Zoantharian specimens from the Azores were obtained from by-catch material caught during scientific longline fishing cruises onboard of the RV “Arquipélago” (ARQDAÇO monitoring programme and DEECON project) and from the local longline fisheries fleet. Samples were collected at depths between 110 and 800 m from 12 locations in the Azores region (Figure 1). One additional zoantharian specimen was collected in the Cap Sicié canyon (Mediterranean Sea) in 2010 with the ROV Achille during the MEDSEACAN cruise (dive SI-ACH- P4) (Fabri et al., 2014). Immediately after collection, each zoantharian specimen was split into two fragments and preserved in 10% formalin for morphological, histological and microanatomy analysis and 96% ethanol for molecular studies. All type material is deposited in the reference collection (COLETA) of the Department of Oceanography and Fisheries, University of the Azores (DOP-UAz).

Genomic DNA was extracted using the commercial kit Mag-Bind® Tissue DNA Kit (Omega Biotek) following the manufacturers' instructions. PCR amplification of the mitochondrial protein-coding gene COI was performed using primers specifically designed for zoantharians, COIZoanF and COIZoanR (Reimer et al., 2007); while for the 16S and 12S rDNA regions, the pairs of primers used were 16Sant1a/16SbmoH and 12S1a/12S3r, respectively (Sinniger et al., 2005). PCR reactions were performed in a 20 μl total-reaction volume with 10 μl of PCR Mastermix solution (Promega), 0.5 μl of each primer (10 μM), 8 μl of pure water and 1 μl of template DNA.

The thermal cycling profile for the three genomic regions started with an initial denaturation at 95°C for 3 min followed by 40 cycles of touch-down PCR (denaturation at 95°C for 30 s, annealing at 52–72°C for 1 min 35 s), and a final extension at 72°C for 7 min on a BioRad Mycycler thermal cycler. Non-template controls were included to all PCR reactions to detect any genomic DNA contamination. Electrophoresis of PCR products on a 1% agarose gel was performed to evaluate the integrity of the products. Finally, all amplified products were purified using ExoSAP-IT (USB Corporation) and sent for sequencing to the BMR Genomics (Padua, Italy) facility using the same set of primers. Newly obtained sequences were deposited in the NCBI database. GenBank accession numbers for all molecular markers and for each species used to reconstruct the phylogeny are reported in Supplementary Table 1.

Additional sequences of closely related species (Supplementary Table 1) were retrieved from the NCBI database to reconstruct the phylogeny of zoantharians from the Azores. All sequences were aligned using MAFFT v. 7 (Katoh and Standley, 2013) and sections with large indels were rearranged by eye. For the 16S, the V5 region was aligned following Sinniger et al's (2013) suggestions. In addition, indel events were coded as binary characters with SeqState (Müller, 2005) using simple-indel coding approach (SIC; Simmons and Ochoterena, 2000) and added to the existing alignment. Moreover, 16S and 12S rDNA alignments were also automatically edited by GBlock v. 0.91b (Castresana, 2000) to eliminate potentially poorly aligned positions; the configuration was set in agreement with the suggestions from Montenegro et al. (2015b), allowing small final blocks, gap positions between final blocks and less strict flanking positions.

For the phylogenetic investigation, the most appropriate nucleotide substitution model was selected from the hierarchical series of likelihood ratio test implemented in MEGA 4 (Tamura et al., 2007). Phylogenetic relationships were reconstructed for all three alignments (SIC, an alignment including indel coding; NoSIC, an alignment with no indel coding; and GBlock, a GBlock edited alignment) using Bayesian inference (BI) in MrBayes v. 3.2.6 (Ronquist et al., 2011), while the Maximum Likelihood (ML) in RAxML v. 8.2.8. (Stamatakis, 2014) approach was used only on the alignment with no indel coding; all phylogenetic reconstructions were run online using the CIPRES Science Gateway portal v. 3.3 (Miller et al., 2010). BI was done implementing the HKY+G model (Hasegawa et al., 1985; Yang, 1993) for DNA sequences and a binary model for the coded indel events, assuming unlinked parameters and rates for each gene and a site-specific rate model for the protein-coding COI gene. BI calculations were conducted with the help of the BEAGLE library (Ayres et al., 2012) and run for 20,000,000 generations until the standard deviation of split frequencies value was less than 0.05. For the ML approach, DNA sequences were analyzed in a partitioned dataset (COI, 16S, and 12S) following a RAxML Workflow interface and using the Maximum Likelihood/Thorough Bootstrap approach and GTR+gamma model, while an autoMRE criterion was used for bootstrapping (660 bootstrap iterations). In both approaches, representatives of the family Epizoanthidae Delage and Hérouard, 1901 were used as an outgroup and inferred trees were viewed with Figtree v. 1.4.2 (Rambaut, 2014).

