Zoologica Scripta Towards a revised Amphinomidae (Annelida, Amphinomida): description and affinities of a new genus and species from the Nile Deep-sea Fan, Mediterranean Sea ELIZABETH BORDA, JERRY D. KUDENOV, CHRISTINA BIENHOLD & GREG W. ROUSE Submitted: 5 September 2011 Accepted: 16 December 2011 doi:10.1111/j.1463-6409.2012.00529.x Borda, E., Kudenov, J.D., Bienhold, C. & Rouse, G.W. (2012). Towards a revised Amphinomidae (Annelida, Amphinomida): description and affinities of a new genus and species from the Nile Deep-sea Fan, Mediterranean Sea. —Zoologica Scripta, 00, 000–000. The discovery of a new amphinomid species from wood falls deployed near cold seeps (1694 m) at the Nile Deep-sea Fan (Mediterranean Sea) highlights the need to revise Amphinomidae to better characterize amphinomid diversity. The phylogenetic affinities of the new amphinomid and 12 other species from nine Amphinomida genera were inferred using data from two nuclear (18S rDNA and 28S rDNA) and two mitochondrial (COI and 16S rDNA) genes. The phylogenetic analyses indicated a close relationship of the new species with other amphinomids associated with temporary pelagic substrata, including Amphinome sensu stricto (emended herein) and Hipponoa. The new species belongs to a distinct lineage and we, here, erect a new genus to accommodate it. Cryptonome gen. n. is the second amphinomid genus established for species from chemosynthetic environments. Cryptonome conclava sp. n. is distinguished morphologically from all previously described rectilinear Amphinomidae by lacking notochaetal hooks, having a reduced caruncle, modified neurochaetae and branchiae on nearly all segments. Taxonomic issues regarding amphinomid species presently assigned to Amphinome and the erroneous placement of related xylophylic taxa in Eurythoe are also outlined. We emend and restrict the five known oceanic flotsam species with stalked heart-shaped caruncles to Amphinome sensu stricto. An additional 15 species previously assigned to Amphinome may belong to other genera (e.g. Linopherus) and are here tentatively considered incertae sedis. Finally, Eurythoe turcica and Eurythoe parvecarunculata are transferred to Cryptonome gen. n. as new combinations. A revised key to a subset of rectilinear amphinomid genera (relevant to this study) is presented. Corresponding Author: Elizabeth Borda, Scripps Institution of Oceanography, University of California San Diego, 9500 Gilman Drive, La Jolla, CA 92034-0202, USA. E-mail: eborda@ ucsd.edu Present address for Elizabeth Borda, Texas A&M University at Galveston, 200 Seawolf Parkway, Ocean and Coastal Science Building 3029, Galveston, TX 77553, USA Jerry D. Kudenov, Biological Sciences, University of Alaska Anchorage, Anchorage, Alaska 99508, USA. E-mail: jdkudenov@uaa.alaska.edu Christina Bienhold, HGF-MPG Group on Deep Sea Ecology and Technology, Max Planck Institute for Marine Microbiology, Celsiusstrasse 1, 28359 Bremen, Germany. E-mail: cbienhol@mpi-bremen.de Greg W. Rouse, Scripps Institution of Oceanography, University of California San Diego, 9500 Gilman Drive, La Jolla, CA 92034-0202, USA. E-mail: grouse@ucsd.edu Introduction Amphinomidae is best known for its colourful large bodied tropical members, the ‘fireworms’, that are equipped with irritating calcareous chaetae serving as a defence mechanism against predators (Kudenov 1995; Nakamura et al. 2008). The family includes approximately 130 recognized ª 2012 The Authors d Zoologica Scripta ª 2012 The Norwegian Academy of Science and Letters species in 21 genera (Hartman 1959, 1965; Fauchald 1977; Kudenov 1994b). Amphinomids are globally distributed and are known from intertidal, continental shelf and shallow reef communities, with comparatively few species recorded from the deep sea (Kudenov 1995; Rouse & Pleijel 2001). To date, only two species have been 1 Wood fall amphinomids and systematics d E. Borda et al. described as being associated with deep ocean chemosynthetic environments. Archinome rosacea (Blake 1985) and Archinome storchi Fiege & Bock 2009 were each described from hydrothermal vents in the east Pacific (>2400 m); though, Archinome spp. have been recorded from vent communities found along ridge systems and oceanic basins in the Atlantic, Indian and west Pacific (Desbruyères et al. 2006). Despite low species numbers at present, ongoing exploration of reducing environments around the world continues to yield new discoveries in amphinomid diversity (Borda et al., in preparation). In 2006, deployments of wood fall experiments were conducted at the Nile Deep-sea Fan (1694 m; Dupré et al. 2007; Foucher et al. 2009) and recovered after 1 year to observe colonization by organisms and the establishment of biogeochemical gradients at wood falls. The recovered wood was heavily occupied by different groups of animals and was highly degraded. The main invertebrate