The genetic identity of dinoflagellate symbionts in Caribbean octocorals

Many cnidarians (e.g., corals, octocorals, sea anemones) maintain a symbiosis with dinoflagellates (zooxanthellae). Zooxanthellae are grouped into clades, with studies focusing on scleractinian corals. We characterized zooxanthellae in 35 species of Caribbean octocorals. Most Caribbean octocoral species (88.6%) hosted clade B zooxanthellae, 8.6% hosted clade C, and one species (2.9%) hosted clades B and C. Erythropodium caribaeorum harbored clade C and a unique RFLP pattern, which, when sequenced, fell within clade C. Five octocoral species displayed no zooxanthella cladal variation with depth. Nine of the ten octocoral species sampled throughout the Caribbean exhibited no regional zooxanthella cladal differences. The exception, Briareum asbestinum, had some colonies from the Dry Tortugas exhibiting the E. caribaeorum RFLP pattern while elsewhere hosting clade B. In the Caribbean, octocorals show more symbiont specificity at the cladal level than scleractinian corals. Both octocorals and scleractinian corals, however, exhibited taxonomic affinity between zooxanthella clade and host suborder.Communicated by R.C. Carpenter     

Symbiodinium diversity among host clionaid sponges from Caribbean and Pacific reefs: Evidence of heteroplasmy and putative host-specific symbiont lineages

Among the Porifera, symbiosis with Symbiodinium spp. (i.e., zooxanthellae) is largely restricted to members of the family Clionaidae. We surveyed the diversity of zooxanthellae associated with sponges from the Caribbean and greater Indo-Pacific regions using chloroplast large subunit (cp23S) domain V sequences. We provide the first report of Clade C Symbiodinium harbored by a sponge (Cliona caesia), and the first report of Clade A Symbiodinium from an Indo-Pacific sponge (C. jullieni). Clade A zooxanthellae were also identified in sponges from the Caribbean, which has been reported previously. Sponges that we examined from the Florida Keys all harbored Clade G Symbiodinium as did C. orientalis from the Indo-Pacific, which also supports earlier work with sponges. Two distinct Clade G lineages were identified in our phylogenetic analysis; Symbiodinium extracted from clionaid sponges formed a monophyletic group sister to Symbiodinium found in foraminiferans. Truncated and 'normal' length variants of 23S rDNA sequences were detected simultaneously in all three morphotypes of C. varians providing the first evidence of chloroplast-based heteroplasmy in a sponge. None of the other sponge species examined showed evidence of heteroplasmy. As in previous work, length variation in cp23S domain V sequences was found to correspond in a highly precise manner to finer resolution of phylogenetic topology among Symbiodinium clades. On a global scale, existing data indicate that members of the family Clionaidae that host zooxanthellae can form symbiotic associations with at least four Symbiodinium clades. The majority of sponge hosts appear to harbor only one cladal type of symbiont, but some species can harbor more than one clade of zooxanthellae concurrently. The observed differences in the number of partners harbored by sponges raise important questions about the degree of coevolutionary integration and specificity of these symbioses. Although our sample sizes are small, we propose that one of the Clade G lineages identified in this study is comprised of sponge-specialist zooxanthellae. These zooxanthellae are common in Caribbean sponges, but additional work in other geographic regions is necessary to test this idea. Sponges from the Indo-Pacific region harbor zooxanthellae from Clades A, C, and G, but more sponges from this region should be examined.     

Patterns of coral-dinoflagellate associations in Acropora: significance of local availability and physiology of Symbiodinium strains and host-symbiont selectivity

Like other reef-building corals, members of the genus Acropora form obligate endosymbioses with dinoflagellates (zooxanthellae) belonging to the genus Symbiodinium. Both Symbiodinium and its hosts are diverse assemblages, and the relationships between host and algal genotypes are unclear. In this study, we determined phylogenetic relationships between Symbiodinium isolates from a wide range of Acropora species and plotted the algal genotypes onto a molecular phylogeny of 28 Acropora species, using the same samples for the host and symbiont genotyping. In addition, we performed a preliminary survey of zooxanthella distribution in Acropora species from the central Great Barrier Reef. Three of the four known major zooxanthellae clades were represented in the 168 samples examined, and within the major clade C, three distinct subclades were identified. No evidence was found for coevolution, but several clear patterns of specificity were identified. Moreover, composition of the zooxanthella pool varied among locales and in one host species we found light-related patterns of zooxanthella distribution.     

