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Coral reefs are threatened by climate change, because it causes increasingly frequent and severe summer heatwaves, resulting in mass coral bleaching and mortality. Coral bleaching is believed to be driven by an excess production of reactive oxygen (ROS) and nitrogen species (RNS), yet their relative roles during thermal stress remain understudied. Here, we measured ROS and RNS net production, as well as activities of key enzymes involved in ROS scavenging (superoxide dismutase and catalase) and RNS synthesis (nitric oxide synthase) and linked these metrics to physiological measurements of cnidarian holobiont health during thermal stress. We did this for both an established cnidarian model, the sea anemone Exaiptasia diaphana, and an emerging scleractinian model, the coral Galaxea fascicularis, both from the Great Barrier Reef (GBR). Increased ROS production was observed during thermal stress in both species, but it was more apparent in G. fascicularis, which also showed higher levels of physiological stress. RNS did not change in thermally stressed G. fascicularis and decreased in E. diaphana. Our findings in combination with variable ROS levels in previous studies on GBR-sourced E. diaphana suggest G. fascicularis is a more suitable model to study the cellular mechanisms of coral bleaching.

Coral reefs house one-third of all marine species and are of high cultural and socioeconomic importance. However, coral reefs are under dire threat from climate change and other anthropogenic stressors. Climate change is causing coral bleaching, the breakdown of the symbiosis between the coral host and its algal symbionts, often resulting in coral mortality and the deterioration of these valuable ecosystems. While it is essential to counteract the root causes of climate change, it remains urgent to develop coral restoration and conservation methods that will buy time for coral reefs. The manipulation of the bacterial microbiome that is associated with corals has been suggested as one intervention to improve coral climate resilience. Early coral microbiome-manipulation studies, which are aimed at enhancing bleaching tolerance, have shown promising results, but the inoculated bacteria did generally not persist within the coral microbiome. Here, we highlight the importance of long-term incorporation of bacterial inocula into the microbiome of target corals, as repeated inoculations will be too costly and not feasible on large reef systems like the Great Barrier Reef. Therefore, coral microbiome-manipulation studies need to prioritise approaches that can provide sustained coral climate resilience.

Microbial symbionts frequently localize within specific body structures or cell types of their multicellular hosts. This spatiotemporal niche is critical to host health, nutrient exchange, and fitness. Measuring host-microbe metabolite exchange has conventionally relied on tissue homogenates, eliminating dimensionality and dampening analytical sensitivity. We have developed a mass spectrometry imaging workflow for a soft- and hard-bodied cnidarian animal capable of revealing the host and symbiont metabolome in situ, without the need for a priori isotopic labelling or skeleton decalcification. The mass spectrometry imaging method provides critical functional insights that cannot be gleaned from bulk tissue analyses or other presently available spatial methods. We show that cnidarian hosts may regulate microalgal symbiont acquisition and rejection through specific ceramides distributed throughout the tissue lining the gastrovascular cavity. The distribution pattern of betaine lipids showed that once resident, symbionts primarily reside in light-exposed tentacles to generate photosynthate. Spatial patterns of these metabolites also revealed that symbiont identity can drive host metabolism.

The symbiotic partnership between corals and dinoflagellate algae is crucial to coral reefs. Corals provide their algal symbionts with shelter, carbon dioxide and nitrogen. In exchange, the symbiotic algae supply their animal hosts with fixed carbon in the form of glucose. But how glucose is transferred from the algal symbiont to the animal host is unknown. We reasoned that a transporter resident in the dinoflagellate cell membrane would facilitate outward transfer of glucose to the surrounding host animal tissue. We identified a candidate transporter in the cnidarian symbiont dinoflagellate Breviolum minutum that belongs to the ubiquitous family of facilitative sugar uniporters known as SWEETs (sugars will eventually be exported transporters). Previous gene expression analyses had shown that BmSWEET1 is upregulated when the algae are living symbiotically in a cnidarian host by comparison to the free-living state [1, 2]. We used immunofluorescence microscopy to localise BmSWEET1 in the dinoflagellate cell membrane. Substrate preference assays in a yeast surrogate transport system showed that BmSWEET1 transports glucose. Quantitative microscopy showed that symbiotic B. minutum cells have significantly more BmSWEET1 protein than free-living cells of the same strain, consistent with export during symbiosis but not during the free-living, planktonic phase. Thus, BmSWEET1 is in the right place, at the right time, and has the right substrate to be the transporter with which symbiotic dinoflagellate algae feed their animal hosts to power coral reefs.

Corals are associated with a variety of bacteria, which occur in the surface mucus layer, gastrovascular cavity, skeleton, and tissues. Some tissue-associated bacteria form clusters, termed cell-associated microbial aggregates (CAMAs), which are poorly studied. Here, we provide a comprehensive characterization of CAMAs in the coral Pocillopora acuta. Combining imaging techniques, laser capture microdissection, and amplicon and metagenome sequencing, we show that (i) CAMAs are located in the tentacle tips and may be intracellular; (ii) CAMAs contain Endozoicomonas (Gammaproteobacteria) and Simkania (Chlamydiota) bacteria; (iii) Endozoicomonas may provide vitamins to its host and use secretion systems and/or pili for colonization and aggregation; (iv) Endozoicomonas and Simkania occur in distinct, but adjacent, CAMAs; and (v) Simkania may receive acetate and heme from neighboring Endozoicomonas. Our study provides detailed insight into coral endosymbionts, thereby improving our understanding of coral physiology and health and providing important knowledge for coral reef conservation in the climate change era.

Global climate change is threatening the persistence of coral reefs as associated summer heatwaves trigger the loss of microalgal endosymbionts (Symbiodiniaceae) from the coral tissues, or coral bleaching. We infected aposymbiotic juveniles of the coral Acropora tenuis with either wildtype (WT10) or heat-evolved (SS1 or SS8) Symbiodiniaceae strains Cladocopium proliferum (formerly referred to as Cladocopium goreaui and Cladocopium C1acro). After 10 months at 27 °C, SS8-juveniles were 2 × larger than SS1- or WT10-juveniles. In response to a simulated heatwave (31 °C for 41 days), the WT10-juveniles bleached and showed a decline in respiration while cell densities and respiration in both SS-juvenile groups remained unchanged compared to the controls. These results reveal that some heat-evolved strains can increase the bleaching tolerance of juvenile corals without a trade-off against growth. This response is opposite to the lower nutrient provisioning often reported for naturally thermotolerant Symbiodiniaceae (e.g. genus Durusdinium), thereby offering enhanced fitness to the host without the ecological consequences of diminished growth.

