Nitrate enrichment & heat stress impacts the physiology of the coral A. kenti and the composition of its associated microbiome

Coral reefs are unarguably under increasing pressures arising from various environmental stressors. Coral survival in the face of environmental change relies heavily on nutrient exchanges between the host and the photosynthetic endosymbionts. While the functional contribution of the coral microbiome remains poorly understood, increasing evidence suggests that associated microorganisms are essential for coral resilience as they are intricately linked to nutrient cycling and energy flows in the ecosystem. Nitrogen underpins many aspects of coral holobiont functioning but the effect of its availability in its most abundant environmental form, nitrate, on the coral response to stress is equivocal: while nitrate sustains symbiont communities, it has also been reported to have adverse effects on the response to oxidative stress and to accentuate bleaching. In this study, using a crossed treatment experimental design in a mesocosm setup, we investigated the responses of the coral Acropora kenti to a nitrate enrichment of 5 µM in combination with a heat stress of 4 DHW over a period of 3 weeks. Corals’ health was monitored throughout the experiment and corals’ physiological response to the different treatments was assessed at the start of the stress and at the end of the experiment by measuring respiration rates, photosynthetic capacity, growth rates, symbiont densities, pigment and protein contents. In addition, corals were sampled to identify the composition of the associated symbiont and microbial communities using high-throughput sequencing of the genes ITS2 and 16S respectively. The heat stress treatment induced moderate to severe bleaching that was not alleviated by the increased nitrate supply. Nuances in the physiological data and the integration with the sequencing data give valuable inputs into the holobiont’s functioning by disentangling the effect of nitrate availability and heat stress on the resilience of the coral and the stability of its associated symbiotic and microbial communities

Nitrate in the coral symbiosis: from the regulation of its assimilation to its impact on the physiology of the holobiont

In oligotrophic reef systems, coral holobionts are remarkably efficient at assimilating nitrogen through heterotrophic feeding or the uptake of dissolved inorganic nitrogen. Symbiodiniaceae are vital partners of the symbiosis for nutrient assimilation. In addition to providing translocated photosynthates, they account for most of the uptake of dissolved inorganic nitrogen. Although NO3- is the most abundant source of nitrogen in the ocean, little is known about the mechanisms regulating its assimilation by the holobiont. Coral hosts are unable to reduce nitrate as they lack the necessary enzymes, whereas Symbiodiniaceae have been shown to express the enzyme nitrate reductase (NR). However, the evidence supporting the active reduction of nitrate by the symbiotic algae during symbiosis is scarce and equivocal. This research aimed at deciphering the pathways of NO3- assimilation in both free-living Symbiodiniaceae and in hospite symbionts while also investigating the relevance of inorganic nitrogen source in physiological responses to stress. We investigated the expression and regulation of NR both in free-living Symbiodiniaceae and in in hospite symbionts using a combined western blot and qRT-PCR approach. We showed that the expression and regulation of NR in free-living Symbiodiniaceae is a dynamic and reversible process impacted by NO3- and NH4+ concentrations. Symbionts from N-depleted corals incubated with NO3- enriched seawater showed an increase in NR synthesis over time. Interestingly, NR protein synthesis did not correlate with NR gene expression, hinting towards a potential post-transcriptional regulation of the enzyme. Additionally, we investigated the impacts of inorganic N source (NO3- vs NH4+ vs N depletion) in combination with stress on the physiology of Symbiodiniaceae (photosynthetic responses, ROS and NO production). The availability of inorganic nitrogen improved photosynthetic capacity while reducing ROS production. Moreover, preliminary experiments showed that NO3- and NH4+ had differential effects on the physiological responses of Symbiodiniaceae subjected to stress

Corals and Sponges Under the Light of the Holobiont Concept: How Microbiomes Underpin Our Understanding of Marine Ecosystems

In the past 20 years, a new concept has slowly emerged and expanded to various domains of marine biology research: the holobiont. A holobiont describes the consortium formed by a eukaryotic host and its associated microorganisms including bacteria, archaea, protists, microalgae, fungi, and viruses. From coral reefs to the deep-sea, symbiotic relationships and host–microbiome interactions are omnipresent and central to the health of marine ecosystems. Studying marine organisms under the light of the holobiont is a new paradigm that impacts many aspects of marine sciences. This approach is an innovative way of understanding the complex functioning of marine organisms, their evolution, their ecological roles within their ecosystems, and their adaptation to face environmental changes. This review offers a broad insight into key concepts of holobiont studies and into the current knowledge of marine model holobionts. Firstly, the history of the holobiont concept and the expansion of its use from evolutionary sciences to other fields of marine biology will be discussed. Then, the ecology and physiology of marine holobionts will be investigated through the examples of corals and sponges. We will discuss the impacts of environmental change on organisms at the holobiont level and how microbiomes contribute to the resilience and/or vulnerability of their host in the face of environmental stressors. Finally, we will conclude with the development of new technologies, holistic approaches, and future prospects for conservation biology surrounding marine holobionts

Deciphering the expression and regulation of nitrate reductase by Symbiodiniaceae in culture and in hospite

