Sympatric populations of the highly cross-fertile coral species Acropora hyacinthus and Acropora cytherea are genetically distinct

High cross-fertilization rates in vitro and non-monophyletic patterns in molecular phylogenies challenge the taxonomic status of species in the coral genus Acropora. We present data from eight polymorphic allozyme loci that indicate small, but significant, differentiation between sympatric populations of Acropora cytherea and Acropora hyacinthus (F(ST) = 0.025-0.068, p < 0.05), a pair of acroporid corals with very high interspecific fertilization rates in vitro. Although no fixed allelic differences were found between these species, the absence of genetic differentiation between widely allopatric populations suggests that allele frequency differences between A. cytherea and A. hyacinthus in sympatry are biologically significant. By contrast, populations of Acropora tenuis, a species which spawns 2-3 hours earlier and shows low cross-fertilization rates with congeners in vitro, were clearly distinct from A. cytherea and A. hyacinthus (F(ST) = 0.427-0.465, p < 0.05). Moreover, allopatric populations of A. tenuis differed significantly, possibly as a consequence of its relatively short period of larval competency. Our results effectively rule out the possibility that A. hyacinthus and A. cytherea are morphotypes within a single species, and indicate that hybridization occurs relatively infrequently between these taxa in nature.     

Spawning times,reproductive compatibilities and genetic structuring in the Acropora aspera group: evidence for natural hybridization and semi-permeable species boundaries in corals

Species boundaries among five sympatric coral species of the Indo-Pacific Acropora aspera group were examined by a combination of in vitro breeding trials, comparisons of spawning times and DNA sequence analysis of ribosomal DNA internal transcribed spacer (rDNA ITS) and 5.8S regions. The breeding trials showed that reproductive compatibility exists between at least some colonies of all the species pairs tested, suggesting a large potential for natural hybridization and introgression. The Acropora ITS regions exhibited extremely high levels of variability (up to approximately 62% for ITS1, approximately 11% for 5.8S and approximately 43% for ITS2), but most of the variation was shared among four of the five species, A. millepora, A. papillare, A. pulchra and A. spathulata, consistent with extensive introgression. Phylogenetic analyses did not resolve these four species as distinct clusters across a wide biogeographic region stretching from the southern Great Barrier Reef to Papua New Guinea. However, most colonies of the fifth species, A. aspera, constituted a distinct clade in phylogenetic analyses. This is consistent with our observations of a semi-permeable temporal barrier involving differences in spawning times between this and the other four species. Although the majority of colonies of all five species generally spawned within 90 min of each other, in two out of four years, gametes were absent prior to mass spawning episodes from at least some A. aspera colonies. Hence, our data suggest that transient reproductive barriers may be the result of year-to-year variation in the date of spawning and that this difference in spawning time contributes to the genetic structure detected among Acropora species in this group. Occasional leakage through the reproductive barrier was confirmed by the observation of A. aspera xA. pulchra F1 hybrids, identified based on additivity of ITS sequences.     

Molecular evolution and phylogenetic implications of internal transcribed spacer sequences of Ribosomal DNA in Winteraceae

The internal transcribed spacers and the 5.8S coding region of nuclear ribosomal DNA were sequenced and analyzed to address questions of generic relationships in Winteraceae. The molecular data generated a single tree that is congruent with one based on morphological data. The sequences of ITS 1 in the family range from 235 to 252 bases in size and of ITS 2 from 213 to 226 bases. The size of the 5.8S coding region is 164 bases. The range of ITS 1 and ITS 2 sequence divergence between pairs of genera within Winteraceae is relatively low in comparison to other plant families. Two types of ITS 1 and ITS 2 sequences were observed in the same individual for some taxa. Sequence variations between the two arrays are 4.7%–6.3% for ITS 1 and 5.1%–7.0% for ITS 2. Both arrays of sequences, however, generate the same phylogenetic relationships. Rates of nucleotide substitutions for the internal transcribed spacers are 3.2–5.2 × 10^-10 substitution per site per year estimated in ITS 1 and 3.6–5.7 × 10^-10 in ITS 2.     

