Collaborative Proposal: Decoding Virus Leviathans

巨型病毒的基因组长度一般超过280，000个碱基对（bp），目前只有不到20个完整的病毒基因组序列在这个范围内。对这一组病毒的粗略分析立即揭示了各种各样的病毒，包括痘病毒、藻DNA病毒（藻类病毒）、疱疹病毒、噬菌体和瘤病毒（虾白色斑点综合征病毒）。迄今为止最大的是多食阿米巴拟病毒，直径为750 nm，基因组为120万bp。来自这些巨型病毒的基因组序列数据可以帮助澄清它们的分类位置。这些巨大的病毒拥有新的和典型的非病毒基因，其中许多编码中央细胞功能和代谢途径。通过流式细胞仪的初步观察和对全球海洋采样数据库的分析表明，巨型病毒在水生环境中极为丰富，但被有偏见的采样技术所忽视。大多数病毒研究着眼于通过0.2微米过滤器的物质。直径达0.75微米的巨型病毒将被这些过滤器捕获并逃避检测。W.威尔逊（毕格罗海洋科学实验室）和S。Wilhelm（Univ Tenn）认为，巨大的海洋病毒无处不在，种类繁多，并含有大量新的遗传信息。该项目正在对目前收集的四种藻类病毒和从海水中分离出的另外16种巨型病毒进行测序。环境中的病毒正在通过采用单细胞分选技术进行分离和测序。一旦分选，巨型病毒通过多重置换扩增（MDA）扩增以产生巨型病毒单扩增基因组（GV-SAG）用于下游测序。为了完成这项工作，PI们使用了最先进的流式细胞仪（Bigelow）和序列分析设备（田纳西大学/橡树岭国家实验室），以及合作者Jean-Michel Claverie教授的生物信息学专业知识，他是巨型病毒研究的世界领导者。这项研究正在一个非正式的Café Scientific计划中进行讨论，该计划旨在向公众传授最新的科学思想。PI还将一个3个数量级（病毒、脂质/蛋白质DNA）模块纳入凯勒-毕格罗实验室数量级（BLOOM）计划，该计划旨在向缅因州高中生传授海洋生态系统知识。PI正在与田纳西大学（UT）的新闻学院合作，将本科生和研究生新闻专业的学生带入实验室，为公众记录这项研究。

Giant viruses are loosely defined as having a genome greater than 280,000 base pairs (bp) and currently there are less than 20 full virus genome sequences in this size range. A cursory analysis of this group immediately reveals a diverse range of viruses including poxviruses, phycodnaviruses (algal viruses), herpesviruses, a bacteriophage and nimaviruses (shrimp white spot syndrome virus). The largest to date is the Acanthamoeba polyphaga Mimivirus at 750 nm diameter with a 1.2-million bp genome. Genome sequence data from these giant viruses can help clarify their taxonomic position. These giant viruses possess novel and typically non-virus genes, many of which encode central cellular functions and metabolic pathways. Preliminary observations by flow cytometry and analysis of the Global Ocean Sampling (GOS) database suggest that giant viruses are extremely abundant in aquatic environments and have been overlooked by biased sampling techniques. Most virus studies look at what passes through a 0.2 micrometer filter. Giant viruses, up to 0.75 micrometers, will be caught on these filters and evade detection. The central hypotheses of this project by Drs. W. Wilson (Bigelow Lab for Ocean Sciences) and S. Wilhelm (Univ Tenn) are that giant marine viruses are ubiquitous, diverse and contain large amounts of novel genetic information. Four algal viruses from current collections and 16 additional giant viruses isolated from seawater are being sequenced in this project. The environmental viruses are being isolated and sequenced by adapting single cell sorting techniques. Once sorted, giant viruses are being amplified by multiple displacement amplification (MDA) to generate Giant Virus Single Amplified Genomes (GV-SAGs) for downstream sequencing. To accomplish this work, the PIs are using state-of-the-art facilities for flow cytometry (Bigelow) and sequence analysis (University of Tennessee/Oak Ridge National Laboratory) coupled with bioinformatics expertise from collaborator Professor Jean-Michel Claverie, a world-leader in giant virus research.This research is being discussed during an informal Café Scientifique program designed to teach the public about the latest ideas in science. The PIs are also incorporating a 3-orders of magnitude (virus, lipids/proteins & DNA) module into the Keller-Bigelow Laboratory Orders Of Magnitude (BLOOM) program, designed to teach Maine high school students about marine ecosystems. The PIs are partnering with the School of Journalism at the University of Tennessee (UT) to bring undergraduate and graduate journalism students into the lab to document this research for the general public.