The electronic edition of this article conforms to the requirements of the amended International Code of Zoological Nomenclature, and hence the new names contained herein are available under that Code from the electronic edition of this article. This published work and the nomenclatural acts it contains have been registered in ZooBank, the online registration system for the ICZN. The ZooBank LSIDs (Life Science Identifiers) can be resolved and the associated information viewed through any standard web browser by appending the LSID to the prefix “http://zoobank.org/.” The LSID for this publication is: urn:lsid:zoobank.org:pub:FED88229-30F9-481F-9155-FF481790AE5C. The electronic edition of this work was published in a journal with an ISSN, and has been archived and is available from the digital repository PubMed Central. The supraspecific nomenclature of the zoantharians followed the recent revision of the order Zoantharia in Low et al. (2016). For valid synonyms of families and genera of the order Zoantharia the reader is referred to Low et al. (2016).

Phylum Cnidaria Hatschek, 1888

    Class Anthozoa Ehrenberg, 1831

    Subclass Hexacorallia Haeckel, 1896

    Order Zoantharia Gray, 1832

Diagnosis: Zoantharians characterized by a complete fifth pair of mesenteries.

Diagnosis: Macrocnemic zoantharians that have an endodermal sphincter muscle. Most species in this family associated with other organisms as substrate.

Type Species: Zibrowius ammophilus Sinniger, Ocaña & Baco, 2013.

Diagnosis: Sand-encrusted, arborescent fan-shaped colonies, golden skeleton, well developed coenenchyme completely covering the host, can be confused with Kulamanamana Sinniger, Ocaña & Baco, 2013, but are easily distinguished by the presence of sand encrustation in the ectoderm, and characteristic insertion/deletion pattern in the 16S V5 region sensu Sinniger et al. (2005).

Figures 2–4.

Isozoanthus primnoidus Carreiro-Silva, Braga-Henriques, Sampaio, de Matos, Porteiro & Ocaña, 2011, pp. 409,410, Figures 2–5, Table 1.

Figures 2–5, Table 1.

urn:lsid:zoobank.org:act:8E186AD4-CA6E-419B-B46A-4C8D11C757DD.

Etymology: This species name is dedicated to Prince Albert I of Monaco for his promotion of the oceanographic sciences in the late 19th to early 20th centuries, and for his oceanographic campaigns in the Azores which contributed to the increased knowledge of CWCs and associated fauna in the region.

Type Species: Hurlizoanthus parrishi Sinniger, Ocaña & Baco, 2013.

Diagnosis: Macrocnemic genus associated with primnoids. Characteristic insertion/deletion pattern in the 16S V5 region sensu Sinniger et al. (2005).

Figures 2–4, Table 1.

urn:lsid:zoobank.org:act:6737B10E-9E87-4BA0-9559-C22D49863732.

Type Species: Palythoa axinella Schimdt, 1862.

Diagnosis: Colonial zoantharians characterized by a mesogleal lacuna and by canals forming a “ring sinus” in distal part of polyp. Fine mineral particles are incorporated in polyps.

Figures 2–4, Table 1.

urn:lsid:zoobank.org:act:3D3AA61D-E5CC-47DF-94F1-A4A2FF59ABEA.

Diagnosis: Characterized by a simple mesogloeal sphincter muscle, this family includes the genera Epizoanthus Gray, 1867, Paleozoanthus Carlgren, 1924, and Thoracactis Gravier, 1918. The genus Palaeozoanthus has not been found or examined in detail since its original description (Carlgren, 1924), while Thoracactis topsenti Gravier, 1918 is the sole representative of its genus and is an epibiont on sponges at 800–1100 meters around the Cape Verde Islands (Gravier, 1918). The type genus of Epizoanthidae, Epizoanthus, includes species that have epibiotic associations with hermit crabs (Ates, 2003; Reimer et al., 2010b; Schejter and Mantelatto, 2011), molluscs (Rees, 1967), eunicid worms (Sinniger et al., 2005; Kise and Reimer, 2016), or the stalks of glass sponges (hexactinellids) (Beaulieu, 2001). There are some cases of free-living species reported (Epizoanthus lindahli Carlgren, 1913, Epizoanthus vagus, Herberts, 1972). Polyps usually strongly encrusted with sand. In colonial species, polyps are linked by stolons or by a continuous coenenchyme. There are no symbioses with Symbiodinium zooxanthellae.