Humans have long sought to restore species but little attention has been directed at how to best select a subset of foundation species for maintaining rich assemblages that support ecosystems, like coral reefs and rainforests, which are increasingly threatened by environmental change. We propose a two-part hedging approach that selects optimized sets of species for restoration. The first part acknowledges that biodiversity supports ecosystem functions and services, and so it ensures precaution against loss by allocating an even spread of phenotypic traits. The second part maximizes species and ecosystem persistence by weighting species based on characteristics that are known to improve ecological persistence—for example abundance, species range and tolerance to environmental change. Using existing phenotypic-trait and ecological data for reef building corals, we identified sets of ecologically persistent species by examining marginal returns in occupancy of phenotypic trait space. We compared optimal sets of species with those from the world's southern-most coral reef, which naturally harbours low coral diversity, to show these occupy much of the trait space. Comparison with an existing coral restoration program indicated that current corals used for restoration only cover part of the desired trait space and programs may be improved by including species with different traits. Synthesis and applications. While there are many possible criteria for selecting species for restoration, the approach proposed here addresses the need to insure against unpredictable losses of ecosystem services by focusing on a wide range of phenotypic traits and ecological characteristics. Furthermore, the flexibility of the approach enables the functional goals of restoration to vary depending on environmental context, stakeholder values, and the spatial and temporal scales at which meaningful impacts can be achieved.

There is overwhelming evidence that tropical coral reefs are severely impacted by human induced climate change. Assessing the capability of reef-building corals to expand their tolerance limits to survive projected climate trajectories is critical for their protection and management. Acclimation mechanisms such as developmental plasticity may provide one means by which corals could cope with projected ocean warming and acidification. To assess the potential of preconditioning to enhance thermal tolerance in the coral Pocillopora acuta, colonies were kept under three different scenarios from settlement to 17 months old: present day (0.9 °C-weeks (Degree Heating Weeks), + 0.75 °C annual, 400 ppm pCO2) mid-century (2.5 °C-weeks, + 1.5 °C annual, 685 ppm pCO2) and end of century (5 °C-weeks, + 2 °C annual, 900 ppm pCO2) conditions. Colonies from the present-day scenario were subsequently introduced to the mid-century and end of century conditions for six weeks during summer thermal maxima to examine if preconditioned colonies (reared under these elevated conditions) had a higher physiological performance compared to naive individuals. Symbiodiniaceae density and chlorophyll a concentrations were significantly lower in mid-century and end of century preconditioned groups, and declines in symbiont density were observed over the six-week accumulated heat stress in all treatments. Maximum photosynthetic rate was significantly suppressed in mid-century and end of century preconditioned groups, while minimum saturating irradiances were highest for 2050 pre-exposed individuals with parents originating from specific populations. The results of this study indicate preconditioning to elevated temperature and pCO2 for 17 months did not enhance the physiological performance in P. acuta. However, variations in trait responses and effects on tolerance found among treatment groups provides evidence for differential capacity for phenotypic plasticity among populations which could have

The sea anemone, Exaiptasia diaphana, is a model of coral-dinoflagellate (Symbiodiniaceae) symbiosis. However, little is known of its potential to form symbiosis with Cladocopium—a key Indo-Pacific algal symbiont of scleractinian corals, nor the host nutritional consequences of such an association. Aposymbiotic anemones were inoculated with homologous algal symbionts, Breviolum minutum, and seven heterologous strains of Cladocopium C1acro (wild-type and heat-evolved) under ambient conditions. Despite lower initial algal cell density, Cladocopium C1acro-anemeones achieved similar cell densities as B. minutum-anemones by week 77. Wild-type and heat-evolved Cladocopium C1acro showed similar colonization patterns. Targeted LC-MS-based metabolomics revealed that almost all significantly different metabolites in the host and Symbiodiniaceae fractions were due to differences between Cladocopium C1acro and B. minutum, with little difference between heat-evolved and wild-type Cladocopium C1acro at week 9. The algal fraction of Cladocopium C1acro-anemones was enriched in metabolites related to nitrogen storage, while the host fraction of B. minutum-anemones was enriched in sugar-related metabolites. Compared to B. minutum, Cladocopium C1acro is likely slightly less nutritionally beneficial to the host under ambient conditions, but more capable of maintaining its own growth when host nitrogen supply is limited. Our findings demonstrate the value of E. diaphana to study experimentally evolved Cladocopium.

Early life stages of most coral species acquire microalgal endosymbionts (Symbiodiniaceae) from the environment, but whether exogenous symbiont uptake is possible in the adult life stage is unclear. Deep sequencing of the Symbiodiniaceae ITS2 genetic marker has revealed novel symbionts in adult corals following bleaching; however these strains may have already been present at densities below detection limits. To test whether acquisition of symbionts from the environment occurs, we subjected adult fragments of corals (six species in four families) to a chemical bleaching treatment (menthol and DCMU). The treatment reduced the native microalgal symbiont abundance to below 2% of their starting densities. The bleached corals were then inoculated with a cultured Cladocopium C1acro strain. Genotyping of the Symbiodiniaceae communities before bleaching and after reinoculation showed that fragments of all six coral species acquired the Cladocopium C1acro strain used for inoculation. Our results provide strong evidence for the uptake of Symbiodiniaceae from the environment by adult corals. We also demonstrate the feasibility of chemical bleaching followed by reinoculation to manipulate the Symbiodiniaceae communities of adult corals, providing an innovative approach to establish new symbioses between adult corals and heat-evolved microalgal symbionts, which could prove highly relevant to coral reef restoration efforts.

