What makes a winner? Symbiont and host dynamics determine Caribbean octocoral resilience to bleaching.

Unlike reef-building, scleractinian corals, Caribbean soft corals (octocorals) have not suffered marked declines in abundance associated with anthropogenic ocean warming. Both octocorals and reef-building scleractinians depend on a nutritional symbiosis with single-celled algae living within their tissues. In both groups, increased ocean temperatures can induce symbiont loss (bleaching) and coral death. Multiple heat waves from 2014 to 2016 resulted in widespread damage to reef ecosystems and provided an opportunity to examine the bleaching response of three Caribbean octocoral species. Symbiont densities declined during the heat waves but recovered quickly, and colony mortality was low. The dominant symbiont genotypes within a host generally did not change, and all colonies hosted symbiont species in the genus Breviolum. Their association with thermally tolerant symbionts likely contributes to the octocoral holobiont's resistance to mortality and the resilience of their symbiont populations. The resistance and resilience of Caribbean octocorals offer clues for the future of coral reefs.

Building consensus around the assessment and interpretation of Symbiodiniaceae diversity.

Within microeukaryotes, genetic variation and functional variation sometimes accumulate more quickly than morphological differences. To understand the evolutionary history and ecology of such lineages, it is key to examine diversity at multiple levels of organization. In the dinoflagellate family Symbiodiniaceae, which can form endosymbioses with cnidarians (e.g., corals, octocorals, sea anemones, jellyfish), other marine invertebrates (e.g., sponges, molluscs, flatworms), and protists (e.g., foraminifera), molecular data have been used extensively over the past three decades to describe phenotypes and to make evolutionary and ecological inferences. Despite advances in Symbiodiniaceae genomics, a lack of consensus among researchers with respect to interpreting genetic data has slowed progress in the field and acted as a barrier to reconciling observations. Here, we identify key challenges regarding the assessment and interpretation of Symbiodiniaceae genetic diversity across three levels: species, populations, and communities. We summarize areas of agreement and highlight techniques and approaches that are broadly accepted. In areas where debate remains, we identify unresolved issues and discuss technologies and approaches that can help to fill knowledge gaps related to genetic and phenotypic diversity. We also discuss ways to stimulate progress, in particular by fostering a more inclusive and collaborative research community. We hope that this perspective will inspire and accelerate coral reef science by serving as a resource to those designing experiments, publishing research, and applying for funding related to Symbiodiniaceae and their symbiotic partnerships.

Nutrient dynamics in coral symbiosis depend on both the relative and absolute abundance of Symbiodiniaceae species.

BACKGROUND: Symbionts provide a variety of reproductive, nutritional, and defensive resources to their hosts, but those resources can vary depending on symbiont community composition. As genetic techniques open our eyes to the breadth of symbiont diversity within myriad microbiomes, symbiosis research has begun to consider what ecological mechanisms affect the identity and relative abundance of symbiont species and how this community structure impacts resource exchange among partners. Here, we manipulated the in hospite density and relative ratio of two species of coral endosymbionts (Symbiodinium microadriaticum and Breviolum minutum) and used stable isotope enrichment to trace nutrient exchange with the host, Briareum asbestinum. RESULTS: The patterns of uptake and translocation of carbon and nitrogen varied with both density and ratio of symbionts. Once a density threshold was reached, carbon acquisition decreased with increasing proportions of S. microadriaticum. In hosts dominated by B. minutum, nitrogen uptake was density independent and intermediate. Conversely, for those corals dominated by S. microadriaticum, nitrogen uptake decreased as densities increased, and as a result, these hosts had the overall highest (at low density) and lowest (at high density) nitrogen enrichment. CONCLUSIONS: Our findings show that the uptake and sharing of nutrients was strongly dependent on both the density of symbionts within the host, as well as which symbiont species was dominant. Together, these complex interactive effects suggest that host regulation and the repression of in hospite symbiont competition can ultimately lead to a more productive mutualism. Video Abstract.

Symbiont genotype influences holobiont response to increased temperature.

