James Davis Reimer

A genetics-based description of Symbiodinium minutum sp. nov. and S. psygmophilum sp. nov. (Dinophyceae), two dinoflagellates symbiotic with cnidaria.

Traditional approaches for describing species of morphologically cryptic and often unculturable forms of endosymbiotic dinoflagellates are problematic. Two new species in the genus Symbiodinium Freudenthal 1962 are described using an integrative evolutionary genetics approach: Symbiodinium minutum sp. nov. are harbored by widespread tropical anemones in the genus Aiptasia; and Symbiodinium psygmophilum sp. nov. are harbored by subtropical and temperate stony corals (e.g., Astrangia, Cladocora, and Oculina) from the Atlantic Ocean and Mediterranean Sea. Both new species are readily distinguished from each other by phylogenetic disparity and reciprocal monophyly of several nucleic acid sequences including nuclear ribosomal internal transcribed spacers 1 and 2, single copy microsatellite flanker Sym15, mitochondrial cytochrome b, and the chloroplast 23S rRNA gene. Such molecular evidence, combined with well‐defined differences in cell size, physiology (thermal tolerance), and ecology (host compatibility) establishes these organisms as distinct species. Future descriptions of Symbiodinium spp. will need to emphasize genetics‐based descriptions because significant morphological overlap in this group obscures large differences in ecology and evolutionary divergence. By using molecular evidence based on conserved and rapidly evolving genes analyzed from a variety of samples, species boundaries are defined under the precepts of Evolutionary and Biological Species Concepts without reliance on an arbitrary genetic distance metric. Because ecological specialization arises through genetic adaptations, the Ecological Species Concept can also serve to delimit many host‐specific Symbiodinium spp.

Reconsidering Zoanthus spp. Diversity: Molecular Evidence of Conspecifity Within Four Previously Presumed Species

Abstract We have conducted the first phylogenetic study to our knowledge of Zoanthus in the northern hemisphere by sequencing and analysing the mitochondrial cytochrome oxidase subunit 1 (COI) gene. Various unidentified Zoanthus specimens and samples of what have been assumed to be four discrete species (Z. pacificus, Z. sansibaricus, Z. gnophodes, Z. erythrochloros) were collected from four field sites in Kagoshima Prefecture, Japan. Based on our obtained COI gene sequences, all but one of our collected Zoanthus samples appear to be conspecific, with nearly 100.00% base pair matching. Genetic results are further backed up by collected polyp diameter, tentacle count, and mesentary count data. These results indicate a need to reconsider and re-analyze current Zoanthus classification and identification. Possible reasons for the large morphological variation in the same genotype in Zoanthus are also discussed.

Potential of DNA Sequences to Identify Zoanthids (Cnidaria: Zoantharia)

Abstract The order Zoantharia is known for its chaotic taxonomy and difficult morphological identification. One method that potentially could help for examining such troublesome taxa is DNA barcoding, which identifies species using standard molecular markers. The mitochondrial cytochrome oxidase subunit I (COI) has been utilized to great success in groups such as birds and insects; however, its applicability in many other groups is controversial. Recently, some studies have suggested that barcoding is not applicable to anthozoans. Here, we examine the use of COI and mitochondrial 16S ribosomal DNA for zoanthid identification. Despite the absence of a clear barcoding gap, our results show that for most of 54 zoanthid samples, both markers could separate samples to the species, or species group, level, particularly when easily accessible ecological or distributional data were included. Additionally, we have used the short V5 region of mt 16S rDNA to identify eight old (13 to 50 years old) museum samples. We discuss advantages and disadvantages of COI and mt 16S rDNA as barcodes for Zoantharia, and recommend that either one or both of these markers be considered for zoanthid identification in the future.

