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通过聚合酶链反应(PCR)检测海水中的牡蛎疱疹病毒1型(OsHV-1)DNA:生物测定中水质参数的影响

Detection of Ostreid Herpesvirus 1 (OsHV-1) DNA in seawater by PCR: influence of water parameters in bioassays.

作者信息

Vigneron Vassilia, Solliec Gaëlle, Montanié Hélène, Renault Tristan

机构信息

Laboratoire de Biologie et Environnement Marins, EA 3168, Université de La Rochelle, Avenue Michel Crépeau, 17042 La Rochelle, France.

出版信息

Dis Aquat Organ. 2004 Nov 23;62(1-2):35-44. doi: 10.3354/dao062035.

Abstract

Since 1991, herpesvirus infections have been reported among larvae and juveniles of various bivalves. Most of the studies focused on detection of viral infections of economically important species. However, the persistence of bivalve herpesviruses in the marine environment is poorly documented. The present study concerns the role of seawater parameters in Ostreid Herpesvirus 1 (OsHV-1) detection by polymerase chain reaction (PCR). Viral DNA extracted from purified particles or virions present in infected oyster larvae were detected by PCR after storage in different media at different temperatures. The lowest detection threshold was found using distilled water or Tris EDTA buffer. In seawater, the threshold was higher. The use of sterile media permitted detection of viral DNA stored over a longer period. Storage temperature also had a significant influence on detection, with lower temperatures promoting DNA detection over a longer period. In summary, water parameters such as temperature influenced detection of OsHV-1 DNA by PCR. However, the PCR technique may also be successfully applied to samples in natural seawater. Indeed, the PCR technique permitted detection of naked viral DNA at 100 ng l(-1) in seawater in bioassays.

摘要

自1991年以来,已报道多种双壳贝类的幼虫和幼体中存在疱疹病毒感染。大多数研究集中在经济重要物种的病毒感染检测上。然而,双壳贝类疱疹病毒在海洋环境中的持久性记录较少。本研究关注海水参数在通过聚合酶链反应(PCR)检测牡蛎疱疹病毒1型(OsHV-1)中的作用。从感染牡蛎幼虫中存在的纯化颗粒或病毒粒子中提取的病毒DNA,在不同温度下于不同培养基中储存后,通过PCR进行检测。使用蒸馏水或Tris EDTA缓冲液时检测阈值最低。在海水中,阈值更高。使用无菌培养基可使储存较长时间的病毒DNA得以检测。储存温度对检测也有显著影响,较低温度可使DNA在更长时间内被检测到。总之,诸如温度等水参数会影响通过PCR检测OsHV-1 DNA。然而,PCR技术也可成功应用于天然海水中的样本。事实上,在生物测定中,PCR技术能够检测出海水中浓度为100 ng l(-1)的裸露病毒DNA。

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