SYMBIODINIUM (DINOPHYTA) DIVERSITY AND STABILITY IN AQUARIUM CORALS1

Indo‐Pacific reef corals growing for years in closed‐system aquaria provide an alternate means to investigate host–symbiont specificity and stability. The diversity of dinoflagellate endosymbionts (Symbiodinium spp.) from coral communities in private and public aquaria was investigated using molecular‐genetic analyses. Of the 29 symbiont types (i.e., species) identified, 90% belonged to the most prevalent group of Symbiodinium harbored by Indo‐Pacific reef corals, Clade C, while the rest belonged to Clade D. Sixty‐five percent of all types were known from field surveys conducted throughout the Pacific and Indian oceans. Because specific coral–dinoflagellate partnerships appear to have defined geographic distributions, correspondence of the same symbionts in aquarium and field‐collected specimens identifies regions where particular colonies must have been collected in the wild. Symbiodinium spp. in clade D, believed to be “stress‐tolerant” and/or “opportunistic,” occurred in a limited number of individual colonies. The absence of a prevalent, or “weedy,” symbiont suggests that conditions under which aquarium corals are grown do not favor competitive replacements of their native symbiont populations. The finding of typical and diverse assemblages of Symbiodinium spp. among aquarium corals living many years under variable chemical/physical conditions, artificial and natural light, while undergoing fragmentation periodically, indicates that individual colonies maintain stable, long‐term symbiotic associations.     

Genetic identity of free‐living Symbiodinium obtained over a broad latitudinal range in the Japanese coast

The dinoflagellate Symbiodinium is well known to engage symbiosis with various marine animals, including corals. Recent records of environmental Symbiodinium (occurring in the environment and separately from host animals; usually referred to as ‘free‐living’ Symbiodinium) are of special interest, since these environmental populations are essential as symbiont sources for many host animals. In the present study, we carried out a phylogenetic analysis of environmental Symbiodinium isolates (culture strains) from sand, tide pools, or macroalgal surfaces, and environmental DNA clones extracted from the water‐column, at numerous sites around Japan. Our phylogenetic analysis based on the nuclear rRNA gene (internal transcribed spacers ‐1, ‐2, and 5.8S), indicated that most of the environmental isolates form monophyletic subclades within the Clade‐A lineage, and separate from a host‐associated Clade‐A population with high bootstrap values. Results of the partial nuclear 28S rDNA phylogeny and thecal‐plate observations revealed that these environmental isolates were closely related to a previously‐described ‘planktonic species’, Symbiodinium natans Gert Hansen et Daugbjerg, which was isolated from a plankton‐net sample from the Northeast‐Atlantic Ocean. On the other hand, the environmental DNA clones were also noted to be mostly nested within host‐associated Symbiodinium groups scattered in various clades. These results led to the assumption that the environmental Symbiodinium can be divided into two groups. One group, as typified by environmental isolates in the present study and previous reports, may be exclusively free‐living; the other group exists transiently in free‐living forms, possibly having been expelled from animal hosts. The populations within the latter group probably represent environmental sources of viable symbionts, because these are normally host‐associated. However, the Symbiodinium in the former group are not expected to engage in stable symbioses with host cnidarians.     

Symbiodinium (Dinophyceae) diversity in reef‐invertebrates along an offshore to inshore reef gradient near Lizard Island,Great Barrier Reef

Despite extensive work on the genetic diversity of reef invertebrate‐dinoflagellate symbioses on the Great Barrier Reef (GBR; Australia), large information gaps exist from northern and inshore regions. Therefore, a broad survey was done comparing the community of inshore, mid‐shelf and outer reefs at the latitude of Lizard Island. Symbiodinium (Freudenthal) diversity was characterized using denaturing gradient gel electrophoresis fingerprinting and sequencing of the ITS2 region of the ribosomal DNA. Thirty‐nine distinct Symbiodinium types were identified from four subgeneric clades (B, C, D, and G). Several Symbiodinium types originally characterized from the Indian Ocean were discovered as well as eight novel types (C1kk, C1LL, C3nn, C26b, C161a, C162, C165, C166). Multivariate analyses on the Symbiodinium species diversity data showed a strong link with host identity, consistent with previous findings. Of the four environmental variables tested, mean austral winter sea surface temperature (SST) influenced Symbiodinium distribution across shelves most significantly. A similar result was found when the analysis was performed on Symbiodinium diversity data of genera with an open symbiont transmission mode separately with chl a and PAR explaining additional variation. This study underscores the importance of SST and water quality related variables as factors driving Symbiodinium distribution on cross‐shelf scales. Furthermore, this study expands our knowledge on Symbiodinium species diversity, ecological partitioning (including host‐specificity) and geographic ranges across the GBR. The accelerating rate of environmental change experienced by coral reef ecosystems emphasizes the need to comprehend the full complexity of cnidarian symbioses, including the biotic and abiotic factors that shape their current distributions.     

