Title: Establishment of a reverse transcription real-time quantitative PCR method for Getah virus detection and its application for epidemiological investigation in Shandong, China

Abstract

Getah virus (GETV) is a mosquito-borne, single-stranded, positive-sense RNA virus belonging to the genus Alphavirus of the family Togaviridae . Natural infections of GETV have been identified in a variety of vertebrate species, with pathogenicity mainly in swine, horses, bovines, and foxes. The increasing spectrum of infection and the characteristic causing abortions in pregnant animals pose a serious threat to public health and the livestock economy. Therefore, there is an urgent need to establish a method that can be used for epidemiological investigation in multiple animals. In this study, a real-time reverse transcription fluorescent quantitative PCR (RT-qPCR) method combined with plaque assay was established for GETV with specific primers designed for the highly conserved region of GETV Nsp1 gene. The results showed that after optimizing the condition of RT-qPCR reaction, the minimum detection limit of the assay established in this study was 7.73 PFU/mL, and there was a good linear relationship between viral load and Cq value with a correlation coefficient ( R ² ) of 0.998. Moreover, the method has good specificity, sensitivity, and repeatability. The established RT-qPCR is 100-fold more sensitive than the conventional RT-PCR. The best cutoff value for the method was determined to be 37.59 by receivermore » operating characteristic (ROC) curve analysis. The area under the curve (AUC) was 0.956. Meanwhile, we collected 2,847 serum specimens from swine, horses, bovines, sheep, and 17,080 mosquito specimens in Shandong Province in 2022. The positive detection rates by RT-qPCR were 1%, 1%, 0.2%, 0%, and 3%, respectively. In conclusion, the method was used for epidemiological investigation, which has extensive application prospects.« less

Authors:

Cao, Xinyu; Qiu, Xiangshu; Shi, Ning; Ha, Zhuo; Zhang, He; Xie, Yubiao; Wang, Peng; Zhu, Xiangyu; Zhao, Wenxin; Zhao, Guanyu; Jin, Ningyi; Lu, Huijun

Publication Date:

Fri Sep 23 00:00:00 EDT 2022

Sponsoring Org.:

USDOE Office of Electricity (OE), Advanced Grid Research & Development. Power Systems Engineering Research

OSTI Identifier:

1889325

Grant/Contract Number:  

2021YFC2301704; 2020-12M-5-001

Resource Type:

Published Article

Journal Name:

Frontiers in Microbiology

Additional Journal Information:

Journal Name: Frontiers in Microbiology Journal Volume: 13; Journal ID: ISSN 1664-302X

Publisher:

Frontiers Media SA

Country of Publication:

Switzerland

Language:

English