Linking microscopy-based identities to molecular identities for problem- or problematic protozoa.

将基于显微镜的身份与问题或有问题的原生动物的分子身份联系起来。

• 批准号: RGPIN-2014-06144
• 负责人: Simpson, Alastair
• 金额: $ 1.97万
• 依托单位: Dalhousie University
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2017
• 资助国家: 加拿大
• 起止时间: 2017-01-01 至 2018-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=632122
• 关键词: Linking; microscopy; based; identities; molecular

Most of the major groups of complex life on Earth are not animals, fungi, plants or algae, but are instead varieties of free-living protozoa. These single celled predators are ecologically important because are often the most important consumers of bacteria and single-celled algae (which form the base of the ocean ecosystem). Some can also cause disease in animals (see below). This application is to support three distinct projects on the biodiversity, distribution and/or evolution of neglected protozoa. The projects all involve connecting DNA data to microscopy-based identification. Project 1: Predatory (phagotrophic) euglenids are important protozoa in sediments, and are of evolutionary interest, but have been difficult to study because there are very few cultures. For example, there are DNA data from <5% of the 100s of described species. Also, they have rapidly evolving gene sequences, which makes them difficult to detect by environmental sequencing (the major tool used to determine which microorganisms are present in a habitat). We recently developed methods for obtaining DNA sequences from uncultured, single euglenid cells that we have identified by microscopy. We will combine this with new technologies to generate multi-gene profiles (’barcodes’) for single euglenids. We will profile about 200 cells from marine and freshwater sediments in Atlantic and West Coast Canada, and Korea. We will use this dataset to estimate a detailed evolutionary history for euglenids that includes all major subtypes (this has been impossible before), and to document the genetic diversity within ‘species’ of predatory euglenids. It will also be used to optimize DNA-based survey methods to better detect the biodiversity of euglenids that is actually present in natural samples. Project 2: Our prior studies show that extremely salty habitats (with >5 times more salt than seawater) harbor many protozoa that are unique to this environment. However, almost all such studies used material from artificial or short-lived habitats (e.g. solar salterns or tiny pools). We will survey the protozoa from the large permanent Great Salt Lake, examining water and shallow sediment samples. This survey will use light microscopy, single cell isolations, and culturing, along with an environmental gene-sequence survey. This will test whether the collection of organisms already cultured from artificial/ephemeral systems represents well the culturable and unculturable diversity from very salty systems in general. We will also be able to compare to species inventories inferred for the strange ‘deep hypersaline anoxic basins’ on the Mediterranean Sea floor, to determine how unusual they actually are. Project 3: Kelp forests are very productive habitats that are important nurseries for fish. Grazing by sea urchins turns large areas of kelp forest on the Atlantic Coast of Nova Scotia into aptly-named ‘barrens’ . The sea urchin population levels are controlled by a disease-causing protozoan, Paramoeba invadens, which is a facultative parasite (i.e. it can also live and grow outside the host). The disease is very temperature-dependent and strikes in late summer, but it is not known where the protozoan comes from, especially how it over-winters. We will develop sensitive DNA-based methods to detect the Paramoeba, and use these to try to detect it in different environmental water column and sediment samples over time. We will also explore whether there are different genetic strains of the protozoan, found in different locations, or that differ in lethality to sea urchins. This data is needed for making accurate predictions of its future extent and influence on kelp forests in the Atlantic region (and thus the ecosystem services they provide) under climate change.

地球上大多数主要的复杂生命群体不是动物、真菌、植物或藻类，而是各种自由生活的原生动物。这些单细胞捕食者在生态学上很重要，因为它们通常是细菌和单细胞藻类（构成海洋生态系统的基础）的最重要消费者。有些也会引起动物疾病（见下文）。这个应用程序是支持三个不同的项目的生物多样性，分布和/或被忽视的原生动物的进化。这些项目都涉及将DNA数据与基于显微镜的识别相连接。项目一：捕食性（食食性）裸藻是沉积物中重要的原生动物，具有进化意义，但由于培养物很少，研究起来比较困难。例如，在100多个被描述的物种中，有不到5%的DNA数据。此外，它们具有快速进化的基因序列，这使得它们难以通过环境测序（用于确定栖息地中存在哪些微生物的主要工具）进行检测。我们最近开发了从未经培养的单个眼虫细胞中获得DNA序列的方法，这些细胞是我们通过显微镜鉴定的。我们将联合收割机与新技术相结合，为单一的裸藻产生多基因图谱（“条形码”）。我们将从大西洋和加拿大西海岸以及韩国的海洋和淡水沉积物中分析大约200个细胞。我们将使用这个数据集来估计包括所有主要亚型的裸藻的详细进化历史（这在以前是不可能的），并记录捕食性裸藻“物种”内的遗传多样性。它还将用于优化基于DNA的调查方法，以更好地检测天然样品中实际存在的裸藻生物多样性。项目二：我们之前的研究表明，极端咸的栖息地（比海水多5倍以上的盐）拥有许多这种环境所特有的原生动物。然而，几乎所有这些研究都使用了来自人工或短期栖息地的材料（例如太阳盐沼或小水池）。我们将调查大型永久性大盐湖的原生动物，检查水和浅层沉积物样品。这项调查将使用光学显微镜，单细胞分离和培养，沿着环境基因序列调查。这将测试已经从人工/短命系统培养的生物体集合是否代表了一般高盐系统的可培养和不可培养的多样性。我们还将能够与地中海海底奇怪的“深高盐缺氧盆地”推断的物种清单进行比较，以确定它们实际上有多不寻常。项目3：海带林是非常多产的生境，是鱼类的重要苗圃。在新斯科舍省的大西洋海岸，海胆的放牧使大片的海藻森林变成了名副其实的“不毛之地”。海胆的数量水平由一种致病的原生动物Paramoeba invadens控制，Paramoeba invadens是一种兼性寄生虫（即它也可以在宿主外生活和生长）。这种疾病是非常依赖于温度和罢工在夏末，但目前还不知道原生动物来自哪里，特别是它如何越冬。我们将开发灵敏的基于DNA的方法来检测Paramoeba，并使用这些方法来尝试随着时间的推移在不同的环境水柱和沉积物样本中检测它。我们还将探索是否有不同的原生动物基因株，发现在不同的位置，或不同的致命海胆。需要这些数据来准确预测其未来的范围和对大西洋地区海带森林的影响（以及它们提供的生态系统服务）。