Phylogenetic relationships among zooxanthellae (Symbiodinium) associated to excavating sponges (Cliona spp.) reveal an unexpected lineage in the Caribbean

Phylogenetic relationships of symbiotic dinoflagellate lineages, distributed in all tropical and subtropical seas, suggest strategies for long distance dispersal but at the same time strong host specialization. Zooxanthellae (Symbiodinium: Dinophyta), which are associated to diverse shallow-water cnidarians, also engage in symbioses with some sponge species of the genus Cliona. In the Caribbean, zooxanthellae-bearing Cliona has recently become abundant due to global warming, overfishing, and algae abundance. Using molecular techniques, the symbionts from five excavating species (Clionacaribbaea, C. tenuis, C. varians, C. aprica and C. laticavicola) from the southern and southwestern Caribbean were surveyed. Several DNA sequence regions were used in order to confirm zooxanthellae identity; 18S rDNA, domain V of chloroplast large subunit (cp23S), internal transcribed spacer 2 (ITS2), and ITS2 secondary structure. Sequence analyses corroborated the presence of three zooxanthellae clades: A, B, and G. Presence of clades A and B in common boring sponges of the Caribbean fit with the general pattern of the province. The discovery of clade G for the first time in any organism of the Atlantic Ocean leads us to consider this unusual finding as a phylogenetic relict through common ancestors of sponge clades or an invasion of the sponge from the Indo-Pacific.     

造礁石珊瑚与其共生藻(Symbiodinium)共生研究进展

对造礁石珊瑚与其共生藻共生研究现状及其在全球变化下的适应能力进行较全面的综述.造礁石珊瑚与遗传和生理功能独特的共生藻组成内共生关系是成功演化的范例.近年来对珊瑚共生体的分子系统学研究表明共生藻遗传多样性极为丰富,当前认为共生藻属至少包括8个(A-H)各自包含亚系群的世系或系群.珊瑚-共生藻共生功能体对诸如全球变化引起的海水温度上升等环境变化十分敏感.由于珊瑚以及珊瑚礁面临气候变化的严峻挑战,对珊瑚与其共生藻共生关系和共生功体适应能力的研究将是未来重要的研究领域之一.     

Fine-scale diversity and specificity in the most prevalent lineage of symbiotic dinoflagellates (Symbiodinium, Dinophyceae) of the Caribbean

The success of coral reefs is due to obligate mutualistic symbioses involving invertebrates and photosynthetic dinoflagellate symbionts belonging to the genus Symbiodinium. In the Caribbean, the vast majority of octocorals and other invertebrate hosts associate with Symbiodinium clade B, and more selectively, with a single lineage of this clade, Symbiodinium B1/B184. Although B1/B184 represents the most prevalent Symbiodinium in the Caribbean, there is little evidence supporting fine-scale diversity and host-alga specificity within this lineage. We explored simultaneously the questions of diversity and specificity in Symbiodinium B1/B184 by sequencing the flanking regions of two polymorphic microsatellites from a series of Symbiodinium clade B cultures along with Symbiodinium B1/B184 populations of the octocorals Pseudopterogorgia elisabethae, P. bipinnata and Gorgonia ventalina. Seven unique sequence variants were identified based on concatenation of the two loci. Phylogenetic analyses of these variants, which we refer to as phylotypes, recognized five as belonging to B1/B184, thus providing the first evidence of distinct taxa within this Symbiodinium lineage. Furthermore, sympatric P. elisabethae and P. bipinnata at San Salvador in the Bahamas were found to harbour distinct Symbiodinium B1/B184 phylotypes, demonstrating unequivocally the existence of fine-scale specificity between Caribbean octocorals and these algae. Taken together, this study exemplifies the complex nature of Symbiodinium biodiversity and specificity.     

