Dimorphic life cycle through transverse division in burrowing hard corals

doi:10.21203/rs.3.rs-1461084/v1

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Dimorphic life cycle through transverse division in burrowing hard corals

https://doi.org/10.21203/rs.3.rs-1461084/v1

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The azooxanthellate solitary scleractinian Deltocyathoides orientalis (Family Turbinoliidae), which has bowl-shaped costate corallites, exhibits burrowing behavior on soft substrates and adapts to an infaunal mode of life. Here, we describe the previously unknown aspects of their life history and asexual mode of reproduction based on morphological and molecular phylogenetic analyses. The findings revealed that (1) D. orientalis exhibits asexual reproduction by transverse division; (2) smaller bowl-shaped costate anthocyathus derived from cylindrical to tympanoid anthocaulus attached to hard substrates, including shell fragments and gravels on soft substrates; (3) the anthocyathus only reproduces sexually after division and the anthocaulus regrew and repeatedly produces anthocyathi through transverse division. The bowl-shaped corallum morphology of the anthocyathus just after division might reduce and time of skeletal formation for infaunal adaptation after transverse division. Immediately after division, D. orientalis is able to smoothly shift to a burrowing lifestyle that efficiently utilizes soft-substrate environments, increasing its survival rate. Thus, morphological formation of prospective anthocyathus in the anthocaulus stage is thought to involve not only increasing clonal individuals but also adaptation to the necessarily burrowing free-living mode of life in the anthocyathus stage.

There are more than 1,500 known species of scleractinian corals in the phylum Cnidaria, which can be roughly divided into two groups: zooxanthellate and azooxanthellate corals^[1-2]. Scleractinian corals exhibit both attached and free-living life modes (e.g., Hoeksema, 1993^[3]). Corals attach to hard substrates such as rocks and reefs, and free-living corals live on soft substrates such as sand and mud. There are two main methods of reproduction in scleractinian corals: sexual and asexual. The asexual reproductive modes include intratentacular budding, extratentacular budding, and autotomy, in which a part of the body detaches to produce a new individual. There are two types of autotomy: transverse division, in which a cut is made perpendicular to the oral-aboral axis of the individual and the individual is divided, and longitudinal division, in which a cut is made parallel to the oral-aboral axis and the body is divided^[4].

Although many azooxanthellate corals have been found in the seas around Japan, small and free-living species are difficult to collect and only a few detailed studies of their populations and ecology have been conducted^[5–7].

The turbinoliids which exclusively are azooxanthellate and solitary show free-living mode of life at least anthocyathus stage (i.e., the upper part of a divided corallum in asexual reproduction)^[8]. The corals typically exhibit bowl-shaped, conical, or cylindrical coralla with smaller calicular diameters less than 1 cm. The family is highly diverse, with 23 extant genera and 6 fossil genera, and the oldest turbinoliid fossils are collected from the Late Cretaceous (Campanian) ^[8–10].

The turbinoliid Deltocyathoides genus is reserved for those species previously placed in Peponocyathus that do not undergo transverse division^[8,11]. The corals are free living and bury themselves in soft substrates by substrate sediment being transported upwards along a diagonal path as well as away from the polyp laterally^[5]. Sentoku et al.^[5] demonstrated that D. orientalis has active burrowing and escaping abilities following burial. The infaunal mode of life and the retraction of the oral side of the polyp into the sediment as a reaction to physical stimuli are considered anti-predator strategies, similar to burrowing sea anemones and tube-dwelling anemones. Although little is known about the predators, predation pressure, or predation frequency of this coral, damage repairs have been well recorded in the skeletons of Deltocyathoides orientalis. Even highly fragmented individuals preserving less than 10% of their original skeleton are able to regenerate and repair^[6].

D. orientalis exhibits slightly flattened bowl-shaped corallites, with an average diameter of 10 mm. There are no juveniles of D. orientalis with a calicular diameter of less than 2 mm or a small number of septa. Therefore, little is known about its life history, especially during its juvenile stage, which is essential for adaptation to soft substrates.

Large numbers of living organisms and skeletons of two coral species (D. orientalis and an undescribed species) were collected during a marine sediment survey off the Pacific and Japan Sea coast, Japan. The morphological characteristics of the calice and septal arrangement of these undescribed attached corals are similar to those of free-living D. orientalis, but differ significantly in terms of their corallum forms (cylindrical or tympanoid vs. bowl shape) and modes of life (attached vs. free-living). However, the skeletal specimens of D. orientalis collected in this study showed basal discoloration characteristic of the decalcification that occurs in autotomic asexual reproduction (e.g., transverse division^[26]).

