Nina Yasuda

Gene flow of Acanthaster planci (L.) in relation to ocean currents revealed by microsatellite analysis

Population outbreaks of the coral‐eating starfish, Acanthaster planci, are hypothesized to spread to many localities in the Indo‐Pacific Ocean through dispersal of planktonic larvae. To elucidate the gene flow of A. planci across the Indo‐Pacific in relation to ocean currents and to test the larval dispersal hypothesis, the genetic structure among 23 samples over the Indo‐Pacific was analysed using seven highly polymorphic microsatellite loci. The F‐statistics and genetic admixture analysis detected genetically distinct groups in accordance with ocean current systems, that is, the Southeast African group (Kenya and Mayotte), the Northwestern Pacific group (the Philippines and Japan), Palau, the North Central Pacific group (Majuro and Pohnpei), the Great Barrier Reef, Fiji, and French Polynesia, with a large genetic break between the Indian and Pacific Oceans. A pattern of significant isolation by distance was observed among all samples (P = 0.001, r = 0.88, n = 253, Mantel test), indicating restricted gene flow among the samples in accordance with geographical distances. The data also indicated strong gene flow within the Southeast African, Northwestern Pacific, and Great Barrier Reef groups. These results suggest that the western boundary currents have strong influence on gene flow of this species and may trigger secondary outbreaks.

Complete mitochondrial genome sequences for Crown-of-thorns starfish Acanthaster planci and Acanthaster brevispinus

BackgroundThe crown-of-thorns starfish, Acanthaster planci (L.), has been blamed for coral mortality in a large number of coral reef systems situated in the Indo-Pacific region. Because of its high fecundity and the long duration of the pelagic larval stage, the mechanism of outbreaks may be related to its meta-population dynamics, which should be examined by larval sampling and population genetic analysis. However, A. planci larvae have undistinguished morphological features compared with other asteroid larvae, hence it has been difficult to discriminate A. planci larvae in plankton samples without species-specific markers. Also, no tools are available to reveal the dispersal pathway of A. planci larvae. Therefore the development of highly polymorphic genetic markers has the potential to overcome these difficulties. To obtain genomic information for these purposes, the complete nucleotide sequences of the mitochondrial genome of A. planci and its putative sibling species, A. brevispinus were determined and their characteristics discussed.ResultsThe complete mtDNA of A. planci and A. brevispinus are 16,234 bp and 16,254 bp in size, respectively. These values fall within the length variation range reported for other metazoan mitochondrial genomes. They contain 13 proteins, 2 rRNA, and 22 tRNA genes and the putative control region in the same order as the asteroid, Asterina pectinifera. The A + T contents of A. planci and A. brevispinus on their L strands that encode the majority of protein-coding genes are 56.3% and 56.4% respectively and are lower than that of A. pectinifera (61.2%). The percent similarity of nucleotide sequences between A. planci and A. brevispinus is found to be highest in the CO2 and CO3 regions (both 90.6%) and lowest in ND2 gene (84.2%) among the 13 protein-coding genes. In the deduced putative amino acid sequences, CO1 is highly conserved (99.2%), and ATP8 apparently evolves faster any of the other protein-coding gene (85.2%).ConclusionThe gene arrangement, base composition, codon usage and tRNA structure of A. planci are similar to those of A. brevispinus. However, there are significant variations between A. planci and A. brevispinus. Complete mtDNA sequences are useful for the study of phylogeny, larval detection and population genetics.

Genetic uniformity and long-distance clonal dispersal in the invasive androgenetic Corbicula clams

The clam genus Corbicula is an interesting model system to study the evolution of reproductive modes as it includes both sexual and asexual (androgenetic) lineages. While the sexual populations are restricted to the native Asian areas, the androgenetic lineages are widely distributed being also found in America and Europe where they form a major aquatic invasive pest. We investigated the genetic diversity of native and invasive Corbicula populations through a worldwide sampling. The use of mitochondrial and nuclear (microsatellite) markers revealed an extremely low diversity in the invasive populations with only four, undiversified, genetic lineages distributed across Europe and America. On the contrary, in the native populations, both sexual and androgenetic lineages exhibited much higher genetic diversity. Remarkably, the most abundant and widely distributed invasive forms, the so‐called form A and form R found in America and Europe respectively, are fixed for the same single COI (cytochrome c oxydase subunit I) haplotype and same multilocus genotype. This suggests that form R, observed in Europe since the 1980s, derived directly from form A found in America since the 1920s. In addition, this form shares alleles with some Japanese populations, indicating a Japanese origin for this invasive lineage. Finally, our study suggests that few androgenetic Corbicula individuals successfully invaded the non‐native range and then dispersed clonally. This is one striking case of genetic paradox raising the issue of invasive and evolutionary success of genetically undiversified populations.