Presence of a short repeat sequence in internal transcribed spacer (ITS) 1 of the rRNA gene of <Emphasis Type="Italic">Sogatella furcifera</Emphasis> (Hemiptera: Delphacidae) from geographically different populations in Asia

Nucleotide sequences of the internal transcribed spacer 1 (ITS1)–5.8S–ITS2 region of the nuclear ribosomal RNA gene were determined in the white-backed planthopper (WBPH) Sogatella furcifera (Horváth) to detect molecular variation among regional populations in Asia. We analyzed 932 sequences from 172 individuals (4–9 clones per individual) of 33 populations collected in 1987–2008 from six countries, Japan, China, Taiwan, Vietnam, Philippines, and Papua New Guinea. WBPH showed intra-individual variation in ITS1, which is mainly attributable to the frequency (0–10) of the 66-bp repeat sequence in ITS1. Among the examined clones, the sequences of 5.8S were mostly identical and those of ITS2 were similar. A single planthopper had a maximum of 6 different variants in the number of ITS1 repeats, suggesting highly varied repeat numbers in individual planthoppers. The ITS1 with four repeats was the most frequently (64%) detected. Such a repeat was not observed in two other economically important planthopper species, Nilaparvata lugens (Stål) and Laodelphax striatellus (Fallén). The ITS nucleotide sequences in the WBPH populations in Asia were genetically close and some variations in the sequences were not related to regional populations, indicating that the nucleotide sequences of the ITS region are not useful for geographical discrimination of the WBPH. This closeness seems to be caused by long distance migration and genetic exchange among populations.     

The ecology of 'Acroporid white syndrome', a coral disease from the southern Great Barrier Reef

Outbreaks of coral disease have increased worldwide over the last few decades. Despite this, remarkably little is known about the ecology of disease in the Indo-Pacific Region. Here we report the spatiotemporal dynamics of a coral disease termed 'Acroporid white syndrome' observed to affect tabular corals of the genus Acropora on the southern Great Barrier Reef. The syndrome is characterised by rapid tissue loss initiating in the basal margins of colonies, and manifests as a distinct lesion boundary between apparently healthy tissue and exposed white skeleton. Surveys of eight sites around Heron Reef in 2004 revealed a mean prevalence of 8.1±0.9%, affecting the three common species (Acropora cytherea, A. hyacinthus, A. clathrata) and nine other tabular Acropora spp. While all sizes of colonies were affected, white syndrome disproportionately affected larger colonies of tabular Acroporids (>80 cm). The prevalence of white syndrome was strongly related to the abundance of tabular Acroporids within transects, yet the incidence of the syndrome appears unaffected by proximity to other colonies, suggesting that while white syndrome is density dependant, it does not exhibit a strongly aggregated spatial pattern consistent with previous coral disease outbreaks. Acroporid white syndrome was not transmitted by either direct contact in the field or by mucus in aquaria experiments. Monitoring of affected colonies revealed highly variable rates of tissue loss ranging from 0 to 1146 cm(-2) week(-1), amongst the highest documented for a coral disease. Contrary to previous links between temperature and coral disease, rates of tissue loss in affected colonies increased threefold during the winter months. Given the lack of spatial pattern and non-infectious nature of Acroporid white syndrome, further studies are needed to determine causal factors and longer-term implications of disease outbreaks on the Great Barrier Reef.     

Variation of nuclear and mitochondrial DNAs in Korean and Chinese isolates of Clonorchis sinensis

We compared the DNA sequence difference of isolates of Clonorchis sinensis from one Korean (Kimhae) and two Chinese areas (Guangxi and Shenyang). The sequences of nuclear rDNA (18S, internal transcribed spacer 1 and 2: ITS1 and ITS2) and mitochondrial DNA (cytochrome c oxidase subunit 1: cox1) were compared. A very few intraspecific nucleotide substitution of the 18S, ITS1, ITS2 and cox1 was found among three isolates of C. sinensis and a few nucleotide insertion and deletion of ITS1 were detected. The 18S, ITS1, ITS2 and cox1 sequences were highly conserved among three isolates. These findings indicated that the Korean and two Chinese isolates are similar at the DNA sequence level.     

Intra-individual internal transcribed spacer 1 (ITS1) and ITS2 ribosomal sequence variation linked with multiple rDNA loci: A case of triploid Atractolytocestus huronensis, the monozoic cestode of common carp

