Award: OCE-1233612

Reef-building corals are in decline worldwide due in part to climate change and other human activities, and it is becoming increasingly important to understand what aspects of coral biology are degraded by environmental stress which then leads to coral mortality. It is now widely known that corals harbor communities of bacteria and archaea that are believed to play important roles in maintaining the health of their hosts, but we lack any appreciable understanding about the identity of the microbes regularly associating with healthy, reef-building corals. The goal of this work was to understand the extent to which reef-building corals harbor fundamental or persistent microbial associates that are symbiotic within their tissues as well as other coral-specific habitats. Our study addressed this goal with direct studies of microbial associates in both Caribbean and Pacific corals as well as using previously collected and publically available data to examine the consistency of microbial associates across species and oceans. This project made significant advances in addressing our goals. For the direct studies, we developed a new approach to separate coral tissue and mucus and identified the most suitable DNA extraction method for these corals. We demonstrated how differentiating between these coral habitats enabled the identification of previously unrecognized tissue and mucus associates of Caribbean corals, including 46 tissue- and 22 mucus-specific and consistent microbial members. Importantly, sequences classifying as "Candidatus Amoebophilus," Bacteroidetes-affiliated intracellular symbionts of amoebae, emerged as previously unrecognized tissue associates of three coral species. This is particularly important because it suggests that corals are multi-symbiotic organisms, and adds a further degree of intricacy to coral holobiont symbioses. This work provides important research directions for understanding the association between corals and their fundamental microbial associates. We also refined coral microscopy methodology to demonstrate for the first time that the common coral bacterium Endozoicomonas is involved in an intimate association, residing within dense aggregations within the tissues of reef building corals. Further, we showed that the location of Endozoicomonas cells within corals does vary with species, and cell aggregates can be located in the tentacles or the gut tissues of their host. This species-specific difference is important because it suggests that coral-Endozoicomonas interactions may differ between coral species. Since Endozoicomonas is such an abundant and common symbiont of corals, we also contributed to a review of the available knowledge on the Endozoicomonas genus, which collectively shows that these bacteria associate with diverse invertebrates from shallow and deep-sea habitats, suggesting that they are an extraordinarily successful group of symbionts that may play a fundamental symbiotic role with much of the invertebrate life in our ocean. We also demonstrated that corals harbor different mucus and tissue-associated bacteria and archaea than are present in the seawater, indicating that corals are highly selective for certain microbes. Additionally, we discovered in this mesocosm-based experiment that corals do heavily influence the community composition and abundance of bacteria in the water. Our results demonstrate that when corals are present in the mesocosms, bacteria are preferentially removed via grazing. When corals were removed, their exudates enhanced the growth of diverse picoplankton. This work provides the first evidence of the multiple influences that corals have on reef picoplankton, which may play an important role in reef productivity. During our analysis of sequence data in this study, we identified a critical problem with commonly utilized molecular primers that excluded an abundant group of marine bacteria (SAR11). Our re-design and improvement of the primers resulted in an important contribution to the ocean microbiome community. Our examination of coral microbial diversity using publically available data has demonstrated that corals harbor one of the most biodiverse microbiomes on Earth, spanning 37 described and candidate phyla of Bacteria and two phyla of Archaea. A number of specific taxonomic groups were also found to be prominent and less-recognized. We constructed two coral microbial sequence databases that house different types of sequence data. These databases as well as the other new sequence data produced during this project are important resources for the scientific community. This project has produced new sequence and supporting oceanographic data which have been shared publically through BCO-DMO. Project data are available at the following url: http://www.bco-dmo.org/project/564442. This project provided training opportunities for four undergraduate students (two females), two female graduate students and a new female tenure-track scientist. We communicated our project results over the course of 12 presentations and 8 scientific publications. These publications are all available online under Open Access. Two studies were featured in press releases, which were re-distributed through a variety of online science news and other news sources. This project also supported the development of a pre-school lesson on coral reefs and microorganisms, which was taught each year to students. Results were also communicated in accessible language to high school, undergraduate and adult audiences. Last Modified: 12/15/2016 Submitted by: Amy Apprill