Symbiotic specificity,association patterns,and function determine community responses to global changes: defining critical research areas for coral‐Symbiodinium symbioses

Climate change‐driven stressors threaten the persistence of coral reefs worldwide. Symbiotic relationships between scleractinian corals and photosynthetic endosymbionts (genus Symbiodinium) are the foundation of reef ecosystems, and these associations are differentially impacted by stress. Here, we couple empirical data from the coral reefs of Moorea, French Polynesia, and a network theoretic modeling approach to evaluate how patterns in coral‐Symbiodinium associations influence community stability under climate change. To introduce the effect of climate perturbations, we simulate local ‘extinctions’ that represent either the loss of coral species or the ability to engage in symbiotic interactions. Community stability is measured by determining the duration and number of species that persist through the simulated extinctions. Our results suggest that four factors greatly increase coral‐Symbiodinium community stability in response to global changes: (i) the survival of generalist hosts and symbionts maximizes potential symbiotic unions; (ii) elevated symbiont diversity provides redundant or complementary symbiotic functions; (iii) compatible symbiotic assemblages create the potential for local recolonization; and (iv) the persistence of certain traits associate with symbiotic diversity and redundancy. Symbiodinium may facilitate coral persistence through novel environmental regimes, but this capacity is mediated by symbiotic specificity, association patterns, and the functional performance of the symbionts. Our model‐based approach identifies general trends and testable hypotheses in coral‐Symbiodinium community responses. Future studies should consider similar methods when community size and/or environmental complexity preclude experimental approaches.     

New insights into the dynamics between reef corals and their associated dinoflagellate endosymbionts from population genetic studies

The mutualistic symbioses between reef‐building corals and micro‐algae form the basis of coral reef ecosystems, yet recent environmental changes threaten their survival. Diversity in host‐symbiont pairings on the sub‐species level could be an unrecognized source of functional variation in response to stress. The Caribbean elkhorn coral, Acropora palmata, associates predominantly with one symbiont species (Symbiodinium ‘fitti’), facilitating investigations of individual‐level (genotype) interactions. Individual genotypes of both host and symbiont were resolved across the entire species’ range. Most colonies of a particular animal genotype were dominated by one symbiont genotype (or strain) that may persist in the host for decades or more. While Symbiodinium are primarily clonal, the occurrence of recombinant genotypes indicates sexual recombination is the source of this genetic variation, and some evidence suggests this happens within the host. When these data are examined at spatial scales spanning the entire distribution of A. palmata, gene flow among animal populations was an order of magnitude greater than among populations of the symbiont. This suggests that independent micro‐evolutionary processes created dissimilar population genetic structures between host and symbiont. The lower effective dispersal exhibited by the dinoflagellate raises questions regarding the extent to which populations of host and symbiont can co‐evolve during times of rapid and substantial climate change. However, these findings also support a growing body of evidence, suggesting that genotype‐by‐genotype interactions may provide significant physiological variation, influencing the adaptive potential of symbiotic reef corals to severe selection.     