Complete sequences of the ribosomal internal transcribed spacers (ITS1 and ITS2) and karyological characters of the monozoic (unsegmented) tapeworm Atractolytocestus huronensis Anthony, 1958 (Cestoda: Caryophyllidea) from Slovakia were analysed, revealing considerable intra-genomic variability and triploidy in all analysed specimens. Analysis of 20 sequences of each ITS1 and ITS2 spacer yielded eight and 10 different sequence types, respectively. In individual tapeworms, two to four ITS1 and three to four ITS2 sequence types were found. Divergent intra-genomic ITS copies were mostly induced by nucleotide substitutions and different numbers of short repetitive motifs within the sequence. In addition, triploidy was found to be a common feature of A. huronensis. The karyotype of Slovakian A. huronensis possesses three sets of chromosomes (3n = 24, n = 4m + 3st + 1 minute chromosome), similar to the previously described triploidy in conspecific tapeworms from North America. Fluorescent in situ hybridisation (FISH) with a ssrDNA probe revealed two distinct rDNA clusters for each homologue of the triplet number 2. To date, A. huronensis is the only cestode species in which intra-individual ITS sequence variants were found in parallel with its triploid nature and multiple rDNA loci. Some of these molecular and genetic features were observed in several other species of basal or nearly basal tapeworms of the orders Caryophyllidea and Diphyllobothriidea, which indicates that the phenomena may be characteristic for evolutionarily lower tapeworms and deserve more attention in future studies.     

A PCR based SNPs marker for specific characterization of English walnut (<Emphasis Type="Italic">Juglans regia</Emphasis> L.) cultivars

English walnut (Juglans regia L.) is the most economically important species from all the 21 species belonging to the genus Juglans and is an important and healthy food as well as base material for timber industry. The aim of this study was to develop a simple technique for specific characterization of English walnut using DNA method. The first and second internal transcribed spacers (ITS1 and ITS2) as well as the intervening 5.8S coding region of the rRNA gene for 18 cultivars of J. regia L. isolated from different geographic origins were characterized. The size of the spacers sequences ranged from 257 to 263 bases for ITS1 and from 217 to 219 bases for ITS2. Variation of GC contents has also been observed and scored as 55–56.7 and 57.1–58.9% for ITS1 and ITS2, respectively. This data exhibited the presence of polymorphism among cultivars. Alignment of the ITS1-5.8S-ITS2 sequences from 18 walnut cultivars showed that there were 244 single nucleotide polymorphisms (SNPs) and 1 short insertion–deletion (indel) at 5′ end ITS1. Amplification refractory mutation system strategy was successfully applied to the SNP markers of the ITS1 and ITS2 sequences for the fingerprinting analysis of 17 on 18 walnut cultivars. The prediction of ITS1 and ITS2 RNA secondary structure from each cultivar was improved by detecting key functional elements shared by all sequences in the alignments. Phylogenetic analysis of the ITS1-5.8S-ITS2 region clearly separated the isolated sequences into two clusters. The results showed that ITS1 and ITS2 region could be used to discriminate these walnut cultivars.     

The highly cross-fertile coral species,Acropora hyacinthus and Acropora cytherea,constitute statistically distinguishable lineages

A major challenge for understanding the evolutionary genetics of mass-spawning corals is to explain the maintenance of discrete morphospecies in view of high rates of interspecific fertilization in vitro and nonmonophyletic patterns in molecular phylogenies. In this study, we focused on Acropora cytherea and A. hyacinthus, which have one of the highest potentials for interspecific fertilization. Using sequences of a nuclear intron, we performed phylogenetic and nested clade analyses (NCA). Both species were polyphyletic in molecular phylogenies, but the NCA indicated that they constitute statistically distinguishable lineages. Phylogenetic analysis using an intergenic region of the mitochondrial DNA (mtDNA), was inconclusive because of low levels of variability in this marker. The position of these two species differed between the nuclear DNA (nDNA) and mtDNA phylogenies and was also at odds with a cladistic analysis based on morphology. We conclude that despite the potential for high levels of hybridization and introgression, A. cytherea and A. hyacinthus constitute statistically distinguishable lineages and their taxonomic status is consistent with the cohesion species concept.     

Genetic differentiation in the African malaria vector, Anopheles gambiae s.s., and the problem of taxonomic status