Heat-tolerant strains of the coral endosymbiont, Cladocopium C1acro (Symbiodiniaceae), have previously been developed via experimental evolution. Here, we examine physiological responses and bacterial community composition (using 16S rRNA gene metabarcoding) in cultures of 10 heat-evolved (SS) and 9 wild-type (WT) strains, which had been exposed for 6 years to 31 °C and 27 °C, respectively. We also examine whether the associated bacterial communities were affected by a three-week reciprocal transplantation to both temperatures. The SS strains had bacterial communities with lower diversities that showed more stability and lower variability when exposed to elevated temperatures compared with the WT strains. Amplicon sequence variants (ASVs) of the bacterial genera Labrenzia, Algiphilus, Hyphobacterium and Roseitalea were significantly more associated with the SS strains compared with the WT strains. WT strains showed higher abundance of ASVs assigned to the genera Fabibacter and Tropicimonas. We hypothesize that these compositional differences in associated bacterial communities between SS and WT strains also contribute to the thermal tolerance of the microalgae. Future research should explore functional potential between bacterial communities using metagenomics to unravel specific genomic adaptations.

AIMS: Fourteen percent of all living coral, equivalent to more than all the coral on the Great Barrier Reef, has died in the past decade as a result of climate change-driven bleaching. Inspired by the 'oxidative stress theory of coral bleaching', we investigated whether a bacterial consortium designed to scavenge free radicals could integrate into the host microbiome and improve thermal tolerance of the coral model, Exaiptasia diaphana. METHODS AND RESULTS: E. diaphana anemones were inoculated with a consortium of high free radical scavenging (FRS) bacteria, a consortium of congeneric low FRS bacteria, or sterile seawater as a control, then exposed to elevated temperature. Increases in the relative abundance of Labrenzia during the first 2 weeks following the last inoculation provided evidence for temporary inoculum integration into the E. diaphana microbiome. Initial uptake of other consortium members was inconsistent, and these bacteria did not persist either in E. diaphana's microbiome over time. Given their non-integration into the host microbiome, the ability of the FRS consortium to mitigate thermal stress could not be assessed. Importantly, there were no physiological impacts (negative or positive) of the bacterial inoculations on the holobiont. CONCLUSIONS: The introduced bacteria were not maintained in the anemone microbiome over time, thus, their protective effect is unknown. Achieving long-term integration of bacteria into cnidarian microbiomes remains a research priority. SIGNIFICANCE AND IMPACT OF THE STUDY: Microbiome engineering strategies to mitigate coral bleaching may assist coral reefs in their persistence until climate change has been curbed. This study provides insights that will inform microbiome manipulation approaches in coral bleaching mitigation research.

Finding coral reefs resilient to climate warming is challenging given the large spatial scale of reef ecosystems. Methods are needed to predict the location of corals with heritable tolerance to high temperatures. Here, we combine Great Barrier Reef-scale remote sensing with breeding experiments that estimate larval and juvenile coral survival under exposure to high temperatures. Using reproductive corals collected from the northern and central Great Barrier Reef, we develop forecasting models to locate reefs harbouring corals capable of producing offspring with increased heat tolerance of an additional 3.4° heating weeks (~3 °C). Our findings predict hundreds of reefs (~7.5%) may be home to corals that have high and heritable heat-tolerance in habitats with high daily and annual temperature ranges and historically variable heat stress. The locations identified represent targets for protection and consideration as a source of corals for use in restoration of degraded reefs given their potential to resist climate change impacts and repopulate reefs with tolerant offspring.

To survive in nutrient-poor waters corals rely on a symbiotic association with intracellular microalgae. However, increased sea temperatures cause algal loss—known as coral bleaching—often followed by coral death. Some of the most compelling evidence in support of the ‘oxidative stress theory of coral bleaching’ comes from studies that exposed corals, cultures of their algal endosymbionts, or the coral modelExaiptasia diaphanato exogenous antioxidants during thermal stress. Here, we replicate these experiments usingE.diaphanawith the addition of the antioxidants ascorbate + catalase, catechin, or mannitol under ambient and elevated temperatures along with an antioxidant-free control. In the absence of exogenous antioxidants,E.diaphanaexposed to elevated temperatures bleached with no change in reactive oxygen species (ROS) levels associated with their microalgal cells. Ascorbate + catalase and mannitol treatments rescued the anemones from bleaching, although microalgal ROS levels increased in these antioxidant treatments under elevated temperature conditions. While bleaching was not associated with changes in net ROS for the intracellular algal symbionts, it is evident from our findings that excess ROS is connected to the bleaching phenotype as exogenous antioxidants were successful in mitigating the effects of thermal stress in cnidarians. This understanding may assist applied research that aims to reduce the impact of climate change on coral reefs.

Reef-building corals form nutritional symbioses with endosymbiotic dinoflagellates (Symbiodiniaceae), a relationship that facilitates the ecological success of coral reefs. These symbionts are mostly acquired anew each generation from the environment during early life stages ("horizontal transmission"). Symbiodiniaceae species exhibit trait variation that directly impacts the health and performance of the coral host under ocean warming. Here, we test the capacity for larvae of a horizontally transmitting coral, Acropora tenuis, to establish symbioses with Symbiodiniaceae species in four genera that have varying thermal thresholds (the common symbiont genera, Cladocopium and Durusdinium, and the less common Fugacium and Gerakladium). Over a 2-week period in January 2018, a series of both no-choice and four-way choice experiments were conducted at three temperatures (27, 30, and 31°C). Symbiont acquisition success and cell proliferation were measured in individual larvae. Larvae successfully acquired and maintained symbionts of all four genera in no-choice experiments, and >80% of larvae were infected with at least three genera when offered a four-way choice. Unexpectedly, Gerakladium symbionts increased in dominance over time, and at high temperatures outcompeted Durusdinium, which is regarded as thermally tolerant. Although Fugacium displayed the highest thermal tolerance in culture and reached similar cell densities to the other three symbionts at 31°C, it remained a background symbiont in choice experiments, suggesting host preference for other symbiont species. Larval survivorship at 1 week was highest in larvae associated with Gerakladium and Fugacium symbionts at 27 and 30°C, however at 31°C, mortality was similar for all treatments. We hypothesize that symbionts that are currently rare in corals (e.g., Gerakladium) may become more common and widespread in early life stages under climate warming. Uptake of such symbionts may function as a survival strategy in t

Coral reef restoration activity is accelerating worldwide in efforts to offset the rate of reef health declines. Many advances have already been made in restoration practices centred on coral biology (coral restoration), and particularly those that look to employ the high adaptive state and capacity of corals in order to ensure that efforts rebuilding coral biomass also equip reefs with enhanced resilience to future stress. We horizon scan the state-of-play for the many coral restoration innovations already underway across the complex life cycle for corals that spans both asexual and sexual reproduction - assisted evolution (manipulations targeted to the coral host and host-associated microbes), biobanking, as well as scalable coral propagation and planting - and how these innovations are in different stages of maturity to support new 21st century reef management frameworks. Realising the potential for coral restoration tools as management aids undoubtedly rests on validating different approaches as their application continues to scale. Whilst the ecosystem service responses to increased scaling still largely remain to be seen, coral restoration has already delivered immense new understanding of coral and coral-associated microbial biology that has long lagged behind advances in other reef sciences.