colonizers included wood-boring bivalves, Xylophaga sp. (T. Haga, personal communication), echinoids (Asterechinus sp. n. Ameziane, personal communication), sipunculids (Phascolosoma sp.; G. Y. Kawauchi, personal communication) and annelids (e.g. Glycera noelae Böggemann et al. 2011), including the discovery of a new species of Amphinomidae, which we herein describe. Initial efforts, however, to describe our new amphinomid were problematic, because specimens keyed out to the genus Amphinome Bruguière 1789 (e.g. Fauchald 1977). The prevailing definition of Amphinome, as revised by Quatrefages (1866), includes species with strikingly divergent caruncular and chaetal morphologies, despite the characteristic ‘stalked’ heart-shaped caruncle with free lateral wings of the type species Amphinome rostrata (Kinberg 1867; Baird 1868; Day 1967). Quatrefages (1866) also continued to include species with reduced ‘sessile’ caruncles entirely fused to the body wall; this category best representing our new amphinomid. Although Baird (1868) suggested restricting Amphinome to forms having small heart-shaped caruncles, he did so without addressing the generic definition or disposition of remaining congeners. To compound matters, other species with ‘sessile’ caruncles have erroneously been placed in other genera (e.g. Eurythoe Kinberg 1857). The continued use of such a muddled construct by subsequent workers (Gravier 1902; Potts 1909; Chamberlin 1919; Fauvel 1923b, 1930, 1953; Treadwell 1926; Hartman 1959, 1965, 1974; Day 1967; Fauchald 1977; Day & Hutchings 1979) and the lack of a phylogenetic context are untenable and obstruct efforts to understand the systematics of Amphinomidae. In this study, we emend and clarify the taxonomic definition of Amphinome sensu stricto, describe a new genus and species from wood falls in the vicinity of cold seeps, corroborated 2 by phylogenetic evidence, and transfer-related xylophilic taxa into this new genus as new combinations. Methods and materials Specimen collections and taxonomic evaluation Specimens were collected from wood colonization experiments near cold seeps in the eastern Mediterranean Sea from a depth of 1694 m (3232¢04"N, 3021¢23"E; Fig. 1A¢). Experiments were deployed at the ‘‘Central Zone 2A’’ in the Nile Deep-sea Fan (Dupré et al. 2007; Foucher et al. 2009) for 1 year. In November 2006, deployments were conducted with the remote operated vehicle (ROV) Quest 4000 (MARUM, Bremen, Germany) during the BIONIL cruise (RV Meteor) and in November 2007, wood recovery took place during the MEDECO cruise (RV Pourquoi Pas?) with ROV Victor 6000 (IFREMER, Toulon, France; Fig. 1B). The wood experiment consisted of a large Douglas fir deployment that comprised one large wood log (length: 200 cm, diameter: 30 cm) with 10 smaller logs (length: 25–30 cm, diameter: 10–15 cm) tied to it (Fig. 1B). Three of the small logs were recovered during the MEDECO cruise. Metadata are stored in the PANGAEA database (http://www.pangaea. de) and corresponding event labels for the recovery of the experiments are MEDECO2-D339-BOX4, MEDECO2-D339-BOX5 and MEDECO2-D339-BOX6. Wood samples were examined in a cold room at an in situ temperature of 13 C and recovered specimens were fixed either in 99% ethanol or in 4% formaldehyde. Live specimens (Fig. 1C,E) were photographed using a Canon Power Shot A630 digital camera mounted on a Zeiss Stemi SV11 binocular. For morphological evaluation and identification, preserved specimens were examined using a stereo-M5-APO Wilde and Leitz DIS Ortholux compound microscopes equipped and illustrated using camera lucidas. Type specimens were deposited to the Scripps Institution of Oceanography (SIO) Benthic Invertebrate Collections (SIO-BIC A2383, A2387, A2388). Primary keys (e.g. Chamberlin 1919; Fauvel 1923b; Day 1967; Fauchald 1977) used to identify the new amphinomid placed it in the genus Amphinome, which in most descriptions is recognized by the characteristic elevated heart-shaped caruncle; a clear mismatch to our species. However, the possession of a ‘well-developed’ caruncle comes with the caveat that ‘it may be difficult to discern in some species’ (Fauchald 1977: 100). The lack of recent taxonomic revision of Amphinomidae [the last major attempt being Gustafson (1930)] and with revisions for some genera (i.e. Amphinome) dating back to the late 19th century, indicated a need to re-evaluate and redelineate the genus Amphinome. In addition, ª 2012 The Authors d Zoologica Scripta ª 2012 The Norwegian Academy of Science and Letters E. Borda et al. 15°0′0″E d 20°0′0″E Wood fall amphinomids and systematics 25°0′0”E 30°0′0″E 35°0′0″E 40°0′0″E 45°0′0″N 40°0′0″N 35°0′0″N 30°0′0″N A′ A 5 mm Fig. 1 —A. Map showing type localities of Cryptonome gen. n. species considered in this study: Cryptonome conclava sp. n. (red star); Cryptonome turcica comb. n. (green star); Cryptonome