Type Species: Dysidea papillosa Johnston, 1842.

Diagnosis: Generally as for the family. Distinct from Paleozoanthus (Gravier, 1918) by its non-fertile micro-mesenteries (Carlgren, 1924, see also Sinniger and Häussermann, 2009). Only one specimen is known from the genus Paleozoanthus and it was not conserved well enough for genus distinction (Sinniger et al., 2005). Furthermore, Epizoanthus is distinct from the genus Thoracactis by presenting polyps joined by a common coenenchyme, in contrast with Thoracactis which shows isolated polyps without apparent coenenchyme connection.

Figures 2–4, 6, Table 1.

Nomenclatural act recorded at Zoobank: urn:lsid:zoobank.org:act:04686BB5-03D7-4132-B52B-CC89DF8EBFA8.

The sequence alignment was 2,067 bp long: 1,114 bp for 16S rDNA, 686 bp for 12S rDNA and 267 bp for COI. Indel coding with SIC approach resulted in an additional 78 binary characters for 12S rDNA and 36 for 16S rDNA that were all added to the existing alignment. After GBlock editing the 16S rDNA alignment shrunk to 50% off its original length (560 bp) and the 12S to 92% (636 bp).

The BI and ML trees had well-supported nodes corresponding to most of the recognized genera (Table 2). One of the four new species of zoantharians reported for the Azores was found to belong to the family Epizoanthidae and three species to the family Parazoanthidae (Figure 7, Supplementary Figures 1, 2). Within the genus Epizoanthus, specimen DOP-3609 from Faial-Pico Channel classified as Epizoanthus martinsae sp. n. was nested within a clade together with the genetically similar Epizoanthus arenaceus (Delle Chiaje, 1823), Epizoanthus vagus Herberts, 1972, and Epizoanthus paxi Abel, 1955.

Inside Parazoanthidae, the first split corresponded to the separation of Isozoanthus Carlgren in Chun, 1903 from the rest of the family representatives. Within Parazoanthidae, all octocoral-associated zoantharians grouped together in a well-supported monophyletic group distinct of Parazoanthus Haddon and Shackleton, 1891, Umimayanthus and Antipathozoanthus species. This clade includes representatives of the genus Savalia Nardo, 1844, Kauluzoanthus Sinniger Ocaña & Baco, 2013, Bullagummizoanthus Sinniger Ocaña & Baco, 2013, Kulamanamana Sinniger Ocaña & Baco, 2013, Corallizoanthus Reimer in Reimer Nonaka Sinniger & Iwase, 2008, Hurlizoanthus Sinniger Ocaña & Baco, 2013, and Zibrowius Sinniger Ocaña & Baco, 2013. Seven of our specimens fell into the Zibrowius clade, where they grouped into two lineages according to the SIC and NoSIC alignments. One of these lineages (specimens: DOP-3242, DOP-804, Si-ACH-P4-1, Si-ACH-P4-2) is classified as Zibrowius primnoidus comb. nov., a recently described species from the Azores that is associated with the gorgonian Callogorgia verticillata and previously placed into the genus Isozoanthus (Carreiro-Silva et al., 2011; for the correct generic placement see also the Systematics results section and the Discussion section), while the second lineage (specimens: DOP-3049, 3050, 5332) is classified as the new species Zibrowius alberti sp. n., closely related to Zibrowius ammophilus Sinniger Ocaña & Baco, 2013. The specimen DOP-3242 of Z. primnoidus differed from its conspecific by a single nucleotide substitution in the 12S rDNA fragment, while the COI and 16S were identical. The genus Isozoanthus represented by the type species Isozoanthus giganteus Carlgren in Chun, 1903 (12S, 16S) and Isozoanthus sulcatus Gosse, 1859 (COI) concatenated in a single sequence (Supplementary Table 1) represented by a well-defined branch formed after a deep first split within Parazoanthidae. This branch is well separated from Z. primnoidus.

