数字农科院2.0

Rapid differentiation of viable and inactivated African swine fever virus by a viability quantitative PCR

文献类型： 外文期刊

作者： Gaocheng Fan;Xiaowei Gao;Hang Xu;Xinying Dong;Hao Song;Yanhui Fu;Jing Li;Yuying Yang;Hua Ji Qiu;Yuzi Luo

作者机构：

关键词： African swine fever virus;EMA;pathogen inactivation;PMA;viability quantitative PCR;viral presence

期刊名称： Veterinary Research

ISSN： 1297-9716

年卷期： 2025 年 56 卷 1 期

页码：

收录情况： SCIE（2025版）

摘要： Real-time quantitative PCR (qPCR) has been widely used for African swine fever virus (ASFV) surveillance, but it cannot differentiate viable from inactivated virions, as inactivated particles often retain intact nucleic acids. This limitation may lead to overestimation of the presence of viruses in tested samples, potentially resulting in unnecessary outbreak responses and increased economic burdens. To address this challenge, a viability qPCR (V-qPCR) assay was established, utilizing nucleic acid intercalating dyes-propidium monoazide (PMA) or ethidium monoazide (EMA), to discriminate viable and inactivated ASFV. These dyes selectively penetrate compromised viral envelopes in inactivated particles, covalently binding viral DNA, and inhibiting PCR amplification. The optimized V-qPCR assay demonstrated a detection limit of 101.5 TCID50/mL for infectious ASFV and effectively discriminated chemically inactivated virus (104.5 TCID50/mL) treated with potassium peroxymonosulfate (PPMS), sodium hydroxide (NaOH), or acetic acid (HAc), showing no amplification signals. In simulated environmental samples (ASFV-spiked swabs from farm walls, rails, feed, floors, urine, and feces), the V-qPCR assay reliably detected inactivation in PPMS-treated samples (walls, rails, urine, feces) and NAOH-treated samples (walls, feces), showing no amplification. In contrast, HAc-inactivated samples yielded residual amplification (Ct ~ 31.5), suggesting superior performance of PPMS and NAOH in complex matrices. Validation using 30 ASFV-positive field environmental swabs revealed three outcomes: (i) 21 samples showed no amplification (confirmed inactivated); (ii) six samples showed similar Ct values before and after heat treatment (ΔCt [Ct(+dye)-Ct(no dye)] < 2.5, indicating inactivation); and (iii) three samples demonstrated detectable Ct values before heat inactivation but no amplification after treatment (confirming the presence of viable virus). This study represents the first application of V-qPCR for field-based assessment of ASFV infectivity, offering a rapid and practical tool to enhance diagnostic accuracy and biosecurity management in pig farms.

分类号：

  • 相关文献

[1]水果PLU编码(PriceLookUpcode)介绍及梨果PLU码汇编. 王文辉,杭博,杜艳民,贾晓辉. 2014

[2]水果PLU编码(Price Look Up code)介绍及梨果PLU码汇编. 王文辉,杭博,杜艳民,贾晓辉. 2014

[3]Isolation and culture of melanocytes from the arctic fox (Alopex lagopus). Bao, Jiarong,Wang, Lei,Wang, Guiwu,Liu, Xueqing,Yang, Fuhe,Bao, Jiarong,Wang, Lei,Wang, Guiwu,Liu, Xueqing,Yang, Fuhe. 2015

[4]PERFORMANCE OF THERMAL PRETREATMENT FOR PATHOGEN INACTIVATION OF FECAL SLUDGE. Yin, Fubin,Dong, Hongmin,Li, Zifu,Wang, Dongling.

[5]Performance of thermal pretreatment and mesophilic fermentation system on pathogen inactivation and biogas production of faecal sludge: Initial laboratory results. Yin, Fubin,Dong, Hongmin,Li, Zifu,Wang, Dongling,Ohlsen, Thomas.

[6]Enhancing pathogen inactivation in pig manure by introducing thermophilic and hyperthermophilic hygienization in a two-stage anaerobic digestion process. Min Lin,Lijuan Ren,Simon Mdondo Wandera,Yi Liu,Renjie Dong,Wei Qiao. 2022

[7]Replication And Virulence In Pigs O.f The First African S wine Fever Virus Isolated In China. Wang, Xijun,Wu, Xinzhou,Bu, Zhigao,Chen, Weiye,Liu, Xing,Wang, Lulu,Bu, Zhigao,Liu, Renqiang,Wang, Jingfei,Zhang, Jiwen,Zhang, Xianfeng,Zhao, Dongming,Li, Fang,Zhang, Jiaoer,Guan, Yuntao,He, Xijun. 2019

[8]Pre-Clinical Evaluation of a Real-Time PCR Assay on a Portable Instrument as a Possible Field Diagnostic Tool: Experiences from the Testing of Clinical Samples for African and Classical Swine Fever Viruses. Liu, L.,Stahl, K.,Belak, S.,Liu, L.,Stahl, K.,Belak, S.,Liu, L.,Luo, Y.,Qiu, H. -J.,Luo, Y.,Qiu, H. -J.,Accensi, F.,Ganges, L.,Rodriguez, F.,Accensi, F.,Rodriguez, F.,Shan, H.,Belak, S..

