PHYLOGENETIC POSITION OF SYMBIODINIUM (DINOPHYCEAE) ISOLATES FROM TRIDACNIDS (BIVALVIA), CARDIIDS (BIVALVIA), A SPONGE (PORIFERA), A SOFT CORAL (ANTHOZOA), AND A FREE-LIVING STRAIN

The genus Symbiodinium is the commonly observed symbiotic dinoflagellate (zooxanthellae) that forms mutual associations with various marine invertebrates. Numerous studies have revealed that the genus is comprised of a group of diverse taxa, and information on the phylogenetic relationships among the genus’ members is increasing. In this study, small subunit (SSU) ribosomal RNA (ssrRNA) gene sequences were determined for 15 more Symbiodinium strains from 12 relatively unstudied host taxa (Indo-Pacific tridacnids, cardiids, sponge, and soft coral), 1 hitherto unreported free-living Symbiodinium strain, and 4 other Symbiodinium strains from four other host taxa (Indo-Pacific zoanthid, foraminifer, jellyfish, and mid-Pacific hard coral). Their respective phylogenetic positions were inferred, and strains that are either closely related to or distinct from previously reported Symbiodinium taxa were revealed. The cultured Symbiodinium strains isolated from individuals of six species of tridacnids and three species of cardiids all had identical ssrRNA gene sequences, are closely related to S. microadriaticum Freudenthal, and are indistinguishable from the RFLP Type A strain previously reported. However, the ssrRNA gene sequences of clam symbionts that were obtained via gene cloning were different from those of the cultured isolates and represent strains that are close to the RFLP Type C strains. The Symbiodinium-like dinoflagellate from the Indo-Pacific sponge Haliclona koremella De Laubenfels is distinct from any of the Symbiodinium taxa studied and may be similar to the symbiont previously isolated from the stony coral Montipora patula Quelch. The isolates from the soft coral Sarcophyton glaucum Quoy et Gaimard and from the zoanthid Zoanthus sp. are both very closely related to S. pilosum Trench et Blank. The free-living Symbiodinium isolate is very closely related to the symbiont isolated from the Indo-Pacific foraminifer Amphisorus hemprichii Ehrenberg, which in turn is distinct from the Red Sea strain isolated from a similar host. Theisolate from Cassiopeia sp. is different from S. microadriaticum F., the type species harbored by Cassiopeia xamachana Bigelow, and is instead very closely related to S. pulchrorum Trench isolated from a sea anemone. The symbiont from the stony coral M. verrucosa Lamarck is a sister taxon to the symbionts isolated from the foraminifera Marginopora kudakajimensis Gudmundsson and Sorites orbiculus Forskål. These data suggest that polymorphic symbioses extend from cnidarians to some bivalve, foraminifer, and jellyfish host species.     

EVALUATING THE MORPHOSPECIES CONCEPT IN THE SELENASTRACEAE (CHLOROPHYCEAE,CHLOROPHYTA)1

Isolates of the genera Monoraphidium Kom.‐Legn., Ankistrodesmus Corda and Raphidocelis Hindák emend. Marvan et al. were cultured from two areas in Minnesota and North Dakota, USA. These isolates were identified to species level (when possible), using light microscopy and standard monographs and then characterized by 18S rDNA sequence analysis. Phylogenetic analyses indicated that in some cases, 18S rDNA sequences from these isolates were very similar, but not identical to the sequences of other isolates of the same morphospecies from different parts of the world. However, some isolates that were identified as the same species actually belong to different lineages within the Selenastraceae, whereas other isolates with identical or nearly identical 18S rDNA sequences possessed rather different morphologies. Overall, our data suggest that the application of a broad morphospecies concept to the Selenastraceae has resulted in an underestimation of the species diversity of this family and probably erroneous conclusions about the distribution of species.     

