Molecular diversity of black corals from the Saudi Arabian Red Sea: a first assessment.

Black corals occur as part of benthic assemblages from shallow to deep waters in all oceans. Despite the importance in many benthic ecosystems, where these act as biodiversity aggregators, antipatharians remain poorly studied, with 75% of the known species occurring below recreational SCUBA diving depth limits. Currently, information regarding the diversity and evolutionary history is limited, with most studies focusing on Hawaii and the South Pacific Ocean. Other regions of the world have received less attention, such as the Red Sea, where only two black coral families and four genera have been recorded. We provide the first analysis of the molecular diversity of black corals in the eastern Gulf of Aqaba and the northern and central Saudi Arabian Red Sea, based on a dataset of 161 antipatharian colonies collected down to 627 m deep. Based on specimen morphology, we ascribed our material to 11 genera belonging to 4 of the 7 known Antipatharia families, i.e. Antipathidae, Aphanipathidae, Myriopathidae and Schizopathidae. The genus level phylogeny of three intergenic mitochondrial regions, the trnW-IGR-nad2 (IgrW ), nad5-IGR-nad1 (IgrN ) and cox3-IGR-cox1 was reconstructed including previously published material. Overall, we recovered six molecular clades that included exclusively Red Sea sequences, with the highest diversity occurring at mesophotic depths. This study highlights that diversity of black corals in the Red Sea is much higher than previously known, with seven new generic records, suggesting that this basin may be a hotspot for antipatharian diversity as is known for other taxa. Our results recovered unresolved relationships within the order at the familial and generic levels. This emphasises the urgent need for an integration of genomic-wide data with a re-examination of informative morphological features necessary to revise the systematics of the order at all taxonomic levels.

Interspecific coral competition does not affect the symbiosis of gall crabs (Decapoda: Cryptochiridae) and their scleractinian hosts.

Coral reefs accommodate a myriad of species, many of which live in association with a host organism. Decapod crustaceans make up a large part of this associated fauna on coral reefs. Among these, cryptochirid crabs are obligately associated with scleractinian corals, in which they create dwellings where they permanently reside. These gall crabs show various levels of host specificity, with the majority of cryptochirids inhabiting a specific coral genus or species. Here, we report the first records of gall crabs living in association with two different Porites species in the Red Sea. Crescent-shaped dwellings were observed in Porites rus and a Porites sp. in situ, and colonies with crabs were collected for further study in the laboratory. Using a combination of morphology and DNA barcoding, the crabs were identified as belonging to Opecarcinus, a genus only known to inhabit Agariciidae corals. The coral skeleton was bleached and studied under a stereo microscope, which revealed that the Porites corals overgrew adjoining agariciid Pavona colonies. We hypothesize that the gall crab originally settled on Pavona, its primary host of choice. Due to coral interspecific competition the Porites colony overgrew the adjacent Pavona colonies, resulting in a secondary and never before reported association of Opecarcinus with Porites. These findings suggest that cryptochirid crabs can adapt to the new microenvironment provided by a different coral host and survive competition for space on coral reefs.

Mitochondrial Genome of Nonmodel Marine Metazoans by Next-Generation Sequencing (NGS).

Mitochondrial genomes (mtgenome) represent an important source of information for addressing fundamental evolutionary, phylogeographic, systematic, and ecological questions in marine organisms. In the last two decades the advent of high-throughput next-generation sequencing (NGS) has provided an unprecedented possibility to access large amount of genomic data and, as such, there has been a rapid growth in mtgenome resources and studies. In particular, NGS strategies represent a great advantage for investigating nonmodel marine organisms for which no or limited genomic resources are available. Here, we describe a routinely used standardized protocol to obtain mtgenome of nonmodel marine organisms by NGS. The protocol is composed of five main steps, including DNA extraction, DNA fragmentation, library preparation, high-throughput sequencing, and bioinformatic analyses. Each of the first three steps is followed by size/quality and concentration validations. The advantages of the described protocol rely on the assumption that no a priori information on mtgenome of the studied organism is needed and on its versatility as researchers may choose several kits for DNA extraction and library preparation and adopt different methods for DNA fragmentation depending on their needs, experience, and suppliers.