In this study, both species were assumed to have dimorphic life cycles of the same species by asexual reproduction, and morphological and molecular phylogenetic analyses were conducted. Additionally, we aimed to reveal the unknown mode of life of D. orientalis.

Corallum morphology

Deltocyathoides orientalis (Anthocyathus)

Ten D. orientalis (Anthocyathus) individuals were collected from St. 1 and 308 individuals from St. 2. D. orientalis inhabits soft-bottom substrates and possesses a conical or bowl-shaped skeleton completely covered by soft tissue (Fig. 1A–C). The mouth is located in the center of the upper part of the corallum (calice) and is encircled by 48 tentacles. The GCD and LCD of D. orientalis without GBSD are 7.03–10.58 mm and 6.72–10.57 mm (Fig. 3A), respectively, HT is up to 5.20 mm. At the thecal lateral faces, costae ridged, granular, very prominent, and separated by deep intercostal grooves. At base, intercostal grooves are shallow. Higher cycles of costae (third and fourth) originate from bi- or trifurcations. A circular decalcification scar was observed on the base of the skeleton of 41/308 D. orientalis specimens collected at St. 2 (Fig. 2A–D). The intercostal in the center of the decalcification scar were indistinct. The center of the basal part of the smaller corallum comprises protuberances and encircled by costae. Within the corallum, costal edge angle is bimodal, and the angle clearly increases (approximately 20–40°) at the periphery of the decalcified part. The GCD and LCD of D. orientalis with GBSD are 3.61–7.75 mm and 3.34–7.66 mm (Fig. 3B). The GBSD and LBSD are 0.91–3.59 mm and 0.91–3.29 mm (Fig. 3C), respectively.

The septa were ordinarily hexamerally arranged in four complete cycles according to the following formula: S1>S2>S4>S3 (48 septa). Sublamellar to styliform pali before all but the last cycle of septa (P1-3). Columella rudimentary is composed of few granulated and interconnected pillars. S1 are independent and almost reaching columella, and bearing a small pali. The inner septal edges of S1 are slightly sinuous. P1 are usually indistinguishable from columellar elements. S2 are approximately 3/4 the width of S1. P2 are three times wider than P1. S3 are approximately 1/2 size of S2. The thinnest and most recessed P3 are present. Axial edge of P3 fuses to distal edge of adjacent P2. S4 dimorphic in size: those adjacent to S1 are wider than S3, and those adjacent to S2 are approximately as wide as S3. Axial edge of each S4 fuses to distal edge of P3.

Undescribed species (Anthocaulus of Deltocyathoides orientalis)

Four individuals of an undescribed species were collected from St. 1 and seven individuals from St. 2 (Fig. 2E–P). These specimens exhibit cylindrical to tympanoid corallum morphology and are attached to the shell fragments. The GCD, LCD, and HT are 2.56–3.95 mm (Fig. 3D), 2.36–3.90 mm, and 1.38–2.64 mm, respectively. The calice was encircled by a thin wall with faint costae of granular ridges on its surface. Two or more white opaque lines on the outside of the wall appeared perpendicular to the growth direction of the corallum (Figs. 2L, O, and 5C). Some specimens showed exerted septa (especially in S1) and distinct costae only in the uppermost part of the corallum (Fig. 5B). In contrast, some specimens exhibited a depressed center of calice with sloped septal edges toward its center without the exerted septa and costae (Fig. 5D). Another wall was occasionally formed within the calice, similar to a marked rejuvenescence (Fig. 5A). The septa were hexamerally arranged in four complete cycles according to the following formula: S1>S2>S4>S3 (48 septa; except for specimen number 501 which exhibited 38 septa). Columella rudimentary composed of few granulated and interconnected pillars. S1 only independent septa, almost reaching columella with slightly sinuous axial edge, bearing small palus usually indistinguishable from columellar elements. S2, which is approximately 3/4 the width of S1. S3 is approximately 1/2 the size of S2, bearing the thinnest. S4 dimorphic in size: those adjacent to S1 were wider than S3, and those adjacent to S2 were approximately as wide as S3. Axial edge of each S4 fuses to distal edge of S3.