Morphological and Molecular Revision of Zoanthus (Anthozoa: Hexacorallia) from Southwestern Japan, with Descriptions of Two New Species

Abstract No clear method of identifying species in the zoanthid genus Zoanthus has been established, due in part to the morphological plasticity of this genus (e.g., in polyp and colony form, oral disk color, tentacle number). Previous research utilizing the mitochondrial cytochrome oxidase I gene (COI) as a phylogenetic marker indicated that Zoanthus spp. in Japan may consist of only one or two species, despite a bewildering variety of observed morphotypes. Here we have utilized not only COI but also mitochondrial 16S ribosomal DNA (mt 16S rDNA) in order to clarify the extent of Zoanthus species diversity in southern Japan. Our molecular genetic results clearly show the presence of three monophyletic Zoanthus species groups with varying levels of morphological plasticity, including the new species Z. gigantus n. sp. and Z. kuroshio n. sp. We describe all three species found in this study, and identify potential morphological characters (coenenchyme and polyp structure as well as polyp external surface pigmentation patterns) useful in Zoanthus species identification. A morphological dichotomous key is provided to assist in field species identification.

The Parazoanthidae (Hexacorallia: Zoantharia) DNA taxonomy: description of two new genera

The taxonomy of the hexacorallian order Zoantharia is very problematic due to the lack of easily accessible and informative morphological taxonomic characters. This is particularly true in the widespread family Parazoanthidae, members of which use a wide variety of different organisms as substrates. Recently, DNA-based studies have proven to be of great use in clarifying relationships among Parazoanthidae. Here we reconsider Parazoanthidae taxonomy based on analyses of multiple molecular markers [mitochondrial cytochrome oxidase subunit 1 (COI), 16S ribosomal DNA (mt 16S rDNA), and the nuclear internal transcribed spacer region (ITS rDNA)], coupled with ecological and morphological characteristics. Two new genera are described in this study: Hydrozoanthus n. gen. within the new family Hydrozoanthidae, and Antipathozoanthus n. gen in the family Parazoanthidae. The genetic data further suggest that the revised genus Parazoanthus is still polyphyletic and is composed of three distinctive subclades. However, as currently these subclades can essentially be differentiated by genetic data, these subclades should remain within Parazoanthus until further molecular, ecological and morphological studies help to clarify their status and relationships to each other.

Molecular Evidence Suggesting Species in the Zoanthid Genera Palythoa and Protopalythoa (Anthozoa: Hexacorallia) Are Congeneric

Abstract Taxonomic status of the zoanthid genera Palythoa and Protopalythoa has been in question for almost a century. Separation of the two genera has been based on traditional morphological methods (colony and polyp form, nematocyst size and form, and number of septa), with Palythoa polyps embedded in a well developed coenenchyme and Protopalythoa polyps standing free and clear of the coenenchyme. Here we sequenced two mitochondrial regions, the cytochrome oxidase I (COI) gene and 16S ribosomal DNA (16S rDNA) genes, from Palythoa and Protopalythoa samples from various parts of the world and performed phylogenetic analyses of the sequence data. The phylogenetic trees for both COI and 16S rDNA from Palythoa and Protopalythoa show four monophyletic groups (designated Palythoa tuberculosa, Palythoa heliodiscus, Palythoa mutuki 1, and Palythoa mutuki 2), with levels of sequence divergence (COI and 16S rDNA divergence approximately 0.0~1.1%) similar to or lower than that previously found among congeneric species within the closely related genus Zoanthus. Surprisingly, sequence differences among Palythoa tuberculosa, Palythoa mutuki 1, and Palythoa mutuki 2 were negligible (0.0~0.2% for both COI and 16S rDNA), potentially indicating relationships below the species level. Our sequences align well with the few Palythoa and Protopalythoa sequences reported to date. These findings strongly indicate that our samples represent a minimum of two and possibly up to four species (the Palythoa tuberculosa -P. mutuki 1 - P. mutuki 2 group, and P. heliodiscus) within the genus Palythoa, and that the genus Protopalythoa is erroneous nomenclature.

