验证用于检测新鲜猪肉渗出物中非洲猪瘟病毒的实时 PCR 法。

IF 2.2 4区 医学 Q3 BIOCHEMICAL RESEARCH METHODS
Marta Cresci , Daria Di Sabatino , Florica Barbuceanu , Paula Tamba , Razvan Motiu , Monica Motiu , Florin Manita , Giacomo Vincifori , Eugenia Ciarrocchi , Barbara Bonfini , Ottavio Portanti , Alessio Lorusso , Doru Hristescu , Paolo Calistri
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引用次数: 0

摘要

非洲猪瘟病毒(ASFV)是非洲猪瘟(ASF)的病原体,这种疾病对家猪、野猪的健康、福利和生产都有不利影响。ASF 的实验室确认基于对血液、血清和器官样本的分析。然而,检测这些样本并不总是方便、经济或可行的。本研究描述了一种基于 PCR 的检测方法的验证过程,该方法以 p72 基因的一部分为靶标,用于从死于 ASFV 的猪的肉类渗出物样本中对 ASFV 进行分子检测。更具体地说,我们研究了一种实时 PCR 检测方法的能力,该方法可以检测从罗马尼亚确诊疫情的 55 头 ASFV 阳性猪和野猪以及从阿布鲁佐地区(意大利)采集的 73 头 ASFV 阴性且定期屠宰的健康猪的膈肌和相应脾脏中获得的肉类渗出物中的 ASFV DNA。该检测能在两种样本中检测到病毒 DNA,脾脏样本的 Ct 值(平均值=21.11,中位值=20.61)低于肉类渗出物样本(平均值=23.08,中位值=22.40)。然而,Ct 值的分布彼此密切相关(R2= 0.83,P-1 稀释可检测出 90% 的阳性样本,而 10-2 稀释则会将更多污染样本的可检测性降低到 78%。由于肉类渗出物可以很容易地从肌肉中获得,而且考虑到这种检测方法可能会用于集中样本,因此它可以作为一种工具来帮助调查 ASV 的传播。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Validation of a real-time PCR assay for the detection of African swine fever virus in fresh pork meat juice

African swine fever virus (ASFV) is the etiological agent of African swine fever (ASF), a disease with detrimental effects on the health, welfare, and production of domestic and wild pigs. The ASF laboratory confirmation is based on the analysis of blood, serum and organ samples. However, testing these samples could not be always convenient, economically feasible or possible. This study describes the validation process of a PCR-based assay targeting a portion of p72 gene, used for the molecular detection of ASFV, from meat juice samples obtained from pigs succumbed to ASFV. More specifically, we investigated the capability of a real-time PCR assay to detect ASFV DNA in meat juices obtained from the diaphragmatic muscle along with the correspondent spleens of 55 ASFV-positive pigs and wild boars sampled from confirmed outbreaks in Romania and from 73 ASFV-negative and regularly slaughtered healthy pigs collected in the Abruzzo region (Italy). The test was able to detect viral DNA in both types of samples, with lower Ct values in spleens (mean=21.11, median=20.61) than meat juices (mean=23.08, median=22.40). However, distributions of Ct values were strongly correlated each other (R2= 0.83, P<0.001). Considering the distribution of the observed Ct values in the 55 positive meat juice samples, a 1:10 dilution would be able to detect 90 % of positive samples, whereas a 1:100 dilution would reduce the detectability to 78 % of more contaminated samples. As meat juice could be obtained easily from muscles and considering the potential use of this test on pooled samples, it could represent a tool to aid the investigation of ASFV spread.

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来源期刊
CiteScore
5.80
自引率
0.00%
发文量
209
审稿时长
41 days
期刊介绍: The Journal of Virological Methods focuses on original, high quality research papers that describe novel and comprehensively tested methods which enhance human, animal, plant, bacterial or environmental virology and prions research and discovery. The methods may include, but not limited to, the study of: Viral components and morphology- Virus isolation, propagation and development of viral vectors- Viral pathogenesis, oncogenesis, vaccines and antivirals- Virus replication, host-pathogen interactions and responses- Virus transmission, prevention, control and treatment- Viral metagenomics and virome- Virus ecology, adaption and evolution- Applied virology such as nanotechnology- Viral diagnosis with novelty and comprehensive evaluation. We seek articles, systematic reviews, meta-analyses and laboratory protocols that include comprehensive technical details with statistical confirmations that provide validations against current best practice, international standards or quality assurance programs and which advance knowledge in virology leading to improved medical, veterinary or agricultural practices and management.
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