The complete mitochondrial genome of Dendrophyllia minuscula (Cnidaria: Scleractinia) from the NEOM region of the Northern Red Sea.

The scleractinian coral family Dendrophylliidae is a major component of shallow and deep-water coral ecosystems worldwide, but our knowledge on the evolutionary history of the family remains scarce. Here, we used ezRAD coupled with Illumina sequencing technology and reconstructed the complete mitochondrial genome of Dendrophyllia minuscula (GenBank accession number OL634845), from mesophotic depths in the Red Sea NEOM area. The mitochondrial genome of D. minuscula consisted of 19,054 bp, organized in 13 protein-coding genes, 2 rRNA genes, and 2 tRNA genes, in agreement with the Scleractinia typical mitogenome organization. This complete mitochondrial genome contributes toward a better knowledge of mesophotic and deep-water coral diversity and evolutionary history.

A new species of Bathypathes (Cnidaria, Anthozoa, Antipatharia, Schizopathidae) from the Red Sea and its phylogenetic position.

A black coral, Bathypathesthermophila Chimienti, sp. nov. is described from the Saudi Arabian coasts of the Gulf of Aqaba and north Red Sea (Neom area) using an integrated taxonomic approach. The morphological distinctiveness of the new species is confirmed by molecular analyses. The species thrives in warm and high salinity waters typical of the Red Sea at bathyal depths. It can form colony aggregations on muddy bottoms with scattered, small hard substrates. Colonies are monopodial, feather-like, and attached to a hard substrate through a thorny basal plate. Pinnules are simple, arranged biserially and alternately, and all the same length (up to approximately 20 cm) except for few, proximal ones. Spines are triangular, laterally compressed, subequal, smooth, and simple or rarely bifurcated. Polyps are elongated transversely, 1.5-2.0 mm in transverse diameter. Large colonies can have one or few branches, whose origin is discussed. The phylogenetic position of B.thermophila sp. nov. within the order Antipatharia, recovered using three mitochondrial markers, shows that it is nested within the family Schizopathidae. It is close to species in the genera Parantipathes, Lillipathes, Alternatipathes, and Umbellapathes rather than to the other available representatives of the genus Bathypathes, as currently defined based on morphology. In agreement with previous findings, our results question the evolutionary significance of morphological characters traditionally used to discriminate Antipatharia at higher taxonomic level.

Phylogenomics of Porites from the Arabian Peninsula.

The advent of high throughput sequencing technologies provides an opportunity to resolve phylogenetic relationships among closely related species. By incorporating hundreds to thousands of unlinked loci and single nucleotide polymorphisms (SNPs), phylogenomic analyses have a far greater potential to resolve species boundaries than approaches that rely on only a few markers. Scleractinian taxa have proved challenging to identify using traditional morphological approaches and many groups lack an adequate set of molecular markers to investigate their phylogenies. Here, we examine the potential of Restriction-site Associated DNA sequencing (RADseq) to investigate phylogenetic relationships and species limits within the scleractinian coral genus Porites. A total of 126 colonies were collected from 16 localities in the seas surrounding the Arabian Peninsula and ascribed to 12 nominal and two unknown species based on their morphology. Reference mapping was used to retrieve and compare nearly complete mitochondrial genomes, ribosomal DNA, and histone loci. De novo assembly and reference mapping to the P. lobata coral transcriptome were compared and used to obtain thousands of genome-wide loci and SNPs. A suite of species discovery methods (phylogenetic, ordination, and clustering analyses) and species delimitation approaches (coalescent-based, species tree, and Bayesian Factor delimitation) suggested the presence of eight molecular lineages, one of which included six morphospecies. Our phylogenomic approach provided a fully supported phylogeny of Porites from the Arabian Peninsula, suggesting the power of RADseq data to solve the species delineation problem in this speciose coral genus.

Evolutionary biogeography of the reef-building coral genus Galaxea across the Indo-Pacific ocean.