Identity and diversity of coral endosymbionts (zooxanthellae) from three Palauan reefs with contrasting bleaching, temperature and shading histories

The potential of corals to associate with more temperature-tolerant strains of algae (zooxanthellae, Symbiodinium) can have important implications for the future of coral reefs in an era of global climate change. In this study, the genetic identity and diversity of zooxanthellae was investigated at three reefs with contrasting histories of bleaching mortality, water temperature and shading, in the Republic of Palau (Micronesia). Single-stranded conformation polymorphism and sequence analysis of the ribosomal DNA internal transcribed spacer (ITS)1 region was used for genotyping. A chronically warm but partly shaded coral reef in a marine lake that is hydrographically well connected to the surrounding waters harboured only two single-stranded conformation polymorphism profiles (i.e. zooxanthella communities). It consisted only of Symbiodinium D in all 13 nonporitid species and two Porites species investigated, with the remaining five Porites harbouring C*. Despite the high temperature in this lake (> 0.5 degrees above ambient), this reef did not suffer coral mortality during the (1998) bleaching event, however, no bleaching-sensitive coral families and genera occur in the coral community. This setting contrasts strongly with two other reefs with generally lower temperatures, in which 10 and 12 zooxanthella communities with moderate to low proportions of clade D zooxanthellae were found. The data indicate that whole coral assemblages, when growing in elevated seawater temperatures and at reduced irradiance, can be composed of colonies associated with the more thermo-tolerant clade D zooxanthellae. Future increases in seawater temperature might, therefore, result in an increasing prevalence of Symbiodinium phylotype D in scleractinian corals, possibly associated with a loss of diversity in both zooxanthellae and corals.     

Diversity of symbiotic algae of the genus Symbiodinium in scleractinian corals of the Xisha Islands in the South China Sea

Symbiotic algae （Symbiodinium sp.） in scleractinian corals are important in understanding how coral reefs will respond to global climate change. The present paper reports on the diversity of Symbiodinium sp. in 48 scleractinian coral species from 25 genera and 10 families sampled from the Xisha Islands in the South China Sea, which were identified with the use of restriction fragment length polymorphism （RFLP） of the nuclear ribosomal DNA large subunit gene （rDNA）. The results showed that： （i） Symbiodinium Clade C was the dominant zooxanthellae in scleractinian corals in the Xisha Islands; （ii） Symbiodinium Clade D was found in the corals Montipora aequituberculata, Galaxea fascicularis, and Plerogyra sinuosa; and （iii） both Symbiodinium Clades C and D were found simultaneously in Montipora digitata, Psammocora contigua, and Galaxeafascicularis. A poor capacity for symbiosis polymorphism, as uncovered by RFLP, in the Xisha Islands indicates that the scleractinian corals have low adaptability to environmental changes. Further studies are needed to investigate zooxanthellae diversity using other molecular markers.     

Diversity of Scleractinia and Octocorallia in the mesophotic zone of the Great Barrier Reef, Australia

Mesophotic coral reefs in the Indo-West Pacific, the most diverse coral reef region on earth, are among the least documented. This study provides the first detailed investigation of the diversity of Scleractinia and Octocorallia of the mesophotic Great Barrier Reef (GBR). Specimens were collected by 100-m rock dredge tows at 47–163 m depth on 23 sites in four regions (15.3°–19.7° latitude South). Twenty-nine hard coral species from 19 families were recorded, with the greatest diversity found at <60 m depth, and no specimen was found >102 m. Many of these species are also commonly observed at shallower depths, particularly in inshore areas. Twenty-seven octocoral genera were collected, 25 of which represented azooxanthellate genera. Generic richness of octocorals was highest at depths >60 m. Sixteen of the 25 azooxanthellate genera were either absent or very rare at <18 m, and only five azooxanthellate genera were common on both shallow and mesophotic reefs. Species-area models indicated that the total diversity of hard corals on the deep mesophotic reefs sampled during this study was ~84 species while octocorals were represented by ~37 genera; however, the wide 95% confidence limits indicates that more intensive sampling effort is required to improve the accuracy of these estimates. Nonetheless, these results show that the taxonomic richness, particularly of hard corals, on mesophotic reefs may be much higher than previously thought, a finding that has implications for the comprehensive and adequate protection of the full range of biodiversity of the GBR.     

不同发育阶段的黄癣蜂巢珊瑚共生虫黄藻多样性

【背景】黄癣蜂巢珊瑚是我国南海主要的造礁石珊瑚之一,但至今未见对其共生虫黄藻的相关研究报道。【目的】研究黄癣蜂巢珊瑚不同发育阶段的共生虫黄藻多样性,探讨其获取途径。【方法】用荧光显微镜观察黄癣蜂巢珊瑚卵子、浮浪幼虫以及珊瑚成体中的共生虫黄藻。基于ITS2序列的克隆文库技术分析珊瑚卵子、浮浪幼虫、成体中共生虫黄藻的多样性。【结果】荧光显微镜观察结果显示,黄癣蜂巢卵子及发育至第4天的浮浪幼虫中均无共生虫黄藻。没有在黄癣蜂巢珊瑚卵子及4 d浮浪幼虫中检测到虫黄藻ITS2序列,但是在黄癣蜂巢珊瑚成体中检测到共生虫黄藻序列,系统发育分析提示为C1型虫黄藻。【结论】成体黄癣蜂巢珊瑚中发现了共生虫黄藻,而卵子、浮浪幼虫阶段均未发现虫黄藻,提示黄癣蜂巢珊瑚中的虫黄藻可能是后期从海水中水平转移而来,而不是由母体垂直遗传的。     

