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Climate change destabilizes the symbiosis between corals and Symbiodiniaceae. The effects of ocean acidification and warming on critical aspects of coral survical such as symbiotic interactions (i.e., carbon and nitrogen assimilation and exchange) during the planula larval stage remain understudied. By combining physiological and stable isotope techniques, here we show that photosynthesis and carbon and nitrogen assimilation (H 13 CO 3 - and 15 ) in Pocillopora damicornis coral larvae is enhanced under acidification (1000 µatm) and elevated temperature (32 °C). Larvae maintain high survival and settlement rates under these treatment conditions with no observed decline in symbiont densities or signs of bleaching. Acidification and elevated temperature both enhance the net and gross photosynthesis of Symbiodiniaceae. This enhances light respiration and elevates C:N ratios within the holobiont. The increased carbon availability is primarily reflected in the 4 + ) in Pocillopora damicornis coral larvae is enhanced under acidification (1000 µatm) and elevated temperature (32 °C). Larvae maintain high survival and settlement rates under these treatment conditions with no observed decline in symbiont densities or signs of bleaching. Acidification and elevated temperature both enhance the net and gross photosynthesis of Symbiodiniaceae. This enhances light respiration and elevates C:N ratios within the holobiont. The increased carbon availability is primarily reflected in the 13 C enrichment of the host, indicating a greater contribution of the algal symbionts to the host metabolism. We propose that this enhanced mutualistic symbiotic nutrient cycling may bolster coral larvae's resistance to future ocean conditions. This research broadens our understanding of the early life stages of corals by emphasizing the significance of symbiotic interactions beyond those of adult corals., Competing Interests: Competing interests The authors declare no competing interests., (© 2024. The Author(s).)

Objectives: The endosymbiosis with Symbiodiniaceae is key to the ecological success of reef-building corals. However, climate change is threatening to destabilize this symbiosis on a global scale. Most studies looking into the response of corals to heat stress and ocean acidification focus on coral colonies. As such, our knowledge of symbiotic interactions and stress response in other stages of the coral lifecycle remains limited. Establishing transcriptomic resources for coral larvae under stress can thus provide a foundation for understanding the genomic basis of symbiosis, and its susceptibility to climate change. Here, we present a gene expression dataset generated from larvae of the coral Pocillopora damicornis in response to exposure to acidification and elevated temperature conditions below the bleaching threshold of the symbiosis., Data Description: This dataset is comprised of 16 samples (30 larvae per sample) collected from four treatments (Control, High pCO 2 , High Temperature, and Combined pCO 2 and Temperature treatments). Freshly collected larvae were exposed to treatment conditions for five days, providing valuable insights into gene expression in this vulnerable stage of the lifecycle. In combination with previously published datasets, this transcriptomic resource will facilitate the in-depth investigation of the effects of ocean acidification and elevated temperature on coral larvae and its implication for symbiosis., (© 2024. The Author(s).)

Background: Global warming is causing large-scale disruption of cnidarian-Symbiodiniaceae symbioses fundamental to major marine ecosystems, such as coral reefs. However, the mechanisms by which heat stress perturbs these symbiotic partnerships remain poorly understood. In this context, the upside-down jellyfish Cassiopea has emerged as a powerful experimental model system., Results: We combined a controlled heat stress experiment with isotope labeling and correlative SEM-NanoSIMS imaging to show that host starvation is a central component in the chain of events that ultimately leads to the collapse of the Cassiopea holobiont. Heat stress caused an increase in catabolic activity and a depletion of carbon reserves in the unfed host, concurrent with a reduction in the supply of photosynthates from its algal symbionts. This state of host starvation was accompanied by pronounced in hospite degradation of algal symbionts, which may be a distinct feature of the heat stress response of Cassiopea. Interestingly, this loss of symbionts by degradation was concealed by body shrinkage of the starving animals, resulting in what could be referred to as "invisible" bleaching., Conclusions: Overall, our study highlights the importance of the nutritional status in the heat stress response of the Cassiopea holobiont. Compared with other symbiotic cnidarians, the large mesoglea of Cassiopea, with its structural sugar and protein content, may constitute an energy reservoir capable of delaying starvation. It seems plausible that this anatomical feature at least partly contributes to the relatively high stress tolerance of these animals in rapidly warming oceans. Video Abstract., (© 2024. The Author(s).)

Medusae of the widely distributed upside-down jellyfish Cassiopea release autonomous, mobile stinging structures. These so-called cassiosomes play a role in predator defense and prey capture, and are major contributors to "contactless" stinging incidents in (sub-)tropical shallow waters. While the presence of endosymbiotic dinoflagellates in cassiosomes has previously been observed, their potential contribution to the metabolism and long-term survival of cassiosomes is unknown. Combining stable isotope labeling and correlative scanning electron microscopy and nanoscale secondary ion mass spectrometry imaging with a long-term in vitro experiment, our study reveals a mutualistic symbiosis based on nutritional exchanges in dinoflagellate-bearing cassiosomes. We show that organic carbon input from the dinoflagellates fuels the metabolism of the host tissue and enables anabolic nitrogen assimilation. This symbiotic nutrient exchange enhances the life span of cassiosomes for at least one month in vitro . Overall, our study demonstrates that cassiosomes, in analogy with Cassiopea medusae, are photosymbiotic holobionts. Cassiosomes, which are easily accessible under aquarium conditions, promise to be a powerful new miniaturized model system for in-depth ultrastructural and molecular investigation of cnidarian photosymbioses.IMPORTANCEThe upside-down jellyfish Cassiopea releases autonomous tissue structures, which are a major cause of contactless stinging incidents in (sub-) tropical coastal waters. These so-called cassiosomes frequently harbor algal symbionts, yet their role in cassiosome functioning and survival is unknown. Our results show that cassiosomes are metabolically active and supported by algal symbionts. Algal photosynthesis enhances the cassiosomes long-term survival in the light. This functional understanding of cassiosomes thereby contributes to explaining the prevalence of contactless stinging incidents and the ecological success of some Cassiopea species. Finally, we show that cassiosomes are miniaturized symbiotic holobionts that can be used to study host-microbe interactions in a simplified system., Competing Interests: The authors declare no conflict of interest.