Genetic diversity, paraphyly and incomplete lineage sorting of mtDNA, ITS2 and microsatellite flanking region in closely related Heliopora species (Octocorallia).

Examining genetic diversity and lineage sorting of different genes in closely related species provide useful information for phylogenetic analyses and ultimately for understanding the origins of biodiversity. In this study, we examined inter- and intraspecific genetic variation in internal transcribed spacer 2 (ITS2), partial mitochondrial gene (mtMutS), and nuclear microsatellite flanking region in two closely related octocoral species (Heliopora coerulea, HC-A and HC-B). These species were recently identified in a population genetic study using microsatellite markers. The two species have different reproductive timing, which ecologically promotes lineage sorting. In this study, we examined whether species boundaries could be detected by the commonly used nuclear ITS2 and mtMutS, as well as by possibly neutral microsatellite flanking sequences. Haplotype network analysis of microsatellite flanking region revealed that a possible ancestral haplotype was still shared between the two species, indicating on-going lineage sorting. Haplotype network analysis of ITS2 and microsatellite flanking region revealed shared haplotypes between the two lineages. The two species shared fewer ITS2 sequences than microsatellite flanking region sequences. The almost fixed point mutation at the tip of helix 3 of ITS2 was not associated with the secondary structure or compensatory base changes (CBCs). The phylogenetic tree of ITS2 showed paraphyly and that of the microsatellite flanking region indicated that lineage sorting for the two species may be incomplete. Much higher intra- and inter-individual variation of ITS2 was observed in HC-B than that in HC-A, highlighting the importance of examining ITS2 from multiple individuals to estimate genetic diversity. The mitochondrial mtMutS gene sequences from 39 individuals, including both species collected from Japan and Taiwan, showed no variation because of slow rates of mitochondrial nucleotide substitution. This study suggests caution is warranted when reciprocal monophyly in a phylogenetic tree is used as the criterion for delimiting closely related octocoral species based on ITS2 or mtMtuS sequences. Detection of boundaries between closely related species requires multi-locus analysis, such as genetic admixture analysis using multiple individuals.

First report of the development of microsatellite markers for a tropical sea cucumber (Stichopus chloronotus)

We isolated new 10 polymorphic microsatellites from the tropical sea cucumber Stichopus chloronotus. These loci provide one class of variable genetic marker as the number of alleles ranged from 2 to 6 and the observed and expected heterozygosity ranged from 0.083 to 1.0 and from 0.081 to 0.724, respectively. We consider that these loci are potentially useful for revealing clones (resulting from asexual repro) and then participate in detailing the genetic structure and gene flow among S. chloronotus populations.

Identification of Genes for Synthesis of the Blue Pigment, Biliverdin IXα, in the Blue Coral Heliopora coerulea

Heliopora coerulea is the only species in the subclass Octocorallia that has a crystalline aragonite skeleton. The skeleton has been reported to contain the blue pigment, biliverdin IXα, which is formed by heme oxygenase (HO) during heme decomposition. There is little information regarding gene expression in H. coerulea; therefore, the biosynthesis pathway for biliverdin IXα is poorly understood. To identify the genes related to heme synthesis and degradation, metatranscripts of H. coerulea and its symbiont Symbiodinium spp. were sequenced and separated from the host- and symbiont-derived sequences. From the metatranscriptome analyses, all genes for heme synthesis and three HOs were isolated from the host and symbiont. From our phylogenetic and amino acid analysis, we noted that one of the HO isoforms in the host coral was predicted to possess HO activity. However, biliverdin reductase, which reduces biliverdin to bilirubin, was not identified in the present study. Similarly, biliverdin reductase was not identified in the transcripts of the red coral Corallium rubrum, a species that also belongs to Octocorallia. However, genes related to heme synthesis and HO were found in C. rubrum. We speculate that Heliopora coerulea can produce biliverdin and accumulate it in the skeleton, while red corals and other Octocorallia species cannot. Further information from molecular studies of H. coerulea will provide insights into the synthesis of biliverdin IXα, the blue pigment in the hard crystalline aragonite skeleton, and will be fundamental to future ecological and physiological studies.

