Establishment and application of a TaqMan-based one-step real-time RT-PCR for the detection of novel goose-origin astrovirus

Dan Yin; Jing Yang; Jiajun Tian; Dalin He; Yi Tang; Youxiang Diao

doi:10.1016/j.jviromet.2019.113757

Establishment and application of a TaqMan-based one-step real-time RT-PCR for the detection of novel goose-origin astrovirus

J Virol Methods. 2020 Jan:275:113757. doi: 10.1016/j.jviromet.2019.113757. Epub 2019 Oct 24.

Authors

Dan Yin¹, Jing Yang¹, Jiajun Tian¹, Dalin He¹, Yi Tang², Youxiang Diao³

Affiliations

¹ College of Animal Science and Technology, Shandong Agricultural University, 61 Daizong Street, Tai'an, Shandong Province, 271018, China; Shandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, Tai'an, Shandong, 271018, China; Shandong Provincial Engineering Technology Research Center of Animal Disease Control and Prevention, Tai'an, Shandong, 271018, China.
² College of Animal Science and Technology, Shandong Agricultural University, 61 Daizong Street, Tai'an, Shandong Province, 271018, China; Shandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, Tai'an, Shandong, 271018, China; Shandong Provincial Engineering Technology Research Center of Animal Disease Control and Prevention, Tai'an, Shandong, 271018, China. Electronic address: tyck288@163.com.
³ College of Animal Science and Technology, Shandong Agricultural University, 61 Daizong Street, Tai'an, Shandong Province, 271018, China; Shandong Provincial Key Laboratory of Animal Biotechnology and Disease Control and Prevention, Tai'an, Shandong, 271018, China; Shandong Provincial Engineering Technology Research Center of Animal Disease Control and Prevention, Tai'an, Shandong, 271018, China. Electronic address: yxdiao@126.com.

PMID: 31669331
DOI: 10.1016/j.jviromet.2019.113757

Abstract

The outbreak of an infectious disease characterized by severe symptom of gout has set great threat to several major goose-producing regions in China since December 2016. The causative agent for the novel infection has been identified was a novel goose-origin astrovirus (GoAstV). Lack of effective detection methods indeed hinders further research, as well as prevention and control of GoAstV. Keep this in mind, a TaqMan-based one-step real-time quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) assay for rapid detection of GoAstV was developed. Primers and probe were targeting the capsid protein gene sequence (ORF2). The method is capable of detecting quite low number of targeting nucleic acid as low as 10 copies/μL. What's more, it is also of great specificity and repeatability for GoAstV detection. No cross-activity was found with other goose-origin viruses. The assay had excellent intra-assay and inter-assay repeatability with the coefficient of variation (CV) value from 0.48% to 0.99%. A total of 340 GoAstV specimens from different regions of China were used in this study to verify the feasibility and effectiveness of this method in clinical diagnosis. The results indicated that qRT-PCR is a highly sensitive, specific and repeatable method for quantitative detection of GoAstV, which can be used to detect this virus, thereby facilitating epidemiological investigations of gout in goslings.

Keywords: Application; Detection; Novel goose-origin astrovirus; TaqMan probe; qRT-PCR.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Astroviridae Infections / veterinary*
Astroviridae Infections / virology
Avastrovirus / genetics
Avastrovirus / isolation & purification*
China
DNA Primers / genetics
Geese / virology*
Kidney / virology
Real-Time Polymerase Chain Reaction / methods*
Sensitivity and Specificity
Viral Load

Substances

DNA Primers