Reef-building corals live very close to their upper thermal limits and their persistence is imperiled by a rapidly warming climate. Human interventions may be used to increase the thermal limits of sensitive corals by cross-breeding with heat-adapted populations. However, the scope of breeding interventions is constrained by regional variation in the annual reproductive cycle of corals. Here we use cryopreservation technology to overcome this barrier and cross-breed conspecific coral populations across ocean basins for the first time. During regional spawning events, sperm samples were cryopreserved from populations of the widespread Indo-Pacific coral, Platygyra daedalea, from the southern Persian Gulf (maximum daily sea surface temperature of 36 °C), the Oman Sea (33 °C), and the central Great Barrier Reef (30 °C). These sperm samples were thawed during a later spawning event to test their ability to fertilize freshly spawned eggs of P. daedalea colonies from the central Great Barrier Reef. Average fertilization success for the Persian Gulf (9%) and Oman Sea (6%) sperm were 1.4-2.5 times lower than those for the native cryopreserved sperm from Great Barrier Reef (13-15%), potentially due to lower sperm quality of the Middle Eastern sperm and/or reproductive incompatibility between these distant populations. Overall, fertilization success with cryopreserved sperm was low compared with fresh sperm (>80%), likely due to the low motility of thawed sperm (≤5%, reduced from 50% to >90% in fresh sperm). To evaluate whether cross-bred offspring had enhanced thermal tolerance, the survival of larvae sired by Persian Gulf cryopreserved sperm, Great Barrier Reef cryopreserved sperm, and Great Barrier Reef fresh sperm was monitored for six days at ambient (27 °C) and elevated (33 °C) temperature. Against expectations of thermal tolerance enhancement, survival of larvae sired by Persian Gulf cryopreserved sperm was 2.6 times lower than larvae sired by Great Barrier Reef fresh

Active restoration or intervention programs will be required in the future to support the resilience and adaptation of coral reef ecosystems in the face of climate change. Selective propagation of corals ex situ can help conserve keystone species and the ecosystems they underpin; cross-disciplinary research and communication between science and industry are essential to this success. Zoos and aquaria have a long history of managing ex situ breed-for-release programs and have led the establishment of wildlife biobanks (collections of cryopreserved living cells) along with the development of associated reproductive technologies for their application to wildlife conservation. Taronga Conservation Society Australia’s CryoDiversity Bank includes cryopreserved coral sperm from the Great Barrier Reef, which represents the largest repository from any reef system around the globe. This paper presents results from an inventory review of the current collection. The review highlighted the skew toward five Acropora species and the necessity to increase the taxonomic diversity of the collection. It also highlighted the need to increase geographic representation, even for the most well represented species. The inventory data will inform Taronga’s future research focus and sampling strategy to maximize genetic variation and biodiversity within the biobank and provide a test case for other practitioners implementing biobanking strategies for coral conservation around the world. Through co-investment and collaboration with research partners over the next decade, Taronga will prioritize and resource critical applied research and expand biobanking efforts to assist interventions for reef recovery and restoration.

Dinoflagellates of the family Symbiodiniaceae are crucial photosymbionts in corals and other marine organisms. Of these, Cladocopium goreaui is one of the most dominant symbiont species in the Indo-Pacific. Here, we present an improved genome assembly of C. goreaui combining new long-read sequence data with previously generated short-read data. Incorporating new full-length transcripts to guide gene prediction, the C. goreaui genome (1.2 Gb) exhibits a high extent of completeness (82.4% based on BUSCO protein recovery) and better resolution of repetitive sequence regions; 45,322 gene models were predicted, and 327 putative, topologically associated domains of the chromosomes were identified. Comparison with other Symbiodiniaceae genomes revealed a prevalence of repeats and duplicated genes in C. goreaui, and lineage-specific genes indicating functional innovation. Incorporating 2,841,408 protein sequences from 96 taxonomically diverse eukaryotes and representative prokaryotes in a phylogenomic approach, we assessed the evolutionary history of C. goreaui genes. Of the 5246 phylogenetic trees inferred from homologous protein sets containing two or more phyla, 35-36% have putatively originated via horizontal gene transfer (HGT), predominantly (19-23%) via an ancestral Archaeplastida lineage implicated in the endosymbiotic origin of plastids: 10-11% are of green algal origin, including genes encoding photosynthetic functions. Our results demonstrate the utility of long-read sequence data in resolving structural features of a dinoflagellate genome, and highlight how genetic transfer has shaped genome evolution of a facultative symbiont, and more broadly of dinoflagellates.

The climate crisis is one of the most significant threats to marine ecosystems. It is leading to severe increases in sea surface temperatures and in the frequency and magnitude of marine heatwaves. These changing conditions are directly impacting coral reef ecosystems, which are among the most biodiverse ecosystems on Earth. Coral-associated symbionts are particularly affected because summer heatwaves cause coral bleaching-the loss of endosymbiotic microalgae (Symbiodiniaceae) from coral tissues, leading to coral starvation and death. Coral-associated Symbiodiniaceae and bacteria have been extensively studied in the context of climate change, especially in terms of community diversity and dynamics. However, data on other microorganisms and their response to climate change are scarce. Here, we review current knowledge on how increasing temperatures affect understudied coral-associated microorganisms such as archaea, fungi, viruses, and protists other than Symbiodiniaceae, as well as microbe-microbe interactions. We show that the coral-microbe symbiosis equilibrium is at risk under current and predicted future climate change and argue that coral reef conservation initiatives should include microbe-focused approaches.