Local endemicity and high diversity characterise high-latitude coral–Symbiodinium partnerships

Obligate symbiotic dinoflagellates (Symbiodinium) residing within the tissues of most reef invertebrates are important in determining the tolerance range of their host. Coral communities living at high latitudes experience wide fluctuations in environmental conditions and thus provide an ideal system to gain insights into the range within which the symbiotic relationship can be sustained. Further, understanding whether and how symbiont communities associated with high-latitude coral reefs are different from their tropical counterparts will provide clues to the potential of corals to cope with marginal or changing conditions. However, little is known of the host and symbiont partnerships at high latitudes. Symbiodinium diversity and specificity of high-latitude coral communities were explored using denaturing gradient gel electrophoresis (PCR-DGGE) analysis of the internal transcribed spacer regions (ITS1 and ITS2) of the ribosomal DNA at Lord Howe Island (31°S; Australia), and the Kermadec Islands (29°S; New Zealand). All but one host associated with clade C Symbiodinium, the exception being a soft coral (Capnella sp.) that contained Symbiodinium B1. Besides ‘host-generalist’ Symbiodinium types C1 and C3, approximately 72% of the Symbiodinium identified were novel C types, and zonation of symbionts in relation to environmental parameters such as depth and turbidity was evident in certain host species. The high-latitude Symbiodinium communities showed little overlap and relatively high diversity compared with communities sampled on the tropical Great Barrier Reef. Although host specificity was maintained in certain species, others shared symbionts and this potential reduction of fidelity at high-latitude locations may be the result of locally challenging and highly variable environmental conditions.     

Different strategies of energy storage in cultured and freshly isolated Symbiodinium sp.

The endosymbiotic relationship between cnidarians and Symbiodinium is critical for the survival of coral reefs. In this study, we developed a protocol to rapidly and freshly separate Symbiodinium from corals and sea anemones. Furthermore, we compared these freshly‐isolated Symbiodinium with cultured Symbiodinium to investigate host and Symbiodinium interaction. Clade B Symbiodinium had higher starch content and lower lipid content than those of clades C and D in both freshly isolated and cultured forms. Clade C had the highest lipid content, particularly when associated with corals. Moreover, the coral‐associated Symbiodinium had higher protein content than did cultured and sea anemone‐associated Symbiodinium. Regarding fatty acid composition, cultured Symbiodinium and clades B, C, and D shared similar patterns, whereas sea anemone‐associated Symbiodinium had a distinct pattern compared coral‐associated Symbiodinium. Specifically, the levels of monounsaturated fatty acids were lower than those of the saturated fatty acids, and the level of polyunsaturated fatty acids (PUFAs) were the highest in all examined Symbiodinium. Furthermore, PUFAs levels were higher in coral‐associated Symbiodinium than in cultured Symbiodinium. These results altogether indicated that different Symbiodinium clades used different energy storage strategies, which might be modified by hosts.     

Host-Specific Interactions with Environmental Factors Shape the Distribution of Symbiodinium across the Great Barrier Reef

Background

The endosymbiotic dinoflagellates (genus Symbiodinium) within coral reef invertebrates are critical to the survival of the holobiont. The genetic variability of Symbiodinium may contribute to the tolerance of the symbiotic association to elevated sea surface temperatures (SST). To assess the importance of factors such as the local environment, host identity and biogeography in driving Symbiodinium distributions on reef-wide scales, data from studies on reef invertebrate-Symbiodinium associations from the Great Barrier Reef (GBR) were compiled.

Methodology/Principal Findings

The resulting database consisted of 3717 entries from 26 studies. It was used to explore ecological patterns such as host-specificity and environmental drivers structuring community complexity using a multi-scalar approach. The data was analyzed in several ways: (i) frequently sampled host species were analyzed independently to investigate the influence of the environment on symbiont distributions, thereby excluding the influence of host specificity, (ii) host species distributions across sites were added as an environmental variable to determine the contribution of host identity on symbiont distribution, and (iii) data were pooled based on clade (broad genetic groups dividing the genus Symbiodinium) to investigate factors driving Symbiodinium distributions using lower taxonomic resolution. The results indicated that host species identity plays a dominant role in determining the distribution of Symbiodinium and environmental variables shape distributions on a host species-specific level. SST derived variables (especially SSTstdev) most often contributed to the selection of the best model. Clade level comparisons decreased the power of the predictive model indicating that it fails to incorporate the main drivers behind Symbiodinium distributions.

Conclusions/Significance

Including the influence of different host species on Symbiodinium distributional patterns improves our understanding of the drivers behind the complexity of Symbiodinium-invertebrate symbioses. This will increase our ability to generate realistic models estimating the risk reefs are exposed to and their resilience in response to a changing climate.     