Morphological comparison between anthocaulus and anthocyathus

The GCD with GBSD (n = 42) and GBSD (n = 42) of the anthocyathi, GCD without GBSD of the anthocyathi (n = 30), and GCD of the anthocauli (n = 11) were measured to compare their size distributions. The size distributions of the GCD with GBSD (mean = 5.95 mm, σ = 1.07), the GBSD (mean = 2.35 mm, σ = 0.63) and the GCD without GBSD (mean = 8.23　mm, σ = 0.96) of the anthocyathi clearly differ (Steel–Dwass's multiple comparison test; p < 0.001, Supplementary Table S2). The GBSD of anthocyathi is slightly smaller than the GCD of anthocauli (mean = 3.32 mm, σ = 0.39; Fig. 3, Steel–Dwass‘s multiple comparison test; p < 0.001, Supplementary Table S2). Figure 4 shows the relationship between GCD/LCD with GBSD, GBSD/LBSD, and GCD/LCD without GBSD of Anthocyathi and GCD/LCD of Anthocauli. The ratios of the greater and lesser diameters at the four measurement points were strongly positively correlated (R² = 0.99, n = 125). The size distributions of the GCD of anthocyathi without GBSD (p = 0.26), GCD of anthocyathi with GBSD (p = 0.83), GBSD of anthocyathi (p = 0.77), and GCD of anthocauli (p = 0.99) also approximated a normal distribution, according to the Kolmogorov–Smirnov test (Supplementary Table S3).　

Molecular phylogenetic analyses

Phylogenetic analysis was carried out using the ML method placed D. orientalis and other undescribed species into a single clade in five regions (mitochondrial 12S ribosomal DNA (Supplementary Fig. S1A), and 16S ribosomal DNA (Supplementary Fig. S1B) and cytochrome c oxidase subunit I (COI) (Fig. 6A) regions and nuclear 28S ribosomal DNA (Fig. 6B) and internal transcribed spacer (ITS) (Supplementary Fig. S1C)).

Turbinoliidae, D. orientalis, and other undescribed species were placed in a single clade by four regions other than ITS. Sequence data for the ITS region of Turbinoliidae were not available in previous studies. In 16S ribosomal DNA (462 bp) and CO1 (646 bp), the sequences of all eight samples of D. orientalis and the undescribed species were identical. In the mitochondrial 12S ribosomal DNA region, sequence data from six samples were used because the sequences of two undescribed species (specimen numbers 501 and 505) were unavailable. There was one variable site among the 906 bp 12S ribosomal DNA sequences. In the nuclear 28S ribosomal DNA region of eight samples, there were 11 variable sites (1.58%) among the 698 bp. In the ITS region of eight samples, there were 19 variable sites (3.51%) among the 541 bp.

Transverse division of Deltocyathoides orientalis

In this study, we conducted a detailed morphological analysis of an undescribed species and D. orientalis. The results showed that the two species were clearly different in terms of their mode of life (attached vs. free-living) and GCD size (Figs. 3 and 4, Supplementary Table S2). However, some morphological characteristics such as granular septa, septal and palar arrangement, and columella were common between the two species.

A circular discoloration was observed on the basal part of the skeleton in 41/308 specimens of free-living D. orientalis collected at St. 2 (Fig.2A–D). Similar discoloration by decalcification for transverse division has been observed in the basal part of almost all anthocyathi of Truncatoflabellum^[12,26]. However, over 80% of the specimens of free-living D. orientalis did not show this basal discoloration (Fig. 4). Soft tissues on the basal part of the anthocyathus of Truncatoflabellum spheniscus are immediately lost after transverse division^[26]. Therefore, the original basal scar and the decalcified skeleton remained without additional skeletal thickening. On the other hand, since D. orientalis has soft parts covering the entire skeleton when it is alive, the outer surface of the corallum, including the discolored basal part, also undergoes additional thickening of the associated skeletal region. Consequently, not all basal parts of the corallum of D. orientalis may exhibit discoloration that is hidden under the new intact skeletons. This consideration corresponds to the smaller size distribution of the GCD of anthocyathus bearing basal scars in D. orientalis, which is thought to have a shorter duration of skeletal precipitation than the GCD of anthocyathus without it (Fig. 3, Table S2). In addition, Sentoku et al.^[6] showed that D. orientalis is capable of self-repair if 10% of the skeleton remains after physical damage, in which the basal discoloration can potentially be lost. Therefore, discoloration restricted to the basal part of D. orientalis is inferred to be generated by decalcification, and almost all D. orientalis may inherently have a discolored basal part.