As coral reefs face warming oceans and increased coral bleaching, a whitening of the coral due to loss of microalgal endosymbionts, the possibility of evolutionary rescue offers some hope for reef persistence. In tightly linked mutualisms, evolutionary rescue may occur through evolution of the host and/or endosymbionts. Many obligate mutualisms are composed of relatively small, fast-growing symbionts with greater potential to evolve on ecologically relevant time scales than their relatively large, slower growing hosts. Numerous jellyfish species harbor closely related endosymbiont taxa to other cnidarian species such as coral, and are commonly used as a model system for investigating cnidarian mutualisms. We examined the potential for adaptation of the upside-down jellyfish Cassiopea xamachana to increased temperature via evolution of its microalgal endosymbiont, Symbiodinium microadriaticum. We quantified trait variation among five algal genotypes in response to three temperatures (26 °C, 30 °C, and 32 °C) and fitness of hosts infected with each genotype. All genotypes showed positive growth rates at each temperature, but rates of respiration and photosynthesis decreased with increased temperature. Responses varied among genotypes but were unrelated to genetic similarity. The effect of temperature on asexual reproduction and the timing of development in the host also depended on the genotype of the symbiont. Natural selection could favor different algal genotypes at different temperatures, affecting host fitness. This eco-evolutionary interaction may be a critical component of understanding species resilience in increasingly stressful environments.

Genetic structure of dinoflagellate symbionts in coral recruits differs from that of parental or local adults.

The symbiotic relationship between dinoflagellate algae in the family Symbiodiniaceae and scleractinian corals forms the base of the tropical reef ecosystem. In scleractinian corals, recruits acquire symbionts either "vertically" from the maternal colony or initially lack symbionts and acquire them "horizontally" from the environment. Regardless of the mode of acquisition, coral species and individual colonies harbor only a subset of the highly diverse complex of species/taxa within the Symbiodiniaceae. This suggests a genetic basis for specificity, but local environmental conditions and/or symbiont availability may also play a role in determining which symbionts within the Symbiodiniaceae are initially taken up by the host. To address the relative importance of genetic and environmental drivers of symbiont uptake/establishment, we examined the acquisition of these dinoflagellate symbionts in one to three-month-old recruits of Orbicella faveolata to compare symbiont types present in recruits to those of parental populations versus co-occurring adults in their destination reef. Variation in chloroplast 23S ribosomal DNA and in three polymorphic microsatellite loci was examined. We found that, in general, symbiont communities within adult colonies differed between reefs, suggesting that endemism is common among symbiont populations of O. faveolata on a local scale. Among recruits, initial symbiont acquisition was selective. O. faveolata recruits only acquired a subset of locally available symbionts, and these generally did not reflect symbiont populations in adults at either the parental or the outplant reef. Instead, symbiont communities within new recruits at a given outplant site and region tended to be similar to each other, regardless of parental source population. These results suggest temporal variation in the local symbiont source pool, although other possible drivers behind the distinct difference between symbionts within O. faveolata adults and new generations of recruits may include different ontogenetic requirements and/or reduced host selectivity in early ontogeny.

Individual variation in growth and physiology of symbionts in response to temperature.

In many cases, understanding species' responses to climate change requires understanding variation among individuals in response to such change. For species with strong symbiotic relationships, such as many coral reef species, genetic variation in symbiont responses to temperature may affect the response to increased ocean temperatures. To assess variation among symbiont genotypes, we examined the population dynamics and physiological responses of genotypes of Breviolum antillogorgium in response to increased temperature. We found broad temperature tolerance across genotypes, with all genotypes showing positive growth at 26, 30, and 32°C. Genotypes differed in the magnitude of the response of growth rate and carrying capacity to increasing temperature, suggesting that natural selection could favor different genotypes at different temperatures. However, the historical temperature at which genotypes were reared (26 or 30°C) was not a good predictor of contemporary temperature response. We found increased photosynthetic rates and decreased respiration rates with increasing contemporary temperature, and differences in physiology among genotypes, but found no significant differences in the response of these traits to temperature among genotypes. In species with such broad thermal tolerance, selection experiments on symbionts outside of the host may not yield results sufficient for evolutionary rescue from climate change.