While coral reefs in Australia have historically been a showcase of conventional management informed by research, recent declines in coral cover have triggered efforts to innovate and integrate intervention and restoration actions into management frameworks. Here we outline the multi-faceted intervention approaches that have developed in Australia since 2017, from newly implemented in-water programs, research to enhance coral resilience and investigations into socio-economic perspectives on restoration goals. We describe in-water projects using coral gardening, substrate stabilisation, coral repositioning, macro-algae removal, and larval-based restoration techniques. Three areas of research focus are also presented to illustrate the breadth of Australian research on coral restoration, (1) the transdisciplinary Reef Restoration and Adaptation Program (RRAP), one of the world's largest research and development programs focused on coral reefs, (2) interventions to enhance coral performance under climate change, and (3) research into socio-cultural perspectives. Together, these projects and the recent research focus reflect an increasing urgency for action to confront the coral reef crisis, develop new and additional tools to manage coral reefs, and the consequent increase in funding opportunities and management appetite for implementation. The rapid progress in trialling and deploying coral restoration in Australia builds on decades of overseas experience, and advances in research and development are showing positive signs that coral restoration can be a valuable tool to improve resilience at local scales (i.e., high early survival rates across a variety of methods and coral species, strong community engagement with local stakeholders). RRAP is focused on creating interventions to help coral reefs at multiple scales, from micro scales (i.e., interventions targeting small areas within a specific reef site) to large scales (i.e., interventions targeting core ecosystem fu

The rapid growth in genomic techniques provides the potential to transform how we protect, manage, and conserve marine life. Further, solutions to boost the resilience of marine species to climate change and other disturbances that characterize the Anthropocene require transformative approaches, made more effective if guided by genomic data. Although genetic techniques have been employed in marine conservation for decades and the availability of genomic data is rapidly expanding, widespread application still lags behind other data types. This Essay reviews how genetics and genomics have been utilized in management initiatives for ocean conservation and restoration, highlights success stories, and presents a pathway forward to enhance the uptake of genomic data for protecting our oceans.

Coral reefs are rapidly declining because of widespread mass coral bleaching causing extensive coral mortality. Elevated seawater temperatures are the main drivers of coral bleaching, and climate change is increasing the frequency and severity of destructive marine heatwaves. Efforts to enhance coral thermal bleaching tolerance can be targeted at the coral host or at coral-associated microorganisms (e.g., dinoflagellate endosymbionts and bacteria). The literature on experimental evolution of bacteria suggests that it has value as a tool to increase coral climate resilience. We provide a workflow on how to experimentally evolve coral-associated bacteria to confer thermal tolerance to coral hosts and emphasize the value of implementing this approach in coral reef conservation and restoration efforts.

Corals house a variety of microorganisms which they depend on for their survival, including endosymbiotic dinoflagellates (Symbiodiniaceae) and bacteria. While cnidarian-microorganism interactions are widely studied, Symbiodiniaceae-bacteria interactions are only just beginning to receive attention. Here, we describe the localization and composition of the bacterial communities associated with cultures of 11 Symbiodiniaceae strains from nine species and six genera. Three-dimensional confocal laser scanning and electron microscopy revealed bacteria are present inside the Symbiodiniaceae cells as well as closely associated with their external cell surface. Bacterial pure cultures and 16S rRNA gene metabarcoding from Symbiodiniaceae cultures highlighted distinct and highly diverse bacterial communities occur intracellularly, closely associated with the Symbiodiniaceae outer cell surface and loosely associated (i.e., in the surrounding culture media). The intracellular bacteria are highly conserved across Symbiodiniaceae species, suggesting they may be involved in Symbiodiniaceae physiology. Our findings provide unique new insights into the biology of Symbiodiniaceae.

Maternal effects have been well documented for offspring morphology and life history traits in plants and terrestrial animals, yet little is known about maternal effects in corals. Further, few studies have explored maternal effects in gene expression. In a previous study, F1 interspecific hybrid and purebred larvae of the coral species Acropora tenuis and Acropora loripes were settled and exposed to ambient or elevated temperature and pCO2 conditions for 7 months. At this stage, the hybrid coral recruits from both ocean conditions exhibited strong maternal effects in several fitness traits. We conducted RNA‐sequencing on these corals and showed that gene expression of the hybrid Acropora also exhibited clear maternal effects. Only 40 genes were differentially expressed between hybrids and their maternal progenitor. In contrast, ~2000 differentially expressed genes were observed between hybrids and their paternal progenitors, and between the reciprocal F1 hybrids. These results indicate that maternal effects in coral gene expression can be long‐lasting. Unlike findings from most short‐term stress experiments in corals, no genes were differentially expressed in the hybrid nor purebred offspring after seven months of exposure to elevated temperature and pCO2 conditions.

Strong population-by-habitat interactions across environmental gradients arise from genetic adaptation or acclimatization and represents phenotypic variation required for populations to respond to changing environmental conditions. As such, patterns of adaptation and acclimatization of reef-building corals are integral to predictions of the future of coral reefs under climate warming. The common brooding coral, Pocillopora damicornis, exhibits extensive differences in host genetic and microbial symbiont community composition between depth habitats at Heron Island in the southern Great Barrier Reef, Australia. An 18-month reciprocal field transplant experiment was undertaken to examine the environmental and genetic drivers behind variation in survival, weight gain, heat tolerance and algal symbiont community between the reef flat and slope habitats. We observed population-by-habitat interactions for in situ partial mortality and weight gain, where trait-related fitness of natives was greater than transplants in most cases, consistent with local adaptation. On average, flat colonies transplanted to the slope had a relatively low partial mortality but minimal weight gain, whereas slope colonies transplanted to the flat had relatively high partial mortality and average weight gain. Experimental heat tolerance was always higher in colonies sourced from the flat, but increased when slope colonies were transplanted to the flat, providing evidence of acclimatization in these colonies. The performance of certain slope to flat transplants may have been driven by each colony’s algal symbiont (Symbiodiniaceae) community, and flat variants were observed in a small number of slope colonies that either had a fixed flat composition before transplantation or shuffled after transplantation. Host genotypes of previously identified genetic outlier loci could not predict survival following transplantation, possibly because of low sample size and/or polygenic basis to the traits examined

