Burge Colleen A, Strenge Robyn E, Friedman Carolyn S
School of Aquatic and Fishery Sciences, University of Washington, Box 355020, Seattle, Washington 98195, USA.
Dis Aquat Organ. 2011 Apr 6;94(2):107-16. doi: 10.3354/dao02314.
The ostreid herpesvirus (OsHV-1) and related oyster herpesviruses (OsHV) are associated with world-wide mortalities of larval and juvenile bivalves. To quantify OsHV viral loads in mollusc tissues, we developed a SYBR Green quantitative PCR (qPCR) based on the A-region of the OsHV-1 genome. Reaction efficiency and precision were demonstrated using a plasmid standard curve. The analytical sensitivity is 1 copy per reaction. We collected Crassostrea gigas, C. sikamea, C. virginica, Ostrea edulis, O. lurida, Mytilus galloprovincialis, and Venerupis phillipinarum from Tomales Bay (TB), and C. gigas from Drakes Estero (DE), California, U.S.A., and initially used conventional PCR (cPCR) to test for presence of OsHV DNA. Subsequently, viral loads were quantified in selected samples of all tested bivalves except O. lurida. Copy numbers were low in each species tested but were significantly greater in C. gigas (p < 0.0001) compared to all other species, suggesting a higher level of infection. OsHV DNA was detected with cPCR and/or qPCR and confirmed by sequencing in C. gigas, C. sikamea, C. virginica, O. edulis, M. galloprovincialis, and V phillipinarum from TB and C. gigas from DE. These data indicate that multiple bivalve species may act as reservoirs for OsHV in TB. A lack of histological abnormalities in potential reservoirs requires alternative methods for their identification. Further investigation is needed to determine the host-parasite relationship for each potential reservoir, including characterization of viral loads and their relationship with infection (via in situ hybridization), assessments of mortality, and host responses.
牡蛎疱疹病毒（OsHV-1）及相关的牡蛎疱疹病毒（OsHV）与全球范围内双壳贝类幼虫和幼体的死亡有关。为了量化软体动物组织中的OsHV病毒载量，我们基于OsHV-1基因组的A区域开发了一种SYBR Green定量PCR（qPCR）方法。使用质粒标准曲线证明了反应效率和精密度。分析灵敏度为每个反应1个拷贝。我们从托马莱斯湾（TB）采集了太平洋牡蛎、蚶、美国牡蛎、欧洲平牡蛎、加州岩牡蛎、地中海贻贝和菲律宾蛤仔，并从美国加利福尼亚州德雷克湾（DE）采集了太平洋牡蛎，最初使用常规PCR（cPCR）检测OsHV DNA的存在。随后，对除加州岩牡蛎外的所有测试双壳贝类的选定样本进行了病毒载量定量。在所测试的每个物种中拷贝数都很低，但与所有其他物种相比，太平洋牡蛎中的拷贝数显著更高（p < 0.0001），表明感染水平更高。在来自TB的太平洋牡蛎、蚶、美国牡蛎、欧洲平牡蛎、地中海贻贝和菲律宾蛤仔以及来自DE的太平洋牡蛎中，通过cPCR和/或qPCR检测到OsHV DNA，并通过测序进行了确认。这些数据表明，多种双壳贝类物种可能是TB中OsHV的储存宿主。潜在储存宿主缺乏组织学异常需要采用替代方法来识别它们。需要进一步研究以确定每个潜在储存宿主的宿主-寄生虫关系，包括病毒载量的特征及其与感染的关系（通过原位杂交）、死亡率评估和宿主反应。