Stony corals (Scleractinia) form the basis for some of the most diverse ecosytems on Earth, but we have much to learn about their evolutionary history and systematic relationships. In order to improve our understanding of species in corals we here investigated phylogenetic relationships between morphologically defined species and genetic lineages in the genus Galaxea (Euphyllidae) using a combined phylogenomic and phylogeographic approach. Previous studies revealed the nominal species G. fascicularis included three genetically well-differentiated lineages (L, S & L+) in the western Pacific, but their distribution and relationship to other species in the genus was unknown. Based on genomic (RAD-seq) and mitochondrial sequence data (non-coding region between cytb and ND2) we investigated whether the morphological taxa represent genetically coherent entities and what is the phylogenetic relationship and spatial distribution of the three lineages of G. fascicularis throughout the observed species range. Using the RAD-seq data, we find that the genus Galaxea is monophyletic and contains three distinct clades: an Indo-Pacific, a Pacific, and a small clade restricted to the Chagos Archipelago. The three lineages of G. fascicularis were associated with different RAD-seq clades, with the 'L' lineage showing some morphological distinction from the other two lineages (larger more asymmetrical polyps). In addition to these, three more genetic lineages in G. fascicularis may be distinguished - a Chagossian, an Ogasawaran, and one from the Indian-Red Sea. Among nominal taxa for which we have multiple samples, G. horrescens was the only monophyletic species. The mitochondrial non-coding region is highly conserved apart of the length polymorphism used to define L, S & L+ lineages and lacks the power to distinguish morphological and genetic groups resolved with genomic RAD-sequencing. The polyphyletic nature of most species warrants a careful examination of the accepted taxonomy of this group with voucher collections and their comparison to type specimens to resolve species boundaries. Further insight to the speciation process in corals will require international cooperation for the sharing of specimens to facilitate scientific discovery.

Uncovering hidden coral diversity: a new cryptic lobophylliid scleractinian from the Indian Ocean.

Extant biodiversity can easily be underestimated owing to the presence of cryptic taxa, even among commonly observed species. Scleractinian corals are challenging to identify because of their ecophenotypic variation and morphological plasticity. In addition, molecular analyses have revealed the occurrence of cryptic speciation. Here, we describe a new cryptic lobophylliid genus and species Paraechinophyllia variabilis gen. nov., sp. nov., which is morphologically similar to Echinophyllia aspera and E. orpheensis. The new taxon occurs in Mayotte Island, Madagascar, the Gulf of Aden and the Red Sea. Six molecular markers (COI, 12S, ATP6-NAD4, NAD3-NAD5, histone H3 and ITS) and 46 morphological characters at three different levels (macromorphology, micromorphology and microstructure) were examined. The resulting molecular phylogenetic reconstruction showed that Paraechinophyllia gen. nov. represents a distinct group within the Lobophylliidae that diverged from the lineage leading to Echinophyllia and Oxypora in the Early Miocene, approximately 21.5 Ma. The morphological phylogenetic reconstruction clustered Paraechinophyllia gen. nov., Echinophyllia and Oxypora together in a single clade. A sole morphological character, calice relief, discriminated Paraechinophyllia gen. nov. from the latter two genera, suggesting that limited morphological variation has occurred over a long period. These results highlight the importance of cryptic taxa in reef corals, with implications for population genetics, ecological studies and conservation.

Using ezRAD to reconstruct the complete mitochondrial genome of Porites fontanesii (Cnidaria: Scleractinia).

Corals in the genus Porites are among the major framework builders of reef structures worldwide, yet the genus has been challenging to study due to a lack of informative molecular markers. Here, we used ezRAD sequencing to reconstruct the complete mitochondrial genome of Porites fontanesii (GenBank accession number MG754069), a widespread coral species endemic to the Red Sea and Gulf of Aden. The gene arrangement of P. fontanesii did not differ from other Scleractinia and consisted of 18,658 bp, organized in 13 protein-coding genes, 2 rRNA genes, and 2 tRNA genes. This mitochondrial genome contributes essential data to work towards a better understanding of evolutionary relationships within Porites.

The complete mitochondrial genome of Porites harrisoni (Cnidaria: Scleractinia) obtained using next-generation sequencing.