Efficient nutrient recycling underpins the ecological success of cnidarian-algal symbioses in oligotrophic waters. In these symbioses, nitrogen limitation restricts the growth of algal endosymbionts in hospite and stimulates their release of photosynthates to the cnidarian host. However, the mechanisms controlling nitrogen availability and their role in symbiosis regulation remain poorly understood. Here, we studied the metabolic regulation of symbiotic nitrogen cycling in the sea anemone Aiptasia by experimentally altering labile carbon availability in a series of experiments. Combining 13 C and 15 N stable isotope labeling experiments with physiological analyses and NanoSIMS imaging, we show that the competition for environmental ammonium between the host and its algal symbionts is regulated by labile carbon availability. Light regimes optimal for algal photosynthesis increase carbon availability in the holobiont and stimulate nitrogen assimilation in the host metabolism. Consequently, algal symbiont densities are lowest under optimal environmental conditions and increase toward the lower and upper light tolerance limits of the symbiosis. This metabolic regulation promotes efficient carbon recycling in a stable symbiosis across a wide range of environmental conditions. Yet, the dependence on resource competition may favor parasitic interactions, explaining the instability of the cnidarian-algal symbiosis as environmental conditions in the Anthropocene shift towards its tolerance limits., (© 2023. The Author(s).)

Background: The development of nanoscale secondary ion mass spectrometry (NanoSIMS) has revolutionized the study of biological tissues by enabling, e.g., the visualization and quantification of metabolic processes at subcellular length scales. However, the associated sample preparation methods all result in some degree of tissue morphology distortion and loss of soluble compounds. To overcome these limitations an entirely cryogenic sample preparation and imaging workflow is required., Results: Here, we report the development of a CryoNanoSIMS instrument that can perform isotope imaging of both positive and negative secondary ions from flat block-face surfaces of vitrified biological tissues with a mass- and image resolution comparable to that of a conventional NanoSIMS. This capability is illustrated with nitrogen isotope as well as trace element mapping of freshwater hydrozoan Green Hydra tissue following uptake of 15 N-enriched ammonium., Conclusion: With a cryo-workflow that includes vitrification by high pressure freezing, cryo-planing of the sample surface, and cryo-SEM imaging, the CryoNanoSIMS enables correlative ultrastructure and isotopic or elemental imaging of biological tissues in their most pristine post-mortem state. This opens new horizons in the study of fundamental processes at the tissue- and (sub)cellular level., Teaser: CryoNanoSIMS: subcellular mapping of chemical and isotopic compositions of biological tissues in their most pristine post-mortem state., (© 2023. The Author(s).)

Symbiotic cnidarians such as corals and anemones form highly productive and biodiverse coral reef ecosystems in nutrient-poor ocean environments, a phenomenon known as Darwin's paradox. Resolving this paradox requires elucidating the molecular bases of efficient nutrient distribution and recycling in the cnidarian-dinoflagellate symbiosis. Using the sea anemone Aiptasia, we show that during symbiosis, the increased availability of glucose and the presence of the algae jointly induce the coordinated up-regulation and relocalization of glucose and ammonium transporters. These molecular responses are critical to support symbiont functioning and organism-wide nitrogen assimilation through glutamine synthetase/glutamate synthase-mediated amino acid biosynthesis. Our results reveal crucial aspects of the molecular mechanisms underlying nitrogen conservation and recycling in these organisms that allow them to thrive in the nitrogen-poor ocean environments.

Nitrogen limitation is the foundation of stable coral-algal symbioses. Diazotrophs, prokaryotes capable of fixing N 2 into ammonia, support the productivity of corals in oligotrophic waters, but could contribute to the destabilization of holobiont functioning when overstimulated. Recent studies on reef-building corals have shown that labile dissolved organic carbon (DOC) enrichment or heat stress increases diazotroph abundance and activity, thereby increasing nitrogen availability and destabilizing the coral-algal symbiosis. However, the (a)biotic drivers of diazotrophs in octocorals are still poorly understood. We investigated diazotroph abundance (via relative quantification of nifH gene copy numbers) in two symbiotic octocorals, the more mixotrophic soft coral Xenia umbellata and the more autotrophic gorgonian Pinnigorgia flava, under (i) labile DOC enrichment for 21 days, followed by (ii) combined labile DOC enrichment and heat stress for 24 days. Without heat stress, relative diazotroph abundances in X. umbellata and P. flava were unaffected by DOC enrichment. During heat stress, DOC enrichment (20 and 40 mg glucose l -1 ) increased the relative abundances of diazotrophs by sixfold in X. umbellata and fourfold in P. flava , compared with their counterparts without excess DOC. Our data suggest that labile DOC enrichment and concomitant heat stress could disrupt the nitrogen limitation in octocorals by stimulating diazotroph proliferation. Ultimately, the disruption of nitrogen cycling may further compromise octocoral fitness by destabilizing symbiotic nutrient cycling. Therefore, improving local wastewater facilities to reduce labile DOC input into vulnerable coastal ecosystems may help octocorals cope with ocean warming., (© 2023 The Authors.)