parvecarunculata comb. n. (yellow star). —A¢. Location of DIWOOD experiment—wood #5 and type locality of C. conclava, at the Nile Deep-Sea Fan, ‘‘Central Zone 2A’’ in the eastern Mediterranean Sea. Map was generated in ArcMap (Arc-GIS Desktop 9.3) with continental margins provided by ESRI (Kranzberg, Germany) and bathymetry obtained from the 2 min Gridded Global Relief Data ETOPO2v2 (2006, http://www.ngdc.noaa.gov/mgg/ fliers/06mgg01.html). —B. DIWOOD experiment—wood #5, showing large Douglas fir log (black arrow) and small logs (5 of 10 visible, white arrows), taken in situ by ROV Victor 6000 (IFREMER, Toulon, France). —C. Live image of C. conclava on wood. —D. Live image of Amphinome rostrata on Lepas from North Stradbroke Island, Queensland, Australia; —E. Live image of C. conclava, showing close-up of mid body chaetigers and dorsal branchiae. Images provided by CB (B, C, E) and GWR (D). B ~2.5 mm D evaluation of the recent description of Eurythoe turcica Çinar 2008 suggested that this species and Eurythoe parvecarunculata Horst 1912 were incorrectly assigned to the genus Eurythoe and were very similar to the new amphinomid described here. Specimens of Eurythoe turcica and Eurythoe parvecarunculata were not examined in this study; therefore, redescriptions and new combinations, proposed here, are based on literature review and associated photo-documentation. In addition to morphological assessments, we inferred whether our new amphinomid should be designated as a new genus and could be unambiguously distinguished phylogenetically from other amphinomids based on molecular data. ª 2012 The Authors d C Zoologica Scripta ª 2012 The Norwegian Academy of Science and Letters E Taxa To explore the phylogenetic affinities and address the taxonomic status of our new species, we followed the taxon sampling presented in studies described by Wiklund et al. (2008), but increased the taxonomic representation, as well as gene data sampling. Molecular data were collected from nuclear 18S rDNA and 28S rDNA (after Wiklund et al. 2008) and mitochondrial cytochrome c oxidase subunit I (COI) and 16S rDNA (this study). Taxa new to this study include the following: Chloeia viridis Schmarda, 1861, Amphinome rostrata, Archinome storchi and our new wood fall amphinomid, described below. Sequence data new to this study were collected for the following taxa: Archinome 3 Wood fall amphinomids and systematics d E. Borda et al. storchi, Amphinome rostrata (Australia, Mariana Is., Mexico), Chloeia flava Quatrefages 1866; Chloeia viridis, Eurythoe complanata (Pallas 1766), Hermodice carunculata (Pallas 1766), Hipponoa gaudichaudi Audouin & Milne-Edwards, 1830 (California), Pareurythoe borealis (M. Sars, 1862) and Euphrosine foliosa Audouin & Milne-Edwards, 1833. Sequences available on GenBank for Archinome rosacea (17ºS, East Pacific Rise), Hipponoa gaudichaudi (Australia), Paramphinome jeffresii McIntosh 1868, Euphrosine armadillo Sars, 1851 and Euphrosine sp. (Lamarck 1818) were also included (Rouse et al. 2004; Struck et al. 2006; Roussset et al. 2007). In most cases, these terminals represent type species for their respective Amphinomidae genus, except for Eurythoe (i.e. Eurythoe capensis Kinberg 1857), Pareurythoe (Pareurythoe japonica Gustafson 1930) and Paramphinome (Paramphinome pulchella Sars, 1869). Euphrosine species (Amphinomida: Euphrosinidae) were used as outgroup taxa and forced to be monophyletic (Wiklund et al. 2008). Table 1 lists collection localities or sequence data source, new GenBank Accession numbers (JN086523–JN086561; JN223394–JN223401) and voucher information (when available) for taxa included in this study. To minimize the amount of missing data introduced into our data matrices, we excluded GenBank data where at least two of the four genes were not available, with the exception for members of Euphrosinidae (outgroup), Archinome and Hipponoa gaudichaudi. Valencia, CA, USA) according to the manufacturer’s protocols. Amplification reaction mixtures (25 lL) included the following: 12.5 lL of GoTaq Green Master Mix (Promega Corporation, Madison, WI, USA.), 9.5–7.5 lL of water (adjusted for DNA template amount used), 1 lL of each primer (10 lM concentration ea.) and 1–3 lL of DNA template. All reaction mixtures were heated to 94 C for 2 min, followed by 35 cycles of 94 C for 30 s; primer pair-specific annealing temperatures for 30 s; and 68 C for 45 s (unless indicated otherwise); followed by a final extension of 7 min at 72 C in an Eppendorf Mastercycler. Three overlapping fragments of 18S rDNA (c. 1750 bp) were amplified using the following primer pairs (Giribet et al. 1996): (i) 18S1F ⁄ 18S5R (c. 820 bp), (ii) 18S3F ⁄ 18Sbi (c. 850 bp) and (iii) 18Sa2.0 ⁄ 18S9R (c. 650 bp). Annealing temperatures for 18S1F ⁄ 18S5R and 18Sa2.0 ⁄ 18S9R were set to 49 C and for 18S3F ⁄ 18Sbi set at 52 C. The 28S rDNA D1–D3 