CONSPECIFICITY AND INDO-PACIFIC DISTRIBUTION OF SYMBIODINIUM GENOTYPES (DINOPHYCEAE) FROM GIANT CLAMS

We previously reported the occurrence of genetically‐diverse symbiotic dinoflagellates (zooxanthellae) within and between 7 giant clam species (Tridacnidae) from the Philippines based on the algal isolates' allozyme and random amplified polymorphic DNA (RAPD) patterns. We also reported that these isolates all belong to clade A of the Symbiodinium phylogeny with identical 18S rDNA sequences. Here we extend the genetic characterization of Symbiodinium isolates from giant clams and propose that they are conspecific. We used the combined DNA sequences of the internal transcribed spacer (ITS)1, 5.8S rDNA, and ITS2 regions (rDNA‐ITS region) because the ITS1 and ITS2 regions evolve faster than 18S rDNA and have been shown to be useful in distinguishing strains of other dinoflagellates. DGGE of the most variable segment of the rDNA‐ITS region, ITS1, from clonal representatives of clades A, B, and C showed minimal intragenomic variation. The rDNA‐ITS region shows similar phylogenetic relationships between Symbiodinium isolates from symbiotic bivalves and some cnidarians as does 18S rDNA, and that there are not many different clade A species or strains among cultured zooxanthellae (CZ) from giant clams. The CZ from giant clams had virtually identical sequences, with only a single nucleotide difference in the ITS2 region separating two groups of isolates. These data suggest that there is one CZ species and perhaps two CZ strains, each CZ strain containing individuals that have diverse allozyme and RAPD genotypes. The CZ isolated from giant clams from different areas in the Philippines (21 isolates, 7 clam species), the Australian Great Barrier Reef (1 isolate, 1 clam species), Palau (8 isolates, 7 clam species), and Okinawa, Japan (1 isolate, 1 clam species) shared the same rDNA‐ITS sequences. Furthermore, analysis of fresh isolates from giant clams collected from these geographical areas shows that these bivalves also host indistinguishable clade C symbionts. These data demonstrate that conspecific Symbiodinium genotypes, particularly clade A symbionts, are distributed in giant clams throughout the Indo‐Pacific.     

SMALL-SUBUNIT RIBOSOMAL DNA SEQUENCE ANALYSES AND A RECONSTRUCTION OF THE INFERRED PHYLOGENY AMONG SYMBIOTIC DINOFLAGELLATES (PYRROPHYTA)1

The small-subunit ribosomal RNA genes (SSU rDNA) from the four symbiotic dinoflagellates, Symbiodinium corculorum Trench isolated from the bivalve mollusc Corculum cardissa (from Belau, Western Caroline Is.), S. meandrinae Trench, from the scleractinian coral Meandrina meandrites (from famaica, W.I.), Gloeodinium viscum Banaszak et al. from the hydrocoral Millepora dichotoma (from the Gulf of Aqaba), and Amphidinium belauense Trench from the acoel flatworm Haplodiscus sp. (from Belau) have been amplified by the polymerase chain reaction, cloned, and sequenced. Following alignment of these complete sequences to homologous sequences from six other dinoflagellates, eight api-complexans, six ciliates, six chromophytes and oomycetes, three ascomycetes, two rhodophytes, two chlorophytes, and two myxomycetes (with Physarum polycephalum as the outgroup), phylogenetic reconstruction was conducted using Fitch and Margoliash distance, DNA maximum likelihood, and Wagner parsimony methods, with bootstrap resampling. All methods generated trees with similar topologies. The inferred “across Kingdom” phylogeny reemphasizes previous reports that show that the dinoflagellates, the apicomplexans, and the ciliates share a common ancestry and that the dinoflagellates are distantly related to the chromophyte-oömycete lineage. The evidence supports the concept of a polyphyletic origin of dinoflagellate-invertebrate symbioses, as symbiotic dinoflagellates represent seven genera in at least four orders. The three symbiotic species, S. corculorum, S. meandrinae, and S. pilosum, consistent with their morphological and biochemical similarities, cluster most closely. Symbiodinium pulchrorum Trench, the symbiontfrom the Hawaiian sea anemone Aiptasia pulchella, is more distantly related to them. Gloeodinium viscum is not closely related to the Symbiodinium species. Amphidinium carterae (free-living) and A. belauense (symbiotic) also appear to be distantly related to Symbiodinium. Some symbionts (e.g. S. corculorum, S. pilosum) from distant geographic locations (the Indo-Pacific and Caribbean, respectively) were found to be very closely related, whereas S. pulchrorum and S. corculorum from the Pacific were found to be distantly related. Analyses of 10 additional symbiotic and nonsymbiotic dinoflagellates, using partial SSU rDNA sequences to generate a tentative dinoflagellate phylogeny, indicate that members of the genus Symbiodinium cluster with most of the other (free-living) dinoflagellates in the genus Gymnodinium. The genus Amphidinium, as represented by A. carterae and A. belauense, appear to be distantly related to the other members of the Gymnodiniaceae. This analysis, combined with morphological and biochemical data, indicates that the symbionts S. pulchrorum (from Aiptasia pulchella) and S. bermudense Trench (from Aiptasia tagetes) from the Indo-Pacific and Caribbean, respectively, are very closely related but are not identical.     