Large-scale mono-clonal structure in the north peripheral population of blue coral, Heliopora coerulea.

We examined the genotypic diversity of the large population of Heliopora coerulea, discovered recently in Ooura Bay, northern Okinawa Main Island Japan, together with another large population in Shiraho Reef, also in southwest Japan, using 9 polymorphic microsatellite markers. From each population, 40 samples were systematically collected along 2 transect lines with 4-m intervals. Surprisingly, all 40 samples from Ooura Bay were mono-genotypic, implying that the huge coral structure (30 m×80 m) originated from a single larva. Conversely, the 40 samples collected from the Shiraho Reef site all had different genotypes; measurements of genetic diversity, H(E) and H(O), were 0.075-0.975 and 0.064-0.655, respectively. At least four factors are considered to make such a huge H. coerulea population with a single genotype in Ooura Bay, (1) origin of single larva or fragment (2) a genetic bottleneck, (3) post settlement selection, and (4) frequent asexual propagation.

Development of 10 novel polymorphic microsatellite markers for the Indo-Pacific horned starfish, Protoreaster nodosus

Ten novel polymorphic nuclear microsatellite simple sequence repeat (SSR) markers were isolated from an Indo-Pacific horned starfish, Protoreaster nodosus. The isolated SSR markers provided polymorphisms of 2-9 alleles per locus in three populations obtained from Indonesia and the Philippines. The expected and observed heterozygosities ranged from 0.049 to 0.691 and from 0.050 to 0.800, respectively. Pairwise FST values among three populations ranged from 0.018 to 0.050 (global FST=0.031). All P. nodosus individuals collected from three populations exhibited different genotypes for the ten identified SSR markers, indicating that P. nodosus reproduces sexually. Then the developed SSR markers will be useful for studying the population and conservation genetics of P. nodosus.

Development of microsatellite loci in the common reef starfish Linckia laevigata and Linckia multifora

We isolated seven polymorphic microsatellite loci from the coral reef starfish Linckia laevigata—a common coral reef sea star distributed widely in the Indo-Pacific. The number of alleles ranged from 4 to 13, with an average of 7.4. The ranges of observed and expected heterozygosities were 0.367/0.933 and 0.332/0.840 for L. laevigata and 0.182/0.818 and 0.329/0.697 for L. multifora, respectively. All of these markers can also be used in the sister species Linckia multifora. These microsatellite loci will be useful for population and conservation genetic studies in coral reef ecosystems.

Integrated evidence reveals a new species in the ancient blue coral genus Heliopora (Octocorallia)

Maintaining the accretion potential and three dimensional structure of coral reefs is a priority but reef-building scleractinian corals are highly threatened and retreating. Hence future reefs are predicted to be dominated by non-constructional taxa. Since the Late Triassic however, other non-scleractinian anthozoans such as Heliopora have contributed to tropical and subtropical reef-building. Heliopora is an ancient and highly conserved reef building octocoral genus within the monospecific Family Helioporidae, represented by a single extant species – H. coerulea, Pallas, 1766. Here we show integrated morphological, genomic and reproductive evidence to substantiate the existence of a second species within the genus Heliopora. Importantly, some individuals of the new species herein described as Heliopora hiberniana sp. nov. feature a white skeleton indicating that the most diagnostic and conserved Heliopora character (the blue skeleton) can be displaced. The new species is currently known only from offshore areas in north Western Australia, which is a part of the world where coral bleaching events have severely impacted the scleractinian community over the last two decades. Field observations indicate individuals of both H. coerulea and H. hiberniana sp. nov. were intact after the 2016 Scott Reef thermal stress event, and we discuss the possibility that bleaching resistant non-scleractinian reef builders such as Heliopora could provide new ecological opportunities for the reconfiguration of future reefs by filling empty niches and functional roles left open by the regression of scleractinian corals.