fragment (c. 996 bp) was amplified using 28SC1’ (Le et al. 1993) and 28SD3 (Vonnemann et al. 2005) with an annealing temperature of 53 C and an extension time of 1 m 30 s. Annealing temperatures for the 16S (c. 342 bp; 16SAnnF ⁄ 16SAnnR; Sjölin et al. 2005) and COI (c. 680 bp; LCO1490 ⁄ HCO2198; Folmer et al. 1994) primer pairs were set to 60 C and 46 C, respectively. PCR amplifications were purified using ExoSAP-IT (Affymetrix, Inc., Santa Clara, CA, USA) and then sequenced and analysed with an ABI PRISM 3730 (Applied Biosystems, Inc. Foster City, CA, USA) sequencer at the Advanced Studies in Genomics, Proteomics and Bioinformatics (University of Hawaii at Manoa). Molecular methods Whole genomic DNA was extracted from tissue samples using the DNeasy Blood and Tissue kit (Qiagen Inc., Table 1 Taxa included in this study, with collection locality data or sequence source, GenBank accession numbers and voucher information Taxon Collection Locality ⁄ Source 18S 28S COI 16S Voucher Archinome storchi Archinome rosacea Amphinome rostrata Amphinome rostrata Amphinome rostrata Amphinome rostrata* Chloeia flava Chloeia viridis Cryptonome conclava sp. n. Eurythoe complanata Hermodice carunculata Hipponoa gaudichaudi Paramphinome jeffreysii Pareurythoe borealis Outgroup Euphrosine armadillo Euphrosine foliosa Euphrosine sp. East Pacific Rise 1725¢S, 11312¢W (c. 2600 m) GENBANK (Wiklund et al. 2008) North Stradbroke Island, Queensland, Australia Uracas Island, Northern Mariana Islands Xahuaychol, Quintana Roo, Mexico 1830¢N, 8745¢W GENBANK (Rouse et al. 2004) Tanabe Bay, Japan 3341¢N, 13519¢E (c. 35 m) Florida Straights, USA 2427¢N, 8311¢W Nile Deep Sea Fan, Egypt 3237¢N, 3021¢E (c. 1700 m) Bocas del Toro, Panama 0921¢N, 8222¢W Carrie Bow Cay, Belize 1648¢N, 8804¢W (1 m) San Diego, CA, USA 3404¢N, 14028¢W GENBANK (Struck et al. 2006) Trondheimsfjord, Norway 6328¢N, 1000¢E (c. 200 m) JN086533 EF076777 JN086534 JN223396 JN223397 AY577888 JN086536 JN086537 JN086535 JN086539 JN086540 JN086542 AY838856 JN086541 JN086523 EF076778 JN086524 JN223400 JN223401 — JN086526 JN086527 JN086525 JN086529 JN086530 JN086532 AY838865 JN086531 JN086543 — JN086544 JN223394 JN223395 — — JN086546 JN086545 JN086548 JN086549 JN086551 AY838875 JN086550 JN086552 — JN086560 JN223398 JN223399 AY577881 JN086554 JN086555 JN086553 JN086557 JN086558 JN086561 AY838840 JN086559 SIO-BIC A2389 SMNH 95029 SIO-BIC A2385 UF_Annelida 427 ECOSUR-OH-P0382 SAM E3362 — UF_Annelida 478 SIO-BIC A2383 SIO-BIC A2380 SIO-BIC A2382 SIO-BIC A2384 — SIO-BIC A2379 GENBANK (Wiklund et al. 2008) Banyuls, France 4228¢N, 0308¢E (0.5 m) GENBANK (Roussset et al. 2007) EF076782 JN086538 DQ779649 EF076783 JN086528 DQ779687 — JN086547 — — JN086556 DQ779613 SMNH 95017 SIO-BIC A2381 — ECOSUR, El Colegio de la Frontera Sur, Chetumal (Mexico); SIO-BICA, Scripps Institution of Oceanography Benthic Invertebrate Collection Annelida (USA); SMNH, Smithsonian National Museum of Natural History; UF, University of Florida Museum of Natural History (USA); SAM, South Australian Museum *Formerly Hipponoe gaudichaudi. 4 ª 2012 The Authors d Zoologica Scripta ª 2012 The Norwegian Academy of Science and Letters E. Borda et al. Sequences were edited and assembled using Sequencher 4.10 (Gene Codes Corporation, Ann Arbor, MI, USA). All data were aligned using Muscle (http://www. ebi.ac.uk/Tools/msa/muscle; Edgar 2004a,b) and trimmed using MESQUITE 2.71 (Maddison & Maddison 2010); COI was also visualized and aligned according to amino acid translation. Substitution saturation of 3rd codon positions of COI was tested using DAMBE (Xia & Xie 2001) via saturation plots of transition and transversion versus corrected (GTR) genetic divergence (GD) and estimation of the index of substitution saturation (Iss; Xia & Lemey 2009; Xia et al. 2003). COI 3rd codon positions were found to be saturated (GD > 20%; 3rd codon position, Iss > Iss.c: Iss = 0.7344; Iss.c = 0.6958; P = 0.2192) and were excluded from all phylogenetic analyses. jModelTest (Posada 2008) was used to infer evolutionary models for the complete dataset and each gene separately, as estimated by the Akaike information criterion. The final dataset of combined COI, 16S, 18S and 28S consisted of 3539 characters. Individual gene datasets (results not shown) and the combined dataset were analysed under the assumptions of maximum parsimony (MP), maximum likelihood (ML) and Bayesian inference (BI). MP analyses were performed using PAUP* v. 4.0a114 (Swofford 2003). Heuristic searches used 100 replicates of random taxon addition and tree-bisection-reconnection branch swapping. Characters were equally weighted and non-additive, and gaps were treated as missing data. Parsimony jackknife (jac) values were obtained with 1000 heuristic pseudoreplicates, using random taxon addition, tree-bisection-reconnection branch swapping and 37% deletion (Farris et al. 