Diversity and distribution of symbiodinium associated with seven common coral species in the Chagos Archipelago, central Indian Ocean

The Chagos Archipelago designated as a no-take marine protected area in 2010, lying about 500 km south of the Maldives in the Indian Ocean, has a high conservation priority, particularly because of its fast recovery from the ocean-wide massive coral mortality following the 1998 coral bleaching event. The aims of this study were to examine Symbiodinium diversity and distribution associated with scleractinian corals in five atolls of the Chagos Archipelago, spread over 10,000 km(2). Symbiodinium clade diversity in 262 samples of seven common coral species, Acropora muricata, Isopora palifera, Pocillopora damicornis, P. verrucosa, P. eydouxi, Seriatopora hystrix, and Stylophora pistillata were determined using PCR-SSCP of the ribosomal internal transcribed spacer 1 (ITS1), PCR-DDGE of ITS2, and phylogenetic analyses. The results indicated that Symbiodinium in clade C were the dominant symbiont group in the seven coral species. Our analysis revealed types of Symbiodinium clade C specific to coral species. Types C1 and C3 (with C3z and C3i variants) were dominant in Acroporidae and C1 and C1c were the dominant types in Pocilloporidae. We also found 2 novel ITS2 types in S. hystrix and 1 novel ITS2 type of Symbiodinium in A. muricata. Some colonies of A. muricata and I. palifera were also associated with Symbiodinium A1. These results suggest that corals in the Chagos Archipelago host different assemblages of Symbiodinium types then their conspecifics from other locations in the Indian Ocean; and that future research will show whether these patterns in Symbiodinium genotypes may be due to local adaptation to specific conditions in the Chagos.     

Specificity is rarely absolute in coral-algal symbiosis: implications for coral response to climate change

Some reef-building corals have been shown to respond to environmental change by shifting the composition of their algal symbiont (genus Symbiodinium) communities. These shifts have been proposed as a potential mechanism by which corals might survive climate stressors, such as increased temperatures. Conventional molecular methods suggest this adaptive capacity may not be widespread because few (～25%) coral species have been found to associate with multiple Symbiodinium clades. However, these methods can fail to detect low abundance symbionts (typically less than 10-20% of the total algal symbiont community). To determine whether additional Symbiodinium clades are present, but are not detected using conventional techniques, we applied a high-resolution, real-time PCR assay to survey Symbiodinium (in clades A-D) from 39 species of phylogenetically and geographically diverse scleractinian corals. This survey included 26 coral species thought to be restricted to hosting a single Symbiodinium clade ('symbiotic specialists'). We detected at least two Symbiodinium clades (C and D) in at least one sample of all 39 coral species tested; all four Symbiodinium clades were detected in over half (54%) of the 26 symbiotic specialist coral species. Furthermore, on average, 68 per cent of all sampled colonies within a given coral species hosted two or more symbiont clades. We conclude that the ability to associate with multiple symbiont clades is common in scleractinian (stony) corals, and that, in coral-algal symbiosis, 'specificity' and 'flexibility' are relative terms: specificity is rarely absolute. The potential for reef corals to adapt or acclimatize to environmental change via symbiont community shifts may therefore be more phylogenetically widespread than has previously been assumed.     