Phylogenetic analyses also indicate that the zoantharian associated with the primnoid octocoral Candidella imbricata constitutes a new species within the recently described genus Hurlizoanthus of the family Parazoanthidae (Sinniger et al., 2013). Hurlizoanthus hirondelleae sp. n. is represented by one specimen (DOP-4098) collected in Voador Seamount.

While the support for and within most of the other genera and lineages that are also analyzed (Antipathozoanthus, Hydrozoanthus, Bergia, Parazoanthus sensu stricto and “Parazoanthus darwini Reimer & Fujii, 2010 + Parazoanthus swiftii Duchassaing de Fonbressin and Michelotti, 1860”) is good in all the phylogenetic trees, the relationships between them could not always be well resolved. Nevertheless, specimen (DOP-4090) from the Voador Seamount clearly falls within the Parazoanthus s. str. clade and represents a new species, Parazoanthus aliceae sp. n., found in association with the stylasterid Errina dabneyi. Furthermore, the genus Umimayanthus was well-supported only in the BI tree constructed from the SIC alignment.

The genus Zibrowius, recently described from Hawaii (Sinniger et al., 2013), also includes Zibrowius alberti sp.n. and Zibrowius primnoidus comb. nov. (Carreiro-Silva et al., 2011). The re-classification of the latter species, which was originally classified as Isozoanthus, is in agreement with the revised morphology, anatomy and molecular phylogeny. This re-classification is supported by the association with a primnoid octocoral (as is also the case with Hurlizoanthus hirondelleae sp. n.), marked morphological differences between this species and other Isozoanthus (large ectodermal nematocysts and the mesogleal structure of the body wall) and the phylogenetic reconstruction, which clearly positions this species within the Zibrowius genus. The identification of Z. primnoidus from Cap Sicié canyon (off Toulon, France) found in association with the octocoral Callogorgia verticillata, also confirmed the presence of this species in the Mediterranean, thus expanding its known geographic range. Unreliability of the morphological characters traditionally used to distinguish Isozoanthus (i.e., the lack or absence of the mesogloeal ring sinus system) from other parazoanthids was also shown to be unreliable in the case of Hydrozoanthus antumbrosus (Swain, 2009a). The latter is a symbiotic zoantharian which was first described as belonging to the genus Isozoanthus but it was more related to Parazoanthus s. str. according to molecular data (Swain, 2009a) and was later transfer to a newly erected genus Hydrozoanthus (Sinniger et al., 2010). In an effort to integrate modern molecular data with traditional morphological research, Swain et al. (2016) reviewed and identified the morphological characters most useful to systematics in Zoanthidea by mapping these traits onto the molecular phylogeny of the group. These authors identified traditionally targeted (fifth mesenteries, marginal muscle arrangement, encircling sinus) and novel (fissure morphology, basal canals of the mesenteries) morphological features, as the most important characters necessary for reunification and revision of Zoanthidea systematics. Useful morphological characters identified for the genus Isozoanthus included fissured capitulum and hypertrophic retractor muscles of the microdirectives, which should be targeted in future studies. In addition, the characterization of the cnidome in Isozoanthus could also aid in the correct identification of the genus.

The newly described zoantharian Zibrowius alberti sp. n. was found associated with the octocorals Paracalyptrophora josephinae and Dentomuricea aff. meteor and it presents unique mesogleal and cnidom features, such as mesogloea with a large number of lacunae with endodermic content and lacunae system, and the presence of large holotrichs in the body wall and p-mastigophores with a well-marked filament. Molecular data indicated that this species is more closely related to Zibrowius ammophilus although in its appearance (polyp size, color, and presence of particles in the body wall) it is more similar to Z. primnoidus.

The genus Hurlizoanthus is characterized by zoantharians associated with the primnoid octocorals. Adding Hurlizoanthus hirondelleae sp. n. has consolidated its monophyly, despite being in contrast with the recommendation of Sinniger et al. (2013), where merging Hurlizoanthus parishi Sinniger, Ocaña & Baco, 2013 and Z. primnoidus (previously Isozoanthus primnoidus) in a single genus was suggested based on their morphology and ecology. Furthermore, the color of H. parrishi and H. hirondelleae sp. n. polyps are different from Zibrowius species (white to light pink vs. orange to brownish, respectively) and Hurlizoanthus holds a thin layer of sand particles on the ectoderm (or not at all) in comparison to heavy encrustations present in Zibrowius. In the original description of the genus Hurlizoanthus the diagnostic character was the association with primnoid corals. This feature is no longer unique as it is also valid for the genus Zibrowius, therefore it is now recommended to use the degree of encrustation and characteristic insertion/deletion pattern in the 16S V5 region (Sinniger et al., 2013) as discriminant characters between these two genera.

