Ecology of diatom viruses: connecting physiology and field dynamics through host transcriptional responses

概述：病毒在水生生态系统中起着关键作用。感染海洋细菌的噬菌体是浮游生物中丰富的成员，它们通过水平基因转移促进细胞死亡，结构种群多样性并驱动基因组进化。感染真核浮游植物的病毒已被证明可以诱导球石藻的生命周期转换和程序性细胞死亡，并在浮游植物，附着植物和针芽植物的开花死亡率的显着代理。然而，对感染最大，最多样化和最具生产力的藻类之一硅藻的病毒知之甚少。迄今为止，仅报道了13种感染硅藻的病毒，但关于它们的感染机制、对宿主代谢的影响以及在该领域的多样性和动态却知之甚少。考虑到硅藻的生态重要性，这是一个显著的知识差距。用克隆病毒感染多系拟菱形藻可在12-16小时内导致宿主完全裂解。多系毕赤酵母病毒（PmDNAV）是具有50 nm的二十面体衣壳的单链DNA病毒。PmDNAV感染任何海洋真核生物感染病毒的最广泛宿主范围，裂解多系毕赤酵母的其它菌株、假菱形藻的其它物种和硅藻的其它属，包括许多中心硅藻。随着最近完成的宿主，P. multiseries的基因组，我们现在有一个模型系统来研究宿主对病毒感染的反应和病毒对硅藻死亡率的潜在影响。本项目的目标是：1。利用已建立的方法，使用各种宿主菌株和田间病毒浓缩物组合，分离和表征其他硅藻病毒2。使用RNA-Seq确定三种硅藻（多系硅藻、刺硅藻和T.在病毒感染的过程中3.在太平洋西北部以硅藻为主的沃茨（P线）的沿海至开放海洋样带上的四个站点确定地表水后生病毒组，重点是ssDNA和ssRNA病毒的多样性和病毒地理学。将通过观察培养物中对感染的基因组反应来鉴定其表达可诊断病毒感染的病毒和宿主基因。这些基因，沿着与病毒组装在metaviromes，将与真核细胞metatranscriptomes已经从相同的沃茨，以评估病毒的活动在field.Intellectual优点：该项目旨在加强模型系统发起的伪菱形多系列DNA病毒的发现。宿主病毒转录组学将为评估病毒对环境中硅藻群落的影响奠定基础。反过来，配对的元病毒组和元转录组将揭示有关硅藻病毒多样性和功能的新问题，然后可以通过受控的、基于培养的实验进一步探索。这项研究将是对硅藻病毒生态学和功能的首次广泛探索，最终将有助于进一步将病毒和硅藻与全球生物地球化学循环联系起来，揭示复杂的生物相互作用，并为与海洋相关的公共健康提供信息。更广泛的影响：硅藻是生产性沿海海洋中占主导地位的初级生产者，并对所有海洋体系的碳输出做出重大贡献。病毒对物种分布和细胞丰度的影响对渔业健康和生态系统对气候变化的反应具有影响。更具体地说，有公共卫生和经济影响与伪菱形藻水华。该项目的另一个目标是扩大从事环境科学研究生工作的个人的人数，为来自代表人数不足的少数群体的学生提供早期研究经验、持续的财政支助、同侪指导和在国家一级接触该学科。来自代表性不足群体的一名研究生和五名本科生将参与这项研究。

Overview: Viruses play critical roles in aquatic ecosystems. Phages infecting marine bacteria are abundant members of the plankton that contribute to cell mortality, structure population diversity and drive genome evolution though horizontal gene transfer. Viruses infecting eukaryotic phytoplankton have been demonstrated to induce both life cycle switching and programmed cell death in coccolithophorids and be significant agents of mortality in blooms of pelagophytes, haptophytes and raphidophytes. However, much less is known about viruses infecting one of the largest, most diverse and most productive groups of algae, the diatoms. Only thirteen diatom infecting viruses have been reported, and little is known about their mechanisms of infection, effects on host metabolism or diversity and dynamics in the field. This is a remarkable knowledge gap considering the ecological importance of the diatoms. Infection with a clonal virus on Pseudo-nitzschia multiseries can result in complete host lysis within 12-16 hours. The P. multiseries virus (PmDNAV) is a single stranded DNA virus with an icosahedral capsid of 50 nm. The PmDNAV infects the widest host range of any marine eukaryote-infecting virus, lysing other strains of P. multiseries, other species of Pseudo-nitzschia, and other genera of diatoms including many centric diatoms. With the recent completion of the genome of the host, P. multiseries, we now have a model system to investigate the response of the host to viral infection and the potential impacts of viruses on diatom mortality in the field. The objectives of this project are to:1. isolate and characterize additional diatom viruses utilizing established methods, using a variety of host strains and field viral concentrate combinations2. use RNA-Seq to determine the transcriptional profiles of three diatoms (P. multiseries, P. pungens and T. pseudonana) during the course of viral infection3. determine a surface water metavirome at four stations on a coastal to open ocean transect in diatom dominated waters in the Pacific Northwest (line P), with an emphasis on diversity and biogeography of ssDNA and ssRNA viruses. Viral and host genes whose expression is diagnostic of viral infection, will be identified by observing genomic responses to infection in culture. These genes, along with viruses assembled in the metaviromes, will be combined with eukaryotic metatranscriptomes already available from the same waters to assess virus activity in the field.Intellectual Merit: This project seeks to strengthen the model system initiated by the discovery of the Pseudo-nitzschia multiseries DNA virus. The host-virus transcriptomics will lay the groundwork for assessing the impact of viruses on diatom communities in the environment. In turn, the paired metaviromes and metatranscriptomes will reveal new questions about both diatom virus diversity and function that can then be further explored by controlled, culture-based experiments. This research will be the first extensive exploration of diatom virus ecology and function and will ultimately help further connect viruses and diatoms to global biogeochemical cycles, unravel complex organismal interactions, and inform ocean-related public health.Broader Impacts: Diatoms are the dominant primary producers in the productive coastal oceans and contribute significantly to carbon export across all oceanic regimes. Viral impacts on species distributions and cell abundances have implications for health of fisheries and ecosystem responses to climate change. More specifically, there are public health and economic impacts associated with Pseudo-nitzschia blooms. Another goal of this project is to broaden the pool of individuals who pursue graduate work in environmental sciences by providing students from underrepresented minority groups early research experience, sustained financial support, peer mentorship and exposure to the discipline at a national level. A graduate student and five undergraduates from underrepresented groups will be involved in the research.