In this study, we sequenced the complete mitochondrial genome of Porites harrisoni using ezRAD and Illumina technology. Genome length consisted of 18,630 bp, with a base composition of 25.92% A, 13.28% T, 23.06% G, and 37.73% C. Consistent with other hard corals, P. harrisoni mitogenome was arranged in 13 protein-coding genes, 2 rRNA, and 2 tRNA genes. nad5 and cox1 contained embedded Group I Introns of 11,133 bp and 965 bp, respectively.

Recent origin and semi-permeable species boundaries in the scleractinian coral genus Stylophora from the Red Sea.

Reticulate evolution, introgressive hybridisation, and phenotypic plasticity have been documented in scleractinian corals and have challenged our ability to interpret speciation processes. Stylophora is a key model system in coral biology and physiology, but genetic analyses have revealed that cryptic lineages concealed by morphological stasis exist in the Stylophora pistillata species complex. The Red Sea represents a hotspot for Stylophora biodiversity with six morphospecies described, two of which are regionally endemic. We investigated Stylophora species boundaries from the Red Sea and the associated Symbiodinium by sequencing seven DNA loci. Stylophora morphospecies from the Red Sea were not resolved based on mitochondrial phylogenies and showed nuclear allele sharing. Low genetic differentiation, weak isolation, and strong gene flow were found among morphospecies although no signals of genetic recombination were evident among them. Stylophora mamillata harboured Symbiodinium clade C whereas the other two Stylophora morphospecies hosted either Symbiodinium clade A or C. These evolutionary patterns suggest that either gene exchange occurs through reticulate evolution or that multiple ecomorphs of a phenotypically plastic species occur in the Red Sea. The recent origin of the lineage leading to the Red Sea Stylophora may indicate an ongoing speciation driven by environmental changes and incomplete lineage sorting.

Species delimitation in the reef coral genera Echinophyllia and Oxypora (Scleractinia, Lobophylliidae) with a description of two new species.

Scleractinian corals are affected by environment-induced phenotypic plasticity and intraspecific morphological variation caused by genotype. In an effort to identify new strategies for resolving this taxonomic issue, we applied a molecular approach for species evaluation to two closely related genera, Echinophyllia and Oxypora, for which few molecular data are available. A robust multi-locus phylogeny using DNA sequence data across four loci of both mitochondrial (COI, ATP6-NAD4) and nuclear (histone H3, ITS region) origin from 109 coral colonies was coupled with three independent putative species delimitation methods based on barcoding threshold (ABGD) and coalescence theory (PTP, GMYC). Observed overall congruence across multiple genetic analyses distinguished two traditional species (E. echinoporoides and O. convoluta), a species complex composed of E. aspera, E. orpheensis, E. tarae, and O. glabra, whereas O. lacera and E. echinata were indistinguishable with the sequenced loci. The combination of molecular species delimitation approaches and skeletal character observations allowed the description of two new reef coral species, E. bulbosa sp. n. from the Red Sea and E. gallii sp. n. from the Maldives and Mayotte. This work demonstrated the efficiency of multi-locus phylogenetic analyses and recently developed molecular species delimitation approaches as valuable tools to disentangle taxonomic issues caused by morphological ambiguities and to re-assess the diversity of scleractinian corals.

Species delimitation in the coral genus Goniopora (Scleractinia, Poritidae) from the Saudi Arabian Red Sea.