Novel restoration methods are currently under consideration worldwide to help coral reefs recover or become more resilient to higher temperature stress. Critical field-based information concerning the paradigm of “local is best” is lacking for many methods; information which is essential to determine the risk and feasibility associated with restoration. One method involves breeding corals from different reef regions with expected variation in heat tolerance and moving those offspring to new locations to enhance offspring survival; thereby augmenting local stock to enhance survival for anticipated warming. In this study, surviving colonies from the 2016 to 2017 mass bleaching events on the Great Barrier Reef (GBR) were reproductively crossed and they included colonies sourced from northern (three) and central (two) reefs. The gravid colonies of Acropora tenuis were collected across 6° of latitude, and they were spawned to produce a total of 17 purebred and hybrid crosses. Juvenile corals (3,748 individual colonies settled on 1,474 terracotta tiles) were deployed to Davies reef in the central GBR after 4 months of aquarium rearing. Survival, growth, and coral colour (as a proxy for bleaching) were assessed after 0, 91, and 217 days of field deployment. Overall, a high percentage of juveniles (17% ± 2.5 SE) survived relative to expected survival at the final census. Survival was significantly higher for central purebred crosses, hybrid crosses had intermediate survival while northern purebreds had the lowest survival. Colour and growth rates (0.001−0.006 mm2 day–1) were not significantly different amongst central, northern, or hybrid crosses but were of a reverse pattern compared to survival. On average, northern purebred crosses grew the fastest, followed by hybrid crosses, and then central purebred crosses. Modelled growth trajectories suggest that northern purebreds would take 8 years to grow to reproductive size, hybrids would take nine, and central purebreds would

Anthropogenic climate change is a rapidly intensifying selection pressure on biodiversity across the globe and, particularly, on the world's coral reefs. The rate of adaptation to climate change is proportional to the amount of phenotypic variation that can be inherited by subsequent generations (i.e., narrow-sense heritability, h2 ). Thus, traits that have higher heritability (e.g., h2 > 0.5) are likely to adapt to future conditions faster than traits with lower heritability (e.g., h2 < 0.1). Here, we synthesize 95 heritability estimates across 19 species of reef-building corals. Our meta-analysis reveals low heritability (h2 < 0.25) of gene expression metrics, intermediate heritability (h2 = 0.25-0.50) of photochemistry, growth, and bleaching, and high heritability (h2 > 0.50) for metrics related to survival and immune responses. Some of these values are higher than typically observed in other taxa, such as survival and growth, while others were more comparable, such as gene expression and photochemistry. There was no detectable effect of temperature on heritability, but narrow-sense heritability estimates were generally lower than broad-sense estimates, indicative of significant non-additive genetic variation across traits. Trait heritability also varied depending on coral life stage, with bleaching and growth in juveniles generally having lower heritability compared to bleaching and growth in larvae and adults. These differences may be the result of previous stabilizing selection on juveniles or may be due to constrained evolution resulting from genetic trade-offs or genetic correlations between growth and thermotolerance. While we find no evidence that heritability decreases under temperature stress, explicit tests of the heritability of thermal tolerance itself-such as coral thermal reaction norm shape-are lacking. Nevertheless, our findings overall reveal high trait heritability for the majority of coral traits, suggesting corals may have a greater potent

Coral reefs are the epitome of species diversity, yet the number of described scleractinian coral species, the framework-builders of coral reefs, remains moderate by comparison. DNA sequencing studies are rapidly challenging this notion by exposing a wealth of undescribed diversity, but the evolutionary and ecological significance of this diversity remains largely unclear. Here, we present an annotated genome for one of the most ubiquitous corals in the Indo-Pacific (Pachyseris speciosa) and uncover, through a comprehensive genomic and phenotypic assessment, that it comprises morphologically indistinguishable but ecologically divergent lineages. Demographic modeling based on whole-genome resequencing indicated that morphological crypsis (across micro- and macromorphological traits) was due to ancient morphological stasis rather than recent divergence. Although the lineages occur sympatrically across shallow and mesophotic habitats, extensive genotyping using a rapid molecular assay revealed differentiation of their ecological distributions. Leveraging "common garden" conditions facilitated by the overlapping distributions, we assessed physiological and quantitative skeletal traits and demonstrated concurrent phenotypic differentiation. Lastly, spawning observations of genotyped colonies highlighted the potential role of temporal reproductive isolation in the limited admixture, with consistent genomic signatures in genes related to morphogenesis and reproduction. Overall, our findings demonstrate the presence of ecologically and phenotypically divergent coral species without substantial morphological differentiation and provide new leads into the potential mechanisms facilitating such divergence. More broadly, they indicate that our current taxonomic framework for reef-building corals may be scratching the surface of the ecologically relevant diversity on coral reefs, consequently limiting our ability to protect or restore this diversity effectively.

The algal cell wall is an important cellular component that functions in defense, nutrient utilization, signaling, adhesion, and cell-cell recognition-processes important in the cnidarian-dinoflagellate symbiosis. The cell wall of symbiodiniacean dinoflagellates is not well characterized. Here, we present a method to isolate cell walls of Symbiodiniaceae and prepare cell-wall-enriched samples for proteomic analysis. Label-free liquid chromatography-electrospray ionization tandem mass spectrometry was used to explore the surface proteome of two Symbiodiniaceae species from the Great Barrier Reef: Breviolum minutum and Cladocopium goreaui. Transporters, hydrolases, translocases, and proteins involved in cell-adhesion and protein-protein interactions were identified, but the majority of cell wall proteins had no homologues in public databases. We propose roles for some of these proteins in the cnidarian-dinoflagellate symbiosis. This work provides the first proteomics investigation of cell wall proteins in the Symbiodiniaceae and represents a basis for future explorations of the roles of cell wall proteins in Symbiodiniaceae and other dinoflagellates.

