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发挥着关键作用。感染海洋细菌的噬菌体是浮游生物中丰富的成员，通过水平基因转移促进细胞死亡、构建种群多样性和推动基因组进化。感染真核浮游植物的病毒可诱导球藻发生生命周期转换和细胞程序性死亡，是导致浮游植物、浮游植物和浮游植物水华死亡的重要因素。然而，人们对感染硅藻这种最大、最多样化、最高产的藻类之一的病毒知之甚少。目前只有13种硅藻感染病毒的报道，对它们的感染机制、对寄主代谢的影响或田间多样性和动态知之甚少。考虑到硅藻的生态重要性，这是一个显著的知识鸿沟。用克隆病毒感染拟菱形藻多系，可在12-16小时内使宿主完全溶解。PmDNAV是一种单链DNA病毒，衣壳二十面体大小为50 nm。PmDNAV感染任何海洋真核生物感染病毒的最广泛的宿主范围，裂解其他多系列P.多系列菌株，其他种类的伪菱形藻，以及其他硅藻属，包括许多中心硅藻。随着多系列寄主基因组的完成，我们现在有了一个模型系统来研究寄主对病毒感染的反应以及病毒对现场硅藻死亡的潜在影响。本项目的目标是：1.利用已建立的方法，利用各种宿主菌株和田间病毒浓缩物组合，分离和鉴定更多的硅藻病毒。利用RNA-Seq技术，测定三种硅藻(多串拟青霉、彭氏假单胞菌和假单胞菌)在病毒感染过程中的转录图谱。在太平洋西北部以硅藻为主的水域(P线)沿海至开阔海洋断面的四个站确定地表水后营养体，重点是单链DNA和单链RNA病毒的多样性和生物地理学。病毒和宿主基因的表达是病毒感染的诊断，将通过观察培养中对感染的基因组反应来鉴定。这些基因，连同在后生生物中组装的病毒，将与从同一水域已有的真核转录本结合起来，以评估病毒在该领域的活性。智力优势：该项目旨在加强因发现伪菱形藻多系列DNA病毒而启动的模型系统。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，成对的后生微体和后转录体将揭示关于硅藻病毒多样性和功能的新问题，然后可以通过基于培养的受控实验来进一步探索。这项研究将是对硅藻病毒生态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。广泛的影响：硅藻是沿海多产海洋的主要初级生产者，对所有海洋制度的碳输出都有重要贡献。病毒对物种分布和细胞丰度的影响对渔业的健康和生态系统对气候变化的反应具有影响。更具体地说，伪菱形藻水华会对公共健康和经济产生影响。该项目的另一个目标是通过在国家一级为代表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、同行指导和接触该学科，来扩大环境科学研究生的人才库。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.