The coral-algal symbiosis is maintained by a constant and limited nitrogen availability in the holobiont. Denitrifiers, i.e., prokaryotes reducing nitrate/nitrite to dinitrogen, could contribute to maintaining the nitrogen limitation in the coral holobiont, however the effect of host and algal identity on their community is still unknown. Using the coral model Aiptasia, we quantified and characterized the denitrifier community in a full-factorial design combining two hosts (CC7 and H2) and two strains of algal symbionts of the family Symbiodiniaceae (SSA01 and SSB01). Strikingly, relative abundance of denitrifiers increased by up to 22-fold in photosymbiotic Aiptasia compared to their aposymbiotic (i.e., algal-depleted) counterparts. In line with this, while the denitrifier community in aposymbiotic Aiptasia was largely dominated by diet-associated Halomonas, we observed an increasing relative abundance of an unclassified bacterium in photosymbiotic CC7, and Ketobacter in photosymbiotic H2, respectively. Pronounced changes in denitrifier communities of Aiptasia with Symbiodinium linucheae strain SSA01 aligned with the higher photosynthetic carbon availability of these holobionts compared to Aiptasia with Breviolum minutum strain SSB01. Our results reveal that the presence of algal symbionts increases abundance and alters community structure of denitrifiers in Aiptasia. Thereby, patterns in denitrifier community likely reflect the nutritional status of aposymbiotic vs. symbiotic holobionts. Such a passive regulation of denitrifiers may contribute to maintaining the nitrogen limitation required for the functioning of the cnidarian-algal symbiosis., (© 2022. The Author(s).)

The skeleton of reef-building coral harbors diverse microbial communities that could compensate for metabolic deficiencies caused by the loss of algal endosymbionts, i.e., coral bleaching. However, it is unknown to what extent endolith taxonomic diversity and functional potential might contribute to thermal resilience. Here we exposed Goniastrea edwardsi and Porites lutea, two common reef-building corals from the central Red Sea to a 17-day long heat stress. Using hyperspectral imaging, marker gene/metagenomic sequencing, and NanoSIMS, we characterized their endolithic microbiomes together with 15 N and 13 C assimilation of two skeletal compartments: the endolithic band directly below the coral tissue and the deep skeleton. The bleaching-resistant G. edwardsi was associated with endolithic microbiomes of greater functional diversity and redundancy that exhibited lower N and C assimilation than endoliths in the bleaching-sensitive P. lutea. We propose that the lower endolithic primary productivity in G. edwardsi can be attributed to the dominance of chemolithotrophs. Lower primary production within the skeleton may prevent unbalanced nutrient fluxes to coral tissues under heat stress, thereby preserving nutrient-limiting conditions characteristic of a stable coral-algal symbiosis. Our findings link coral endolithic microbiome structure and function to bleaching susceptibility, providing new avenues for understanding and eventually mitigating reef loss., (© 2022. The Author(s).)

Endozoicomonas are prevalent, abundant bacterial associates of marine animals, including corals. Their role in holobiont health and functioning, however, remains poorly understood. To identify potential interactions within the coral holobiont, we characterized the novel isolate Endozoicomonas marisrubri sp. nov. 6c and assessed its transcriptomic and proteomic response to tissue extracts of its native host, the Red Sea coral Acropora humilis. We show that coral tissue extracts stimulated differential expression of genes putatively involved in symbiosis establishment via the modulation of the host immune response by E. marisrubri 6c, such as genes for flagellar assembly, ankyrins, ephrins, and serpins. Proteome analyses revealed that E. marisrubri 6c upregulated vitamin B1 and B6 biosynthesis and glycolytic processes in response to holobiont cues. Our results suggest that the priming of Endozoicomonas for a symbiotic lifestyle involves the modulation of host immunity and the exchange of essential metabolites with other holobiont members. Consequently, Endozoicomonas may play an important role in holobiont nutrient cycling and may therefore contribute to coral health, acclimatization, and adaptation., (© 2022. The Author(s).)

Efficient nutrient cycling in the coral-algal symbiosis requires constant but limited nitrogen availability. Coral-associated diazotrophs, i.e., prokaryotes capable of fixing dinitrogen, may thus support productivity in a stable coral-algal symbiosis but could contribute to its breakdown when overstimulated. However, the effects of environmental conditions on diazotroph communities and their interaction with other members of the coral holobiont remain poorly understood. Here we assessed the effects of heat stress on diazotroph diversity and their contribution to holobiont nutrient cycling in the reef-building coral Stylophora pistillata from the central Red Sea. In a stable symbiotic state, we found that nitrogen fixation by coral-associated diazotrophs constitutes a source of nitrogen to the algal symbionts. Heat stress caused an increase in nitrogen fixation concomitant with a change in diazotroph communities. Yet, this additional fixed nitrogen was not assimilated by the coral tissue or the algal symbionts. We conclude that although diazotrophs may support coral holobiont functioning under low nitrogen availability, altered nutrient cycling during heat stress abates the dependence of the coral host and its algal symbionts on diazotroph-derived nitrogen. Consequently, the role of nitrogen fixation in the coral holobiont is strongly dependent on its nutritional status and varies dynamically with environmental conditions., (© 2021. The Author(s).)

Over this century, coral reefs will run the gauntlet of climate change, as marine heatwaves (MHWs) become more intense and frequent, and ocean acidification (OA) progresses. However, we still lack a quantitative assessment of how, and to what degree, OA will moderate the responses of corals to MHWs as they intensify throughout this century. Here, we first projected future MHW intensities for tropical regions under three future greenhouse gas emissions scenario (representative concentration pathways, RCP2.6, RCP4.5 and RCP8.5) for the near-term (2021-2040), mid-century (2041-2060) and late-century (2081-2100). We then combined these MHW intensity projections with a global data set of 1,788 experiments to assess coral attribute performance and survival under the three emissions scenarios for the near-term, mid-century and late-century in the presence and absence of OA. Although warming and OA had predominately additive impacts on the coral responses, the contribution of OA in affecting most coral attributes was minor relative to the dominant role of intensifying MHWs. However, the addition of OA led to greater decreases in photosynthesis and survival under intermediate and unrestricted emissions scenario for the mid- and late-century than if intensifying MHWs were considered as the only driver. These results show that role of OA in modulating coral responses to intensifying MHWs depended on the focal coral attribute and extremity of the scenario examined. Specifically, intensifying MHWs and OA will cause increasing instances of coral bleaching and substantial declines in coral productivity, calcification and survival within the next two decades under the low and intermediate emissions scenario. These projections suggest that corals must rapidly adapt or acclimatize to projected ocean conditions to persist, which is far more likely under a low emissions scenario and with increasing efforts to manage reefs to enhance resilience., (© 2021 The Authors. Global Change Biology published by John Wiley & Sons Ltd.)