Symbiodinium clade C dominates zooxanthellate corals (Scleractinia) in the temperate region of Japan

Endosymbiotic algae of the genus Symbiodinium have been divided into nine clades (A-I) following genetic classification; some clades are known to have physiological properties that enable the coral hosts to adapt to different environmental conditions. To understand the relationships of coral-alga symbioses, we focused on Symbiodinium diversity in zooxanthellate corals living under the severe environmental conditions of the temperate region (30°-35°N) of Japan. We investigated Symbiodinium clades in 346 colonies belonging to 58 coral species from six locations. We then selected three coral species-Acropora hyacinthus, Acropora japonica, and Cyphastrea chalcidicum-to investigate whether Symbiodinium clades changed during winter or summer over the course of year (May 2009-Apr 2010) in Tanabe Bay, Japan. Three Symbiodinium clades (C, D, and F) were detected in corals in the temperate region. Notably, 56 coral species contained Symbiodinium clade C. Oulastrea crispata predominantly contained clade D, but traces of clade C were also detected in all samples. The temperate-specific species Alveopora japonica contained clades C and F simultaneously. Seasonal change of symbiont clades did not occur in the three coral species during the investigation period where SSTs range on 12.5-29.2°C. However, we found Acropora (2 spp.) and Cyphastrea (1 sp.) contained different subcladal types of clade C. These results reveal that most coral species harbored Symbiodinium clade C stably throughout the year, suggesting that Symbiodinium clade C shows low-temperature tolerance, and that two hypothetical possibilities; genetic differences of subcladal types generating physiological differences or wide physiological flexibility in the clade C.     

Variation in Symbiodinium ITS2 sequence assemblages among coral colonies

Endosymbiotic dinoflagellates in the genus Symbiodinium are fundamentally important to the biology of scleractinian corals, as well as to a variety of other marine organisms. The genus Symbiodinium is genetically and functionally diverse and the taxonomic nature of the union between Symbiodinium and corals is implicated as a key trait determining the environmental tolerance of the symbiosis. Surprisingly, the question of how Symbiodinium diversity partitions within a species across spatial scales of meters to kilometers has received little attention, but is important to understanding the intrinsic biological scope of a given coral population and adaptations to the local environment. Here we address this gap by describing the Symbiodinium ITS2 sequence assemblages recovered from colonies of the reef building coral Montipora capitata sampled across Kāne'ohe Bay, Hawai'i. A total of 52 corals were sampled in a nested design of Coral Colony(Site(Region)) reflecting spatial scales of meters to kilometers. A diversity of Symbiodinium ITS2 sequences was recovered with the majority of variance partitioning at the level of the Coral Colony. To confirm this result, the Symbiodinium ITS2 sequence diversity in six M. capitata colonies were analyzed in much greater depth with 35 to 55 clones per colony. The ITS2 sequences and quantitative composition recovered from these colonies varied significantly, indicating that each coral hosted a different assemblage of Symbiodinium. The diversity of Symbiodinium ITS2 sequence assemblages retrieved from individual colonies of M. capitata here highlights the problems inherent in interpreting multi-copy and intra-genomically variable molecular markers, and serves as a context for discussing the utility and biological relevance of assigning species names based on Symbiodinium ITS2 genotyping.     

Functional significance of genetically different symbiotic algae Symbiodinium in a coral reef symbiosis

The giant sea anemone Condylactis gigantea associates with members of two clades of the dinoflagellate alga Symbiodinium, either singly or in mixed infection, as revealed by clade-specific quantitative polymerase chain reaction of large subunit ribosomal DNA. To explore the functional significance of this molecular variation, the fate of photosynthetically fixed carbon was investigated by (14)C radiotracer experiments. Symbioses with algae of clades A and B released ca. 30-40% of fixed carbon to the animal tissues. Incorporation into the lipid fraction and the low molecular weight fraction dominated by amino acids was significantly higher in symbioses with algae of clade A than of clade B, suggesting that the genetically different algae in C. gigantea are not functionally equivalent. Symbioses with mixed infections yielded intermediate values, such that this functional trait of the symbiosis can be predicted from the traits of the contributing algae. Coral and sea anemone symbioses with Symbiodinium break down at elevated temperature, a process known as 'coral bleaching'. The functional response of the C. gigantea symbiosis to heat stress varied between the algae of clades A and B, with particularly depressed incorporation of photosynthetic carbon into lipid of the clade B algae, which are more susceptible to high temperature than the algae of clade A. This study provides a first exploration of how the core symbiotic function of photosynthate transfer to the host varies with the genotype of Symbiodinium, an algal symbiont which underpins corals and, hence, coral reef ecosystems.     

Repopulation of Zooxanthellae in the Caribbean corals Montastraea annularis and M. faveolata following experimental and disease-associated bleaching.

