Development of a Visible Reverse Transcription-Loop-Mediated Isothermal Amplification Assay for the Detection of Rift Valley Fever Virus

Qiuxue Han; Shengnan Zhang; Dongping Liu; Feihu Yan; Hualei Wang; Pei Huang; Jinhao Bi; Hongli Jin; Na Feng; Zengguo Cao; Yuwei Gao; Hang Chi; Songtao Yang; Yongkun Zhao; Xianzhu Xia

doi:10.3389/fmicb.2020.590732

Development of a Visible Reverse Transcription-Loop-Mediated Isothermal Amplification Assay for the Detection of Rift Valley Fever Virus

Front Microbiol. 2020 Nov 13:11:590732. doi: 10.3389/fmicb.2020.590732. eCollection 2020.

Authors

Qiuxue Han^{1

2}, Shengnan Zhang², Dongping Liu³, Feihu Yan^{2

4}, Hualei Wang⁵, Pei Huang^{2

6}, Jinhao Bi^{2

6}, Hongli Jin^{2

5}, Na Feng², Zengguo Cao^{2

5}, Yuwei Gao^{2

4}, Hang Chi^{2

4}, Songtao Yang^{2

4}, Yongkun Zhao^{2

4}, Xianzhu Xia^{1

2

4}

Affiliations

¹ Institute of Laboratory Animal Science, Chinese Academy of Medical Sciences (CAMS) and Comparative Medicine Center, Peking Union Medical College (PUMC), Beijing, China.
² Key Laboratory of Jilin Province for Zoonosis Prevention and Control, Institute of Military Veterinary Medicine, Academy of Military Medical Sciences, Changchun, China.
³ The Nanjing Unicorn Academy of Innovation, Institute Pasteur of Shanghai, Chinese Academy of Sciences, Nanjing, China.
⁴ Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Disease and Zoonoses, Yangzhou University, Yangzhou, China.
⁵ College of Veterinary Medicine, Jilin University, Changchun, China.
⁶ Animal Science and Technology College, Jilin Agricultural University, Changchun, China.

Abstract

Rift Valley fever (RVF) is a severe infectious disease, which can through mosquito bites, direct contact and aerosol transmission infect sheep, goats, people, camels, cattle, buffaloes, and so on. In this paper, a conserved region of the S RNA segment of Rift Valley fever virus (RVFV) ZH501 strain was used as target sequence. The RVFV RT-LAMP-VF assay was successfully established combined reverse transcription-loop-mediated isothermal amplification with a vertical flow visualization strip. The detection limit is up to 1.94 × 10⁰ copies/μl of synthesized RVFV-RNA. RNA extracted from cell culture of an inactivated RVFV-BJ01 strain was also used as templates, and the detection limit is 1.83 × 10³ copies/μl. In addition, there was no cross-reactivity with other viruses that can cause similar fever symptoms. The RVFV-LAMP-VF assay exhibited very high levels of diagnostic sensitivity, which had 100-fold more sensitive than RVFV real-time RT-PCR assay. Accordingly, the RVFV RT-LAMP-VF assay developed in this study is suitable for the rapid and sensitive diagnosis of RVFV without specialized equipment and can rapidly complete detection within 60 min, and the results are visible by vertical flow visualization strip within 5 min.

Keywords: Rift Valley fever virus; inactivated RVFV-BJ01 strain; nucleic acid visualization; reverse transcription-loop-mediated isothermal amplification; visual detection.