New Species of Closely Related Endosymbiotic Dinoflagellates in the Greater Caribbean have Niches Corresponding to Host Coral Phylogeny

Symbiotic dinoflagellates in the genus Breviolum (formerly Symbiodinium Clade B) dominate coral communities in shallow waters across the Greater Caribbean. While some formally described species exist, mounting genetic, and ecological evidence indicate that numerous more comprise this genus, many of which are closely related. To test this, colonies of common reef‐building corals were sampled across a large geographical range. Phylogenetic and population genetic markers then used to examine evolutionary divergence and delineate boundaries of genetic recombination. Three new candidate species were distinguished by fixed differences in nucleotide sequences from nuclear and chloroplast DNA. Population connectivity was evident within each lineage over thousands of kilometers, however, substantial genetic structure persisted between lineages co‐occurring within sampling locations, signifying reproductive isolation. While geographically widespread with overlapping distributions, each species is ecologically distinct, exhibiting specific mutualisms with phylogenetically distinct coral hosts. Moreover, significant differences in mean cell sizes provide some morphological evidence substantiating formal species distinctions. In providing evidence that satisfies the biological, phylogenetic, ecological, and morphological species concepts, we classify and formally name Breviolum faviinorum n. sp., primarily associated with Caribbean corals belonging to the Caribbean subfamily Faviinae; B. meandrinium n. sp., associated with corals belonging to the family Meandrinidae; and B. dendrogyrum n. sp., a symbiont harbored exclusively by the threatened coral Dendrogyra cylindrus. These findings support the primary importance of niche diversification (i.e. host habitat) in the speciation of symbiotic dinoflagellates.     

Many corals host thermally resistant symbionts in high-temperature habitat

Physiologically distinct lines of dinoflagellate symbionts, Symbiodinium spp., may confer distinct thermal tolerance thresholds on their host corals. Therefore, if a coral can alternately host distinct symbionts, changes in their Symbiodinium communities might allow corals to better tolerate increasing environmental temperatures. However, researchers are currently debating how commonly coral species can host different symbiont types. We sequenced chloroplast 23 s rDNA from the Symbiodinium communities of nine reef-building coral species across two thermally distinct lagoon pools separated by ~500 m. The hotter of these pools reaches 35°C in the summer months, while the other pool’s maximum temperature is 1.5°C cooler. Across 217 samples from nine species, we found a single haplotype in both Symbiodinium clades A and D, but four haplotypes in Symbiodinium clade C. Eight of nine species hosted a putatively thermally resistant member of clade D Symbiodinium at least once, one of which hosted this clade D symbiont exclusively. Of the remaining seven that hosted multiple Symbiodinium types, six species showed higher proportions of the clade D symbiont in the hotter pool. Average percentage rise in the frequency of the clade D symbiont from the hotter to cooler pool was 52% across these six species. Even though corals hosted members of both the genetically divergent clades D and C Symbiodinium, some showed patterns of host–symbiont specificity within clade C. Both Acropora species that hosted clade C exclusively hosted a member of sub-clade C2, while all three Pocillopora species hosted a member of sub-clade C1 (sensu van Oppen et al. 2001). Our results suggest that coral–algal symbioses often conform to particular temperature environments through changes in the identity of the algal symbiont.     

CORRESPONDENCE BETWEEN COLD TOLERANCE AND TEMPERATE BIOGEOGRAPHY IN A WESTERN ATLANTIC SYMBIODINIUM (DINOPHYTA) LINEAGE1

Many corals form obligate symbioses with photosynthetic dinoflagellates of the genus Symbiodinium Freudenthal (1962). These symbionts vary genotypically, with their geographical distribution and abundance dependent upon host specificity and tolerance to temperature and light variation. Despite the importance of these mutualistic relationships, the physiology and ecology of Symbiodinium spp. remain poorly characterized. Here, we report that rDNA internal transcribed spacer region 2 (ITS2) defined Symbiodinium type B2 associates with the cnidarian hosts Astrangia poculata and Oculina arbuscula from northerly habitats of the western Atlantic. Using pulse‐amplitude‐modulated (PAM) fluorometry, we compared maximum photochemical efficiency of PSII of type B2 to that of common tropical Symbiodinium lineages (types A3, B1, and C2) under cold‐stress conditions. Symbiont cultures were gradually cooled from 26°C to 10°C to simulate seasonal temperature declines. Cold stress decreased the maximum photochemical efficiency of PSII and likely the photosynthetic potential for all Symbiodinium clades tested. Cultures were then maintained at 10°C for a 2‐week period and gradually returned to initial conditions. Subsequent to low temperature stress, only type B2 displayed rapid and full recovery of PSII photochemical efficiency, whereas other symbiont phylotypes remained nonfunctional. These findings indicate that the distribution and abundance of Symbiodinium spp., and by extension their cnidarian hosts, in temperate climates correspond significantly with the photosynthetic cold tolerance of these symbiotic algae.     