According to the most recently accepted definition of the genus Parazoanthus (Low et al., 2016), the genus includes zoantharians often associated with sponges but not Hydrozoa and lacking skeletal secretion. However, the new species Parazoanthus aliceae sp. n., observed growing on the hydrocoral Errina dabneyi (Hydrozoa, family Stylasteridae), falls into the clade Parazoanthus s. str. (a.k.a. clade A, in Sinniger et al., 2010) together with Parazoanthus elongatus McMurrich, 1904, Parazoanthus juan-fernandezii Carlgren, 1922, Parazoanthus capensis Duerden, 1907, Parazoanthus anguicomus Norman, 1869, and Parazoanthus axinellae (Schmidt, 1862), species associated to sponges and rocks (Sinniger et al., 2010). The reason for this apparent incongruity may be related to the fact that P. aliceae sp. n. was found to colonize dead portions of E. dabneyi skeleton. Thus P. aliceae sp. n. appears to use the hydrocoral solely as substrata, instead of forming a symbiotic association with hydrozoans as described by Sinniger et al. (2005), Swain (2009b) and Sinniger et al. (2005), but this may need further examination. Indeed, P. aliceae sp. n. differs greatly from its closest relatives in terms of morphology to such a degree that one could suggest placing it in a new genus. However, molecular phylogenetics and especially short patristic distances clearly indicate a very close relationship with P. juan-fernandezii and P. elongatus (Figure 7). Such a discrepancy between molecular and morphological data for the genus Parazoanthus further emphasizes the need to identify and standardize the morphological characters used in Zoantharia systematics (Swain et al., 2016). The new species presents a large variety of nematocysts as the main character, (including several b-mastigophore categories absent from the other species in the same clade) large holotrichs in all tissues and at least two different categories of homotrich in all the analyzed tissues. In addition, P. elongatus and P. juan-fernandezii are reported at shallow depths (10–40 m) in the Pacific, while similar depths are reported for P. axinellae, P. capensis, and P. swiftii in the Mediterranean Sea, Indian and Atlantic Oceans. P. anguicomus is a North Atlantic species growing on sponges and other substrata (stones, worm tubes, coral debris) at depths between 20 and 400 m (see Manuel, 1981). However, it differs from P. aliceae sp. n. by having two categories of holotrichs in its body wall—one larger and the other broader (40–53 × 19–25 μm; 23–35 × 13–19 μm, length × width), and by having more encrustations in the ectoderm.

There are also other deep-sea Parazoanthus species for which sequences are not yet available, but which could possibly belong to this Parazoanthus s. str. clade. One such species is Parazoanthus haddoni Carlgren, 1913, a zoantharian that grows on sponges and presents larger holotrichs (41–46 × 17–18 μm length × width) in the body wall in comparison with P. aliceae sp. n., which has developed two categories of smaller holotrichs in the same tissue (25–30 × 10–15 μm; 19–23 × 7–10 μm length × width) but also other cnidae categories. Even though cnidae show great promise for taxonomic delimitation, they should be used with caution as some types of cnidae found in P. aliceae sp. n. were also reported from Antipathozoanthus macaronesicus (Ocaña and Brito, 2004). However, the latter grows on hydroids (Ocaña et al., 2007) and it was never found on hydrocorals.

There has been continuous effort with the aim of resolving the paraphyly of Parazoanthus with the description of new genera: Antipathozoanthus, Hydrozoanthus, Bergia, and Umimayanthus (Sinniger et al., 2010; Montenegro et al., 2015a,b), and the discovery of new species is providing stronger support to the structure of the groupings. In our phylogenetic reconstruction there is a monophyletic clade (also observed by Montenegro et al., 2015b) formed by P. swiftii and P. darwini, a basal cluster of the Parazoanthus s. str. clade. This clade is well supported by the molecular data, but the morphology needs further investigation before proper taxonomic reassessment can be conducted for this clade. However, according to the literature (Reimer and Fujii, 2010), P. swiftii and P. darwini share similarities in both polyp and coenenchyme color but P. swiftii has shorter and fewer tentacles; and while P. darwini presents special spirulae (not holotrich sensu Reimer and Fujii, 2010) no detailed information on the morphology of cnidae of P. swiftii is available. In addition, P. swiftii differs from P. darwini by being found more often associated with sponges, while P. darwini can sometimes cover rock surfaces. Moreover, these species occur in geographic distinct areas (P. swiftii in the Caribbean and P. darwini in the Galapagos).

