Development of a triplex quantitative reverse transcription-polymerase chain reac-tion for the detection of porcine epidemic diarrhea virus,transmissible gastroenter-itis virus,andporcine delta coronavirus
10.16303/j.cnki.1005-4545.2025.05.03
- VernacularTitle:猪流行性腹泻病毒、猪传染性胃肠炎病毒和猪δ冠状病毒三重RT-qPCR检测方法的建立
- Author:
Qianlin CHEN
1
;
Shaomei LI
;
Yifan ZHANG
;
Hao MU
;
Mingni LIU
;
Liu YANG
;
Qingyong GUO
;
Lizhi FU
Author Information
1. 国家生猪技术创新中心疫病防控研究院,重庆荣昌 402460;新疆农业大学动物医学学院,新疆乌鲁木齐 830052
- Publication Type:Journal Article
- Keywords:
porcine epidemic diarrhea virus;
transmissible gastroenteritis virus;
porcine delta corona-virus;
RT-qPCR;
detection methods
- From:
Chinese Journal of Veterinary Science
2025;45(5):905-912
- CountryChina
- Language:Chinese
-
Abstract:
Swine enteric coronaviruses(SeCoV),such as porcine epidemic diarrhea virus(PEDV),transmissible gastroenteritis virus(TGEV),and porcine delta coronavirus(PDCoV),cause severe diarrhea in piglets,resulting in substantial losses in pig farming.In this study we establish a triple fluorescence reverse transcription-quantitative PCR(RT-qPCR)method for the simultaneous de-tection of PEDV,TGEV,and PDCoV.The specific primers and probes for each target virus were designed based on conserved sequences from the PEDV M gene,the TGEV ORF 1b gene,and the PDCoV ORF 1b gene respectively.Following the optimization of parameters and conditions,a triple RT-qPCR method was successfully established to simultaneously detect PEDV,TGEV,and PD-CoV.The developed assay exhibits strong specificity for these three pathogens without any cross-reaction with other common porcine viruses like CSFV,PCV2,PoRVA,PRV,and PRRSV.The de-tection limit of linear templates for pTOPO-PEDV 128,pTOPO-TGEV 116,and pTOPO-PDCoV 125 recombinant plasmids were 16.835,17.610 and 17.020 copies/μL,respectively.The intra group and inter group coefficients of variation were less than 5％,with no significant differences observed(P＞0.05).Moreover,the detection consistency rate of the developed RT-qPCR was compared with standard method and showed 100％agreement.Out of 35 small intestine tissue samples,17 tested positive for PEDV,resulting in a positive rate of 48.57％(17/35).The tests for TGEV and PDCoV yielded negative results,and no mixed infections were detected.Based on the above results,the tri-ple RT-qPCR method established is specific,sensitive,stable,and rapid,and can be used for clinical detection and differential diagnosis of PEDV,TGEV,and PDCoV simultaneously,providing a method for the detection and epidemiological investigation of porcine diarrhea coronaviruses.