Of the seven recognized species of the Anopheles gambiae complex, A. gambiae s.s. is the most widespread and most important vector of malaria. It is becoming clear that, in parts of West Africa, this nominal species is not a single panmictic unit. We found that the internal transcribed spacer (ITS) of the X-linked rDNA has two distinct sequences with three fixed nucleotide differences; we detected no heterozygotes at these three sites, even in areas of sympatry of the two ITS types. The intergenic spacer (IGS) of this region also displays two distinct sequences that are in almost complete linkage disequilibrium with the distinct ITS alleles. We have designated these two types as S/type I and M/type II. These rDNA types correspond at least partly to the previously recognized chromosomal forms. Here we expand the geographic range of sampling to 251 individuals from 38 populations. Outside of West Africa, a single rDNA type, S/type I, corresponds to the Savanna chromosomal form. In West Africa, both types are often found in a single local sample. To understand if these findings might be due to unusual behavior of the rDNA region, we sequenced the same region for 46 A. arabiensis, a sympatric sibling species. No such distinct discontinuity was observed for this species. Autosomal inversions in one chromosome arm (2R), an insecticide resistance gene on 2L, and this single X-linked region indicate at least two genetically differentiated subpopulations of A. gambiae. Yet, rather extensive studies of other regions of the genome have failed to reveal genetic discontinuity. Evidently, incomplete genetic isolation exists within this single nominal species.     

Variation in the ribosomal internal transcribed spacers (ITS1 and ITS2) among eight taxa of the Mimulus guttatus species complex

The internal transcribed spacer region (ITS1 and ITS2) of the 18S-25S nuclear ribosomal DNA sequence and the intervening 5.8S region were sequenced from three individuals in each of eight taxa of the Mimulus guttatus species complex. Three discrete variants, or "types," of ITS sequences were found, among which 30%-40% of sites differed, compared with 1%-2% within types. Dot plots indicate that these types were not related by conspicuous rearrangements or inversions. More than one ITS type was often found in the same taxon, and two of three ITS types span species boundaries, indicating their presence prior to speciation. These ITS sequences showed essentially no positional homology with the nearest sequenced relative, tomato. In contrast, the 5.8S region was relatively unvaried, with 8 of 162 sites varied in the sample among all eight taxa. The phylogeny inferred by the most common ITS sequence type, rooted by the two other ITS types, agreed with isozymes in showing the distinctness of M. nudatus, M. laciniatus, and M. tilingii from the other five taxa.      

Intraspecific invariability of the internal transcribed spacer region of rDNA of the truffleTerfezia terfezioides in Europe

ITS regions (internal transcribed spacers—ITS1 andITS2—with the 5.8S gene of the nuclear rDNA) of 25 fruit body samples ofTerfezia terfezioides, originating from Hungary and Italy, were compared. The amplification and sequencing of the ITS region was successful with both theITS1-ITS4 andITSIF-ITS4 primer pairs. No differences of the restriction fragment length polymorphism profiles were detected among 19 samples collected in one place at the same time. The sequences of the ITS region of 9 samples collected in different localities were highly invariable, differing in only two bases. Thus the intraspecific homogeneity of the ITS region seems to be an important species-specific characteristic ofT. terfezioides in contrast to otherTerfezia species. As the samples of the species were collected from different and distant localities of Europe, the ITS sequence ofT. terfezioides can be considered a very conservative, reliable molecular marker of the fungus. *** DIRECT SUPPORT *** A00EN076 00008     

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.     

Genetic diversity of nuclear ITS1-5.8S-ITS2 rDNA sequence in Clonorchis sinensis Cobbold, 1875 (Trematoda: Opisthorchidae) from the Russian Far East

The present study examined the molecular organisation and sequence variation in the nuclear ribosomal DNA (rDNA) region, including the two internal transcribed spacers (ITS1 and ITS2) and the 5.8S gene of the Clonorchis sinensis from the Russian Far East. The relevant sequences from other parts of this species' area were downloaded from GenBank. The results showed 100% identity for all investigated 5.8S-ITS2 rDNA sequences. In contrast, two levels of intraspecific variations were revealed in the complete ITS1 sequences. The intra-genomic variation resulted from a C/T polymorphism in a single position. The inter-individual differences between the ITS1 sequences were both due to nucleotide and size polymorphisms resulting from a varying number of five-nucleotide repeats and followed by two ITS1 length variants. These variant frequencies correlate with the clonorchiasis level in some geographical localities. ITS1 differences, both in the mutation profile and mutation localisation, were revealed between northern and southern geographical samples. The presence of GC boxes that are identical to known regulatory motifs in eukaryotes was detected within the ITS1 sub-repeats. The predicted secondary structures for ITS1 consist of two large branches, one of which was invariable, while another depended on ITS1 length. The predicted secondary structure for ITS2 includes four helices around the core. The main differences between C. sinensis and other opisthorchids were localised on the tops of helices 2, 3, and 4. A phylogenetic MST reconstruction subdivided all ITS1 sequences into two well differentiated clusters, each with the major widespread ribotype, and showed that ribotype diversity in both Russia and Korea is much lower than in China. The results obtained demonstrate the feasibility of complete ITS1 sequences in C. sinensis population genetics and can be considered as a basis for further studies of the parasite infection because they may help to elucidate the molecular mechanisms of pathogen evolution and adaptation.     