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.     

USE OF RIBOSOMAL DNA INTERNAL TRANSCRIBED SPACERS FOR PHYLOGENETIC STUDIES IN DIATOMS1

Sequence variation of ribosomal DNA internal transcribed spacers (ITS) among populations, species, and genera of the diatom genus Stephanodiscus was investigated. ITS 1 and ITS 2, including the 5.8S gene, were sequenced from geographically distant and nearby populations of S. niagarae Ehrenberg. In addition, repeats from S. hantzschii Grunow and Cyclotella meneghiniana Kützing were sequenced to determine the taxonomic range over which the ITS region could be used for diatom systematics. The morphologically distinct S. yellowstonensis Theriot & Stoermer, thought to have evolved from S. niagarae in Yellowstone Lake between 12,000 and 8000 years ago, also was sequenced to assess its relationship to nearby S. niagarae populations. The organization and relative sizes of ITS 1 and ITS 2 in Stephanodiscus species were similar to those reported for other eukaryotes. In general, ITS 2 was slightly larger and more variable than ITS 1. Cladistic analysis of ITS sequences did not resolve relationships of nearby S. niagarae and S. yellowstonensis populations. However, central North American S. niagarae populations were in a clade supported by two nucleotide changes. For Cyclotella, much of the ITS region was not alignable with that for Stephanodiscus species; therefore, generic-level comparison within the Thalassiosiraceae may not be possible. The variation (95–96% similarity) between S. hantzschii and other Stephanodiscus species suggests that interspecific relationships could be assessed with ITS sequences. Although S. yellowstonensis is morphologically distinct from S. niagarae, no autapomorphic nucleotide sites were identified. Two S. niagarae populations (Heart and Lewis Lakes), however, did possess autapomorphic ITS sites.     

Improved resolution of reef-coral endosymbiont (Symbiodinium) species diversity, ecology, and evolution through psbA non-coding region genotyping

Ribosomal DNA sequence data abounds from numerous studies on the dinoflagellate endosymbionts of corals, and yet the multi-copy nature and intragenomic variability of rRNA genes and spacers confound interpretations of symbiont diversity and ecology. Making consistent sense of extensive sequence variation in a meaningful ecological and evolutionary context would benefit from the application of additional genetic markers. Sequences of the non-coding region of the plastid psbA minicircle (psbA(ncr)) were used to independently examine symbiont genotypic and species diversity found within and between colonies of Hawaiian reef corals in the genus Montipora. A single psbA(ncr) haplotype was recovered in most samples through direct sequencing (~80-90%) and members of the same internal transcribed spacer region 2 (ITS2) type were phylogenetically differentiated from other ITS2 types by substantial psbA(ncr) sequence divergence. The repeated sequencing of bacterially-cloned fragments of psbA(ncr) from samples and clonal cultures often recovered a single numerically common haplotype accompanied by rare, highly-similar, sequence variants. When sequence artifacts of cloning and intragenomic variation are factored out, these data indicate that most colonies harbored one dominant Symbiodinium genotype. The cloning and sequencing of ITS2 DNA amplified from these same samples recovered numerically abundant variants (that are diagnostic of distinct Symbiodinium lineages), but also generated a large amount of sequences comprising PCR/cloning artifacts combined with ancestral and/or rare variants that, if incorporated into phylogenetic reconstructions, confound how small sequence differences are interpreted. Finally, psbA(ncr) sequence data from a broad sampling of Symbiodinium diversity obtained from various corals throughout the Indo-Pacific were concordant with ITS lineage membership (defined by denaturing gradient gel electrophoresis screening), yet exhibited substantially greater sequence divergence and revealed strong phylogeographic structure corresponding to major biogeographic provinces. The detailed genetic resolution provided by psbA(ncr) data brings further clarity to the ecology, evolution, and systematics of symbiotic dinoflagellates.     

