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利用磁珠快速捕获和检测太平洋牡蛎和海水中的牡蛎疱疹病毒 1 型。

Rapid capture and detection of ostreid herpesvirus-1 from Pacific oyster Crassostrea gigas and seawater using magnetic beads.

机构信息

IRTA, Ctra., Sant Carles de la Ràpita, Tarragona, Spain.

出版信息

PLoS One. 2018 Oct 3;13(10):e0205207. doi: 10.1371/journal.pone.0205207. eCollection 2018.

Abstract

Ostreid herpesvirus-1 (OsHV-1) has been involved in mass mortality episodes of Pacific oysters Crassostrea gigas throughout the world, causing important economic losses to the aquaculture industry. In the present study, magnetic beads (MBs) coated with an anionic polymer were used to capture viable OsHV-1 from two types of naturally infected matrix: oyster homogenate and seawater. Adsorption of the virus on the MBs and characterisation of the MB-virus conjugates was demonstrated by real-time quantitative PCR (qPCR). To study the infective capacity of the captured virus, MB-virus conjugates were injected in the adductor muscle of naïve spat oysters, using oyster homogenate and seawater without MBs as positive controls, and bare MBs and sterile water as negative controls. Mortalities were induced after injection with MB-virus conjugates and in positive controls, whereas no mortalities were recorded in negative controls. Subsequent OsHV-1 DNA and RNA analysis of the oysters by qPCR and reverse transcription qPCR (RT-qPCR), respectively, confirmed that the virus was the responsible for the mortality event and the ability of the MBs to capture viable viral particles. The capture of viable OsHV-1 using MBs is a rapid and easy isolation method and a promising tool, combined with qPCR, to be applied to OsHV-1 detection in aquaculture facilities.

摘要

牡蛎疱疹病毒 1(OsHV-1)已被证实与世界各地太平洋牡蛎(Crassostrea gigas)的大规模死亡事件有关,给水产养殖业造成了重大的经济损失。在本研究中,使用带负电荷聚合物涂层的磁珠(MBs)从两种天然感染基质:牡蛎匀浆和海水中捕获有活力的 OsHV-1。通过实时定量 PCR(qPCR)证明了病毒在 MBs 上的吸附和 MB-病毒缀合物的特性。为了研究捕获病毒的感染能力,将 MB-病毒缀合物注入到幼稚的牡蛎苗的闭壳肌中,用未带 MBs 的牡蛎匀浆和海水作为阳性对照,以及裸 MBs 和无菌水作为阴性对照。注射 MB-病毒缀合物和阳性对照后会引起死亡,而阴性对照则没有记录到死亡。随后通过 qPCR 和反转录 qPCR(RT-qPCR)对牡蛎进行 OsHV-1 DNA 和 RNA 分析,分别证实了病毒是导致死亡事件的原因,以及 MBs 捕获有活力的病毒颗粒的能力。使用 MBs 捕获有活力的 OsHV-1 是一种快速简便的分离方法,结合 qPCR 是在水产养殖设施中检测 OsHV-1 的有前途的工具。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/1d39/6169968/a9fda6252c79/pone.0205207.g001.jpg

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