Mutualistic nutrient cycling in the coral-algae symbiosis depends on limited nitrogen (N) availability for algal symbionts. Denitrifying prokaryotes capable of reducing nitrate or nitrite to dinitrogen could thus support coral holobiont functioning by limiting N availability. Octocorals show some of the highest denitrification rates among reef organisms; however, little is known about the community structures of associated denitrifiers and their response to environmental fluctuations. Combining 16S rRNA gene amplicon sequencing with nirS in-silico PCR and quantitative PCR, we found differences in bacterial community dynamics between two octocorals exposed to excess dissolved organic carbon (DOC) and concomitant warming. Although bacterial communities of the gorgonian Pinnigorgia flava remained largely unaffected by DOC and warming, the soft coral Xenia umbellata exhibited a pronounced shift toward Alphaproteobacteria dominance under excess DOC. Likewise, the relative abundance of denitrifiers was not altered in P. flava but decreased by 1 order of magnitude in X. umbellata under excess DOC, likely due to decreased proportions of Ruegeria spp. Given that holobiont C:N ratios remained stable in P. flava but showed a pronounced increase with excess DOC in X. umbellata , our results suggest that microbial community dynamics may reflect the nutritional status of the holobiont. Hence, denitrifier abundance may be directly linked to N availability. This suggests a passive regulation of N cycling microbes based on N availability, which could help stabilize nutrient limitation in the coral-algal symbiosis and thereby support holobiont functioning in a changing environment. IMPORTANCE Octocorals are important members of reef-associated benthic communities that can rapidly replace scleractinian corals as the dominant ecosystem engineers on degraded reefs. Considering the substantial change in the (a)biotic environment that is commonly driving reef degradation, maintaining a dynamic and metabolically diverse microbial community might contribute to octocoral acclimatization. Nitrogen (N) cycling microbes, in particular denitrifying prokaryotes, may support holobiont functioning by limiting internal N availability, but little is known about the identity and (a)biotic drivers of octocoral-associated denitrifiers. Here, we show contrasting dynamics of bacterial communities associated with two common octocoral species, the soft coral Xenia umbellata and the gorgonian Pinnigorgia flava after a 6-week exposure to excess dissolved organic carbon under concomitant warming conditions. The specific responses of denitrifier communities of the two octocoral species aligned with the nutritional status of holobiont members. This suggests a passive regulation based on N availability in the coral holobiont.

Our understanding of the response of reef-building corals to changes in their physical environment is largely based on laboratory experiments, analysis of long-term field data, and model projections. Experimental data provide unique insights into how organisms respond to variation of environmental drivers. However, an assessment of how well experimental conditions cover the breadth of environmental conditions and variability where corals live successfully is missing. Here, we compiled and analyzed a globally distributed dataset of in-situ seasonal and diurnal variability of key environmental drivers (temperature, pCO 2 , and O 2 ) critical for the growth and livelihood of reef-building corals. Using a meta-analysis approach, we compared the variability of environmental conditions assayed in coral experimental studies to current and projected conditions in their natural habitats. We found that annual temperature profiles projected for the end of the 21st century were characterized by distributional shifts in temperatures with warmer winters and longer warm periods in the summer, not just peak temperatures. Furthermore, short-term hourly fluctuations of temperature and pCO 2 may regularly expose corals to conditions beyond the projected average increases for the end of the 21st century. Coral reef sites varied in the degree of coupling between temperature, pCO 2 , and dissolved O 2 , which warrants site-specific, differentiated experimental approaches depending on the local hydrography and influence of biological processes on the carbonate system and O 2 availability. Our analysis highlights that a large portion of the natural environmental variability at short and long timescales is underexplored in experimental designs, which may provide a path to extend our understanding on the response of corals to global climate change., (© 2021 The Authors. Global Change Biology published by John Wiley & Sons Ltd.)

Ocean acidification is posing a threat to calcifying organisms due to the increased energy requirements of calcification under high CO 2 conditions. The ability of scleractinian corals to cope with future ocean conditions will thus depend on their ability to fulfil their carbon requirement. However, the primary productivity of coral holobionts is limited by low nitrogen (N) availability in coral reef waters. Here, we employed CO 2  seeps of Tutum Bay (Papua New Guinea) as a natural laboratory to understand how coral holobionts offset their increased energy requirements under high CO 2 conditions. Our results demonstrate for the first time that under high pCO 2 conditions, N assimilation pathways of Pocillopora damicornis are jointly modified. We found that diazotroph-derived N assimilation rates in the Symbiodiniaceae were significantly higher in comparison to an ambient CO 2 control site, concomitant with a restructured diazotroph community and the specific prevalence of an alpha-proteobacterium. Further, corals at the high CO 2  site also had increased feeding rates on picoplankton and in particular exhibited selective feeding on Synechococcus sp., known to be rich in N. Given the high abundance of picoplankton in oligotrophic waters at large, our results suggest that corals exhibiting flexible diazotrophic communities and capable of exploiting N-rich picoplankton sources to offset their increased N requirements may be able to cope better in a high pCO 2 world., (© 2021 The Authors. Molecular Ecology published by John Wiley & Sons Ltd.)