Caribbean corals of the Montastraea annularis species complex associate with four taxa of symbiotic dinoflagellates (zooxanthellae; genus Symbiodinium) in ecologically predictable patterns. To investigate the resilience of these host-zooxanthella associations, we conducted field experiments in which we experimentally reduced the numbers of zooxanthellae (by transplanting to shallow water or by shading) and then allowed treated corals to recover. When depletion was not extreme, recovering corals generally contained the same types of zooxanthellae as they did prior to treatment. After severe depletion, however, recovering corals were always repopulated by zooxanthellae atypical for their habitat (and in some cases atypical for the coral species). These unusual zooxanthellar associations were often (but not always) established in experimentally bleached tissues even when adjacent tissues were untreated. Atypical zooxanthellae were also observed in bleached tissues of unmanipulated Montastraea with yellow-blotch disease. In colonies where unusual associations were established, the original taxa of zooxanthellae were not detected even 9 months after the end of treatment. These observations suggest that zooxanthellae in Montastraea range from fugitive opportunists and stress-tolerant generalists (Symbiodinium A and E) to narrowly adapted specialists (Symbiodinium B and C), and may undergo succession.     

New insights into the genetic diversity of zooxanthellae in Mediterranean anthozoans

Symbiotic dinoflagellates of the genus Symbiodinium, also called zooxanthellae, are found in association with a wide diversity of shallow-water anthozoans. The Symbiodinium genus includes numerous lineages, also referred to as clades or phylotypes, as well as a wide diversity of genetic sub-clades and sub-phylotypes. There are few studies characterizing the genetic diversity of zooxanthellae in Mediterranean anthozoans. In this study, we included anthozoans from the Western Mediterranean Sea and by means of internal transcriber (ITS) and large sub-unit (LSU) rRNA markers we corroborate what has been previously identified, demonstrating that phylotype “Temperate A” is very common among host Cnidaria in this basin. Our finding of fixed differences in ITS and LSU markers that correspond to different host taxa, indicate that this clade may comprise several closely-related species. Previous studies have reported the occurrence of Symbiodinium psygmophilum (formerly sub-clade B2) associated with Oculina patagonica and Cladocora caespitosa in the Eastern Mediterranean. Here, we identify this association in O. patagonica from the Western Mediterranean but not in C. caespitosa, suggesting some differences in symbiotic combinations between the Western and Eastern Mediterranean Basins.     

Isolation and characterization of microsatellite loci in Symbiodinium B1/B184, the dinoflagellate symbiont of the Caribbean sea fan coral, Gorgonia ventalina

Here we report primers targeting 10 microsatellite loci of dinoflagellates in the genus Symbiodinium (clade B1/B184) symbiotic with the Caribbean sea fan coral, Gorgonia ventalina. Primers were tested on 12 Symbiodinium B1/B184 cultures, as well as 40 genomic DNA extracts of G. ventalina tissue samples. All loci were polymorphic with allelic richness ranging from 4-16. Gene diversity ranged from 0.15 to 0.91. These primers provide powerful tools for examining the fine-scale population structure and dynamics of Symbiodinium within a single host species.     

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.     

Azooxanthellate? Most Hawaiian black corals contain Symbiodinium

The ecological success of shallow-water reef-building corals (Hexacorallia: Scleractinia) is framed by their intimate endosymbiosis with photosynthetic dinoflagellates in the genus Symbiodinium (zooxanthellae). In contrast, the closely related black corals (Hexacorallia: Anthipatharia) are described as azooxanthellate (lacking Symbiodinium), a trait thought to reflect their preference for low-light environments that do not support photosynthesis. We examined 14 antipatharian species collected between 10 and 396 m from Hawai'i and Johnston Atoll for the presence of Symbiodinium using molecular typing and histology. Symbiodinium internal transcribed spacer-2 (ITS-2) region sequences were retrieved from 43 per cent of the antipatharian samples and 71 per cent of the examined species, and across the entire depth range. The ITS-2 sequences were identical or very similar to those commonly found in shallow-water scleractinian corals throughout the Pacific. Histological analyses revealed low densities of Symbiodinium cells inside antipatharian gastrodermal tissues (0-92 cells mm(-3)), suggesting that the Symbiodinium are endosymbiotic. These findings confirm that the capacity to engage in endosymbiosis with Symbiodinium is evolutionarily conserved across the cnidarian subclass Hexacorallia, and that antipatharians associate with Symbiodinium types found in shallow-water scleractinians. This study represents the deepest record for Symbiodinium to date, and suggests that some members of this dinoflagellate genus have extremely diverse habitat preferences and broad environmental ranges.