Competition and succession among coral endosymbionts.

Host species often support a genetically diverse guild of symbionts, the identity and performance of which can determine holobiont fitness under particular environmental conditions. These symbiont communities are structured by a complex set of potential interactions, both positive and negative, between the host and symbionts and among symbionts. In reef-building corals, stable associations with specific symbiont species are common, and we hypothesize that this is partly due to ecological mechanisms, such as succession and competition, which drive patterns of symbiont winnowing in the initial colonization of new generations of coral recruits. We tested this hypothesis using the experimental framework of the de Wit replacement series and found that competitive interactions occurred among symbionts which were characterized by unique ecological strategies. Aposymbiotic octocoral recruits within high- and low-light environments were inoculated with one of three Symbiodiniaceae species as monocultures or with cross-paired mixtures, and we tracked symbiont uptake using quantitative genetic assays. Priority effects, in which early colonizers excluded competitive dominants, were evidenced under low light, but these early opportunistic species were later succeeded by competitive dominants. Under high light, a more consistent competitive hierarchy was established in which competitive dominants outgrew and limited the abundance of others. These findings provide insight into mechanisms of microbial community organization and symbiosis breakdown and recovery. Furthermore, transitions in competitive outcomes across spatial and temporal environmental variation may improve lifetime host fitness.

Genetic variation in Breviolum antillogorgium, a coral reef symbiont, in response to temperature and nutrients.

Symbionts within the family Symbiodiniaceae are important on coral reefs because they provide significant amounts of carbon to many different reef species. The breakdown of this mutualism that occurs as a result of increasingly warmer ocean temperatures is a major threat to coral reef ecosystems globally. Recombination during sexual reproduction and high rates of somatic mutation can lead to increased genetic variation within symbiont species, which may provide the fuel for natural selection and adaptation. However, few studies have asked whether such variation in functional traits exists within these symbionts. We used several genotypes of two closely related species, Breviolum antillogorgium and B. minutum, to examine variation of traits related to symbiosis in response to increases in temperature or nitrogen availability in laboratory cultures. We found significant genetic variation within and among symbiont species in chlorophyll content, photosynthetic efficiency, and growth rate. Two genotypes showed decreases in traits in response to increased temperatures predicted by climate change, but one genotype responded positively. Similarly, some genotypes within a species responded positively to high-nitrogen environments, such as those expected within hosts or eutrophication associated with global change, while other genotypes in the same species responded negatively, suggesting context-dependency in the strength of mutualism. Such variation in traits implies that there is potential for natural selection on symbionts in response to temperature and nutrients, which could confer an adaptive advantage to the holobiont.

Comparative growth rates of cultured marine dinoflagellates in the genus Symbiodinium and the effects of temperature and light.

Many dinoflagellate microalgae of the genus Symbiodinium form successful symbioses with a large group of metazoans and selected protists. Yet knowledge of growth kinetics of these endosymbionts and their ecological and evolutionary implications is limited. We used a Bayesian biphasic generalized logistic model to estimate key parameters of the growth of five strains of cultured Symbiodinium, S. microadriaticum (cp-type A194; strain 04-503), S. microadriaticum (cp-type A194; strain CassKB8), S. minutum (cp-type B184; strain Mf 1.05b.01.SCI.01), S. psygmophilum (cp-type B224; strain Mf 11.05b.01) and S. trenchii (cp-type D206; strain Mf 2.2b), grown in four different combinations of temperature and light. Growth kinetics varied among Symbiodinium strains and across treatments. Biphasic growth was especially evident for S. minutum and S. psygmophilum across all treatments. Monophasic growth was more common when final asymptotic densities were relatively low (~ 200 million cells ml-1). All species tended to grow faster and / or reached a higher asymptote at 26°C than at 18°C. The fastest growth was exhibited by S. minutum, with an approximate four-fold increase in estimated cell density after 60 days. The strongest effect of light was seen in S. trenchii, in which increasing light levels resulted in a decrease in initial growth rate, and an increase in asymptotic density, time when growth rate was at its maximum, final growth rate, and maximum growth rate. Results suggest that Symbiodinium species have different photokinetic and thermal optima, which may affect their growth-related nutritional physiology and allow them to modify their response to environmental changes.