From all the specimens' sequences included into the phylogenetic reconstruction, placement of representatives belonging to Umimayanthus varied the most when comparing phylogenetic trees built from the different alignments. While all the representatives from these genera formed a well-supported clade according to the SIC alignment (0.96 posterior probability), their monophyly was not supported according to the NoSIC or the GBlock alignment (Supplementary Figures 1, 2). As the formation of the genus Umimayanthus was mostly based on the presence of highly conservative and unique indels in the V5 region (Montenegro et al., 2015b) such discrepancies are not unexpected. Namely, the SIC alignment also included indel events coded as binary characters, the NoSIC alignment did not utilize phylogenetic information carried by the indel events in such a way, while the Gblock alignment had such potentially poorly aligned positions excluded altogether.

The zoantharian Bergia puertoricense (West, 1979) (formerly known as Parazoanthus clade C sensu Sinniger et al., 2010) forms a well-defined lineage and stands out as a sister species to the Hydrozoanthus clade. The genus Hydrozoanthus was described on the basis of substrate specificity and characteristic patterns in DNA sequences (specific insertions and deletions in the 16S rDNA, especially in the V5 region as defined in Sinniger et al., 2005, 2010). However, our phylogenetic reconstructions of all three alignments (SIC, NoSIC, and Gblock, Table 2) clearly indicate that this genus is well-nested within the Parazoanthidae at least according to the phylogeny of mitochondrial DNA. While limited support for such positioning also comes from the Gblocks alignment (0.84 posterior probabilities; Table 2) which excludes poorly aligned positions (16S rDNA alignment was shrunk to only 50% off its original length) such positioning of the Hydrozoanthus is not supported by the phylogenetic reconstruction based on COI, ITS, and 16S rDNA in the original description of this genus (Sinniger et al., 2010). However, reconstruction of higher level phylogeny within Zoantharia is beyond the scope of this study and should also include representatives from other families accompanied with a detailed revision of their morphology and anatomy. Furthermore, to resolve the relationships within Parazoanthidae and to identify true paraphylies future efforts should be focused on establishing the correct species tree using a multilocus approach with a multispecies coalescent model and not just relying on mtDNA and limited number of nuclear loci.

The zoantharian Epizoanthus martinsae sp. n. growing on portions of dead skeletons of black coral Leiopathes sp. clustered as a sister species of Epizoanthus arenaceus, Epizoanthus paxi and Epizoanthus vagus, which are zoantharians that form no specific relationship with living organisms, and are commonly found attached to rocky bottoms and gastropod shells (Costello et al., 2001). E. arenaceus is morphologically distinct from these species by the presence of a strong sphincter and, among other less important cnidome differences, the absence of an extra category of wide b-mastigophores in the tentacles and the body wall (see Herberts, 1972). E. paxi is quite different from E. martinsae sp. n. by the presence of strong sphincter and smaller holotrichs in its tissues, especially in the tentacles (see Pax and Müller, 1962; Gili et al., 1987). E. vagus presents an extended sphincter along the column and disposed in the center of the mesoglea, meanwhile E. martinsae's sphincter is shorter, covering almost all the mesogloea width. E. vagus has smaller holotrichs in the tentacles and pharynx than those present in E. martinsae sp. n. and does not have two or three different b-mastigophore categories in the tissues (see Herberts, 1972).

Substrate specificity corroborates the structure of the tree associating a unique type of host (or typology of substrate) to each of the monophyletic clade of zoantharians: antipatharians to Antipathozoanthus, sponges to Umimayanthus, hydrozoans to Hydrozoanthus, octocorals to the large clade housing Zibrowius, Hurlizoanthus, Corallizoanthus, Kulamanamana, Bullagummizoanthus, Kauluzoanthus, and Savalia (Sinniger et al., 2005, 2010, 2013; Reimer et al., 2008a; Montenegro et al., 2015b). In contrast, species within the genus Epizoanthus do not appear to form monophyletic groups according to the type of substrate used, being often associated with gastropod shells (regularly inhabited by pagurid crustaceans), tube worms, stalks of glass sponges or living on rocks or dead organisms (e.g., Reimer et al., 2010a; Kise and Reimer, 2016).

