The Development of Microbial Associations in Major Reef Building Corals of the Pacific Ocean

珊瑚-微生物相关性研究的基础是了解何时以及如何建立这种关系，以及它们的特殊性。最近，研究人员首次提供了珊瑚(Pocillopora Meandrina)早期生活史阶段与一组特定细菌(甲型变形杆菌罗斯杆菌分支的Jannaschia血统)之间存在特定联系的证据。他们将通过研究北太平洋夏威夷和南太平洋法属波利尼西亚莫雷亚的关键造礁珊瑚中的微生物组合来扩大这项工作。了解与这些珊瑚物种相关的微生物群落的开始、特性和功能对于了解和预测珊瑚对环境变化的反应是必要的。这项建议的主要目标是：1.使用独立于栽培的技术来鉴定和量化与北太平洋夏威夷几种主要造礁珊瑚相关的微生物，这些微生物代表了各种繁殖策略(孵化和投射产卵)和不同的虫黄菌共生体传播方式(垂直与水平)，贯穿生殖周期、早期发育阶段和每个珊瑚的定居后阶段。收集和分析在法属波利尼西亚莫雷亚南太平洋发现的相同(或类似)珊瑚物种的类似样本，以评估目标1中记录的联系是否局限于夏威夷，或广泛分布于太平洋各地，可能代表珊瑚发育的共同特征。使用荧光原位杂交法对采集于夏威夷的米氏假单胞菌的整个发育周期内的罗氏杆菌分支的贾纳希氏菌谱系的细胞进行计数。PIS将扩大这一目标，以包括其他珊瑚物种、目标细菌和/或地理位置，因为他们确定了更多的联系。分离在与栽培无关的调查中流行的微生物(例如Jannaschia sp.)通过应用新的养殖技术，以开发研究珊瑚-微生物相互作用的模式系统。由于人类活动造成的环境压力增加，珊瑚礁的智力正在下降，目前有相当多的证据表明，它们受到海洋表面温度上升和海洋酸化的影响。与珊瑚相关的微生物被认为在维持珊瑚宿主的健康和弹性方面发挥着各种潜在的重要作用，主要由分子生物学领域的发展推动的方法学进步正在促进对这种联系的深入了解。许多珊瑚微生物的研究都集中在微生物对疾病和漂白的贡献上，而且几乎完全集中在成年珊瑚群体上。这项研究将提供关于微生物在其整个发育周期中与健康珊瑚相互作用的方式、这些关系的特殊性、它们是如何启动的以及它们在自然界中的分布和频率的独特信息。(1)这项建议将支持一名博士后学者在先进的分子微生物生态学技术、新的基于栽培的方法学和珊瑚生物学方面的培训。(2)研究人员将开发一个网站，该网站将包含珊瑚-微生物相互作用的信息，其内容旨在作为K-12学生的教育资源，并作为对珊瑚-微生物协会感兴趣的其他科学家的资源。这项研究产生的数据，包括出版物、介绍、微生物菌株、基因序列、显微照片和系统发育分析，将通过该网站向公众公布。(3)博士后和私人助理将设计并每年提供为期五天的强化培训讲习班，介绍ARB序列分析程序的使用，包括将荧光原位显微镜应用于海洋细菌和珊瑚样本。(4)该提案将支持在博士后学者的指导下对本科生进行培训，并将利用现有的项目和关系，专门为夏威夷大学未被充分代表的学生，如夏威夷原住民和太平洋岛民提供这些机会。

Fundamental to the study of coral-microbial associations is an understanding of when and how the relationships are established, and their specificity. Recently, the investigators provided the first evidence of a specific association between an early life history stage of a coral (Pocillopora meandrina) and a particular group of bacteria (Jannaschia lineage of the Roseobacter clade of Alphaproteobacteria). They will be expanding this work by examining the onset of microbial associations in key reef building corals from Hawaii in the North Pacific Ocean and Moorea, French Polynesia, in the South Pacific Ocean. Understanding the onset, specificity and function of the microbial community associated with these coral species is necessary to understand and predict the coral holobiont response to a changing environment.The main objectives of this proposal are to: 1. Use cultivation independent techniques to identify and quantify microorganisms associated with several major reef building corals of Hawaii in the North Pacific Ocean that represent a variety of reproductive strategies (brooding and broadcast spawning) and differing modes of zooxanthellae symbiont transmission (vertical vs. horizontal), throughout the reproductive cycle, early developmental stages, and post-settlement stages of each.2. Collect and analyze similar samples from the same (or similar) species of coral found in the South Pacific Ocean in Moorea, French Polynesia, in order to assess whether the associations documented in objective 1 are localized to Hawaii, or broadly distributed across the Pacific and likely to represent common features of coral development.3. Use fluorescence in situ hybridization to enumerate cells of the Jannaschia lineage of the Roseobacter clade throughout the development cycle of P. meandrina collected in Hawaii. The PIs will expand this objective to include other coral species, target bacteria, and/or geographic location as they identify additional associations.4. Isolate microorganisms prevalent in cultivation-independent surveys of P. meandrina-associated microbial communities (e.g. Jannaschia sp.) by the application of novel culturing techniques, in order to develop model systems for the investigation of coral-microbe interactions.Intellectual merit Coral reefs are in decline as a result of increasing environmental stress due to anthropogenic activity, and there is now considerable evidence indicating that they are under threat from the effects of rising sea surface temperature and ocean acidification. Microorganisms associated with corals are thought to play a variety of potentially important roles in maintaining the health and resiliency of the coral host, and advances in methodology primarily driven by developments in the field of molecular biology are facilitating growing insight into this association. Much of coral-microbial research is focused on the contribution of microorganisms to disease and bleaching, and is focused almost exclusively on adult coral colonies. This study will provide unique information on the manner in which microorganisms interact with healthy corals throughout their developmental cycle, the specificity of these relationships, how they are initiated, and their distribution and frequency in nature.Broader impacts (1) This proposal will support the training of one postdoctoral scholar in advancing molecular microbial ecology techniques, novel cultivation-based methodology, and coral biology. (2) The investigators will develop a website that will contain information on coral-microbial interactions, with content designed to serve as an educational resource for K-12 students, and to serve as a resource for other scientists interested in coral-microbial associations. Data generated from this study, including publications, presentations, microbial strains, gene sequences, photomicrographs and phylogenetic analyses, will be made publicly available via the website. (3) The postdoc and PI will design and annually offer a five-day intensive training workshop on the use of ARB sequence analysis program, including the application of fluorescence in situ microscopy to marine bacteria and coral samples. (4) The proposal will support the training of undergraduate students under the mentorship of the postdoctoral scholar, and will take advantage of existing programs and relationships to specifically offer these opportunities to underrepresented students at the University of Hawaii such as Native Hawaiians and Pacific Islanders.