Insertions or Deletions (Indels) in the rrn 16S-23S rRNA Gene Internal Transcribed Spacer Region (ITS) Compromise the Typing and Identification of Strains within the Acinetobacter calcoaceticus-baumannii (Acb) Complex and Closely Related Members

To determine whether ITS sequences in the rrn operon are suitable for identifying individual Acinetobacter Acb complex members, we analysed length and sequence differences between multiple ITS copies within the genomes of individual strains. Length differences in ITS reported previously between A. nosocomialis BCRC15417^T (615 bp) and other strains (607 bp) can be explained by presence of an insertion (indel 13i/1) in the longer ITS variant. The same Indel 13i/1 was also found in ITS sequences of ten strains of A. calcoaceticus, all 639 bp long, and the 628 bp ITS of Acinetobacter strain BENAB127. Four additional indels (13i/2–13i/5) were detected in Acinetobacter strain c/t13TU 10090 ITS length variants (608, 609, 620, 621 and 630 bp). These ITS variants appear to have resulted from horizontal gene transfer involving other Acinetobacter species or in some cases unrelated bacteria. Although some ITS copies in strain c/t13TU 10090 are of the same length (620 bp) as those in Acinetobacter strains b/n1&3, A. pittii (10 strains), A. calcoaceticus and A. oleivorans (not currently acknowledged as an Acb member), their individual ITS sequences differ. Thus ITS length by itself can not by itself be used to identify Acb complex strains. A shared indel in ITS copies in two separate Acinetobacter species compromises the specificity of ITS targeted probes, as shown with the Aun-3 probe designed to target the ITS in A. pitti. The presence of indel 13i/5 in the ITS of Acinetobacter strain c/t13TU means it too responded positively to this probe. Thus, neither ITS sequencing nor the currently available ITS targeted probes can distinguish reliably between Acb member species.     

Internal transcribed spacer polymorphism in Pseudo-nitzschia multistriata (Bacillariophyceae) in the Gulf of Naples: recent divergence or intraspecific hybridization?

The planktonic diatom Pseudo-nitzschia multistriata is a potentially toxic species recorded during late summer-autumn in the Gulf of Naples (Tyrrhenian Sea, Italy). We analysed the genetic structure by amplifying the internal transcribed spacer (ITS-1-5.8S-ITS-2) region of the ribosomal DNA of 44 strains isolated along 2 years. Polymorphism in the ITS region was detected by direct sequencing and the PCR-products from selected strains were thus cloned to assess intra-strain ITS variability. Strains clustered into three main types: type A and B - differing by 0.6% sequence divergence - and type A/B, showing both A and B variants within the same genome. The three types showed no differences in the large subunit sequences (LSU) of the rDNA, ultrastructure of the valve, secondary structure of ITS-1 and ITS-2, ploidy level and they were sexually compatible. Based on the results of these multiple approaches, we can state that the three ITS-types belong to the same reproductive unit (or "species" sensu Mayr [(1942). Systematics and the Origin of Species. Columbia University Press, New York]). We suggest that ITS polymorphism in P. multistriata may be related to the contemporary occurrence of different but still inbreeding populations which either diverged recently or originated in different geographic areas and became sympatric in the studied area.     

Genotypic diversity and gene flow in brooding and spawning corals along the Great Barrier Reef, Australia