Marine microalgae are a diverse group of microscopic eukaryotic and prokaryotic organisms capable of photosynthesis. They are important primary producers and carbon sinks but their physiology and persistence are severely affected by global climate change. Powerful experimental evolution technologies are being used to examine the potential of microalgae to respond adaptively to current and predicted future conditions, as well as to develop resources to facilitate species conservation and restoration of ecosystem functions. This review synthesizes findings and insights from experimental evolution studies of marine microalgae in response to elevated temperature and/or pCO2 . Adaptation to these environmental conditions has been observed in many studies of marine dinoflagellates, diatoms and coccolithophores. An enhancement in traits such as growth and photo-physiological performance and an increase in upper thermal limit have been shown to be possible, although the extent and rate of change differ between microalgal taxa. Studies employing multiple monoclonal replicates showed variation in responses among replicates and revealed the stochasticity of mutations. The work to date is already providing valuable information on species' climate sensitivity or resilience to managers and policymakers but extrapolating these insights to ecosystem- and community-level impacts continues to be a challenge. We recommend future work should include in situ experiments, diurnal and seasonal fluctuations, multiple drivers and multiple starting genotypes. Fitness trade-offs, stable versus plastic responses and the genetic bases of the changes also need investigating, and the incorporation of genome resequencing into experimental designs will be invaluable.

BACKGROUND: Coral reefs are among the most diverse and productive ecosystems on Earth. This success relies on the coral's association with a wide range of microorganisms, including dinoflagellates of the family Symbiodiniaceae that provide coral hosts with most of their organic carbon requirements. While bacterial associates have long been overlooked, research on these microorganisms is gaining traction, and deciphering bacterial identity and function is greatly enhancing our understanding of cnidarian biology. Here, we investigated bacterial communities in defensive tissues (acontia) of the coral model, the sea anemone Exaiptasia diaphana. Acontia are internal filaments that are ejected upon detection of an external threat and release toxins to repel predators. RESULTS: Using culturing techniques and 16S rRNA gene metabarcoding we identified bacterial communities associated with acontia of four Great Barrier Reef-sourced E. diaphana genotypes. We show that bacterial communities are similar across genotypes, and dominated by Alteromonadaceae, Vibrionaceae, Rhodobacteraceae, and Saprospiraceae. By analyzing abundant amplicon sequence variants (ASVs) from metabarcoding data from acontia and comparing these to data from whole anemones, we identified five potentially important bacterial genera of the acontia microbiome: Vibrio, Sulfitobacter, Marivita, Alteromonas, and Lewinella. The role of these bacteria within the acontia remains uninvestigated but could entail assistance in defense processes such as toxin production. CONCLUSIONS: This study provides insight into potential bacterial involvement in cnidarian defense tissues and highlights the need to study bacterial communities in individual compartments within a holobiont.

The global decline of coral reefs heightens the need to understand how corals may persist under changing environmental conditions. Restructuring of the coral-associated bacterial community, either through natural or assisted strategies, has been suggested as a means of adaptation to climate change. A low complexity microbial system would facilitate testing the efficacy of microbial restructuring strategies. We used the model organism for corals, Exaiptasia diaphana, and determined that short-term (3 weeks) exposure to filter-sterilized seawater conditions alone reduced the complexity of the microbiome. Metabarcoding of the V5–V6 region of the bacterial 16S rRNA gene revealed that alpha diversity was approximately halved in anemones reared in filter-sterilized seawater compared to controls reared in unfiltered seawater and that the composition (beta diversity) differed significantly between the two. By reducing the complexity of the E. diaphana microbiome, the development of a system for testing assisted strategies such as probiotics, is more feasible.

Corals are colonized by symbiotic microorganisms that profoundly influence the animal's health. One noted symbiont is a single-celled alga (in the dinoflagellate family Symbiodiniaceae), which provides the coral with most of its fixed carbon. Thermal stress increases the production of reactive oxygen species (ROS) by Symbiodiniaceae during photosynthesis. ROS can both damage the algal symbiont's photosynthetic machinery and inhibit its repair, causing a positive feedback loop for the toxic accumulation of ROS. If not scavenged by the antioxidant network, excess ROS may trigger a signaling cascade ending with the coral host and algal symbiont disassociating in a process known as bleaching. We use Exaiptasia diaphana as a model for corals and constructed a consortium comprised of E. diaphana-associated bacteria capable of neutralizing ROS. We identified six strains with high free radical scavenging (FRS) ability belonging to the families Alteromonadaceae, Rhodobacteraceae, Flavobacteriaceae and Micrococcaceae. In parallel, we established a consortium of low FRS isolates consisting of genetically related strains. Bacterial whole genome sequences were used to identify key pathways that are known to influence ROS.

Corals are the main primary producers of coral reefs and build the three-dimensional reef structure that provides habitat to more than 25% of all marine eukaryotes. They harbor a complex consortium of microorganisms, including bacteria, archaea, fungi, viruses, and protists, which they rely on for their survival. The symbiosis between corals and bacteria is poorly studied, and their symbiotic relationships with intracellular bacteria are only just beginning to be acknowledged. In this review, we emphasize the importance of characterizing intracellular bacteria associated with corals and explore how successful approaches used to study such microorganisms in other systems could be adapted for research on corals. We propose a framework for the description, identification, and functional characterization of coral-associated intracellular bacterial symbionts. Finally, we highlight the possible value of intracellular bacteria in microbiome manipulation and mitigating coral bleaching.

Reef-building corals form associations with a huge diversity of microorganisms, which are essential for the survival and well-being of their host. While the acquisition patterns of Symbiodiniaceae microalgal endosymbionts are strongly linked to the coral's reproductive strategy, few studies have investigated the transmission mode of bacteria, especially in brooding species. Here, we relied on 16S rRNA gene and Internal Transcribed Spacer 2 marker metabarcoding in conjunction with fluorescence in situ hybridisation microscopy to describe the onset of microbial associations in the common brooding coral Pocillopora acuta. We analysed the bacterial and Symbiodiniaceae community composition in five adult colonies, their larvae, and 4-day old recruits. Larvae and recruits inherited Symbiodiniaceae, as well as a small number of bacterial strains, from their parents. Rhodobacteraceae and Endozoicomonas were among the most abundant taxa that were likely maternally transmitted to the offspring. The presence of bacterial aggregates in newly released larvae was observed with confocal microscopy, confirming the occurrence of vertical transmission of bacteria in P. acuta. We concluded that host factors, as well as the environmental bacterial pool influenced the microbiome of P. acuta.

