Chemosynthetic symbioses are partnerships between invertebrate animals and chemosynthetic bacteria. The latter are the primary producers, providing most of the organic carbon needed for the animal host's nutrition. We sequenced genomes of the chemosynthetic symbionts from the lucinid bivalve Loripes lucinalis and the stilbonematid nematode Laxus oneistus. The symbionts of both host species encoded nitrogen fixation genes. This is remarkable as no marine chemosynthetic symbiont was previously known to be capable of nitrogen fixation. We detected nitrogenase expression by the symbionts of lucinid clams at the transcriptomic and proteomic level. Mean stable nitrogen isotope values of Loripes lucinalis were within the range expected for fixed atmospheric nitrogen, further suggesting active nitrogen fixation by the symbionts. The ability to fix nitrogen may be widespread among chemosynthetic symbioses in oligotrophic habitats, where nitrogen availability often limits primary productivity.S ymbioses between animals and chemosynthetic bacteria are widespread in Earth's oceans 1 . Animals from at least seven phyla have formed such symbioses, and even more chemosynthetic bacterial lineages have evolved symbioses with animal hosts 1 . Chemosynthetic symbionts can use a range of chemicals, such as sulfide, methane, hydrogen and carbon monoxide, to power their metabolism 2-4 . The hosts of chemosynthetic symbionts dominate some animal communities. For example, shallow-water lucinid bivalves, which host sulfur-oxidizing symbionts, often dominate the macrobenthic infaunal community in seagrass meadows, where they can reach densities greater than 3,500 individuals per square metre 5,6 . Their diversity in nature, their persistence over evolutionary timescales and their dominance in many habitats attest to the success of these symbiotic partnerships 1 .Chemosynthetic symbionts are primarily considered 'nutritional symbionts', meaning their primary role is to provide nutrition for their hosts 1,7 . So far, most studies have focused on inorganic carbon fixation by the symbionts and the transfer of fixed organic carbon compounds to the hosts. In addition to organic carbon, all animals require a source of fixed nitrogen. However, nitrogen metabolism in chemosynthetic symbioses has received far less attention. Chemosynthetic symbionts have been shown to gain their nitrogen from ammonium or nitrate in their environment 8-10 and co-occurring nitrogen-fixing and chemosynthetic symbionts have been found in cold-water corals 11 . Nitrogen fixation by chemosynthetic symbionts has long been hypothesized, but so far not yet shown [12][13][14] .Our study focused mainly on the endosymbiosis between bivalves of the family Lucinidae and sulfur-oxidizing bacteria. Lucinids are by far the most diverse and widespread group of bivalves that host chemosynthetic symbionts 15 . There are at least 400 living species, occupying a range of habitats including mangrove sediments, seagrass beds, coral reef sediments and coastal mud and sand 16 . In seagrass...

Functional traits define species by their ecological role in the ecosystem. Animals themselves are host–microbe ecosystems (holobionts), and the application of ecophysiological approaches can help to understand their functioning. In hard coral holobionts, communities of dinitrogen (N 2 )-fixing prokaryotes (diazotrophs) may contribute a functional trait by providing bioavailable nitrogen (N) that could sustain coral productivity under oligotrophic conditions. This study quantified N 2 fixation by diazotrophs associated with four genera of hermatypic corals on a northern Red Sea fringing reef exposed to high seasonality. We found N 2 fixation activity to be 5- to 10-fold higher in summer, when inorganic nutrient concentrations were lowest and water temperature and light availability highest. Concurrently, coral gross primary productivity remained stable despite lower Symbiodinium densities and tissue chlorophyll a contents. In contrast, chlorophyll a content per Symbiodinium cell increased from spring to summer, suggesting that algal cells overcame limitation of N, an essential element for chlorophyll synthesis. In fact, N 2 fixation was positively correlated with coral productivity in summer, when its contribution was estimated to meet 11% of the Symbiodinium N requirements. These results provide evidence of an important functional role of diazotrophs in sustaining coral productivity when alternative external N sources are scarce.

Tropical coral reefs are among the most productive and diverse ecosystems, despite being surrounded by ocean waters where nutrients are in short supply. Benthic dinitrogen (N2) fixation is a significant internal source of “new” nitrogen (N) in reef ecosystems, but related information appears to be sparse. Here, we review the current state (and gaps) of knowledge on N2 fixation associated with coral reef organisms and their ecosystems. By summarizing the existing literature, we show that benthic N2 fixation is an omnipresent process in tropical reef environments. Highest N2 fixation rates are detected in reef-associated cyanobacterial mats and sea grass meadows, clearly showing the significance of these functional groups, if present, to the input of new N in reef ecosystems. Nonetheless, key benthic organisms such as hard corals also importantly contribute to benthic N2 fixation in the reef. Given the usually high coral coverage of healthy reef systems, these results indicate that benthic symbiotic associations may be more important than previously thought. In fact, mutualisms between carbon (C) and N2 fixers have likely evolved that may enable reef communities to mitigate N limitation. We then explore the potential effects of the increasing human interferences on the process of benthic reef N2 fixation via changes in diazotrophic populations, enzymatic activities, or availability of benthic substrates favorable to these microorganisms. Current knowledge indicates positive effects of ocean acidification, warming, and deoxygenation and negative effects of increased ultraviolet radiation on the amount of N fixed in coral reefs. Eutrophication may either boost or suppress N2 fixation, depending on the nutrient becoming limiting. As N2 fixation appears to play a fundamental role in nutrient-limited reef ecosystems, these assumptions need to be expanded and confirmed by future research efforts addressing the knowledge gaps identified in this review.