Marine organisms exhibit great variation in reproductive modes, larval types, and other life-history traits that may have major evolutionary consequences. We measured local and regional patterns of genetic variation in corals along Australia's Great Barrier Reef to determine the relative contributions of sexual and asexual reproduction to recruitment and to infer levels of gene flow both locally (among adjacent sites, < 5 km apart) and regionally (among reefs separated by 500-1,200 km). We selected five common brooding species (Acropora cuneata, A. palifera, Pocillopora damicornis, Seriatopora hystrix, and Stylophora pistillata) and four broadcast spawners (Acropora hyacinthus, A. cytherea, A. millepora, and A. valida), which encompassed a wide range of larval types and potential dispersal capabilities. We found substantial genotypic diversity at local scales in six of the nine species (four brooders, two spawners). For these six, each local population displayed approximately the levels of multilocus genotypic diversity (Go) expected for outcrossed sexual reproduction (mean values of Go:Ge ranged from 0.85 to 1.02), although consistent single-locus heterozygous deficits indicate that inbreeding occurs at the scale of whole reefs. The remaining three species, the brooder S. hystrix and the spawners A. valida and A. millepora displayed significantly less multilocus genotypic diversity (Go) than was expected for outcrossed sexual reproduction (Ge) within each of several sites. Acropora valida and A. millepora showed evidence of extensive localized asexual replication: (1) a small number of multilocus (clonal) genotypes were numerically dominant within some sites (Go:Ge values were as low as 0.17 and 0.20): (2) single-locus genotype frequencies were characterized by both excesses and deficits of heterozygotes (cf. Hardy-Weinberg expectations), and (3) significant linkage disequilibria occurred. For the brooding S. hystrix Go:Ge values were also low within each of four sites (x = 0.48). However, this result most likely reflects the highly restricted dispersal of gametes or larvae, because levels of genetic variation among sites within reefs were extremely high (FSR = 0.28). For all species, we detected considerable genetic subdivision among sites within each reef (high FSR-values), and we infer that larval dispersal is surprisingly limited (i.e., Nem among sites ranging from 0.6 to 3.3 migrants per generation), even in species that have relatively long planktonic durations. Nevertheless, our estimates of allelic variation among reefs (FRT) also imply that for all four broadcast spawning species and three of the brooders, larval dispersal is sufficient to maintain moderate to high levels of gene flow along the entire Great Barrier Reef (i.e., Nem among reefs ranged from 5 to 31). In contrast, widespread populations of S. hystrix and S. pistilata (the two remaining brooders) are relatively weakly connected (Nem among reefs was 1.4 and 2.5, respectively). We conclude that most recruitment by corals is very local, particularly in brooders, but that enough propagules are widely dispersed to ensure that both broadcast spawning and brooding species form vast effectively panmictic populations on the Great Barrier Reef.     

Symbiodinium diversity in the sea anemone Entacmaea quadricolor on the east Australian coast

The diversity of Symbiodinium spp. in Entacmaea quadricolor was analysed from five locations along ~2,100 km on the east coast of Australia using denaturing gradient gel electrophoresis (DGGE) of the internal transcribed spacer 2 region (ITS2) combined with bacterial cloning. DGGE revealed that E. quadricolor predominantly associated with six types of clade C (four of which are novel) and that most anemones harboured multiple types simultaneously. Anemones from southern locations associated with a mixed assemblage of C25 and a variant of C3. This assemblage also dominated the central location, but was absent at the northern location. At central and northern sites, two novel variants of C42(type2) and C1 were found. Anemones hosting C42(type2) also showed a low abundance of variants of C3 and C1, and E1 was found in one sample, as revealed by bacterial cloning. The occurrence of geographically distinct ITS2 types or a consortium of types might reflect a need to optimise physiological performance of the symbiosis at different latitudes.     

Molecular evidence for the hybrid origin of species in the soft coral genus Alcyonium (Cnidaria: Anthozoa: Octocorallia)

Several recent studies have suggested that hybridization may play a previously unrecognized and important role in the evolution of corals. Our observations of polymorphic and recombinant sequences in the multicopy ribosomal internal transcribed spacer (ITS) region suggested the possible hybrid origin of two European soft coral species, Alcyonium hibernicum and Bellonella bocagei. To examine this possibility further we cloned and sequenced ITS-1 from multiple individuals and populations of these two species as well as two sympatric congeners, A. coralloides and A. sp. M2. Phylogenetic analyses separated the observed sequence variants into two distinct clades. All A. coralloides sequences belonged to clade A, while A. sp. M2 had only clade B sequences. A majority of A. hibernicum individuals, however, contained both clade A and B sequences that were identical to the predominant sequence variants found in A. coralloides and A. sp. M2, respectively. This pattern of additivity suggests that A. hibernicum originated from a hybrid cross between A. coralloides and A. sp. M2, a hypothesis that is supported by its unusual mode of reproduction (meiotic parthenogenesis). The predominant sequence variant found in B. bocagei was a unique, derived clade B sequence; in addition, however, most individuals of this species also had copies of a sequence identified as a recombinant between clade A and clade B sequence types. The presence of this recombinant sequence in the B. bocagei genome suggests that this species may also be the product of past hybridization events within the clade. Reticulate evolution may explain the failure of several previous studies to resolve the phylogeny of these four species.