[9]A comparative analysis on the synonymous codon usage pattern in viral functional genes and their translational initiation region of ASFV. Zhou, Jian-hua,Gao, Zong-liang,Sun, Dong-jie,Ding, Yao-zhong,Zhang, Jie,Liu, Yong-sheng,Stipkovits, Laszlo,Szathmary, Susan,Pejsak, Zygmunt. 2013

[10]Recombinant Newcastle disease virus expressing African swine fever virus protein 72 is safe and immunogenic in mice. Chen, Xinxin,Yang, Jifei,Ji, Yanhong,Liu, Bin,Li, Xiaoyang,Yin, Hong,Zhu, Qiyun,Yin, Hong,Okoth, Edward. 2016

[11]Genome Sequences Derived From Pig A.nd Dried Blood Pig F eed Samples Provide Important Insights Into The Transmission Of African Swine Fever Virus In China In 2018. Wen, XX, He, XJ, Zhang, X, Zhang, XF, Liu, LL, Guan, YT, Zhang, Y, Bu, ZG. 2019

[12]Pre-Clinical Evaluation Of A Real-Time P.cr Assay On A P ortable Instrument As A Possible Field Diagnostic Tool: Experiences From The Testing Of Clinical Samples For African And Classical Swine. Liu, L, Luo, Y, Accensi, F, Ganges, L, Rodriguez, F, Shan, H, Stahl, K, Qiu, HJ, Belak, S. 2017

[13]Roles Of African Swine Fever V.irus Structural Proteins In V iral Infection. Jia, N, Ou, YW, Pejsak, Z, Zhang, YG, Zhang, J. 2017

[14]CD1d facilitates African swine fever virus entry into the host cells via clathrin-mediated endocytosis. Xin Chen,Jun Zheng,Chuanxia Liu,Tingting Li,Xiao Wang,Xuewen Li,Miaofei Bao,Jiangnan Li,Li Huang,Zhaoxia Zhang,Zhigao Bu,Changjiang Weng. 2023

[15]Recombinant porcine interferon cocktail delays the onset and lessens the severity of African swine fever. Pengtao Jiao,Shuchao Wang,Wenhui Fan,He Zhang,Hongyan Yin,Yingli Shang,Hongfei Zhu,Wenjun Liu,Rongliang Hu,Lei Sun. 2023

[16]The African swine fever virus I10L protein inhibits the NF-κB signaling pathway by targeting IKKβ. Xing Chen,Lian Feng Li,Zhong Yuan Yang,Meilin Li,Shuai Fan,Lan Fang Shi,Zi Yu Ren,Xue Jing Cao,Yuhang Zhang,Shichong Han,Bo Wan,Hua Ji Qiu,Gaiping Zhang,Wen Rui He. 2023

[17]Development of a TaqMan-Probe-Based Multiplex Real-Time PCR for the Simultaneous Detection of African Swine Fever Virus, Porcine Circovirus 2, and Pseudorabies Virus in East China from 2020 to 2022. Huaicheng Liu,Jianwen Zou,Rongchao Liu,Jing Chen,Xiaohan Li,Haixue Zheng,Long Li,Bin Zhou. 2023

[18]Development of an indirect ELISA to specifically detect antibodies against African swine fever virus: bioinformatics approaches. Gao Zhan,Shao Jun-Jun,Zhang Guang-Lei,Ge Su-Dan,Chang Yan-Yan,Xiao Lei,Chang Hui-Yun. 2021

[19]Mechanism of interaction between virus and host is inferred from the changes of gene expression in macrophages infected with African swine fever virus CN/GS/2018 strain. Bo Yang,Chaochao Shen,Dajun Zhang,Ting Zhang,Xijuan Shi,Jinke Yang,Yu Hao,Dengshuai Zhao,Huimei Cui,Xingguo Yuan,Xuehui Chen,Keshan Zhang,Haixue Zheng,Xiangtao Liu. 2021

[20]Development and in vivo evaluation of MGF100-1R deletion mutant in an African swine fever virus Chinese strain. Yingnan Liu,Yao Li,Zhenhua Xie,Qingying Ao,Dongdong Di,Wanqi Yu,Lu Lv,Qiuping Zhong,Yingying Song,Xinxin Liao,Qingqing Song,Heng Wang,Hongjun Chen. 2021

作者其他论文 更多>>