1996). ML analyses were performed using RAxML-VI-HPC (Stamatakis 2006) using the resources at Cyberinfrastructure for Phylogenetic Research (CIPRES, Miller et al. 2010; http://www.phylo.org) or RaxML GUI (Silvestro & Michalak 2010). ML analyses of the combined data, partitioned by gene, were run with the thorough bootstrap (boot; 10 runs; 1000 pseudoreplicates) under the GTR+I+G model of evolution. BI analyses were performed in MRBAYES v.3.1.2 (Ronquist & Huelsenbeck 2003) using the resources at CIPRES. Posterior probabilities (pp) of combined data, partitioned by gene, were estimated with GTR+I+G. The default prior distribution of parameters were used for MCMCMC analyses, with one cold chain and three heated chains for two independent runs for 306 generations and sampled every 1000th generation, discarding the first 25% of the burnin. Tracer 1.5 (Rambaut & Drummond 2007) and AWTY (Nylander et al. 2008) were used to check for stationarity and convergence of tree topologies, chains, log likelihoods and model parameters. ª 2012 The Authors d Zoologica Scripta ª 2012 The Norwegian Academy of Science and Letters d Wood fall amphinomids and systematics Results Phylogenetics The phylogenetic position of Cryptonome conclava gen. n. sp. n., described later, within Amphinomidae was inferred from COI, 16S, 18S and 28S, analysed in combination (Fig. 2). Excluding the 3rd codon position for the combined dataset reduced the number of characters from 3765 to 3539, of which 2977 were constant, with 174 and 388 characters being parsimony uninformative and informative, respectively. Analyses of the combined data resulted in five MP trees (L = 1150; CI = 0.677; RI = 0.654). Overall, the resulting topologies from MP, ML and BI analyses were congruent, except for the positions of Eurythoe, Hermodice and Pareurythoe. The expanded taxonomic and genetic sampling (Fig. 2A) corroborated the basal relationships reported by Wiklund et al. (2008) including: (i) paraphyly of Amphinomidae (i.e. Archinomidae nested within); (ii) monophyly of Archinome (Archinomidae) + Chloeia (Amphinomidae), here designated as Clade I (jac = 100; boot = 100; pp = 1.00); and (iii) monophyly of Paramphinome + Pareurythoe + Hermodice + Eurythoe + Hipponoa, here designated as Clade II (jac = 100; boot = 84; pp = 1.00). Within Clade II, Paramphinome was sister to the remaining representative genera, which also included Cryptonome gen. n. and Amphinome. Cryptonome gen. n. received moderate to high support as sister (jac = 99; boot = 82; pp = 0.94) to a monophyletic Amphinome + Hipponoa (jac = 98; boot = 97; pp = 100); however, the positions of Hermodice, Eurythoe and Pareurythoe were poorly supported across all analyses (jac < 70; boot < 70; pp < 0.70). Analyses of individual 18S and 16S (results not shown) and all data combined (Fig. 2) indicated that voucher specimen Hipponoa gaudichauldi (SAM E3362) for which 18S (AY577888) and 16S (AY577881) were sequenced was misidentified by Rouse et al. (2004). The collection locality and sequences are also identical to Amphinome rostrata (SIO-BIC A2385); therefore, the species name for these sequences have been corrected as Amphinome rostrata on GenBank. Systematics FAMILY Amphinomidae GENUS Amphinome Bruguière 1789, emended. Amphinome Bruguière 1789; 48; pl. 61, figs 8–12; Quatrefages 1866: 392; Kinberg 1867: 87–89; Baird 1868: 217–218; Horst 1886: 158–160; Fauvel 1923b: 127; fig. 46a–g, 1953: 81–82, fig. 37; Day 1967: 123; fig. 3.1f–k; Orensanz 1972: 493– 494; fig. 4; Linero-Arana & Diaz 2010:108–109, fig. 1a–e. Pleione Lamarck 1818: 60. Asloegia Kinberg 1867: 89. Colonianella Kinberg 1867: 89. 5 Wood fall amphinomids and systematics d E. Borda et al. Euphrosine foliosa GB --/--/* Euphrosinidae Euphrosine sp. GB */*/* Euphrosine armadillo Archinomidae Archinome storchi */*/* Archinome rosacea GB I */*/* Chloeia flava */--/* Amphinomidae */*/* Chloeia viridis Paramphinome jeffreysi GB II Hermodice carunculata */84/* Pareurythoe borealis Eurythoe complanata */--/-- Amphinome rostrata AU Amphinome rostrata MAR */*/* Amphinome rostrata MEX Hipponoa gaudichaudi GB */*/* Hipponoa gaudichaudi */82/0.94 0.05 Cryptonome conclava gen. n., sp. n. Fig. 2 Phylogenetic hypotheses of the position of Cryptonome gen. n. within Clade II, based on the analyses of the combined dataset (COI, 16S, 18S, 28S). Topology based on BI analyses. Support values at nodes include: MP jackknife (jac), ML bootstrap (boot) and BI posterior probabilities (pp); represented as jac ⁄ boot ⁄ pp). *Nodes of high support: jac > 95%; boot > 95%; boot > 0.98. Values < 85 (jac; boot) and <0.98 (pp) not shown. Not Lenora Grube 1878: 2, pl. 1, figs 1–2; Hartman 1959: 135; Fauchald 1977: 103. Type species. Amphinome rostrata (Kinberg 1867) Material examined. North Stradbroke Island, Queensland Australia: on Lepas found on driftwood (SIO-BIC A2385; SAM E3362; Fig. 