ANALYSIS OF ALEXANDRIUM (DINOPHYCEAE) SPECIES USING SEQUENCES OF THE 5.8S RIBOSOMAL DNA AND INTERNAL TRANSCRIBED SPACER REGIONS1

The 5.8S ribosomal RNA gene (rDNA) and flanking internal transcribed spacers 1 and 2 (ITS1 and ITS2) from 7 isolates of Alexandrium catenella (Wedon et Kofoid) Taylor, 13 isolates of A. tamarense (Lebour) Balech, 2 isolates of A. affine (Fukuyo et Inoue) Balech, and single isolates of A. fundyense Balech, A. insuetum Balech, and A. pseudogonyaulax (Biecheler) Horiguchi ex Yuki et Fukuyo comb. nov. from Japan, Thailand, and the United States were amplified using the polymerase chain reaction (PCR), sequenced, and subjected to phylogenetic analysis. The sequences ranged from 518 to 535 base pairs (bp) exclusive of the 18S and 28S rDNA coding regions. Sequence comparisons revealed seven divergent “ITS types” designated as follows: 1) catenella type, 2) tamarense type, 3) WKS-1 type, 4) Thai type, 5) affine type, 6) insuetum type, and 7) pseudogonyaulax type. Isolates of the tamarense type from various locations in Japan and the United States and of A. fundyense from the United States were closely related to each other and were clearly divergent from isolates of A. tamarense WKS-1 (WKS-I type) or A. tamarense CU-15 (Thai type). These latter two strains carried unique ITS types, although they were not distinguishable from isolates of the tamarense type by morphological criteria. Distance values between isolates of the tamarense type and the WKS-1 or Thai type were quite high (about 0.21 and 0.39, respectively). Seven isolates of A. catenella from Japan (catenella type) clearly diverged from the other ITS types already mentioned. Distance values between isolates of the catenella type were extremely low (<0.01), whereas distance values of ITS between the catenella type and the tamarense, WKS-1, or Thai type were 0.17, 0.18, and 0.40, respectively. Isolates of A. affine, A. insuetum, and A. pseudogonyaulax all carried unique ITS types. The ITSs of the tamarense type exhibited two distinct ITS sets, the “A gene” and the “B gene.” The two sequences occurred in a 1:1 ratio in PCR products. In contrast, the ITSs of all other isolates appeared homogeneous. Sequence comparisons also showed that the variations in the 3′ end of ITS1 (150-177 bp) were low within each ITS type but extremely high between ITS types. The number of different nucleotides among the seven Alexandrium types in this 28-bp region is more than 10. High diversity of this region may facilitate the design of DNA probes specific for each ITS type/species of Alexandrium.     

Genetic characterization of Strongyloides spp. from captive, semi-captive and wild Bornean orangutans (Pongo pygmaeus) in Central and East Kalimantan, Borneo, Indonesia

Orangutans (Pongo spp.), Asia's only great apes, are threatened in their survival due to habitat loss, hunting and infections. Nematodes of the genus Strongyloides may represent a severe cause of death in wild and captive individuals. In order to better understand which Strongyloides species/subspecies infect orangutans under different conditions, larvae were isolated from fecal material collected in Indonesia from 9 captive, 2 semi-captive and 9 wild individuals, 18 captive groups of Bornean orangutans and from 1 human working with wild orangutans. Genotyping was done at the genomic rDNA locus (part of the 18S rRNA gene and internal transcribed spacer 1, ITS1) by sequencing amplicons. Thirty isolates, including the one from the human, could be identified as S. fuelleborni fuelleborni with 18S rRNA gene identities of 98·5-100%, with a corresponding published sequence. The ITS1 sequences could be determined for 17 of these isolates revealing a huge variability and 2 main clusters without obvious pattern with regard to attributes of the hosts. The ITS1 amplicons of 2 isolates were cloned and sequenced, revealing considerable variability indicative of mixed infections. One isolate from a captive individual was identified as S. stercoralis (18S rRNA) and showed 99% identity (ITS1) with S. stercoralis sequences from geographically distinct locations and host species. The findings are significant with regard to the zoonotic nature of these parasites and might contribute to the conservation of remaining orangutan populations.     