In the attached specimens, white opaque discoloration lines on the outside of the wall perpendicular to the growth direction and double-walled structures, such as rejuvenescences, were observed (Fig. 5A–C). Anthocauli of Pliocene fossils of Truncatoflabellum carinatum and extant Truncatoflabellum spp. show successive rejuvenescences that are derived from a temporal decrease in polyp diameter due to transverse division injury ^[12,26]. Moreover, the anthocauli of extant Truncatoflabellum spp. exhibit traces of decalcification at the uppermost periphery of the outer wall portion of the rejuvenescence. A similar trace of decalcification of anthocaulus has been reported in Fungia fungites ^[27]. The discoloration and double-walled structures in the attached specimens are thought to have been formed by transverse division with decalcification.

Furthermore, the nucleotide sequences of the undescribed species and D. orientalis were completely consistent in the 16S-rDNA region (462 bp) and CO1 region (646 bp). In addition, because of creating phylogenetic trees using the nucleotide sequences obtained from the five gene regions, the undescribed species and D. orientalis showed a single clade in all gene regions (Fig. 6A–B, Supplementary Fig. S1A–C).

Based on the results of both the morphological and molecular phylogenetic analyses, we concluded that the undescribed species was the anthocaulus of D. orientalis. Moreover, the free-living anthocyathus of D. orientalis reproduces asexually by transverse division of the attached anthocaulus.

Control of anthocyathus morphology by transverse division

Two calical morphologies were observed in the attached coralla: the depressed calice and the exerted and bulged calice with distinct costae (Fig. 5). Particularly, the skeletal characteristics of the uppermost part of the latter attached specimens closely resemble those of the anthocyathus of D. orientalis (Fig. 5B). The smaller corallum of the anthocyathus of D. orientalis shows a slightly protruding base with decalcification. The center of the basal part is composed of tubercular fragments of divided columella of the anthocaulus and is encircled by costae. In the side view, the costal edge angle is bimodal, and the angle clearly increases (from approximately 20–40°) at the periphery of the decalcification part. The protruding morphology of the lower section of the anthocyathus of D. orientalis fits into the depressed calice morphology of the anthocauli. Thus, the exerted and bulged calical part with the deeper part of the inner calice of the anthocaulus without the outer wall of D. orientalis corresponding to slightly bowl-shaped anthocyathus may be scooped from anthocaulus by transverse division. Since only the inside of the calice was extracted from the anthocaulus, the GBSD of anthocyathus of D. orientalis was, therefore, slightly smaller than the GCD of anthocaulus (Fig. 3).

The bowl-shaped corallum and distinct costae of D. orientalis play important roles in burrowing into soft substrates^[5,6]. The corallum morphology of the anthocyathus with a protruding base immediately after division might have an ecological function that facilitates free living on soft substrates. The anthocyathi of Truncatoflabellum show truncated basal scars that are almost horizontal or along the arcuate growth line on its wall surface^[4,26]. In addition, the increased thickening deposits in the lower parts of the anthocyathus of Truncatoflabellum might contribute toward an increase in stability in the substrate, as well as keeping the calice oriented toward the sea surface, which would be advantageous for purposes of food acquisition^[26]. Thus, the morphological formation patterns of prospective anthocyathus by skeletal growth and carving by decalcification in the anthocaulus stage are thought to involve not only increasing clonal individuals but also adaptation to the free-living mode of life after the transverse division in the anthocyathus stage.

Alternation of generations by transverse division

Statistical examination with Steel-Dwass's multiple comparison test showed that the size distributions of GCD and GBSD of the anthocyathi in D. orientalis were distinctly different (Fig. 3, Table S2). The size distribution of the basal scars approximates a normal distribution (Table S3), suggesting that transverse divisions occur at the coralla that reach a certain diameter, not randomly. The size distribution of GCD of the anthocyathi are significantly different from GBSD (Table S2). The difference clearly indicates that the anthocyathus of D. orientalis does not show asexual reproduction by transverse division. In contrast, plural white opaque horizontal lines of decalcification on the outside of the wall of the anthocauli of D. orientalis indicate repeated transverse divisions (Figs. 2L, O, and 5C). The present statistical analysis supports that the anthocauli of D. orientalis repeatedly reproduce asexually through transverse division, whereas the anthocyathi only reproduces sexually. D. orientalis thus exhibits a distinct alternation of generations (sexual in anthocyathi vs. asexual in anthocauli), which is also well known in Fungia and Truncatoflabellum ^[4,12,26,28]. Differences were observed in the number of anthocauli (seven individuals) and anthocyathi (384 individuals) of D. orientalis collected at St. 2. The small number of the anthocauli of D. orientalis is similar to it of anthocauli of Truncatoflabellum ^[12]. These larger increasing rates of anthocyathus suggest that asexual reproduction may play an important role in increasing coral population size in soft-substrate environments, such as sand and mud.