The nature of the symbiotic association between zoantharians and their hosts varies along the mutualism-parasitism continuum. While the association between zoantharians and hydrozoans is often thought to be commensal or even potentially mutualistic (e.g., Swain, 2009a,b; Di Camillo et al., 2010), zoantharians associated with gorgonians and black corals are mostly considered as parasitic (Ocaña and Brito, 2004; Reimer and Fujii, 2010; Sinniger et al., 2010, 2013; Carreiro-Silva et al., 2011; Bo et al., 2012). In many cases, coral colonization by zoantharians may result in the complete death of the host. This is the case of the zoantharians Antipathozoanthus cf. hickmani Reimer & Fujii, 2010, and Terrazoanthus onoi Reimer & Fujii, 2010 that colonizes the black corals Myriopathes panamensis (Verrill, 1869) and Antipathes galapagensis Deichmann, 1941 in Ecuador in the Pacific Ocean (Bo et al., 2012). Reaching the organic axis of the host for mechanical support is the final colonization effect for most parasitic zoantharians, benefiting the zoantharians by raising them well above the substratum into faster flowing water that can be filtered, while avoiding the investment of energy to build their own skeleton (Ocaña et al., 1995; Ocaña and Brito, 2004). The parasitic behavior of the zoantharian Savalia savaglia (Bertoloni, 1819) toward their host octocorals (Paramuricea clavata, Eunicella spp.) represents an extreme case of parasitism (Ocaña and Brito, 2004). In this case, when the host is completely engulfed by the zoantharian, S. savaglia becomes able to produce a hard layered proteinaceous skeleton deposited on the host skeleton that can reach large sizes (up to 2 m high, with a main trunk diameter up to 14 cm; Bell, 1891) and attain 2,700 years of age (Roark et al., 2006), becoming itself an important component of coral gardens where it provides structural habitat for a large number of associated fauna species (Cerrano et al., 2010).

The symbiotic association between zoantharians and octocorals observed in the present study and reported in Carreiro-Silva et al. (2011) for Zibrowius primnoidus (previously known as Isozoanthus. primnoidus) and Callogorgia verticillata, suggests parasitic relationships with their hosts. Support for a parasitic behavior is based on the following observations: (1) zoantharians covering gorgonian polyps and coenenchyme damaging the host tissue and forcing the gorgonian polyps to live only on the axis; and (2) incorporation of gorgonian sclerites in the zoantharian tissue (Figure 3, see also Figure 3 in Carreiro-Silva et al., 2011). Therefore, these observations suggest that zoantharians progressively eliminate gorgonian tissue, keeping the gorgonian axis as support, while coral sclerites are used for protection, although the complete overgrowth and death of the octocoral host was never observed. Nevertheless, observations made in aquaria confirm that these zoantharians are able to survive in the absence of their coral host, feeding on particles in the water column.

In contrast, zoantharians associated with the antipatharian Leiopathes sp. and the hydrocoral Errina dabneyi reported in the present study appear to use the coral host only as support, often colonizing dead portions of their skeleton, with no visible damage to the host tissue.

A similar parasitic relationship between a zoantharian and a deep-sea gorgonian has been described for Epizoanthus sp. and the primnoid octocoral Primnoa resedaeformis (Gunnerus, 1763) in the Northeast Channel (Buhl-Mortensen and Mortensen, 2005; Mortensen et al., 2005). Those authors observed what they classified as Epizoanthus sp. gradually overgrowing and killing P. resedaeformis. Mortensen et al. (2005) suggested that the degree of incidence of this zoantharian is related to gorgonian damage by fishing, with Epizoanthus sp. colonizing tissue-abraded areas and taking over large parts of the gorgonian skeleton.

We found the greatest incidence of zoantharians to occur on C. verticillata and Paracalyptrophora josephinae, two octocorals that are most often captured or damaged during long line fishing operations in the Azores (Sampaio et al., 2012). However, based on our data, we cannot determine whether a similar relationship exists between gorgonian damage caused by fishing and the degree of colonization by zoantharians. All octocoral specimens examined in this study were collected within fishing grounds, so we have no comparison with areas closed to fishing. Future studies comparing number of colonized C. verticillata colonies inside and outside fishing areas will help to clarify the effect of fisheries on the degree of colonization by zoantharians.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.