IDENTIFICATION OF ALEXANDRIUM (DINOPHYCEAE) SPECIES USING PCR AND rDNA-TARGETED PROBES

A PCR (polymerase chain reaction)-based assay for the detection of Alexandrium species in cultured samples using rDNA-targeted probes was developed. The internal transcribed spacers 1 and 2 (ITS1 and ITS2) and the 5.8S ribosomal RNA gene (rDNA) from cultured isolates of A. tamarense (Lebour) Taylor, A. catenella (Whedon et Kofoid) Balech, A. fundyense Balech and A. lusitanicum Balech were amplified using PCR and sequenced. Sequence comparisons showed that the 5.8S and ITS1-ITS2 regions contain sequences specific for the Alexandrium genus, especially at the 3' end of the 5.8S coding region. PCR primers and a radioactive ³²P-labeled DNA probe were devised for this region. The cross-reactivity of the PCR primers and probe was tested against cultured isolates of Alexandrium and other dinoflagellates and diatoms. All the Alexandrium isolates screened reacted toward the genus-specific probe; in contrast, the other groups of microalgae (dinoflagellates and diatoms) did not react with the probe. Furthermore, the PCR amplification technique combined with the use of the rDNA-target probe allowed us to develop a method for the detection of Alexandrium cells in cultured samples. This PCR method might offer a new approach for the identification and enumeration of the HAB (harmful algal bloom) species present in natural phytoplankton populations.     

The role of deep reefs in shallow reef recovery: an assessment of vertical connectivity in a brooding coral from west and east Australia

Approximately one quarter of zooxanthellate coral species have a depth distribution from shallow waters (<30 m) down to mesophotic depths of 30-60 m. The deeper populations of such species are less likely to be affected by certain environmental perturbations, including high temperature/high irradiance causing coral bleaching. This has led to the hypothesis that deep populations may serve as refuges and a source of recruits for shallow reef habitats. The extent of vertical connectivity of reef coral species, however, is largely unquantified. Using 10 coral host microsatellite loci and sequences of the host mtDNA putative control region, as well as ribosomal DNA (rDNA) ITS2 sequences of the coral's algal endosymbionts (Symbiodinium), we examine population structure, connectivity and symbiont specificity in the brooding coral Seriatopora hystrix across a depth profile in both northwest (Scott Reef) and northeast Australia (Yonge Reef). Strong genetic structuring over depth was observed in both regions based on the microsatellite loci; however, Yonge Reef exhibited an additional partitioning of mtDNA lineages (associated with specific symbiont ITS2 types), whereas Scott Reef was dominated by a single mtDNA lineage (with no apparent host-symbiont specificity). Evidence for recruitment of larvae of deep water origin into shallow habitats was found at Scott Reef, suggesting that recovery of shallow water habitats may be aided by migration from deep water refuges. Conversely, no migration from the genetically divergent deep slope populations into the shallow habitats was evident at Yonge Reef, making recovery of shallow habitats from deeper waters at this location highly unlikely.     

Effect of temperature and light on growth of and photosynthesis by Synechococcus isolates typical of those predominating in the octopus spring microbial mat community of Yellowstone National Park

Previous molecular analysis of the Octopus Spring cyanobacterial mat revealed numerous genetically distinct 16S rRNA sequences from predominant Synechococcus populations distantly related to the readily cultivated unicellular cyanobacterium Synechococcus lividus. Patterns in genotype distribution relative to temperature and light conditions suggested that the organisms contributing these 16S rRNA sequences may fill distinct ecological niches. To test this hypothesis, Synechococcus isolates were cultivated using a dilution and filtration approach and then shown to be genetically relevant to natural mat populations by comparisons of similarities of 16S rRNA genes and 16S-23S internal transcribed spacer (ITS) regions. Most isolates were identical or nearly identical at both loci to predominant mat genotypes; others showed 1- to 2-nucleotide differences at the 16S rRNA locus and even greater difference in ITS sequences. Isolates with predominant mat genotypes had distinct temperature ranges and optima for growth that were consistent with their distributions in the mat. Isolates with genotypes not previously detected or known to be predominant in the mat exhibited temperature ranges and optima that were not representative of predominant mat populations and also grew more slowly. Temperature effects on photosynthesis did not reflect temperature relations for growth. However, the isolate with the highest temperature optimum and upper limit was capable of performing photosynthesis at a higher temperature than other isolates. Growth rate and photosynthetic responses provided evidence for light acclimation but evidence of, at best, only subtle light adaptation.     