Caribbean massive corals not recovering from repeated thermal stress events during 2005-2013.

Massive coral bleaching events associated with high sea surface temperatures are forecast to become more frequent and severe in the future due to climate change. Monitoring colony recovery from bleaching disturbances over multiyear time frames is important for improving predictions of future coral community changes. However, there are currently few multiyear studies describing long-term outcomes for coral colonies following acute bleaching events. We recorded colony pigmentation and size for bleached and unbleached groups of co-located conspecifics of three major reef-building scleractinian corals (Orbicella franksi, Siderastrea siderea, and Stephanocoenia michelini; n = 198 total) in Bocas del Toro, Panama, during the major 2005 bleaching event and then monitored pigmentation status and changes live tissue colony size for 8 years (2005-2013). Corals that were bleached in 2005 demonstrated markedly different response trajectories compared to unbleached colony groups, with extensive live tissue loss for bleached corals of all species following bleaching, with mean live tissue losses per colony 9 months postbleaching of 26.2% (±5.4 SE) for O. franksi, 35.7% (±4.7 SE) for S. michelini, and 11.2% (±3.9 SE) for S. siderea. Two species, O. franksi and S. michelini, later recovered to net positive growth, which continued until a second thermal stress event in 2010. Following this event, all species again lost tissue, with previously unbleached colony species groups experiencing greater declines than conspecific sample groups, which were previously bleached, indicating a possible positive acclimative response. However, despite this beneficial effect for previously bleached corals, all groups experienced substantial net tissue loss between 2005 and 2013, indicating that many important Caribbean reef-building corals will likely suffer continued tissue loss and may be unable to maintain current benthic coverage when faced with future thermal stress forecast for the region, even with potential benefits from bleaching-related acclimation.

The effects of Symbiodinium (Pyrrhophyta) identity on growth, survivorship, and thermal tolerance of newly settled coral recruits.

For many coral species, the obligate association with phylogenetically diverse algal endosymbiont species is dynamic in time and space. Here, we used controlled laboratory inoculations of newly settled, aposymbiotic corals (Orbicella faveolata) with two cultured species of algal symbiont (Symbiodinium microadriaticum and S. minutum) to examine the role of symbiont identity on growth, survivorship, and thermal tolerance of the coral holobiont. We evaluated these data in the context of Symbiodinium photophysiology for 9 months post-settlement and also during a 5-d period of elevated temperatures Our data show that recruits that were inoculated with S. minutum grew significantly slower than those inoculated with S. microadriaticum (occasionally co-occurring with S. minutum), but that there was no difference in survivorship of O. faveolata polyps infected with Symbiodinium. However, photophysiological metrics (∆Fv/F'm, the efficiency with which available light is used to drive photosynthesis and α, the maximum light utilization coefficient) were higher in those slower growing recruits containing S. minutum. These findings suggest that light use (i.e., photophysiology) and carbon acquisition by the coral host (i.e., host growth) are decoupled, but did not distinguish the source of this difference. Neither Symbiodinium treatment demonstrated a significant negative effect of a 5-d exposure to temperatures as high as 32°C under low light conditions similar to those measured at settlement habitats.

Taxonomic and environmental variation of metabolite profiles in marine dinoflagellates of the genus symbiodinium.

