus (JEV) and blank control were all negative. Tenfold successive dilutions of positive WNV DNA were used to measure the sensitivity of RT-PCR. The assay system showed high reproducibility with coefficient of variation (CV) <2%. Thus the newly established RT-PCR assay was shown to be a rapid, sensitive and specific test for detecting WNV.

Keywords

West Nile viruscapsid protein genereal-time PCR
Type
Research Papers
Information
Chinese Journal of Agricultural Biotechnology , Volume 6 , Issue 1 , April 2009 , pp. 55 - 59
Copyright
Copyright © China Agricultural University 2009

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

Bakonyi, T and Ivanics, E (2006) Lineage 1 and Lineage 2 strains of encephalitis West Nile Virus, Central Europe. Emerging Infectious Diseases 12: 618–623.CrossRefGoogle ScholarPubMed
Bakonyi, T, Hubalek, Z, Rudolf, I, et al. (2005) Novel flavivirus or new lineage of West Nile Virus, Central Europe. Emerging Infectious Diseases 11: 225–231.CrossRefGoogle ScholarPubMed
Cui, F, Raymond, M, Berthomieu, A, Alout, H, et al. (2006) Recent emergence of insensitive acetylcholinesterase in Chinese populations of the mosquito Culex pipiens. Journal of Medical Entomology 43: 878–883.CrossRefGoogle ScholarPubMed
Hadfield, TL, Turell, M, Dempsey, MP, et al. (2001) Detection of West Nile Virus in mosquitoes by RT-PCR. Molecular and Cellular Probes 15: 147–150.CrossRefGoogle ScholarPubMed
Lo, MK, Tilgner, M and Shi, PY (2003) Potential high-throughput assay for screening inhibitors of West Nile Virus replication. Journal of Virology 77: 12901–12906.CrossRefGoogle ScholarPubMed
Nicolle, LE, Gutkin, A, Smart, G, et al. (2004) Serological studies of West Nile Virus in a liver transplant population. Canadian Journal of Infectious Diseases and Medical Microbiology 15: 271–274.CrossRefGoogle Scholar
Parida, M, Posadas, G, Inoue, S, Hasebe, F and Morita, K (2004) Real-time reverse transcription loop-mediated isothermal amplification for rapid detection of West Nile Virus. Journal of Clinical Microbiology 42: 257–263.CrossRefGoogle ScholarPubMed
Petersen, LR and Roehrig, JT (2001) West Nile Virus: a reemerging global pathogen. Emerging Infectious Diseases 7: 611–614.CrossRefGoogle ScholarPubMed
Zhang, JG (2004) Blood screening of West Nile Virus and transfusion-associated transmission. Journal of Clinical Transfusion and Laboratory Medicine 6: 308–310 (in Chinese).Google Scholar