Variable skeletal morphology, genotype induced plasticity, and homoplasy of skeletal structures have presented major challenges for scleractinian coral taxonomy and systematics since the 18th century. Although the recent integration of genetic and micromorphological data is helping to clarify the taxonomic confusion within the order, phylogenetic relationships and species delimitation within most coral genera are still far from settled. In the present study, the species boundaries in the scleractinian coral genus Goniopora were investigated using 199 colonies from the Saudi Arabian Red Sea and sequencing of four molecular markers: the mitochondrial intergenic spacer between CytB and NAD2, the nuclear ribosomal ITS region, and two single-copy nuclear genes (ATPsß and CalM). DNA sequence data were analyzed using a variety of methods and exploratory species-delimitation tools. The results were broadly congruent in identifying five distinct molecular lineages within the sequenced Goniopora samples: G. somaliensis/G. savignyi, G. djiboutiensis/G. lobata, G. stokesi, G. albiconus/G. tenuidens, and G. minor/G. gracilis. Although the traditional macromorphological characters used to identify these nine morphospecies were not able to discriminate the obtained molecular clades, informative micromorphological and microstructural features (such as the micro-ornamentation and the arrangement of the columella) were recovered among the five lineages. Moreover, unique in vivo morphologies were associated with the genetic-delimited lineages, further supporting the molecular findings. This study represents the first attempt to identify species boundaries within Goniopora using a combined morpho-molecular approach. The obtained data establish a basis for future taxonomic revision of the genus, which should include colonies across its entire geographical distribution in the Indo-Pacific.

Scleractinian corals (Fungiidae, Agariciidae and Euphylliidae) of Pulau Layang-Layang, Spratly Islands, with a note on Pavonamaldivensis (Gardiner, 1905).

Layang-Layang is a small island part of an oceanic atoll in the Spratly Islands off Sabah, Malaysia. As the reef coral fauna in this part of the South China Sea is poorly known, a survey was carried out in 2013 to study the species composition of the scleractinian coral families Fungiidae, Agariciidae and Euphylliidae. A total of 56 species was recorded. The addition of three previously reported coral species brings the total to 59, consisting of 32 Fungiidae, 22 Agariciidae, and five Euphylliidae. Of these, 32 species are new records for Layang-Layang, which include five rarely reported species, i.e., the fungiids Lithophyllonranjithi, Podabaciasinai, Sandalolithaboucheti, and the agariciids Leptoseriskalayaanensis and Leptoseristroglodyta. The coral fauna of Layang-Layang is poor compared to other areas in Sabah, which may be related to its recovery from a crown-of-thorns seastar outbreak in 2010, and its low habitat diversity, which is dominated by reef slopes consisting of steep outer walls. Based on integrative molecular and morphological analyses, a Pavona variety with small and extremely thin coralla was revealed as Pavonamaldivensis. Since specimens from Sabah previously identified as Pavonamaldivensis were found to belong to Pavonaexplanulata, the affinities and distinctions of Pavonamaldivensis and Pavonaexplanulata are discussed.

Pachyseris inattesa sp. n. (Cnidaria, Anthozoa, Scleractinia): a new reef coral species from the Red Sea and its phylogenetic relationships.

A new scleractinian coral species, Pachyseris inattesa sp. n., is described from the Red Sea. Despite a superficial resemblance with some species in the agariciid genus Leptoseris with which it has been previously confused, P. inattesa sp. n. has micro-morphological characters typical of the genus Pachyseris. This genus, once part of the Agariciidae, is comprised of five extant species and is widely distributed throughout the tropical Indo-Pacific. It is currently incertae sedis as a result of recent molecular analysis and appears to be closely related to the Euphylliidae. A molecular phylogenetic reconstruction including P. inattesa sp. n., the genus type species P. rugosa, and P. speciosa, all present in the Red Sea, was performed using the mitochondrial intergenic spacer between COI and 16S-rRNA. The results confirm that P. inattesa sp. n. is a monophyletic lineage closely related to the other Pachyseris species examined.

Lobophylliidae (Cnidaria, Scleractinia) reshuffled: pervasive non-monophyly at genus level.

The Indo-Pacific scleractinian coral family Lobophylliidae was recently described on the basis of molecular data and micromorphological and microstructural characters. We present the most comprehensive molecular phylogeny reconstruction of the family to date based on COI and rDNA including 9 genera and 32 species, 14 of which were investigated for the first time. The monophyly of the family is now strongly supported, with the inclusion of the genera Acanthastrea and Micromussa, whereas previously it was based on uncertain molecular relationships. Nevertheless, these and the other lobophylliid genera Echinophyllia, Micromussa, Oxypora, and Symphyllia, are not themselves monophyletic and need to be investigated from a morphological point of view. Acanthastrea faviaformis is nested within the family Merulinidae. This study highlights the need for further analyses at species level and of formal taxonomic actions.