The red coral Corallium rubrum is a habitat-forming species with a prominent and structural role in mesophotic habitats, which sustains biodiversity hotspots. This precious coral is threatened by both over-exploitation and temperature driven mass mortality events. We report here that biocalcification, growth rates and polyps' (feeding) activity of Corallium rubrum are significantly reduced at pCO2 scenarios predicted for the end of this century (0.2 pH decrease). Since C. rubrum is a long-living species (>200 years), our results suggest that ocean acidification predicted for 2100 will significantly increases the risk of extinction of present populations. Given the functional role of these corals in the mesophotic zone, we predict that ocean acidification might have cascading effects on the functioning of these habitats worldwide.

Biological dinitrogen (N 2 ) fixation (diazotrophy, BNF) relieves marine primary producers of nitrogen (N) limitation in a large part of the world oceans. N concentrations are particularly low in tropical regions where coral reefs are located, and N is therefore a key limiting nutrient for these productive ecosystems. In this context, the importance of diazotrophy for reef productivity is still not resolved, with studies up to now lacking organismal and seasonal resolution. Here, we present a budget of gross primary production (GPP) and BNF for a highly seasonal Red Sea fringing reef, based on ecophysiological and benthic cover measurements combined with geospatial analyses. Benthic GPP varied from 215 to 262 mmol C m . Planktonic GPP and BNF rates were respectively approximately 60-and 20-fold lower than those of the benthos, emphasizing the importance of the benthic compartment in reef biogeochemical cycling. BNF showed higher sensitivity to seasonality than GPP, implying greater climatic control on reef BNF. Up to about 20% of net reef primary production could be supported by BNF during summer, suggesting a strong biogeochemical coupling between diazotrophy and the reef carbon cycle.

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Coral holobionts (i.e., coral-algal-prokaryote symbioses) exhibit dissimilar thermal sensitivities that may determine which coral species will adapt to global warming. Nonetheless, studies simultaneously investigating the effects of warming on all holobiont members are lacking. Here we show that exposure to increased temperature affects key physiological traits of all members (herein: animal host, zooxanthellae and diazotrophs) of both Stylophora pistillata and Acropora hemprichii during and after thermal stress. S. pistillata experienced severe loss of zooxanthellae (i.e., bleaching) with no net photosynthesis at the end of the experiment. Conversely, A. hemprichii was more resilient to thermal stress. Exposure to increased temperature (+ 6°C) resulted in a drastic increase in daylight dinitrogen (N2 ) fixation, particularly in A. hemprichii (threefold compared with controls). After the temperature was reduced again to in situ levels, diazotrophs exhibited a reversed diel pattern of activity, with increased N2 fixation rates recorded only in the dark, particularly in bleached S. pistillata (twofold compared to controls). Concurrently, both animal hosts, but particularly bleached S. pistillata, reduced both organic matter release and heterotrophic feeding on picoplankton. Our findings indicate that physiological plasticity by coral-associated diazotrophs may play an important role in determining the response of coral holobionts to ocean warming.

Since productivity and growth of coral-associated dinoflagellate algae is nitrogen (N)-limited, dinitrogen (N 2 ) fixation by coral-associated microbes is likely crucial for maintaining the coral−dinoflagellate symbiosis. It is thus essential to understand the effects future climate change will have on N 2 fixation by the coral holobiont. This laboratory study is the first to investigate short-term effects of ocean acidification on N 2 fixation activity associated with the tropical, hermatypic coral Seriatopora hystrix using the acetylene reduction assay in combination with calcification measurements. Findings reveal that simulated ocean acidification ( pCO 2 1080 µatm) caused a rapid and significant decrease (53%) in N 2 fixation rates associated with S. hystrix compared to the present day scenario ( pCO 2 486 µatm). In addition, N 2 fixation associated with the coral holobiont showed a positive exponential relationship with its calcification rates. This suggests that even small declines in calcification rates of hermatypic corals under high CO 2 conditions may result in decreased N 2 fixation activity, since these 2 processes may compete for energy in the coral holobiont. Ultimately, an intensified N limitation in combination with a decline in skeletal growth may trigger a negative feedback loop on coral productivity exacerbating the negative long-term effects of ocean acidification.