The Role of Fungi in Biogeochemical Transformations of Mn and Other Micro- and Macro-nutrients Along Chemoclines of the Baltic Sea

越来越多的证据表明，真菌构成了栖息在海洋环境中的微生物群落的一个活跃而多样的部分。其中，全球分布的低氧和低盐度公海和沿海沃茨预计将随着气候变化而扩大和加剧。锰（Mn）是一种分布在低氧海洋系统中的营养物质，在主要元素循环中起着至关重要的作用，包括微生物进行的循环。因此，锰与海洋微生物组的健康、代谢和功能有着错综复杂的联系。尽管活性真菌在低氧和半咸水生态系统中对锰和其他营养物质循环做出重大贡献的潜力，但对真菌在这些过程中的作用和影响知之甚少。子囊菌门和担子菌门真菌物种被确定为在各种海洋环境中重要的营养物质和碳循环，但是很少有真菌在化跃层设置（水柱与氧浓度的转换）的研究。已知真菌分离株将Mn的化学转化（Mn（II）氧化）与有机碳降解和活性氧物质（ROS）的产生联系起来，并且ROS的真菌产生可能在金属如Mn以及低氧/缺氧（零氧）海洋环境中的碳和氮的循环和生物利用度中发挥核心作用。富锰波罗的海是一个理想的模式生态系统，研究未来的沿海海洋由于人为影响的经验，其盐度梯度。该项目旨在帮助了解真菌多样性和锰，氮和硫循环中的作用，以及低氧和半咸水海洋生态系统中活性氧的产生。这些生境中的真菌活动可能影响重要的全球海洋生物地球化学循环，了解它们的作用和影响有助于更准确地预测未来海洋和气候的生物地球化学。这项研究产生的培养物收集预计将恢复许多新的真菌菌株，其生态学，新特性和潜在的医学相关的生物活性化合物可以由感兴趣的研究人员探索。本研究以每年六位本科生、一位研究生及二位高中生为对象，并与当地高中美术教师合作，教授“科学中的艺术”单元，并于社区图书馆展出其产品，沿着海洋真菌及其生态角色的教材。该项目涉及国际合作，以及培训早期职业和代表性不足的少数民族科学家。该提案利用2024年初预定巡航期间的采样机会，在波罗的海两个站点沿化学跃层沿着3个深度采集水样，并具有不同的O2浓度分布以及N和Mn物种。该项目的总体目标是表征这些样品中存在的原核生物（细菌和古细菌）和真菌类群的多样性，以及表达与锰，碘，氧和氮的转化相关的生物地球化学循环的基因的那些，特别强调真菌的作用。 该工作计划包括分析真菌和微型真核生物及原核生物的分类标记基因，以及催化报告沉积荧光原位杂交（CARD-FISH），以估计主要真菌群体的原位丰度。多聚腺苷酸和非多聚腺苷酸转录组学的水样品提供了一个表达的代谢基因的一般概述，并特别确定参与锰氧化的基因。高通量培养的努力，将激光散射比浊法用于收集最广泛的代表性的可培养的海洋真菌在这些化跃层栖息地，并确定那些携带感兴趣的基因参与耦合锰（II）氧化有机碳降解。船上孵育研究纳入真菌和原核生物抑制剂用于确定真菌在多大程度上有助于锰转化过程。孵育研究的耦合RT-qPCR和元转录组分析用于阐明与Mn转化相关的真菌基因的表达（例如，Mn过氧化物酶）、氮循环（例如，该奖项反映了NSF的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Mounting evidence suggests that fungi constitute an active and diverse fraction of the microbial community inhabiting marine environments. Among these, globally-distributed low-oxygen, and low salinity open-ocean and coastal waters are expected to expand and intensify with climate change. Manganese (Mn) is a nutrient that is distributed throughout low-oxygen marine systems that plays an essential role in major elemental cycles, including those performed by microorganisms. Manganese is thus intricately linked to the health, metabolism, and function of the ocean microbiome. Despite the potential for active fungi in low-oxygen and brackish ecosystems to make significant contributions to Mn and other nutrient cycling, little is known about the roles and impacts of fungi on those processes. Ascomycota and Basidiomycota fungal species are identified as important nutrient and carbon recyclers in various marine settings, however there are few studies of fungi in chemocline settings (water columns with transitions in oxygen concentration). Fungal isolates are known that link chemical transformation of Mn (Mn(II) oxidation) to organic carbon degradation and production of reactive oxygen species (ROS), and fungal production of ROS may play a central role in the cycling and bioavailability of metals like Mn, as well as carbon and nitrogen in low-oxygen/anoxic (zero oxygen) marine environments. The Mn-rich Baltic Sea is an ideal model ecosystem for studying the future coastal ocean due to anthropogenic impacts experiences, and its salinity gradients. This project aims to contribute to understanding of fungal diversity and roles in manganese, nitrogen and sulfur cycling, as well as production of reactive oxygen species in low oxygen and brackish marine ecosystems. Fungal activities in these habitats may influence important global marine biogeochemical cycles, and knowledge of their role(s) and impacts allows more accurate predictions of the biogeochemistry of a future ocean and climate. The culture collection generated by this study is anticipated to recover many new fungal strains, whose ecology, novel properties, and potential medically-relevant bioactive compounds can be explored by interested researchers. Six undergraduate students, one graduate student, and 2 high school students per year are included in this research, and a collaboration is established with a local high school art teacher to teach an art-in-science unit, and to displayed its product at the community library, along with education materials on marine fungi and their ecological roles. The project involves international collaboration, as well as training of early career and under-represented minority scientists. This proposal leverages a sampling opportunity during a scheduled cruise in early 2024 to collect water samples from 3 depths along the chemocline at two stations in the Baltic Sea with distinct profiles of O2 concentration, and N and Mn species. The overall goal of this project is to characterize the diversity of prokaryotic (bacteria and archaea) and fungal taxa present in these samples, and those expressing genes involved in biogeochemical cycles related to transformations of manganese, iodine, oxygen, and nitrogen, with a specific emphasis on the role of fungi. The workplan incorporates analysis of taxonomic marker genes for fungi and micro-eukaryotes, and prokaryotes, as well as catalyzed-reporter deposition fluorescence in situ hybridization (CARD-FISH) to estimate the in situ abundance of major fungal groups. Poly-A and non polyA transcriptomics of water samples provide a general overview of expressed metabolic genes, and specifically identify genes involved in Mn oxidation. High-throughput culturing efforts incorporating laser nephelometry are used to gather the broadest possible representation of culturable marine fungi in these chemocline habitats, and to identify those that carry genes of interest involved in coupling Mn(II) oxidation to organic carbon degradation. Shipboard incubation studies incorporating fungal and prokaryotic inhibitors are used to determine the extent to which fungi contribute to Mn transformation processes. Coupled RT-qPCR and metatranscriptome analyses of incubation studies are used to elucidate expression of fungal genes related to Mn transformations (e.g., Mn peroxidases), nitrogen cycling (e.g., key fungal denitrification genes p450nor, nirK), and ROS production/decay (SOD1, NOXA).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.