Small increases in ocean temperature can disrupt the obligate symbiosis between corals and dinoflagellate microalgae, resulting in coral bleaching. Little is known about the genes that drive the physiological and bleaching response of algal symbionts to elevated temperature. Moreover, many studies to-date have compared highly divergent strains, making it challenging to accredit specific genes to contrasting traits. Here, we compare transcriptional responses at ambient (27°C) and bleaching-relevant (31°C) temperatures in a monoclonal, wild-type (WT) strain of Symbiodiniaceae to those of a selected-strain (SS), derived from the same monoclonal culture and experimentally evolved to elevated temperature over 80 generations (2.5 years). Thousands of genes were differentially expressed at a log fold-change of >8 between the WT and SS over a 35 days temperature treatment period. At 31°C, WT cells exhibited a temporally unstable transcriptomic response upregulating genes involved in the universal stress response such as molecular chaperoning, protein repair, protein degradation and DNA repair. Comparatively, SS cells exhibited a temporally stable transcriptomic response and downregulated many stress response genes that were upregulated by the WT. Among the most highly upregulated genes in the SS at 31°C were algal transcription factors and a gene probably of bacterial origin that encodes a type II secretion system protein, suggesting interactions with bacteria may contribute to the increased thermal tolerance of the SS. Genes and functional pathways conferring thermal tolerance in the SS could be targeted in future genetic engineering experiments designed to develop thermally resilient algal symbionts for use in coral restoration and conservation.

The rate of coral reef degradation from climate change is accelerating and, as a consequence, a number of interventions to increase coral resilience and accelerate recovery are under consideration. Acropora spathulata coral colonies that survived mass bleaching in 2016 and 2017 were sourced from a bleaching-impacted and warmer northern reef on the Great Barrier Reef (GBR). These individuals were reproductively crossed with colonies collected from a recently bleached but historically cooler central GBR reef to produce pure and crossbred offspring groups (warm-warm, warm-cool and cool-warm). We tested whether corals from the warmer reef produced more thermally tolerant hybrid and purebred offspring compared with crosses produced with colonies sourced from the cooler reef and whether different symbiont taxa affect heat tolerance. Juveniles were infected with Symbiodinium tridacnidorum, Cladocopium goreaui and Durusdinium trenchii and survival, bleaching and growth were assessed at 27.5°C and 31°C. The contribution of host genetic background and symbiont identity varied across fitness traits. Offspring with either both or one parent from the northern population exhibited a 13- to 26-fold increase in survival odds relative to all other treatments where survival probability was significantly influenced by familial cross identity at 31°C but not 27.5°C (Kaplan-Meier P=0.001 versus 0.2). If in symbiosis with D. trenchii, a warm sire and cool dam provided the best odds of juvenile survival. Bleaching was predominantly driven by Symbiodiniaceae treatment, where juveniles hosting D. trenchii bleached significantly less than the other treatments at 31°C. The greatest overall fold-benefits in growth and survival at 31°C occurred in having at least one warm dam and in symbiosis with D. trenchii Juveniles associated with D. trenchii grew the most at 31°C, but at 27.5°C, growth was fastest in juveniles associated with C. goreaui In conclusion, selective breeding with warmer GBR cor

Coral reefs worldwide are suffering mass mortalities from marine heat waves. With the aim of enhancing coral bleaching tolerance, we evolved 10 clonal strains of a common coral microalgal endosymbiont at elevated temperatures (31°C) for 4 years in the laboratory. All 10 heat-evolved strains had expanded their thermal tolerance in vitro following laboratory evolution. After reintroduction into coral host larvae, 3 of the 10 heat-evolved endosymbionts also increased the holobionts' bleaching tolerance. Although lower levels of secreted reactive oxygen species (ROS) accompanied thermal tolerance of the heat-evolved algae, reduced ROS secretion alone did not predict thermal tolerance in symbiosis. The more tolerant symbiosis exhibited additional higher constitutive expression of algal carbon fixation genes and coral heat tolerance genes. These findings demonstrate that coral stock with enhanced climate resilience can be developed through ex hospite laboratory evolution of their microalgal endosymbionts.

Genetic signatures caused by demographic and adaptive processes during past climatic shifts can inform predictions of species’ responses to anthropogenic climate change. To identify these signatures in Acropora tenuis, a reef-building coral threatened by global warming, we first assembled the genome from long reads and then used shallow whole-genome resequencing of 150 colonies from the central inshore Great Barrier Reef to inform population genomic analyses. We identify population structure in the host that reflects a Pleistocene split, whereas photosymbiont differences between reefs most likely reflect contemporary (Holocene) conditions. Signatures of selection in the host were associated with genes linked to diverse processes including osmotic regulation, skeletal development, and the establishment and maintenance of symbiosis. Our results suggest that adaptation to post-glacial climate change in A. tenuis has involved selection on many genes, while differences in symbiont specificity between reefs appear to be unrelated to host population structure.

BACKGROUND: Coral reefs have sustained damage of increasing scale and frequency due to climate change, thereby intensifying the need to elucidate corals' biological characteristics, including their thermal tolerance and microbial symbioses. The sea anemone, Exaiptasia diaphana, has proven an ideal coral model for many studies due to its close phylogenetic relationship and shared traits, such as symbiosis with algae of the family Symbiodiniaceae. However, established E. diaphana clonal lines are not available in Australia thus limiting the ability of Australian scientists to conduct research with this model. To help address this, the bacterial and Symbiodiniaceae associates of four Great Barrier Reef (GBR)-sourced E. diaphana genotypes established in laboratory aquaria and designated AIMS1-4, and from proxies of wild GBR E. diaphana were identified by metabarcoding of the bacterial 16S rRNA gene and eukaryotic rRNA gene ITS2 region. The relationship between AIMS1-4 and their bacterial associates was investigated, as was bacterial community phenotypic potential. Existing data from two existing anemone clonal lines, CC7 and H2, were included for comparison. RESULTS: Overall, 2238 bacterial amplicon sequence variants (ASVs) were observed in the AIMS1-4 bacterial communities, which were dominated by Proteobacteria and Bacteroidetes, together comprising > 90% relative abundance. Although many low abundance bacterial taxa varied between the anemone genotypes, the AIMS1-4 communities did not differ significantly. A significant tank effect was identified, indicating an environmental effect on the microbial communities. Bacterial community richness was lower in all lab-maintained E. diaphana compared to the wild proxies, suggesting a reduction in bacterial diversity and community phenotypic potential due to culturing. Seventeen ASVs were common to every GBR lab-cultured anemone, however five were associated with the Artemia feedstock, making their specific association to E. d