Latitudinal and intracolony ITS-rDNA sequence variation in the symbiotic dinoflagellate genus Symbiodinium (Dinophyceae) in Zoanthus sansibaricus (Anthozoa : Hexacorallia)

We sequenced the internal transcribed spacer of ribosomal DNA (ITS‐rDNA) of Symbiodinium spp. (Freudenthal) from conspecific Zoanthus sansibaricus (Carlgren) colonies along a latitudinal gradient in Japan. Phylogenetic analysis reveals that Zoanthus in the two northern sites of Kokubu and Sakurajima harbor exclusively Symbiodinium subclade C1, whereas Yakushima Zoanthus harbors Symbiodinium subclades C1 and C15, and southernmost Amami Zoanthus Symbiodinium subclades A1 and C1, indicating holobiont flexibility. Individual Zoanthus colonies associated exclusively with one single subclade, but unexpectedly there was small variation between Symbiodinium ITS‐rDNA clone sequences obtained from within individual Zoanthus colonies. There was also a large deletion in the ITS‐2/28S rDNA boundary region in one clone sequence, and another large deletion in the 5.8S rDNA region in another clone. Our intracolony sequence heterogeneity might be a result of the presence of multiple copies of the ITS‐rDNA region within individual Symbiodinium genomes, or result from the possible presence of closely related Symbiodinium genotypes in the host.

DNA barcoding reveals the coral “laboratory-rat”, Stylophora pistillata encompasses multiple identities

Stylophora pistillata is a widely used coral “lab-rat” species with highly variable morphology and a broad biogeographic range (Red Sea to western central Pacific). Here we show, by analysing Cytochorme Oxidase I sequences, from 241 samples across this range, that this taxon in fact comprises four deeply divergent clades corresponding to the Pacific-Western Australia, Chagos-Madagascar-South Africa, Gulf of Aden-Zanzibar-Madagascar, and Red Sea-Persian/Arabian Gulf-Kenya. On the basis of the fossil record of Stylophora, these four clades diverged from one another 51.5-29.6 Mya, i.e., long before the closure of the Tethyan connection between the tropical Indo-West Pacific and Atlantic in the early Miocene (16–24 Mya) and should be recognised as four distinct species. These findings have implications for comparative ecological and/or physiological studies carried out using Stylophora pistillata as a model species, and highlight the fact that phenotypic plasticity, thought to be common in scleractinian corals, can mask significant genetic variation.

Diversity and evolution in the zoanthid genus Palythoa (Cnidaria: Hexacorallia) based on nuclear ITS-rDNA

Previous phylogenetic studies based on mitochondrial DNA markers have suggested that the zoanthid genus Palythoa may consist of both Palythoa species (Palythoa tuberculosa) and species formerly assigned to the genus Protopalythoa (Palythoa mutuki, Palythoa heliodiscus). In the present study various Palythoa spp. samples collected primarily from southern Japan with additional samples from the Indo-Pacific and Caribbean Sea were examined. The nuclear internal transcribed spacer of ribosomal DNA (ITS-rDNA) was sequenced and aligned for phylogenetic analyses to further investigate the relationship between P. tuberculosa, P. mutuki, and P. heliodiscus. ITS-rDNA analyses showed species groups forming monophylies with similar topology but with much higher resolution than seen for mitochondrial phylogenetic analyses. The results also confirmed the very close relationship of P. tuberculosa and P. mutuki. Some specimens appeared to be a potentially undescribed Palythoa species (designated Palythoa sp. sakurajimensis). Additionally, ITS-rDNA sequences of P. mutuki and P. tuberculosa showed additive polymorphic site, demonstrating for the first time a potential history of reticulate evolution in Palythoa.

Fast-Evolving Mitochondrial DNA in Ceriantharia: A Reflection of Hexacorallia Paraphyly?

The low evolutionary rate of mitochondrial genes in Anthozoa has challenged their utility for phylogenetic and systematic purposes, especially for DNA barcoding. However, the evolutionary rate of Ceriantharia, one of the most enigmatic “orders” within Anthozoa, has never been specifically examined. In this study, the divergence of mitochondrial DNA of Ceriantharia was compared to members of other Anthozoa and Medusozoa groups. In addition, nuclear markers were used to check the relative phylogenetic position of Ceriantharia in relation to other Cnidaria members. The results demonstrated a pattern of divergence of mitochondrial DNA completely different from those estimated for other anthozoans, and phylogenetic analyses indicate that Ceriantharia is not included within hexacorallians in most performed analyses. Thus, we propose that the Ceriantharia should be addressed as a separate clade.