Coral holobionts are highly efficient in the assimilation of nitrogen through heterotrophic feeding or the uptake of dissolved inorganic nitrogen. Although NO3- is the most abundant nitrogen source in the ocean, corals preferably uptake NH4+ due to its reduced state and energetically favorable assimilation. However, in conditions of low availability of environmental NH4+, coral holobionts are capable of depleting environmental NO3-. Symbiodiniaceae are vital partners of the symbiosis for nutrient assimilation. In addition to providing translocated photosynthates, they facilitate the acquisition of environmental nitrogen and account for most of the uptake of dissolved inorganic nitrogen. While the uptake of NH4+ by the coral host and its symbiotic partners is a well-known process, NO3- assimilation remains poorly studied. Coral hosts are incapable of nitrate reduction as they do not possess the necessary enzymes, while Symbiodiniaceae have been shown to express the enzyme nitrate reductase (NR). However, the evidence supporting the active reduction of nitrate by the symbiotic algae during symbiosis is scarce and equivocal. Rigorous studies on nitrate assimilation by Symbiodiniaceae are lacking yet essential for the understanding of coral holobiont functioning. We studied the expression and regulation of NR in monoclonal and axenic Symbiodiniaceae cultures (SSB01 Breviolum minutum and Symbiodinium microadriaticum) on different N sources (NO3-, NH4+, NO3- + NH4+). The algae expressed NR in culture, but the enzyme expression was repressed in the presence of NH4+ even when NO3- was available. Protein expression kinetics as well as NH4+ concentration threshold for NO3- assimilation inhibition were investigated using increasing NH4+ concentrations on NO3- medium. Following the addition of NH4+ to the medium, NR was actively degraded under 6 hours. In the absence of NH4+ and following the addition of NO3-, NR was synthesized over 3 to 6 hours. This illustrates the inhibition effect of NH4+ on NR expression. Symbiodiniaceae in culture are capable of utilizing environmental NO3-. However, the symbiotic algae could see their activity hindered by the host cellular environment and cellular NH4+ concentrations. These results raise questions about the occurrence of the enzyme NR in symbiosis along with the process of NO3- assimilation by coral holobionts

Specific lipid and carotenoid oxidation products, and preconditioning to oxidative stress in Symbiodiniaceae

In symbiotic cnidarians, living in symbiosis with photosynthetic dinoflagellates of the Symbiodiniaceae family causes daily local hyperoxia and promotes the generation of Reactive Oxygen Species (ROS) that, under some particular environmental conditions, may cause the breakdown of the symbiotic interaction (i.e. bleaching). Despite much effort, there are still large gaps in our understanding of ROS signaling and how the antioxidant network is modulated in Symbiodiniaceae. It is known from plants and green algae that the nature of ROS signaling depends on the chemical identity of ROS. Therefore, to understand the mechanisms by which cells sense and respond to oxidative stress, it is necessary to investigate responses to individual ROS. Under oxidative stress, oxidants such as free radicals attack polyunsaturated fatty acids (PUFAs) and carotenoids containing C-C double bonds. During this study we determined signatures that discriminate between 1O2 and hydroxyl radical-dependent lipid and carotenoid oxidation. First, we characterized the fatty acid and the carotenoid compositions in cultured and freshly isolated Symbiodinium microadriaticum by using chromatographic methods. Then we oxidized the predominant PUFAs and carotenoids in vitro by exposing them to photosynthesizer and light to generate 1O2, and H2O2 in presence of Fe3+ to generate hydroxyl radical. Finally, knowing the pattern of carotenoid and lipid oxidation products produced in vitro by 1O2 or free radicals, we looked for those oxidation products in cultured and symbiotic S. microadriaticum cells subjected to various stressful conditions such as conditions involving endogenously produced 1O2 or superoxide ion/H2O2 ; exposure to high light intensity; and high light exposure combined to elevated temperature incubation. This work also aimed to find conditions in which cells of S. microadriaticum could be preconditioned or acclimatized by ROS. To this end, different prooxidant molecules were tested

Impacts of oxidized fatty acids on the physiology of the coral endosymbiont Breviolum minutum

peer reviewe

Crude Oil in the Deep: Investigating the Responses of a Deep-Sea Sponge Holobiont to Hydrocarbon Exposure

peer reviewedThe deep-sea is increasingly considered for extraction of petroleum reserves as offshore oil and gas development is expanding with the identification of new reservoirs. This raises considerable concerns about accidental releases of hydrocarbons into deep-sea ecosystems and the subsequent cascading effects on associated fauna. Sponges are abundant and ecologically valuable in these ecosystems. They are known to be efficient filter feeders, contributing to benthic-pelagic coupling and providing a habitat for a suit of organisms. Surprisingly, there is a dearth of studies on the impacts of oil pollution on sponges. Using a mesocosm setup, we investigated the responses of a deep-sea sponge from the Northern Atlantic, Geodia barretti, exposed to three ecologically-relevant oil concentrations for a duration of 8 days, followed by a recovery period of 30 days. Changes in physiology and cellular stress were assessed through respiration rates and lysosomal membrane stability, respectively. The structure of the sponge associated microbiome was investigated using high-throughput sequencing of 16s rRNA gene amplicons. Acute crude oil exposure did not induce strong sub-lethal stress effects in G. barretti. Respiration showed varying patterns of both increased and decreased rates, with no significant effect from the treatments, while lysosomes were significantly impacted by oil, displaying destabilization of lysosomal membranes. After the recovery period, sponges were observed to return to their initial physiological and cellular pre-experimental state. Sponge microbiome however, showed no significant changes with either treatment or exposure time. Such studies are essential to advance our understanding of the vulnerability and resilience of deep-sea sponges to hydrocarbon exposure, providing useful data for managing risks associated with oil and gas exploration in the Northern Atlantic