Facultad de Ciencias Médicas, Instituto de Virología "Dr. J. M. Vanella", Universidad Nacional de Córdoba, Enfermera Gordillo Gómez S/N, Ciudad Universitaria, 5016, Córdoba, Argentina.
Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Córdoba, Argentina.
Food Environ Virol. 2024 Nov 19;17(1):1. doi: 10.1007/s12560-024-09617-z.
Hepatitis E virus (HEV) is primarily transmitted via the fecal-oral route and is considered an anthropozoonosis. Genotypes with zoonotic potential (mainly HEV-3 and HEV-4) can be transmitted through the consumption of raw or undercooked pork, wild boar, deer meat, or processed products. This study aims to explore methodologies for processing meat samples to establish a protocol for HEV detection in meat. The analysis of pre-analytical conditions involved comparing homogenization with PBS versus TRIzol, comparing tissue disruption methods (ultra-turrax versus mortar and pestle), and assessing nucleic acid extraction techniques (spin columns and magnetic beads) across three types of artificially contaminated meat matrices: pork, salmon (fish-meat), and salami. Each test included a process control virus (PP7) and an HEV transcript. Molecular detection was performed via RT-qPCR. Results indicated that TRIzol provided better recovery rates for homogenization, while spin columns were the most effective option for RNA extraction. Both the ultra-turrax homogenizer and the mortar-pestle methods were effective for pork and fish-meat homogenization, while the use of the UT yielded superior results for salami. HEV recovery rates were 36.7%, 26.3%, and 34.1% for salami, salmon, and pork meat, respectively. In conclusion, we reached a simple and reliable protocol for the detection of RNA-HEV from three meat matrices. This method, which includes homogenization with TRIzol, mechanical tissue disruption, and RNA extraction using spin columns followed by real-time PCR, can be applied in future studies to evaluate HEV prevalence in food sources and contribute to the discussion about HEV detection methodologies.
戊型肝炎病毒（HEV）主要通过粪-口途径传播，被认为是一种人畜共患病。具有人畜共患潜力的基因型（主要是 HEV-3 和 HEV-4）可通过食用生的或未煮熟的猪肉、野猪、鹿肉或加工产品传播。本研究旨在探索处理肉样的方法，以建立检测肉中 HEV 的方法。分析了预分析条件，包括 PBS 与 TRIzol 相比的匀浆、组织破碎方法（超声破碎仪与研钵和杵相比）以及评估核酸提取技术（离心柱和磁珠）在三种人为污染肉基质中的效果：猪肉、三文鱼（鱼肉）和萨拉米香肠。每种测试都包括一个过程控制病毒（PP7）和一个 HEV 转录本。分子检测通过 RT-qPCR 进行。结果表明，TRIzol 提供了更好的匀浆回收率，而离心柱是 RNA 提取最有效的选择。超声破碎仪和研钵-杵法都能有效地用于猪肉和鱼肉的匀浆，而使用超声破碎仪对萨拉米香肠的效果更好。HEV 的回收率分别为萨拉米香肠、三文鱼和猪肉的 36.7%、26.3%和 34.1%。总之，我们为三种肉基质中 RNA-HEV 的检测制定了一个简单可靠的方案。该方法包括用 TRIzol 匀浆、机械组织破碎以及使用离心柱提取 RNA 后进行实时 PCR，可以在未来的研究中应用，以评估食源 HEV 的流行情况，并为 HEV 检测方法的讨论做出贡献。