Do introduced endosymbiotic dinoflagellates ‘take’ to new hosts?

In a recent communication by Stat and Gates (Biol Invasions 10: 579–583, 2008), discovery of a symbiotic combination involving the coral Acropora cytherea and the dinoflagellate endosymbiont, Symbiodinium A1 (Symbiodinium microadriaticum, Freudenthal sensu stricto) in the Northwest Hawaiian Islands was interpreted to be the result of a ‘recent’ introduction. While introductions of symbiotic dinoflagellates have occurred and are occurring, the authors’ conclusion was made without sufficient information about the geographic range and host specificity exhibited by A1. The only direct genetic analysis of symbionts from the putative host vector, a jellyfish in the genus Cassiopeia sp., from Kaneohe Bay on the Island of Oahu, found that it contained a different symbiont species, A3. Furthermore, Stat and Gates (Biol Invasions 10: 579–583, 2008) did not consider the importance of host-symbiont specificity in preventing the establishment of a foreign symbiont species. In comparison to A. cytherea, A. longicyathus on the southern most Great Barrier Reef also hosts Symbiodinium A1 and a closely related endemic, A1a. Instead of assuming that A. cytherea has an unnatural association, a practical explanation is that long-term ecological and evolutionary processes influenced by local environments underlie the unusual, but not unprecedented finding of a Pacific acroporid associating with Clade A Symbiodinium spp.     

Methods for sampling free-living Symbiodinium (zooxanthellae) and their distribution and abundance at Lizard Island (Great Barrier Reef)

The abundance and distribution of free-living dinoflagellates in the genus Symbiodinium have important implications for the ecology of coral reefs, determining both the symbionts available to newly recruited corals and symbiont types available for uptake by adult corals during environmental stress. However, little is known about where symbiotic dinoflagellates reside outside the host, due to the difficulty of capturing and detecting unicellular organisms in the marine environment. This study presents a successful protocol for sampling Symbiodinium from both the benthos and the water column. Comparisons of two detection methods for enumerating Symbiodinium indicated that conventional microscope analysis is accurate and more efficient when estimating Symbiodinium densities in sediment samples, while an automated particle counter (FlowCAM) was more efficient in detecting cells in the water column where densities are low. Symbiodinium densities were found to be relatively high (1000–4000 cells/mL) in sediment samples and much lower (up to 80 cells/mL) in the water column, indicating that the free-living form resides mainly in the benthos. Symbiodinium densities were found to be highly variable spatially, differing significantly between two reef locations. Within sites, elevated densities of Symbiodinium along reef margins combined with significant decreases in densities one meter away from the reef, suggest that cells aggregate within the reef habitat.     

Symbiont acquisition strategy drives host–symbiont associations in the southern Great Barrier Reef

Coral larvae acquire populations of the symbiotic dinoflagellate Symbiodinium from the external environment (horizontal acquisition) or inherit their symbionts from the parent colony (maternal or vertical acquisition). The effect of the symbiont acquisition strategy on Symbiodinium-host associations has not been fully resolved. Previous studies have provided mixed results, probably due to factors such as low sample replication of Symbiodinium from a single coral host, biogeographic differences in Symbiodinium diversity, and the presence of some apparently host-specific symbiont lineages in coral with either symbiont acquisition strategies. This study set out to assess the effect of the symbiont acquisition strategy by sampling Symbiodinium from 10 coral species (five with a horizontal and five with a vertical symbiont acquisition strategy) across two adjacent reefs in the southern Great Barrier Reef. Symbiodinium diversity was assessed using single-stranded conformational polymorphism of partial nuclear large subunit rDNA and denaturing gradient gel electrophoresis of the internal transcribed spacer 2 region. The Symbiodinium population in hosts with a vertical symbiont acquisition strategy partitioned according to coral species, while hosts with a horizontal symbiont acquisition strategy shared a common symbiont type across the two reef environments. Comparative analysis of existing data from the southern Great Barrier Reef found that the majority of corals with a vertical symbiont acquisition strategy associated with distinct species- or genus-specific Symbiodinium lineages, but some could also associate with symbiont types that were more commonly found in hosts with a horizontal symbiont acquisition strategy.     