Anthocyathi of Peponocyathus duncani ^[29] and Bourneotrochus stellulatus^[30]show repeated asexual reproduction by transverse division (e.g.,^[31–33]). Zibrowius^[35] and Stolarski^[32] discussed importance of the transverse divisions of anthocyathi for adaptive strategies on soft substrates, as well as the increase in clonal individuals^[3²^{, 35]}. Corallum size and weight reduction by transverse division of the anthocyathus could lead to efficient automobility in Peponocyathus duncani having a smaller cylindrical corallum with a maximum calicular diameter of 3.7 mm^[32]. However, the morphological analysis of D. orientalis revealed that the anthocyathi did not undergo transverse division. The acquisition of automobility in anthocyathi of D. orientalis, including burrowing, escape from burial, and righting behaviors, implies that the species is actively utilizing habitats under the seafloor. The ability of D. orientalis to retract the oral side of the polyp into the sediment is considered to be an anti-predator response, similar to burrowing sea anemones and tube-dwelling anemones^[5]. The asexual reproduction of division that divides after the formation of a bowl-shaped anthocyathus with costae revealed in this study reduces the cost and time of skeletal formation for infaunal adaptation after transverse division. Immediately after division, D. orientalis is able to smoothly shift to a burrowing lifestyle that efficiently utilizes the soft-substrate environments, increasing its survival rate (Fig. 7).

In this study, we clarified the life history of D. orientalis based on morphological and molecular phylogenetic analyses. In the future, it will be necessary to clarify the frequency of sexual and asexual reproduction and the detailed growth pattern of the species by observing the development of the gonads of anthocyathi and the ecology of anthocauli. Furthermore, since Deltocyathoides has been classified based on the presence or absence of transverse division with the closely related genus Peponocyathus, it is necessary to systematically revise both genera in the future.

Specimens

We collected samples at depths of 104 m off the Pacific coast of Ohakozaki (39°21.966110 N, 142°01.59220 E; St.1) and at depths of 195 m off Hachinohe (40°30.08620 N, 141°50.85440 E; St.2), Japan. We examined 318 individuals of D. orientalis and 11 individuals of an undescribed species of solitary coral attached to shells (Fig. 1). Of these, four individuals with soft parts were selected for molecular phylogenetic analyses (Fig. 2). Additionally, individuals with basal scars formed at the bottom of D. orientalis were selected for morphological analysis. Details of the macro-skeletal features were observed using stereomicroscopes (Leica M165C) and a digital microscope (Keyence VHX-7000). The relevant corallae were photographed at various angles and magnifications for documentation purposes. Where necessary, measurements were performed using Adobe Photoshop (Adobe Inc., CA, USA) and ImageJ (NIH, MA, USA)^[13] and electronic calipers. These data were further statistically verified using Steel–Dwass’s multiple comparison test. The size distributions of GCD and GBSD were assessed for the goodness of fit of the normal distribution using the Kolmogorov–Smirnov test (Supplementary Table S3). The following abbreviations of morphological terms are used in the text ^[14,15]: GCD, greater calicular diameter; LCD, lesser calicular diameter; GBSD, greater basal scar diameter; LBSD, lesser basal scar diameter; HT, height of corallum; Sx, Px, cycle of septa and pali.

DNA preparation, amplification, and sequence analyses

Four individuals each with soft parts of D. orientalis (specimen numbers: 502, 504, 506, and 508) and undescribed species (specimen numbers: 501, 503, 505, and 507) were selected for DNA extraction. Specimens of the entire system (including the skeleton) were extracted and immersed in a lysis buffer. Genomic DNA was extracted using a DNeasy Tissue Kit (QIAGEN Germany, Hilden). DNA concentrations were determined using a Nanodrop 1000 (Thermo Scientific) prior to Polymerase Chain Reaction (PCR) amplification under the following conditions:

(i) 16S rDNA, the primers developed by Le Goff-Vitry et al.^[16] (LP16SF 5′ -TTGACCGGTATGAATGGTGT and LP16SR 5′ -TCCCCAGGGTAACTTTTATC) were used to amplify a fragment of 462 bp.

(ii) COX1, the universal primers developed by Folmer et al.^[17] (LCO1 490 5′ -GGTCAACAAATCATAAAGATATTGG and HCO2 198 5′ -TAAACTTCAGGGTGACCAAAAAATCA) were used to amplify a fragment of 646 bp.

