Simultaneous detection and differentiation of dengue virus serotypes 1-4, Japanese encephalitis virus, and West Nile virus by a combined reverse-transcription loop-mediated isothermal amplification assay

Virology Journal

Guangwen Cao¹  Guozhang Xu⁵  Wenjun Chang¹  Jianhua Yin¹  Yifang Han¹  Hongwei Zhang¹  Qiuxia Shen¹  Hongxia Ni⁵  Bin Zhou⁴  Meiyu Fang²  Shuhua Li³ 

[1] Department of Epidemiology, Shanghai Key Laboratory of Medical Biodefense, Second Military Medical University, Shanghai, China;Department of Microbiology, Center of Disease Control and Prevention of Guangzhou Military Region, Guangzhou, China;Department of Comprehensive Administration, District Center for Diseases Control and Prevention of Hongkou, Shanghai, China;Department of Emergency Medicine, First Affiliated Hospital, Second Military Medical University, Shanghai, China;Department of Infectious Diseases, Center of Disease Control and Prevention of Ningbo, Ningbo, China

关键词: Identification;    West Nile virus;    Japanese encephalitis virus;    Dengue virus;   

Others  :  1156420
DOI  :  10.1186/1743-422X-8-360

 received in 2011-05-03, accepted in 2011-07-21,  发布年份 2011

【 摘 要 】

Background

Rapid identification and differentiation of mosquito-transmitted flaviviruses in acute-phase sera of patients and field-caught vector mosquitoes are important for the prediction and prevention of large-scale epidemics.

Results

We developed a flexible reverse-transcription loop-mediated isothermal amplification (RT-LAMP) unit for the detection and differentiation of dengue virus serotypes 1-4 (DENV1-4), Japanese encephalitis virus (JEV), and West Nile virus (WNV). The unit efficiently amplified the viral genomes specifically at wide ranges of viral template concentrations, and exhibited similar amplification curves as monitored by a real-time PCR engine. The detection limits of the RT-LAMP unit were 100-fold higher than that of RT-PCR in 5 of the six flaviviruses. The results on specificity indicated that the six viruses in the assay had no cross-reactions with each other. By examining 66 viral strains of DENV1-4 and JEV, the unit identified the viruses with 100% accuracy and did not cross-react with influenza viruses and hantaviruses. By screening a panel of specimens containing sera of 168 patients and 279 pools of field-caught blood sucked mosquitoes, results showed that this unit is high feasible in clinical settings and epidemiologic field, and it obtained results 100% correlated with real-time RT-PCR.

Conclusions

The RT-LAMP unit developed in this study is able to quickly detect and accurately differentiate the six kinds of flaviviruses, which makes it extremely feasible for screening these viruses in acute-phase sera of the patients and in vector mosquitoes without the need of high-precision instruments.

【 授权许可】

   
2011 Li et al; licensee BioMed Central Ltd.

【 图 表 】

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