Microorganisms in terrestrial and marine ecosystems are essential to environmental sustainability. In the marine environment, invertebrates often depend on metabolic cooperation with their endosymbionts. Coral reefs, one of the most important marine ecosystems, are based on the symbiosis between a broad diversity of dinoflagellates of the genus Symbiodinium and a wide phyletic diversity of hosts (i.e., cnidarian, molluscan, poriferan). This diversity is reflected in the ecology and physiology of the symbionts, yet the underlying biochemical mechanisms are still poorly understood. We examined metabolite profiles of four cultured species of Symbiodinium known to form viable symbioses with reef-building corals, S. microadriaticum (cp-type A194), S. minutum (cp-type B184), S. psygmophilum (cp-type B224) and S. trenchii (cp-type D206). Metabolite profiles were shown to differ among Symbiodinium species and were found to be affected by their physiological response to growth in different temperatures and light regimes. A combined Random Forests and Bayesian analysis revealed that the four Symbiodinium species examined primarily differed in their production of sterols and sugars, including a C29 stanol and the two sterols C28Δ5 and C28Δ5,22, as well as differences in metabolite abundances of a hexose and inositol. Inositol levels were also strongly affected by changes in temperature across all Symbiodinium species. Our results offer a detailed view of the metabolite profile characteristic of marine symbiotic dinoflagellates of the genus Symbiodinium, and identify patterns of metabolites related to several growth conditions.

New species of Clade B Symbiodinium (Dinophyceae) from the greater Caribbean belong to different functional guilds: S. aenigmaticum sp. nov., S. antillogorgium sp. nov., S. endomadracis sp. nov., and S. pseudominutum sp. nov.

Molecular approaches have begun to supersede traditional morphometrics in the species delineation of micro-eukaryotes. In addition to fixed differences in DNA sequences, recent genetics-based descriptions within the dinoflagellate genus Symbiodinium have incorporated confirmatory morphological, physiological, and ecological evidence when possible. However, morphological and physiological data are difficult to collect from species that have not been cultured, while the natural ecologies of many cultured species remain unknown. Here, we rely on genetic evidence-the only data consistently available among all taxa investigated-to describe four new Clade B Symbiodinium species. The 'host-specialized' species (S. antillogorgium sp. nov. and S. endomadracis sp. nov.) engage in mutualisms with specific cnidarian hosts, but exhibit differences in our ability to culture them in vitro. The ecologically 'cryptic' species (S. aenigmaticum sp. nov. and S. pseudominutum sp. nov.) thrive in culture, but their roles or functions in the ecosystem (i.e., niches) are yet to be documented. These new species call further attention to the spectrum of ecological guilds among Symbiodinium.

Cryptic diversity hides host and habitat specialization in a gorgonian-algal symbiosis.

Shallow water anthozoans, the major builders of modern coral reefs, enhance their metabolic and calcification rates with algal symbionts. Controversy exists over whether these anthozoan-algae associations are flexible over the lifetimes of individual hosts, promoting acclimative plasticity, or are closely linked, such that hosts and symbionts co-evolve across generations. Given the diversity of algal symbionts and the morphological plasticity of many host species, cryptic variation within either partner could potentially confound studies of anthozoan-algal associations. Here, we used ribosomal, organelle and nuclear sequences, along with microsatellite variation, to study the relationship between lineages of a common Caribbean gorgonian and its algal symbionts. The gorgonian Eunicea flexuosa is a broadcast spawner, composed of two recently diverged, genetically distinct lineages largely segregated by depth. We sampled colonies of the two lineages across depth gradients at three Caribbean locations. We find that each host lineage is associated with a unique Symbiodinium B1/184 phylotype. This relationship between host and symbiont is maintained when host colonies are reciprocally transplanted, although cases of within phylotype switching were also observed. Even when the phylotypes of both partners are present at intermediate depths, the specificity between host and symbiont lineages remained absolute. Unrecognized cryptic diversity may mask host-symbiont specificity and change the inference of evolutionary processes in mutualistic associations. Symbiotic specificity thus likely contributes to the ecological divergence of the two partners, generating species diversity within coral reefs.

Tracking transmission of apicomplexan symbionts in diverse Caribbean corals.