Nitrogen cycling in coral reefs may be affected by nutrient availability, but knowledge about concentration-dependent thresholds that modulate dinitrogen fixation and denitrification is missing. We determined the effects of different nitrate concentrations (ambient, 1, 5, 10 μM nitrate addition) on both processes under two light scenarios (i.e., light and dark) using a combined acetylene assay for two common benthic reef substrates, i.e., turf algae and coral rubble. For both substrates, dinitrogen fixation rates peaked at 5 μM nitrate addition in light, whereas denitrification was highest at 10 μM nitrate addition in the dark. At 10 μm nitrate addition in the dark, a near-complete collapse of dinitrogen fixation concurrent with a 76-fold increase in denitrification observed for coral rubble, suggesting potential threshold responses linked to the nutritional state of the community. We conclude that dynamic nitrogen cycling activity may help stabilise nitrogen availability in microbial communities associated with coral reef substrates., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Ecosystem services provided by coral reefs may be susceptible to the combined effects of benthic species shifts and anthropogenic nutrient pollution, but related field studies are scarce. We thus investigated in situ how dissolved inorganic nutrient enrichment, maintained for two months, affected community-wide biogeochemical functions of intact coral- and degraded algae-dominated reef patches in the central Red Sea. Results from benthic chamber incubations revealed 87% increased gross productivity and a shift from net calcification to dissolution in algae-dominated communities after nutrient enrichment, but the same processes were unaffected by nutrients in neighboring coral communities. Both community types changed from net dissolved organic nitrogen sinks to sources, but the increase in net release was 56% higher in algae-dominated communities. Nutrient pollution may, thus, amplify the effects of community shifts on key ecosystem services of coral reefs, possibly leading to a loss of structurally complex habitats with carbonate dissolution and altered nutrient recycling., (Copyright © 2021 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Coral reefs experience phase shifts from coral- to algae-dominated benthic communities, which could affect the interplay between processes introducing and removing bioavailable nitrogen. However, the magnitude of such processes, i.e., dinitrogen (N 2 ) fixation and denitrification levels, and their responses to phase shifts remain unknown in coral reefs. We assessed both processes for the dominant species of six benthic categories (hard corals, soft corals, turf algae, coral rubble, biogenic rock, and reef sands) accounting for > 98% of the benthic cover of a central Red Sea coral reef. Rates were extrapolated to the relative benthic cover of the studied organisms in co-occurring coral- and algae-dominated areas of the same reef. In general, benthic categories with high N 2 fixation exhibited low denitrification activity. Extrapolated to the respective reef area, turf algae and coral rubble accounted for > 90% of overall N 2 fixation, whereas corals contributed to more than half of reef denitrification. Total N 2 fixation was twice as high in algae- compared to coral-dominated areas, whereas denitrification levels were similar. We conclude that algae-dominated reefs promote new nitrogen input through enhanced N 2 fixation and comparatively low denitrification. The subsequent increased nitrogen availability could support net productivity, resulting in a positive feedback loop that increases the competitive advantage of algae over corals in reefs that experienced a phase shift.

Recent research suggests that nitrogen (N) cycling microbes are important for coral holobiont functioning. In particular, coral holobionts may acquire bioavailable N via prokaryotic dinitrogen (N 2 ) fixation or remove excess N via denitrification activity. However, our understanding of environmental drivers on these processes in hospite remains limited. Employing the strong seasonality of the central Red Sea, this study assessed the effects of environmental parameters on the proportional abundances of N cycling microbes associated with the hard corals Acropora hemprichii and Stylophora pistillata. Specifically, we quantified changes in the relative ratio between nirS and nifH gene copy numbers, as a proxy for seasonal shifts in denitrification and N 2 fixation potential in corals, respectively. In addition, we assessed coral tissue-associated Symbiodiniaceae cell densities and monitored environmental parameters to provide a holobiont and environmental context, respectively. While ratios of nirS to nifH gene copy numbers varied between seasons, they revealed similar seasonal patterns in both coral species, with ratios closely following patterns in environmental nitrate availability. Symbiodiniaceae cell densities aligned with environmental nitrate availability, suggesting that the seasonal shifts in nirS to nifH gene abundance ratios were probably driven by nitrate availability in the coral holobiont. Thereby, our results suggest that N cycling in coral holobionts probably adjusts to environmental conditions by increasing and/or decreasing denitrification and N 2 fixation potential according to environmental nitrate availability. Microbial N cycling may, thus, extenuate the effects of changes in environmental nitrate availability on coral holobionts to support the maintenance of the coral-Symbiodiniaceae symbiosis., (© 2021 The Authors.)

Recurrent mass bleaching events are pushing coral reefs worldwide to the brink of ecological collapse. While the symptoms and consequences of this breakdown of the coral-algal symbiosis have been extensively characterized, our understanding of the underlying causes remains incomplete. Here, we investigated the nutrient fluxes and the physiological as well as molecular responses of the widespread coral Stylophora pistillata to heat stress prior to the onset of bleaching to identify processes involved in the breakdown of the coral-algal symbiosis. We show that altered nutrient cycling during heat stress is a primary driver of the functional breakdown of the symbiosis. Heat stress increased the metabolic energy demand of the coral host, which was compensated by the catabolic degradation of amino acids. The resulting shift from net uptake to release of ammonium by the coral holobiont subsequently promoted the growth of algal symbionts and retention of photosynthates. Together, these processes form a feedback loop that will gradually lead to the decoupling of carbon translocation from the symbiont to the host. Energy limitation and altered symbiotic nutrient cycling are thus key factors in the early heat stress response, directly contributing to the breakdown of the coral-algal symbiosis. Interpreting the stability of the coral holobiont in light of its metabolic interactions provides a missing link in our understanding of the environmental drivers of bleaching and may ultimately help uncover fundamental processes underpinning the functioning of endosymbioses in general., Competing Interests: The authors declare no competing interest., (Copyright © 2021 the Author(s). Published by PNAS.)