CHARACTERIZATION OF OSTREOPSIS AND COOLIA (DINOPHYCEAE) ISOLATES IN THE WESTERN MEDITERRANEAN SEA BASED ON MORPHOLOGY,TOXICITY AND INTERNAL TRANSCRIBED SPACER 5.8S rDNA SEQUENCES1

Several isolates of epiphytic dinoflagellates belonging to the genera Ostreopsis Schmidt and Coolia Meunier from the western Mediterranean Sea were examined by LM and EM, toxicity assays, and internal transcribed spacer (ITS) regions of nuclear rDNA, and 5.8S rDNA were sequenced. Morphological comparisons based on the analyses of cell shape, size, thecal plates, and surface ornamentation revealed two distinct species in the western Mediterranean: O. cf. siamensis Schmidt from the Catalan, Andalusian, and Sicilian coasts and O. ovata Fukuyo from the Ligurian coast, southern Tyrrhenian Sea, and Balearic Islands. Both Ostreopsis species were toxic; however, no differences in toxicity were detected between the two Ostreopsis species. Coolia monotis Meunier was nontoxic. The morphological studies were supported by phylogenetic analyses; all western Mediterranean isolates of O. cf. siamensis showed ITS and 5.8S rDNA sequences identical to each other and so did those of O. ovata, whereas high genetic diversity was detected between the western Mediterranean and Asian isolates of O. ovata. The nucleotide sequence analyses of the C. monotis strains showed that all C. monotis isolates from Europe formed a homogeneous clade. Further, the genetic diversity was high between the European and Asian C. monotis isolates. In this study, genetic markers combined with morphology and toxicity analyses was useful in the taxonomic and phylogenetic studies of the Ostreopsidaceae in a temperate area.     

NUCLEAR RIBOSOMAL DNA INTERNAL TRANSCRIBED SPACER REGIONS (ITS1 AND ITS2) DEFINE DISCRETE BIOGEOGRAPHIC GROUPS IN CLADOPHORA ALBIDA (CHLOROPHYTA)1

Nucleotide sequences of the nuclear ribosomal DNA internal transcribed spacers (ITS1 and ITS2), the 5.8S, and short stretches of the adjacent 18S and 26S coding regions were determined in isolates from four disjunct Cladophora albida (Huds.) Kütz. populations (NE-America, W-Europe, Japan, and W-Australia). The two Pacific isolates share nearly identical ITS sequences as do the two Atlantic isolates. In contrast, interoceanic comparisons exhibit a 21% sequence difference. Variation within ITS regions is useful for identification of population groups on a regional or oceanic scale. However, both spacers are characterized by numerous repeat motifs as well as point mutations, which result in alignment problems at the interspecific level within Cladophora.     

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.     

金耳与其近似种的rDNA-ITS序列分析

对金耳(Tremella aurantialba)的担子果、酵母状分生孢子培养物和菌丝培养物的核糖体DNA内部转录间隔区(ITS)序列进行PCR扩增和测序。结果表明金耳担子果的ITS区PCR产物均为碱基数不同的两条带,片段长度和序列与酵母状分生孢子培养物、菌丝培养物一致。通过对ITS1和ITS2联合进行系统发育分析表明金耳酵母状分生孢子培养物归属于银耳属的金耳,参与组成担子果的寄主菌丝为毛韧革菌(Stereum hirsutum)。结合GenBank中登录的金黄银耳、脑状银耳、橙黄银耳等近似种构建了的系统发育树,结果支持形态学证据,表明金耳是一个独立种。     

Additional data for a new Theileria sp. from China based on the sequences of ribosomal RNA internal transcribed spacers

Theileria sinensis was recently isolated and named as an independent Theileria species that infects cattle in China. To date, this parasite has been described based on its morphology, transmission and molecular studies, indicating that it should be classified as a distinct species. To test the validity of this taxon, the two internal transcribed spacers (ITS1 and ITS2) and the 5.8S rRNA gene were cloned and sequenced from three T. sinensis isolates. The complete ITS sequences were compared with those of other Theileria sp. available in GenBank. Phylogenetic analyses based on sequence data for the complete ITS sequences indicate that T. sinensis lies in a distinct clade that is separate from that of T. buffeli/orientalis and T. annulata. Sequence comparisons indicate that different T. sinensis isolates possess unique sizes of ITS1 and ITS2 as well as species-specific nucleotide sequences. This analysis provides new molecular data to support the classification of T. sinensis as a distinct species from other known Theileria spp. based on ITS sequences.