Symbionts in each generation are transmitted to new host individuals either vertically (parent to offspring), horizontally (from exogenous sources), or a combination of both. Scleractinian corals make an excellent study system for understanding patterns of symbiont transmission since they harbor diverse symbionts and possess distinct reproductive modes of either internal brooding or external broadcast spawning that generally correlate with vertical or horizontal transmission, respectively. Here, we focused on the under-recognized, but apparently widespread, coral-associated apicomplexans (Protista: Alveolata) to determine if symbiont transmission depends on host reproductive mode. Specifically, a PCR-based assay was utilized towards identifying whether planula larvae and reproductive adults from brooding and broadcast spawning scleractinian coral species in Florida and Belize harbored apicomplexan DNA. Nearly all (85.5%; n = 85/89) examined planulae of five brooding species (Porites astreoides, Agaricia tenuifolia, Agaricia agaricites, Favia fragum, Mycetophyllia ferox) and adults of P. astreoides were positive for apicomplexan DNA. In contrast, no (n = 0/10) apicomplexan DNA was detected from planulae of four broadcast spawning species (Acropora cervicornis, Acropora palmata, Pseudodiploria strigosa, and Orbicella faveolata) and rarely in gametes (8.9%; n = 5/56) of these species sampled from the same geographical range as the brooding species. In contrast, tissue samples from nearly all (92.0%; n = 81/88) adults of the broadcast spawning species A. cervicornis, A. palmata and O. faveolata harbored apicomplexan DNA, including colonies whose gametes and planulae tested negative for these symbionts. Taken together, these data suggest apicomplexans are transmitted vertically in these brooding scleractinian coral species while the broadcast spawning scleractinian species examined here acquire these symbionts horizontally. Notably, these transmission patterns are consistent with those of other scleractinian coral symbionts. While this study furthers knowledge regarding these symbionts, numerous questions remain to be addressed, particularly in regard to the specific interaction(s) between these apicomplexans and their hosts.

Draft assembly of the Symbiodinium minutum nuclear genome reveals dinoflagellate gene structure.

BACKGROUND: Dinoflagellates are known for their capacity to form harmful blooms (e.g., "red tides") and as symbiotic, photosynthetic partners for corals. These unicellular eukaryotes have permanently condensed, liquid-crystalline chromosomes and immense nuclear genome sizes, often several times the size of the human genome. Here we describe the first draft assembly of a dinoflagellate nuclear genome, providing insights into its genome organization and gene inventory. RESULTS: Sequencing reads from Symbiodinium minutum were assembled into 616 Mbp gene-rich DNA regions that represented roughly half of the estimated 1,500 Mbp genome of this species. The assembly encoded ∼42,000 protein-coding genes, consistent with previous dinoflagellate gene number estimates using transcriptomic data. The Symbiodinium genome contains duplicated genes for regulator of chromosome condensation proteins, nearly one-third of which have eukaryotic orthologs, whereas the remainder have most likely been acquired through bacterial horizontal gene transfers. Symbiodinium genes are enriched in spliceosomal introns (mean = 18.6 introns/gene). Donor and acceptor splice sites are unique, with 5' sites utilizing not only GT but also GC and GA, whereas at 3' sites, a conserved G is present after AG. All spliceosomal snRNA genes (U1-U6) are clustered in the genome. Surprisingly, the Symbiodinium genome displays unidirectionally aligned genes throughout the genome, forming a cluster-like gene arrangement. CONCLUSIONS: We show here that a dinoflagellate genome exhibits unique and divergent characteristics when compared to those of other eukaryotes. Our data elucidate the organization and gene inventory of dinoflagellates and lay the foundation for future studies of this remarkable group of eukaryotes.

Evaluating the ribosomal internal transcribed spacer (ITS) as a candidate dinoflagellate barcode marker.