The upside-down jellyfish Cassiopea engages in symbiosis with photosynthetic microalgae that facilitate uptake and recycling of inorganic nutrients. By contrast to most other symbiotic cnidarians, algal endosymbionts in Cassiopea are not restricted to the gastroderm but are found in amoebocyte cells within the mesoglea. While symbiont-bearing amoebocytes are highly abundant, their role in nutrient uptake and cycling in Cassiopea remains unknown. By combining isotopic labelling experiments with correlated scanning electron microscopy, and Nano-scale secondary ion mass spectrometry (NanoSIMS) imaging, we quantified the anabolic assimilation of inorganic carbon and nitrogen at the subcellular level in juvenile Cassiopea medusae bell tissue. Amoebocytes were clustered near the sub-umbrella epidermis and facilitated efficient assimilation of inorganic nutrients. Photosynthetically fixed carbon was efficiently translocated between endosymbionts, amoebocytes and host epidermis at rates similar to or exceeding those observed in corals. The Cassiopea holobionts efficiently assimilated ammonium, while no nitrate assimilation was detected, possibly reflecting adaptation to highly dynamic environmental conditions of their natural habitat. The motile amoebocytes allow Cassiopea medusae to distribute their endosymbiont population to optimize access to light and nutrients, and transport nutrition between tissue areas. Amoebocytes thus play a vital role for the assimilation and translocation of nutrients in Cassiopea , providing an interesting new model for studies of metabolic interactions in photosymbiotic marine organisms.

Coral bleaching is one of the main drivers of reef degradation. Most corals bleach and suffer mortality at just 1-2°C above their maximum monthly mean temperatures, but some species and genotypes resist or recover better than others. Here, we conducted a series of 18-hr short-term acute heat stress assays side-by-side with a 21-day long-term heat stress experiment to assess the ability of both approaches to resolve coral thermotolerance differences reflective of in situ reef temperature thresholds. Using a suite of physiological parameters (photosynthetic efficiency, coral whitening, chlorophyll a, host protein, algal symbiont counts, and algal type association), we assessed bleaching susceptibility of Stylophora pistillata colonies from the windward/exposed and leeward/protected sites of a nearshore coral reef in the central Red Sea, which had previously shown differential mortality during a natural bleaching event. Photosynthetic efficiency was most indicative of the expected higher thermal tolerance in corals from the protected reef site, denoted by an increased retention of dark-adapted maximum quantum yields at higher temperatures. These differences were resolved using both experimental setups, as corroborated by a positive linear relationship, not observed for the other parameters. Notably, short-term acute heat stress assays resolved per-colony (genotype) differences that may have been masked by acclimation effects in the long-term experiment. Using our newly developed portable experimental system termed the Coral Bleaching Automated Stress System (CBASS), we thus highlight the potential of mobile, standardized short-term acute heat stress assays to resolve fine-scale differences in coral thermotolerance. Accordingly, such a system may be suitable for large-scale determination and complement existing approaches to identify resilient genotypes/reefs for downstream experimental examination and prioritization of reef sites for conservation/restoration. Development of such a framework is consistent with the recommendations of the National Academy of Sciences and the Reef Restoration and Adaptation Program committees for new intervention and restoration strategies., (© 2020 The Authors. Global Change Biology published by John Wiley & Sons Ltd.)

Reef-building corals harbour an astonishing diversity of microorganisms, including endosymbiotic microalgae, bacteria, archaea, and fungi. The metabolic interactions within this symbiotic consortium are fundamental to the ecological success of corals and the unique productivity of coral reef ecosystems. Over the last two decades, scientific efforts have been primarily channelled into dissecting the symbioses occurring in coral tissues. Although easily accessible, this compartment is only 2-3 mm thick, whereas the underlying calcium carbonate skeleton occupies the vast internal volume of corals. Far from being devoid of life, the skeleton harbours a wide array of algae, endolithic fungi, heterotrophic bacteria, and other boring eukaryotes, often forming distinct bands visible to the bare eye. Some of the critical functions of these endolithic microorganisms in coral health, such as nutrient cycling and metabolite transfer, which could enable the survival of corals during thermal stress, have long been demonstrated. In addition, some of these microorganisms can dissolve calcium carbonate, weakening the coral skeleton and therefore may play a major role in reef erosion. Yet, experimental data are wanting due to methodological limitations. Recent technological and conceptual advances now allow us to tease apart the complex physical, ecological, and chemical interactions at the heart of coral endolithic microbial communities. These new capabilities have resulted in an excellent body of research and provide an exciting outlook to further address the functional microbial ecology of the "overlooked" coral skeleton.

Denitrification may potentially alleviate excess nitrogen (N) availability in coral holobionts to maintain a favourable N to phosphorous ratio in the coral tissue. However, little is known about the abundance and activity of denitrifiers in the coral holobiont. The present study used the nirS marker gene as a proxy for denitrification potential along with measurements of denitrification rates in a comparative coral taxonomic framework from the Red Sea: Acropora hemprichii, Millepora dichotoma, and Pleuractis granulosa. Relative nirS gene copy numbers associated with the tissues of these common corals were assessed and compared with denitrification rates on the holobiont level. In addition, dinitrogen (N 2 ) fixation rates, Symbiodiniaceae cell density, and oxygen evolution were assessed to provide an environmental context for denitrification. We found that relative abundances of the nirS gene were 16- and 17-fold higher in A. hemprichii compared to M. dichotoma and P. granulosa, respectively. In concordance, highest denitrification rates were measured in A. hemprichii, followed by M. dichotoma and P. granulosa. Denitrification rates were positively correlated with N 2 fixation rates and Symbiodiniaceae cell densities. Our results suggest that denitrification may counterbalance the N input from N 2 fixation in the coral holobiont, and we hypothesize that these processes may be limited by photosynthates released by the Symbiodiniaceae.