(iii) For 12S rDNA, the primers developed by Chen and Yu (2000)^[18](ANTMT12SF 5′-AGCCACACTTTCACTGAAACAAGG and ANTMT12SR 5′ -GTTCCCYYWCYCTYACYATGTTACGAC) were used to amplify a fragment of approximately 905 bp.

(iv) 28S rDNA primers developed by Medina et al.^[19] (28S.F63sq 5′ -AATAAGCGGAGGAAAAGAAAC and 28S.R635sq 5′ -GGTCCGTGTTTCAAGACGG) was used to amplify a fragment of approximately 700 bp.

(v) ITS1-5.8rRNA-ITS2 – the primers developed by McFadden^[20] (1S-GGTACCCTTTGTACACACCGCCCGTCGCT and 2SS-GCTTTGGGCGGCAGTCCCAAGCAACCCGACTC）were used to amplify a fragment of approximately 540 bp.

PCR cycling conditions were as follows: 95°C for 4 min, followed by four cycles of 30 s at 94°C, 60 s at 50°C, 120 s at 72°C, and 30 cycles of 30 s at 94°C, 60 s at 55°C, 120 s at 72°C, and 4 min at 72°C. The PCR products were run on 1.5% agarose gel, extracted from the gel, and purified using the Wizard SV Gel and PCR Clean-Up System (Promega, Madison, WI, USA). The fragments were directly sequenced on a 3130 Genetic Analyzer (Applied Biosystems, Foster City, CA, USA) with a BigDye Terminator v1.1 or v3.1 Cycle Sequence Kit (Applied Biosystems). The sequences were deposited in GenBank (accession numbers, see Table S1).

Six to eight sequences from the five regions gathered in this study were combined with 137 sequences derived from previous studies (Supplementary Fig. S1, Supplementary Table S1). The sequences were aligned using the online version of MAFFT (v7.503; http://mafft.cbrc.jp/alignment/server/, last accessed February 2^nd, 2022^[21,22]). The gap regions were trimmed using TrimAl (1.2rev59)^[23]. Maximum likelihood trees were constructed using RaxML^[24] under the GTRCAT model^[25] with 1000 bootstrap replications. The sequences of Gardineria hawaiiensis were used as outgroups, except for the ITS region analysis. All phylogenetic trees were visualized and edited using FigTree v1.4.4 (http://tree.bio.ed.ac.uk/software/figtree/, last accessed February 2^nd, 2022)

Acknowledgments

We are most grateful to the captains and crew members of the JAMSTEC (KS14-5) for their generous help in collecting specimens. We thank K. Endo, T. Sasaki for providing us with specimens. This study was supported by grants from the Scientific Research Fund of the Japan Society for the Promotion of Science (18K13649, 20K04147, B18H03366A, 21K14032). This work was also supported by Tottori University of Environmental Studies Grant-in-Aid for Special Research.

Author Contributions

A.S. designed and performed all the experiments, A.S., K.S. and T.N. analyzed the results. A.S. and Y.T. wrote the manuscript. All authors gave final approval for publication.

Data Availability Statement

The data underlying this article are available in the GenBank/EMBL/DDBJ database at https://www.ddbj.nig.ac.jp/index-e.html, and can be accessed with accession numbers LC685944-LC685965 and LC686129-LC686144.

e.g.,

http://getentry.ddbj.nig.ac.jp/getentry/na/LC685944/?format=flatfile&filetype=html&trace=true&show_suppressed=false&limit=10

Additional Information

Competing interests: The authors declare no competing interests.

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No competing interests reported.

FigureS1A.ai
Supplementary Figure S1A-C. Results of phylogenetic analyses of the scleractinian corals based on 3 regions the mitochondrial 12S ribosomal DNA (A), 16S ribosomal DNA (B) and ITS (C) regions. All maximum likelihood trees were constructed using RAxML under GTRCAT model with 1000 bootstrap replicates. Numbers on nodes indicate the bootstrap values. Asterisks indicate 100% bootstrap support.
FigureS1B.ai
FigureS1C.ai
TableS1.xlsx
Supplementary Table S1. A list of GenBank accession numbers of genes used in our phylogenetic analyses.
TableS2.xlsx
Supplementary Table S2. A result table of Steel-Dwass's multiple comparison test
TableS3.xlsx

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Dimorphic life cycle through transverse division in burrowing hard corals

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