Collaborative Research: Diversity and Biogeography of Marine Oligotrich and Choreotrich Ciliates Assessed by Morphological and Molecular Markers

海洋中的微型浮游动物组合通常由寡毛类和舞毛类纤毛虫主导。纤毛虫群落是多样和动态的，在短时间尺度上丰度和种类组成变化很快。如果我们要将一个纤毛虫群落或几个养殖物种的测量结果(例如放牧、生长)外推到区域或全球食物网模型，了解纤毛虫群落多样性的性质和影响至关重要。此外，这些纤毛虫在空间和时间尺度上的种间和种内遗传变异性的程度是未知的。最后，仅对形态标记的分析就引起了研究人员之间的相当大的争论，他们认为所有的纤毛虫都是世界性的，还有一些人认为地理上孤立的纤毛虫种群之间的基因流动有限。解决所有这些问题的一个关键是量化种群内部和种群之间的分子标记和形态之间的对应关系。然而，目前这些生态重要群体的分子数据还很稀少。本项目结合分子和形态方法阐明了编毛虫亚纲和寡毛亚纲海洋纤毛虫的多样性。这一大群浮游食草动物构成了浮游植物和细菌生产以及更高消费者之间的重要营养联系。本研究的重点是收集编毛类和寡毛类自然种群的DNA序列数据，并将这些序列与传统方法确定的形态种相关联。一种技术组合将应用于从自然群落中手工挑选的细胞，以评估从几米到数千公里的范围内的多样性，并对新英格兰南部海岸和英国西海岸的纤毛虫进行密集采样。种内遗传变异性也将被量化为种群内在短时间和空间尺度上的变化。这项研究有两个目标：1)利用形态和分子标记，量化从近岸环境中采集的海洋寡毛类和编毛类物种的一个子集的多样性；2)评估几种丰富的海洋寡毛类和编毛类纤毛虫在短空间和时间尺度上的种内变异模式。为了这两个目标，我们将挑选单个细胞进行遗传分析，并保存下来进行物种鉴定。对于目标一，我们将比较小亚基核糖体基因序列。对于第二个目标，将分析更多的可变标记，包括核糖体基因的内部转录间隔区(ITS1和ITS2)以及组蛋白H3和H4之间的中间序列(H3H4IS)。结合分子和形态方法，这项研究将产生关于寡毛类和编毛类种群遗传多样性的全面数据。这些数据是将时间和空间采样的个别物种丰度信息与整个纤毛虫群落联系起来的重要第一步。此外，这项工作将提供第一次使用分子数据进行海洋纤毛虫特有的测试，并将培训研究生相关的形态和分子技术。由此产生的数据将对有关全球原生生物多样性和生物地理的问题产生更广泛的影响，对于充分探讨纤毛虫在海洋系统的能量流动和营养循环中的作用是必要的。

Microzooplankton assemblages in the sea are often dominated by oligotrich and choretrich ciliates. Ciliate communities are diverse and dynamic, with rapid changes in abundance and species composition over short time scales. Understanding the nature and implications of diversity in ciliate communities is critical if we are to extrapolate measurements (e.g. grazing, growth) obtained with one ciliate community or a few species in culture to regional or global models of food webs. In addition, the degree of inter- and intraspecific genetic variability across spatial and temporal scales is unknown for these ciliates. Finally, analysis of morphological markers alone has generated considerable debate between researchers who argue that all ciliates are cosmopolitan versus those who believe that there is limited gene flow between geographically isolated ciliate populations. One key to addressing all of these issues is to quantify the correspondence between molecular markers and morphology within and between populations. At present, however, molecular data are sparse for these ecologically important groups. This project combines molecular and morphological methods to elucidate the diversity of marine ciliates in the subclasses Choreotrichia and Oligotrichia (Cl: Spirotrichea). This large group of planktonic grazers comprises an important trophic link between phytoplankton and bacterial production and higher consumers. The main focus of the study is to collect DNA sequence data from natural populations of choreotrichs and oligotrichs, and to relate these sequences to morphospecies determined by traditional methods. A combination of techniques will be applied to handpicked cells from natural assemblages to evaluate diversity across a range of scales from meters to thousands of kilometers, with intensive sampling of ciliates along the southern coast of New England and the western coast of the United Kingdom. Intraspecific genetic variability will also be quantified as changes within populations over short temporal and spatial scales. The study has two objectives: 1) to quantify diversity within a subset of marine oligotrich and choreotrich species sampled from nearshore environments using both morphology and molecular markers, and 2) to assess patterns of intraspecific variation on short spatial and temporal scales in a few abundant species of oligotrich and choreotrich ciliates. For both objectives, individual cells will be picked for genetic analysis and preserved for species identification. For objective one, small subunit ribosomal gene sequences will be compared. For the second objective, more variable markers will be analyzed, including the internally transcribed spacer regions (ITS1 and ITS2) of the ribosomal genes and an intervening sequence between histones H3 and H4 (H3H4IS). Combining molecular and morphological approaches, this study will produce comprehensive data on genetic diversity of populations of oligotrichs and choreotrichs. These data are an essential first step for connecting information on individual species abundances sampled across time and space to whole communities of ciliates. In addition, this work will provide the first test using molecular data on marine ciliate endemism and will train graduate students in relevant morphological and molecular techniques. The resulting data will have broader implications for questions regarding global protist diversity and biogeography and will be necessary to fully explore the role of ciliates in energy flow and nutrient cycling in marine systems.