BACKGROUND: DNA barcoding offers an efficient way to determine species identification and to measure biodiversity. For dinoflagellates, an ancient alveolate group of about 2000 described extant species, DNA barcoding studies have revealed large amounts of unrecognized species diversity, most of which is not represented in culture collections. To date, two mitochondrial gene markers, Cytochrome Oxidase I (COI) and Cytochrome b oxidase (COB), have been used to assess DNA barcoding in dinoflagellates, and both failed to amplify all taxa and suffered from low resolution. Nevertheless, both genes yielded many examples of morphospecies showing cryptic speciation and morphologically distinct named species being genetically similar, highlighting the need for a common marker. For example, a large number of cultured Symbiodinium strains have neither taxonomic identification, nor a common measure of diversity that can be used to compare this genus to other dinoflagellates. METHODOLOGY/PRINCIPAL FINDINGS: The purpose of this study was to evaluate the Internal Transcribed Spacer units 1 and 2 (ITS) of the rDNA operon, as a high resolution marker for distinguishing species dinoflagellates in culture. In our study, from 78 different species, the ITS barcode clearly differentiated species from genera and could identify 96% of strains to a known species or sub-genus grouping. 8.3% showed evidence of being cryptic species. A quarter of strains identified had no previous species identification. The greatest levels of hidden biodiversity came from Scrippsiella and the Pfiesteriaceae family, whilst Heterocapsa strains showed a high level of mismatch to their given species name. CONCLUSIONS/SIGNIFICANCE: The ITS marker was successful in confirming species, revealing hidden diversity in culture collections. This marker, however, may have limited use for environmental barcoding due to paralogues, the potential for unidentifiable chimaeras and priming across taxa. In these cases ITS would serve well in combination with other markers or for specific taxon studies.

Symbiodinium transcriptomes: genome insights into the dinoflagellate symbionts of reef-building corals.

Dinoflagellates are unicellular algae that are ubiquitously abundant in aquatic environments. Species of the genus Symbiodinium form symbiotic relationships with reef-building corals and other marine invertebrates. Despite their ecologic importance, little is known about the genetics of dinoflagellates in general and Symbiodinium in particular. Here, we used 454 sequencing to generate transcriptome data from two Symbiodinium species from different clades (clade A and clade B). With more than 56,000 assembled sequences per species, these data represent the largest transcriptomic resource for dinoflagellates to date. Our results corroborate previous observations that dinoflagellates possess the complete nucleosome machinery. We found a complete set of core histones as well as several H3 variants and H2A.Z in one species. Furthermore, transcriptome analysis points toward a low number of transcription factors in Symbiodinium spp. that also differ in the distribution of DNA-binding domains relative to other eukaryotes. In particular the cold shock domain was predominant among transcription factors. Additionally, we found a high number of antioxidative genes in comparison to non-symbiotic but evolutionary related organisms. These findings might be of relevance in the context of the role that Symbiodinium spp. play as coral symbionts.Our data represent the most comprehensive dinoflagellate EST data set to date. This study provides a comprehensive resource to further analyze the genetic makeup, metabolic capacities, and gene repertoire of Symbiodinium and dinoflagellates. Overall, our findings indicate that Symbiodinium possesses some unique characteristics, in particular the transcriptional regulation in Symbiodinium may differ from the currently known mechanisms of eukaryotic gene regulation.

Development of microsatellite markers as a molecular tool for conservation studies of the Mediterranean reef builder coral Cladocora caespitosa (Anthozoa, Scleractinia).

Cladocora caespitosa is a reef-building zooxanthellate scleractinian coral in the Mediterranean Sea. Mortality events have recurrently affected this species during the last decade. Thus, knowledge of its genetic structure, population diversity, and connectivity is needed to accomplish suitable conservation plans. In order to obtain a better understanding of the population genetics of this species, 13 highly variable microsatellites markers were developed from a naturally bleached colony. The developed primers failed to amplify zooxanthella DNA, isolated from C. caespitosa, verifying that these markers were of the coral and not algal symbiont origin. The degree of polymorphism of these loci was tested on tissue samples from 28 colonies. The allele number for each loci ranged from 2 to 13 (mean N(a) = 5.4), with an average observed heterozygosity of 0.42 (H(e) = 0.43) and all loci were in Hardy-Weinberg equilibrium. These new markers should be useful in future conservation genetic studies and will help to improve the resolution of the individual identification within this coral species. Primers were also tested in Oculina patagonica, with successful amplifications of several loci.