Coral reefs are in global decline mainly due to increasing sea surface temperatures triggering coral bleaching. Recently, high salinity has been linked to increased thermotolerance and decreased bleaching in the sea anemone coral model Aiptasia. However, the underlying processes remain elusive. Using two Aiptasia host--endosymbiont pairings, we induced bleaching at different salinities and show reduced reactive oxygen species (ROS) release at high salinities, suggesting a role of osmoadaptation in increased thermotolerance. A subsequent screening of osmolytes revealed that this effect was only observed in algal endosymbionts that produce 2-O-glycerol-α-D-galactopyranoside (floridoside), an osmolyte capable of scavenging ROS. This result argues for a mechanistic link between osmoadaptation and thermotolerance, mediated by ROS-scavenging osmolytes (e.g., floridoside). This sheds new light on the putative mechanisms underlying the remarkable thermotolerance of corals from water bodies with high salinity such as the Red Sea or Persian/Arabian Gulf and holds implications for coral thermotolerance under climate change.This article has an associated First Person interview with the first author of the paper., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2019. Published by The Company of Biologists Ltd.)

[This corrects the article on p. 214 in vol. 9, PMID: 29615919.].

From protists to humans, all animals and plants are inhabited by microbial organisms. There is an increasing appreciation that these resident microbes influence the fitness of their plant and animal hosts, ultimately forming a metaorganism consisting of a uni- or multicellular host and a community of associated microorganisms. Research on host-microbe interactions has become an emerging cross-disciplinary field. In both vertebrates and invertebrates a complex microbiome confers immunological, metabolic and behavioural benefits; conversely, its disturbance can contribute to the development of disease states. However, the molecular and cellular mechanisms controlling the interactions within a metaorganism are poorly understood and many key interactions between the associated organisms remain unknown. In this perspective article, we outline some of the issues in interspecies interactions and in particular address the question of how metaorganisms react and adapt to inputs from extreme environments such as deserts, the intertidal zone, oligothrophic seas, and hydrothermal vents., (Copyright © 2018 The Authors. Published by Elsevier GmbH.. All rights reserved.)

The symbiosis between cnidarian hosts and microalgae of the genus Symbiodinium provides the foundation of coral reefs in oligotrophic waters. Understanding the nutrient-exchange between these partners is key to identifying the fundamental mechanisms behind this symbiosis, yet has proven difficult given the endosymbiotic nature of this relationship. In this study, we investigated the respective contribution of host and symbiont to carbon and nitrogen assimilation in the coral model anemone Aiptaisa. For this, we combined traditional measurements with nanoscale secondary ion mass spectrometry (NanoSIMS) and stable isotope labeling to investigate patterns of nutrient uptake and translocation both at the organismal scale and at the cellular scale. Our results show that the rate of carbon and nitrogen assimilation in Aiptasia depends on the identity of the host and the symbiont. NanoSIMS analysis confirmed that both host and symbiont incorporated carbon and nitrogen into their cells, implying a rapid uptake and cycling of nutrients in this symbiotic relationship. Gross carbon fixation was highest in Aiptasia associated with their native Symbiodinium communities. However, differences in fixation rates were only reflected in the δ 13 C enrichment of the cnidarian host, whereas the algal symbiont showed stable enrichment levels regardless of host identity. Thereby, our results point toward a "selfish" character of the cnidarian- Symbiodinium association in which both partners directly compete for available resources. Consequently, this symbiosis may be inherently instable and highly susceptible to environmental change. While questions remain regarding the underlying cellular controls of nutrient exchange and the nature of metabolites involved, the approach outlined in this study constitutes a powerful toolset to address these questions.

The importance of Symbiodinium algal endosymbionts and a diverse suite of bacteria for coral holobiont health and functioning are widely acknowledged. Yet, we know surprisingly little about microbial community dynamics and the stability of host-microbe associations under adverse environmental conditions. To gain insight into the stability of coral host-microbe associations and holobiont structure, we assessed changes in the community structure of Symbiodinium and bacteria associated with the coral Pocillopora verrucosa under excess organic nutrient conditions. Pocillopora -associated microbial communities were monitored over 14 days in two independent experiments. We assessed the effect of excess dissolved organic nitrogen (DON) and excess dissolved organic carbon (DOC). Exposure to excess nutrients rapidly affected coral health, resulting in two distinct stress phenotypes: coral bleaching under excess DOC and severe tissue sloughing (>90% tissue loss resulting in host mortality) under excess DON. These phenotypes were accompanied by structural changes in the Symbiodinium community. In contrast, the associated bacterial community remained remarkably stable and was dominated by two Endozoicomonas phylotypes, comprising on average 90% of 16S rRNA gene sequences. This dominance of Endozoicomonas even under conditions of coral bleaching and mortality suggests the bacterial community of P. verrucosa may be rather inflexible and thereby unable to respond or acclimatize to rapid changes in the environment, contrary to what was previously observed in other corals. In this light, our results suggest that coral holobionts might occupy structural landscapes ranging from a highly flexible to a rather inflexible composition with consequences for their ability to respond to environmental change.

Coastal pollution and algal cover are increasing on many coral reefs, resulting in higher dissolved organic carbon (DOC) concentrations. High DOC concentrations strongly affect microbial activity in reef waters and select for copiotrophic, often potentially virulent microbial populations. High DOC concentrations on coral reefs are also hypothesized to be a determinant for switching microbial lifestyles from commensal to pathogenic, thereby contributing to coral reef degradation, but evidence is missing. In this study, we conducted ex situ incubations to assess gene expression of planktonic microbial populations under elevated concentrations of naturally abundant monosaccharides (glucose, galactose, mannose, and xylose) in algal exudates and sewage inflows. We assembled 27 near-complete (>70%) microbial genomes through metagenomic sequencing and determined associated expression patterns through metatranscriptomic sequencing. Differential gene expression analysis revealed a shift in the central carbohydrate metabolism and the induction of metalloproteases, siderophores, and toxins in Alteromonas, Erythrobacter, Oceanicola, and Alcanivorax populations. Sugar-specific induction of virulence factors suggests a mechanistic link for the switch from a commensal to a pathogenic lifestyle, particularly relevant during increased algal cover and human-derived pollution on coral reefs. Although an explicit test remains to be performed, our data support the hypothesis that increased availability of specific sugars changes net microbial community activity in ways that increase the emergence and abundance of opportunistic pathogens, potentially contributing to coral reef degradation.

