PurSUiT: Discovery and phylogenetic analysis of chytrid fungi using cultures and single cells

壶菌是一组研究很少的水生真菌，普遍栖息在沼泽，池塘和海洋。这些真菌是营养物质循环的基础，例如通过花粉，角蛋白的降解，或通过藻类或青蛙等生物体的寄生。关于壶菌多样性的大部分知识都是基于上个世纪的显微镜观察或过去几十年建立的纯培养。然而，基于多样性的评估，使用最近开发的DNA为基础的环境方法的挑战，我们是否真正知道壶菌的多样性，因为以这种方式记录的物种通常无法匹配使用传统的方法确定的物种。该项目将实施多种方法，以提供壶菌多样性和分类的整体图景，避免任何特定方法的偏见。了解这一鲜为人知的群体和其他真菌的分类对于编目生物多样性和预测其环境影响至关重要。该项目将支持一名博士后研究员，一名研究生，并将为本科生提供培训。将开发关于壶菌生物学、分类和生物学的网络内容，以便向更广泛的受众传播信息。为期一周的关于壶菌方法的实践研讨会将在密歇根大学生物站举行，以鼓励对这一未充分研究的群体继续进行研究。 该项目利用现有的和不断增长的壶菌培养物的历史收集和最近开发的方法来对单细胞DNA进行测序。拟议的研究包括在密歇根大学（CZEUM）的游动孢子真真菌集合中生成或完成至少840个核糖体RNA序列和20个壶菌基因组序列。它还将采用单细胞方法对未培养的多样性进行遗传学表征。具体地说，壶菌将在现场鉴定，拍照，并进行基因组扩增的单细胞。从成功扩增的基因组中，100个不同的样品将被靶向用于使用长读段、单分子DNA测序的全核糖体RNA操纵子测序。另外20份样本将用于基因组测序。一个主要的成果将是一个系统发育分析的壶菌结合核糖体RNA和基因组测序成一个超级树。该项目的最终目的是筛选和测序CZEUM收集的真菌病毒的新多样性，并确定生态和形态对病毒流行的影响。这个项目将作为一个模型，解决系统学在真核细胞谱系中，基于文化的方法产生的多样性的不完整的图片。这个奖项反映了NSF的法定使命，并已被认为是值得通过评估使用基金会的知识价值和更广泛的影响审查标准的支持。

Chytrids are a poorly studied group of aquatic fungi that ubiquitously inhabit bogs, ponds, and oceans. These fungi are fundamental to the cycling of nutrients, such as through the degradation of pollen, keratin, or through parasitism of organisms such as algae or frogs. Most of what is known about the diversity of chytrids is based on microscopic observations over the last century or from pure cultures established in the last few decades. However, diversity based assessments using the more recently developed DNA-based environmental approaches challenge whether we truly know chytrid diversity, because the species documented in this way typically fail to match species identified using traditional approaches. This project will implement multiple approaches to provide a holistic picture of chytrid diversity and classification that avoids biases of any particular method. Understanding taxonomy of this poorly known group and other fungi is vital for cataloging biodiversity and predicting its environmental impacts. This project will support one Postdoctoral Fellow, a graduate student, and will provide training to undergraduates. Web-based content on the chytrid biology, classification, and will be developed in order to disseminate information to a broader audience. A week-long, hands on workshop on chytrid methods will be held at the University of Michigan Biological Station in order to encourage continued research on this understudied group. This project leverages an existing and growing historical collection of chytrid cultures and recently developed methods to sequence DNA from single cells. The proposed research includes generating or completing at least 840 ribosomal RNA sequences and 20 genome sequences of chytrids in the Collection of Zoosporic Eufungi at the University of Michigan (CZEUM). It will also employ a single cell approach to phylogenetically characterize uncultured diversity. Specifically, chytrid fungi will be identified in situ in from field sites, photographed, and single cells subjected to genome amplification. From the successfully amplified genomes, 100 diverse samples will be targeted for whole ribosomal RNA operon sequencing using long-read, single molecule DNA sequencing. An additional 20 samples will be used for genome sequencing. A major outcome will be a phylogenetic analysis of the chytrids combining both ribosomal RNA and genome sequencing into a supertree. A final aim of the project is to screen and sequence novel diversity of fungal viruses from the CZEUM collection and to determine the effect of ecology and morphology on viral prevalence. This project will serve as a model for addressing systematics in eukaryotic lineages where culture-based approaches produce an incomplete picture of diversity.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.