Sibling species of mutualistic Symbiodinium clade G from bioeroding sponges in the western Pacific and western Atlantic oceans

Dinoflagellates in the genus Symbiodinium associate with a broad array of metazoan and protistian hosts. Symbiodinium‐based symbioses involving bioeroding sponge hosts have received less attention than those involving popular scleractinian hosts. Certain species of common Cliona harbor high densities of an ecologically restricted group of Symbiodinium, referred to as Clade G. Clade G Symbiodinium are also known to form stable and functionally important associations with Foraminifera and black corals (Antipatharia) Analyses of genetic evidence indicate that Clade G likely comprises several distinct species. Here, we use nucleotide sequence data in combination with ecological and geographic attributes to formally describe Symbiodinium endoclionum sp. nov. obtained from the Pacific boring sponge Cliona orientalis and Symbiodinium spongiolum sp. nov. from the congeneric western Atlantic sponge Cliona varians. These species appear to be part of an adaptive radiation comprising lineages of Clade G specialized to the metazoan phyla Porifera and Cnidaria, which began prior to the separation of the Pacific and Atlantic Oceans.     

Stability of coral–endosymbiont associations during and after a thermal stress event in the southern Great Barrier Reef

Shifts in the community of symbiotic dinoflagellates to those that are better suited to the prevailing environmental condition may provide reef-building corals with a rapid mechanism by which to adapt to changes in the environment. In this study, the dominant Symbiodinium in 10 coral species in the southern Great Barrier Reef was monitored over a 1-year period in 2002 that coincided with a thermal stress event. Molecular genetic profiling of Symbiodinium communities using single strand conformational polymorphism of the large subunit rDNA and denaturing gradient gel electrophoresis of the internal transcribed spacer 2 region did not detect any changes in the communities during and after this thermal-stress event. Coral colonies of seven species bleached but recovered with their original symbionts. This study suggests that the shuffling or switching of symbionts in response to thermal stress may be restricted to certain coral species and is probably not a universal feature of the coral–symbiont relationship. Communicated by Biology Editor Dr. Clay Cook     

Assessing Symbiodinium diversity in scleractinian corals via next‐generation sequencing‐based genotyping of the ITS2 rDNA region

The persistence of coral reef ecosystems relies on the symbiotic relationship between scleractinian corals and intracellular, photosynthetic dinoflagellates in the genus Symbiodinium. Genetic evidence indicates that these symbionts are biologically diverse and exhibit discrete patterns of environmental and host distribution. This makes the assessment of Symbiodinium diversity critical to understanding the symbiosis ecology of corals. Here, we applied pyrosequencing to the elucidation of Symbiodinium diversity via analysis of the internal transcribed spacer 2 (ITS2) region, a multicopy genetic marker commonly used to analyse Symbiodinium diversity. Replicated data generated from isoclonal Symbiodinium cultures showed that all genomes contained numerous, yet mostly rare, ITS2 sequence variants. Pyrosequencing data were consistent with more traditional denaturing gradient gel electrophoresis (DGGE) approaches to the screening of ITS2 PCR amplifications, where the most common sequences appeared as the most intense bands. Further, we developed an operational taxonomic unit (OTU)‐based pipeline for Symbiodinium ITS2 diversity typing to provisionally resolve ecologically discrete entities from intragenomic variation. A genetic distance cut‐off of 0.03 collapsed intragenomic ITS2 variants of isoclonal cultures into single OTUs. When applied to the analysis of field‐collected coral samples, our analyses confirm that much of the commonly observed Symbiodinium ITS2 diversity can be attributed to intragenomic variation. We conclude that by analysing Symbiodinium populations in an OTU‐based framework, we can improve objectivity, comparability and simplicity when assessing ITS2 diversity in field‐based studies.