The endosymbiosis between dinoflagellate algae of the genus Symbiodinium and stony corals provides the foundation of coral reef ecosystems. Coral bleaching, the expulsion of endosymbionts from the coral host tissue as a consequence of heat or light stress, poses a threat to reef ecosystem functioning on a global scale. Hence, a better understanding of the factors contributing to heat stress susceptibility and tolerance is needed. In this regard, some of the most thermotolerant corals live in particularly saline habitats, but possible effects of high salinity on thermotolerance in corals are anecdotal. Here we test the hypothesis that high salinity may lead to increased thermotolerance. We conducted a heat stress experiment at low, intermediate, and high salinities using a set of host-endosymbiont combinations of the coral model Aiptasia. As expected, all host-endosymbiont combinations showed reduced photosynthetic efficiency and endosymbiont loss during heat stress, but the severity of bleaching was significantly reduced with increasing salinities for one of the host-endosymbiont combinations. Our results show that higher salinities can convey increased thermotolerance in Aiptasia, although this effect seems to be dependent on the particular host strain and/or associated symbiont type. This finding may help explain the extraordinarily high thermotolerance of corals in high salinity environments, such as the Red Sea and the Persian/Arabian Gulf, and provides novel insight regarding factors that contribute to thermotolerance. Since our results are based on a salinity effect in symbiotic sea anemones, it remains to be determined whether this salinity effect can also be observed in stony corals., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2017. Published by The Company of Biologists Ltd.)

The disruption of the coral-algae symbiosis (coral bleaching) due to rising sea surface temperatures has become an unprecedented global threat to coral reefs. Despite decades of research, our ability to manage mass bleaching events remains hampered by an incomplete mechanistic understanding of the processes involved. In this study, we induced a coral bleaching phenotype in the absence of heat and light stress by adding sugars. The sugar addition resulted in coral symbiotic breakdown accompanied by a fourfold increase of coral-associated microbial nitrogen fixation. Concomitantly, increased N:P ratios by the coral host and algal symbionts suggest excess availability of nitrogen and a disruption of the nitrogen limitation within the coral holobiont. As nitrogen fixation is similarly stimulated in ocean warming scenarios, here we propose a refined coral bleaching model integrating the cascading effects of stimulated microbial nitrogen fixation. This model highlights the putative role of nitrogen-fixing microbes in coral holobiont functioning and breakdown., (© 2017 The Authors. Global Change Biology Published by John Wiley & Sons Ltd.)

The productivity of coral reefs in oligotrophic tropical waters is sustained by an efficient uptake and recycling of nutrients. In reef-building corals, the engineers of these ecosystems, this nutrient recycling is facilitated by a constant exchange of nutrients between the animal host and endosymbiotic photosynthetic dinoflagellates (zooxanthellae), bacteria, and other microbes. Due to the complex interactions in this so-called coral holobiont, it has proven difficult to understand the environmental limitations of productivity in corals. Among others, the micronutrient iron has been proposed to limit primary productivity due to its essential role in photosynthesis and bacterial processes. Here, we tested the effect of iron enrichment on the physiology of the coral Pocillopora verrucosa from the central Red Sea during a 12-day experiment. Contrary to previous reports, we did not see an increase in zooxanthellae population density or gross photosynthesis. Conversely, respiration rates were significantly increased, and microbial nitrogen fixation was significantly decreased. Taken together, our data suggest that iron is not a limiting factor of primary productivity in Red Sea corals. Rather, increased metabolic demands in response to iron enrichment, as evidenced by increased respiration rates, may reduce carbon (i.e., energy) availability in the coral holobiont, resulting in reduced microbial nitrogen fixation. This decrease in nitrogen supply in turn may exacerbate the limitation of other nutrients, creating a negative feedback loop. Thereby, our results highlight that the effects of iron enrichment appear to be strongly dependent on local environmental conditions and ultimately may depend on the availability of other nutrients.

Microbial nitrogen fixation (diazotrophy) is a functional trait widely associated with tropical reef-building (scleractinian) corals. While the integral role of nitrogen fixation in coral nutrient dynamics is recognized, its ecological significance across different coral functional groups remains yet to be evaluated. Here we set out to compare molecular and physiological patterns of diazotrophy (i.e., nifH gene abundance and expression as well as nitrogen fixation rates) in two coral families with contrasting trophic strategies: highly heterotrophic, free-living members of the family Fungiidae ( Pleuractis granulosa , Ctenactis echinata ), and mostly autotrophic coral holobionts with low heterotrophic capacity (Pocilloporidae: Pocillopora verrucosa , Stylophora pistillata ). The Fungiidae exhibited low diazotroph abundance (based on nifH gene copy numbers) and activity (based on nifH gene expression and the absence of detectable nitrogen fixation rates). In contrast, the mostly autotrophic Pocilloporidae exhibited nifH gene copy numbers and gene expression two orders of magnitude higher than in the Fungiidae, which coincided with detectable nitrogen fixation activity. Based on these data, we suggest that nitrogen fixation compensates for the low heterotrophic nitrogen uptake in autotrophic corals. Consequently, the ecological importance of diazotrophy in coral holobionts may be determined by the trophic functional group of the host.

Corals are animals that form close mutualistic associations with endosymbiotic photosynthetic algae of the genus Symbiodinium. Together they provide the calcium carbonate framework of coral reef ecosystems. The importance of the microbiome (i.e., bacteria, archaea, fungi, and viruses) to holobiont functioning has only recently been recognized. Given that growth and density of Symbiodinium within the coral host is highly dependent on nitrogen availability, nitrogen-cycling microbes may be of fundamental importance to the stability of the coral-algae symbiosis and holobiont functioning, in particular under nutrient-enriched and -depleted scenarios. We summarize what is known about nitrogen cycling in corals and conclude that disturbance of microbial nitrogen cycling may be tightly linked to coral bleaching and disease., (Copyright © 2015 Elsevier Ltd. All rights reserved.)