Biodiversity, phylogeny, and ecology of large naked ramose/reticulose amoebae: barely studied but diverse and common components of many ecosystems

分子生物学技术和分析的最新进展极大地推动了微生物真核生物(原生生物)的进化和生态学研究。因为它们的外表往往不能反映它们的进化关系，通常只有当我们同时拥有形态和遗传数据并能够使用系统发育分析来重建它们的进化关系时，才能正确地理解原始类群。最近，许多原生生物类群都是这样被阐明的。此外，相关的分子技术已经被用来表明，在大多数环境中，原生生物的遗传多样性是巨大的--等待发现的生物比形态上已知的要多得多。然而，有一组原生生物在很大程度上没有得到基于培养和分子研究的结果，那就是大的、无壳的阿米巴，它们的细胞具有长的、通常是分枝的延伸(分枝伪足)，其中一些融合形成大小不一的细胞质‘网络’(网足)。我们称它们为NRRA-裸枝/网状阿米巴。NRRA可能几乎是静止的，附着在表面或在土壤或沉积物颗粒之间延伸伪足，或者它们可能是活跃的捕食者，穿过表面或穿过颗粒物质寻找猎物。培养的NRRA并不多，因为它们很难生长，但所有已知的菌株都以广泛的食物来源为食--其他原生动物、单细胞和多细胞藻类、许多类型的真菌，包括那些已知会导致农作物或其他植物疾病的真菌，甚至还有线虫和轮虫等小动物。有些细菌可能会攻击更大的生物体。因此，NRRA很可能是生物防治的重要因素，影响其他微生物的活动，包括致病微生物和参与分解和营养循环的微生物。NRRA仍然鲜为人知，因为它们微妙的细胞形状意味着它们经常被采样技术摧毁。它们需要很长时间才能出现在实验室的环境样本中，而且经常需要特别的(但通常是未知的！)生长的条件。此外，分子研究相对较少检测到它们，因为用于此类研究的它们的基因版本往往与许多其他生物如此不同，以至于标准的分子探针无法检测到它们。我们项目的目的是在广泛的生境中发现更多的NRRA，并描述它们的形态和生命周期，并测量它们的食欲有多大--包括它们饮食的多样性和它们吃的量(包括已知会导致高等植物疾病的真菌和其他有机体)。我们将对NRRA细胞的基因进行测序，以了解它们的进化关系和历史。我们将测量它们在许多不同环境条件下的多样性(有多少物种，尽管这在许多原生动物中通常是一个不清楚的概念)。为了进一步探讨这一问题，我们将与三个研究项目合作，这三个项目在1)森林土壤、2)农业土壤与未耕作和未耕作土壤以及3)淡水湖上进行长期实验。这些实验已经建立了很好的抽样策略和从样本站点收集的其他数据，这将为我们关于NRRA的新数据提供一个非常有用的背景。通过对所有三个实验的采样点进行密集采样，我们将调查NRRA多样性和丰度的变化是否与其他微生物(以及小动物和植物)种群的变化相关。如果他们这样做了，我们将为这种相关性提出可能的原因，这可以在未来的研究中得到检验。我们也会计算出环境条件(温度、酸碱值、死亡有机物的数量等)。在这种情况下，NRRA是最多样和最丰富的。这些结果综合起来将有助于科学家和环境机构评估土壤和淡水环境的“健康”和碳收支。

Recent advances in molecular biology techniques and analyses have greatly advanced the evolutionary and ecological study of microbial eukaryotes (protists). Because their appearance often does not reflect their evolutionary relationships, protist groups can generally only be properly understood when we have both morphological and genetic data and can use phylogenetic analyses to reconstruct their evolutionary relationships. Many protist groups have recently been elucidated in this way. In addition, related molecular techniques have been used to show that the genetic diversity of protists in most environments is massive - many more organisms await discovery than are known morphologically. However, one group of protists that has largely eluded both culture-based and molecular studies is that of large, non-shelled amoebae with long, often branching extensions to their cells (ramose pseudopodia), some of which fuse to form cytoplasmic 'networks' (reticulopodia) of varying sizes. We call them NRRA - naked ramose/reticulose amoebae. NRRA may be almost stationary, attached to surfaces or extending their pseudopodia between soil or sediment particles, or they can be active predators, moving across surfaces or through particulate material in search of prey. There are not many cultured NRRA because they are difficult to grow, but all known strains feed voraciously on a wide range of food sources - other protozoa, unicellular and multicellular algae, many types of fungi including those known to cause diseases of crops or other plants, and even small animals like nematode worms and rotifers. It is possible that some attack even larger organisms. Therefore, NRRA are likely to be important agents of biological control, influencing other microbial activity including disease-causing organisms and those involved in decomposition and nutrient cycling. NRRA remain very poorly known because their delicate cell shape means they are often destroyed by sampling techniques. They take a long time to appear in environmental samples in the laboratory and often need particular (but usually unknown!) conditions to grow. Furthermore, they are relatively rarely detected by molecular studies because their versions of the genes used for such studies are often so different from many other organisms that the standard molecular probes fail to detect them. The aim of our project is to find more NRRA in a wide range of habitats and describe their morphologies and lifecycles, and to measure how big their appetites are - both in terms of the diversity of their diet and how much of it they eat (including fungi and other organisms which are known to cause diseases of higher plants). We will sequence genes from NRRA cells in order to understand their evolutionary relationships and histories. We will measure how diverse they are (how many 'species', although this is often an unclear concept in many protists) under many different environmental conditions. Taking this line of enquiry further, we will collaborate with three research programmes that run long-term experiments on 1) forest soils, 2) agricultural vs. unploughed and uncultivated soils, and 3) freshwater lakes. These experiments have well established sampling strategies and other data collected from the sample sites, which will provide a very informative context for our new data on NRRA. By intensively sampling sites contributing to all three experiments we will investigate whether changes in NRRA diversity and abundance correlate with changes in other microbial (and small animal and plant) groups. If they do we will suggest possible reasons for the correlations, which can be tested in future studies. We will also work out the environmental conditions (temperature, pH, amount of dead organic material, etc.) under which NRRA are most diverse and abundant. These results combined will help scientists and environmental agencies to assess the 'health' and carbon budgets of soil and freshwater environments.