1D); Uracas Island, Northern Mariana Islands: 30 August 2003 (UF-Annelida 427); Xahuaychol, Quintana Roo, Mexico: 1830¢N, 8745¢W, 27 November 2005 (ECOSUR-OH-P0382). Emended diagnosis. Body elongate, rectilinear, with indeterminant number of chaetigers. Prostomium reduced, with anterior and posterior lobes present. Five cephalic appendages present on prostomium: paired antennae and palps on anterior lobe; median antenna on posterior lobe (Figs 1D and 3A). Caruncle heart-shaped, with free lateral wings (Figs 1D and 3A), marginally notched to accommodate median antenna; suspended above body wall by stalk, latter attached and confined to chaetiger 1. Chaetiger 1 complete dorsally and incomplete ventrally. Parapodia biramous; notopodia and neuropodia raised above body wall, encompassed by conspicuous collars. Notochaetae include capillaries lacking spurs or prongs and harpoons 6 with two rows of barbs (Fig. 3A¢, 1). Neurochaetae spinous, resilient, tapering distally to curved unidentate hooks in adults (bidentate in juveniles); internally solid, non-retractile. Noto- and neuroaciculae hastate. Branchiae dichotomously branching tufts from chaetiger 3 and continuing almost to end of body. Dorsal cirri cirriform. Ventral cirri subulate, distally pointed. Dorsal anus opening on terminal two to three chaetigers. Pygidial cirrus medial, unpaired. Ecology. Inhabitants of oceanic flotsam and driftwood; commensal of pelagic Spirula spirula Linnaeus, 1758 (Mollusca: Spirulidae) and Lepas Linnaeus, 1758 (Crustacea: Lepadidae) (Day 1967; Rouse, personal observation; Carrera-Parra, personal observation). Remarks. The genus Amphinome was established by Bruguière (1789) to represent four species initially described and assigned to Aphrodita Linnaeus, 1758. Bruguière first listed Amphinome (Aphrodita) flava Pallas 1766 as a nominal type of Amphinome. Lamarck (1818), however, later transferred Amphinome (Aphrodita) flava to his newly established genus Chloeia Lamarck 1818; which later became the type species of this genus (i.e. Chloeia flava Quatrefages 1866). Lamarck (1818) also transferred ª 2012 The Authors d Zoologica Scripta ª 2012 The Norwegian Academy of Science and Letters E. Borda et al. d Wood fall amphinomids and systematics ntf A A′ b dc 1) ca B vc ntp ppl 2) 0.5 mm dc cr I II ma apl e a vc p dc b apl a p C ntf e I ma ca II py pe E ntp an b ntf F′ cr nrf dc 0.5 mm I II III IV vc 0.5 mm vc dc nrp 0.5 mm ppl 0.5 mm D F dc pc 0.5 mm m vc M L G 0.05 mm N O N′ nrp nrp 0.5 mm 0.05 mm 0.1 mm nrf vc 0.1 mm 0.05 mm H′ I 0.05 mm H J P K Q I′ 0.1 mm 0.1 mm 0.05 mm 0.05 mm 0.05 mm 0.05 mm Fig. 3 Amphinome rostrata. —A. Prostomium and anterior segments, dorsolateral view; A¢. (1) Harpoon notochaeta and (2) Neurochaeta (insert after Gustafson 1930). Cryptonome conclava sp. n. —B. Prostomium and anterior segments, Holotype, dorsal view; (C) Prostomium and chaetigers 1(I)–2(II), Holotype, excluding chaetae; (D) Chaetigers 21–23, left side of body, Holotype, lateral view; arrow indicating direction of anterior. —E. Peristomium and anterior segments, Paratype (SIO-BIC A2387F), anteroventral view, I–IV indicate chaetigers; (F) Pygidium and posterior segments, Holotype, dorsal view; (F¢) Pygidium and bilobed anal papilla, Holotype, ventral view; (G) Chaetiger 20, left side of body, Holotype, posterior view; (H) Spurred capillary notochaetae, Holotype, lateral view; (H¢) Variation detail, spurs of notochaetal capillaries; (I) Bifurcate capillary notochaetae, Holotype, lateral view. —I¢. Variation detail, short prongs of bifurcate notochaetal capillaries; (J) Spurred capillary neurochaetae, Holotype, lateral view; (K) Harpoon notochaeta, left chaetiger 43, Holotype, lateral view; (L) Bifurcate notochaetae, left chaetiger 2, Holotype, lateral view; (M) Notoacicula, lateral view; N, P. Bifurcate neurochaetae, left chaetiger 8, lateral view; (O) Neuroacicula, Holotype, lateral view; (Q) Bifurcate neurochaetae, left chaetiger 20, Holotype, lateral view. Abbreviations: a, antenna; ap, anal papilla; apl, anterior prostomial lobe; b, branchia; ca, caruncle; cr, cirriphore; dc, dorsal cirrus; e, eye; ma, median antenna; m, mouth; nrf, neurofascicular lobe; nrp, neuropodium; ntf, notofascicular lobe; ntp, notopodium; p, palp; pc, pygidial cirrus; pe, peristomium; ppl, posterior prostomial lobe; py, pygidium; vc, ventral cirrus. Illustrations by JDK and EB. ª 2012 The Authors d Zoologica Scripta ª 2012 The Norwegian Academy of Science and Letters 7 Wood fall amphinomids and systematics d E. Borda et al. Bruguière’s remaining three Amphinome species (Amphinome carunculata (Pallas 1766), Amphinome complanata (Pallas 1766), Amphinome tetraedra Bruguière 1789) to Pleione, first listing Pleione (Amphinome) tetraedra. Kinberg (1867) commented on the confusion between Pleione (Amphinome) tetraedra Savigny in Lamarck 1818 and Amphinome (Aphrodita) rostrata, cynically suggesting that Pallas and Savigny seemed to have described two species with the same name. Kinberg (1867) noted that Amphinome rostrata and Amphinome (Pleione) tetraedra were previously considered to be synonyms by previous authors and thus elected Amphinome rostrata, the older named species, as the type of Amphinome while providing a diagnosis so broadly defined so it could be used for individuals from the Americas. Kinberg (1867) additionally proposed Asloegia capillata Kinberg 1867 and Colonianella rostrata Kinberg 1867 as new genera and species. Hartman (1948) determined both genera to be synonyms of Amphinome, where she found Colonianella rostrata to be the junior homonym and Asloegia capillata a synonym of Amphinome rostrata sensu Kinberg (1867; Hartman 1965; Fauchald 1977; Bellan 2010). Quatrefages (1866) is the most recent taxonomic study of Amphinome and the genus remains a confusing grouping. Baird (1868) informally presented the earliest restricted definition of Amphinome, highlighting the heartshaped caruncle of Amphinome rostrata. Though, this simply facilitated his newly described Amphinome jukesi Baird 1868; he did not extend his efforts to address the generic status of the remaining Amphinome species. Horst (1886) conveniently followed Baird’s lead in restricting Amphinome, with his description of Amphinome longosetosa Horst 1886. Baird’s definition made such compelling taxonomic sense that subsequent workers such as Fauvel (1923b), Day (1967), Fauchald (1977) and others recognized it. Although neither Baird (1868), nor Horst (1886) re-evaluated species assigned to Amphinome, all subsequent workers continued embracing the generic ambiguity of Amphinome sensu Quatrefages (1866), until now. Amphinome sensu stricto (proposed here) is one of the few genera that includes members in which chaetiger 1 (I; Fig. 3A) forms a dorsally complete ring (best seen in living specimens), and although reduced, it is not dramatically so when compared to chaetiger 2 (II; Fig. 3A). The stalked caruncle is heart-shaped (chordate) with free lateral wings and is physically attached middorsally to chaetiger 1 (Figs 1D and 3A). As proposed in the emended generic definition for Amphinome, we formally restrict this genus to species possessing a characteristic stalked heart-shaped caruncle, with chaetiger 1 dorsally complete and have solid, non-retractile unidentate neurohooks in adults (Fig. 3A¢, 2); bidentate hooks in juveniles. There are 15 species currently attributed to Amphinome that do not 8 align with the emended definition and their generic affiliations are unclear. For example, Amphinome djiboutiensis Gravier 1902; Amphinome maldivensis Potts 1909; Amphinome nigrobranchiata Horst 1912; Amphinome pallida Quatrefages 1866 and Amphinome (Lenora) phillippensis (Grube 1878) have reduced ‘sessile’ caruncles (fused to the body wall) confined to chaetiger 2, but have serrated neurochaetae somewhat reminiscent of those described for Pherecardia Horst 1886. Based on Grube’s (1878) description of the caruncle and chaetal morphology, Lenora Grube 1878 is not a junior synonym of Amphinome sensu stricto (Amphinome philippensis). Amphinome anatifera Krishnamoorthi & Daniel 1950 is most likely based on misidentified Hipponoa specimens from the brood chamber of Lepas (Krishnamoorthi & Daniel 1950) and Amphinome microcarunculata Treadwell, 1901 is a species of Benthoscolex Horst 1912 (Kudenov unpublished). In the future, we will likely need to transfer some of these 15 species to other genera with sessile caruncles (e.g. Linopherus), depending on neurochaetal morphology and branchial distributions. We here propose the removal of the following species from Amphinome sensu stricto, as emended earlier, and declare them incertae sedis: Amphinome abhortoni Quatrefages 1866 Amphinome alba Baird in McIntosh 1895 Amphinome anatifera Krishnamoorthi & Daniel 1950 Amphinome coccinea Renier 1804 Amphinome denudata Quatrefages 1866 Amphinome djiboutiensis Gravier 1902 Amphinome eolides Lamarck 1818 Amphinome latissima Schmarda 1861 Amphinome maldivensis Potts 1909 Amphinome microcarunculata Treadwell 1901 Amphinome nigrobranchiata Horst 1912 Amphinome pallida Quatrefages 1865 Amphinome philippensis Grube 1878 Amphinome stylifera Grube 1860 Amphinome umbo Grube 1870. The type species, Amphinome rostrata, is well illustrated and described morphologically (Baird 1868; McIntosh 1885; Horst 1886; Fauvel 1953; Orensanz 1972; SalazarVallejo 1992; Linero-Arana & Diaz 2010); but the number of barbed rows on its characteristic harpoon notochaetae appears to depend on body size. While most workers describe and illustrate two rows of barbs per harpoon chaeta for specimens measuring c. 50 mm long, Kinberg (1867) recorded the presence of four rows (i.e. Colonianella rostrata), Baird (1868: fig. 1a) illustrated three rows, and Horst (1886: 158) noted 3–5 rows in specimens measuring 175 mm in length. The barbs appear to be smaller and more delicate compared with those present in other amphinomids, such as Eurythoe or Hermodice (Gustafson 1930). ª 2012 The Authors d Zoologica Scripta ª 2012 The Norwegian Academy of Science and Letters E. Borda et al. As such, barbs are not always conspicuous in preserved material, leading to the early surmise and later confirmation that they are particularly susceptible to chemical erosion in acidic solutions (Gustafson 1930; McIntosh 1885). Finally, juvenile specimens possess bidentate neurochaetae superficially similar to those present in Hipponoa gaudichaudi (Kudenov 1994a). In contrast to profligate descriptions and records of Amphinome rostrata, several major taxonomic issues concerning the definition of Amphinome sensu stricto remain. In the first instance, Pallas (1766) types of Amphinome (Aphrodita) rostrata are lost (von Egmond & ten Hove, personal communication) and a neotype from the type locality (Indian Ocean per Pallas 1766: 106) is being described separately as part of a larger revision of Amphinome (Kudenov, unpublished). In addition, Kinberg’s types must be re-examined and all other genus-level synonymies of Amphinome sensu stricto also needs to be scrutinized. The morphological features historically used to establish the other species of Amphinome sensu stricto are based on single records or atypical specimens. For example, four additional species besides Amphinome rostrata are technically assigned to Amphinome sensu stricto. However, we question the validity of each of the latter four species: Amphinome carnea Grube 1856 from St. Croix, US Virgin Islands may be a juvenile specimen and is known only from Grube’s original description, to which Quatrefages (1866) mistakenly refers Amphinome rosea Quatrefages 1866 as a junior synonym (sensu Baird (1868: 219); Amphinome jukesi Baird 1868 is also based on a single juvenile worm from Queensland, Australia, that was likely exposed to acidic solutions that rendered its harpoon notochaetae featureless (note that Amphinome nitida Haswell 1878 and Amphinome pulchra Horst 1912 are surmised to be its junior synonyms and both have welldefined harpoon chaetae; Hartman 1959); Amphinome longosetosa Horst 1886; with its distinctive swimming capillary chaetae, is based on a single adult specimen that is probably a sexually mature swimming stage of Amphinome rostrata; and Amphinome praelonga Haswell 1878 is based on a single, poorly preserved specimen from Papua New Guinea that seems to lack features distinguishing it from Amphinome rostrata and Amphinome jukesi. In essence, the number of recognized species currently assigned to Amphinome sensu stricto is likely misrepresented and their synonymies are likely also confused. Amphinome sensu stricto species and several of the synonymies in the context of Kinberg’s (1867) broad diagnosis, as applied by Hartman (1959), appear to overlap both morphologically and geographically with Amphinome rostrata, which further confounds the current definition of Amphinome sensu stricto. With the possible exceptions ª 2012 The Authors d Zoologica Scripta ª 2012 The Norwegian Academy of Science and Letters d Wood fall amphinomids and systematics of Bruguière (1789) and Kinberg’s (1867) records of Amphinome (Plerone) tetraedra and Amphinome (Pleione) tetraedra and Colonianella rostrata, respectively, we question both the remaining synonymies of Amphinome rostrata (sensu Kinberg 1867; Hartman 1959) and those presented for the other four species cited below. At the moment it appears as if the genus Amphinome is morphologically represented by a single species, namely Amphinome rostrata, as the remaining four Amphinome species may eventually be referred to it as junior synonyms, pending examination of type material or acquisition of fresh material. Amphinome rostrata (Kinberg 1867); Type locality: Indian Ocean Aphrodita rostrata Pallas 1766: 106–109, pl. 8, figs 14–18. Amphinome tetraedra Bruguiere, 1789; Type locality: Indian Ocean ?Amphinome lepadis Verrill, 1885; Type locality: off New England, Atlantic Ocean ?Amphinome luzoniae Kinberg 1867; Type locality: Luzon, Philippines ?Amphinome natans Kinberg 1867; Type locality: ‘Spanish Seas’, Atlantic Ocean ?Amphinome pallasii Quatrefages 1866; Type locality: Antilles, Caribbean Sea ?Amphinome quatrefagesi Kinberg 1867; Type locality: Brazil, Atlantic Ocean The remaining four species below are morphologically consistent with the newly restricted definition of Amphinome, but are here considered incertae sedis: Amphinome carnea Grube 1856; Type locality: St. Croix, US Virgin Islands, Caribbean Sea ?Amphinome rosea Quatrefages 1866; Type locality: West Indies, Caribbean Sea (Hartman 1959) Amphinome jukesi Baird 1868; Type locality: Raine Island, Australia, Coral Sea ?Amphinome nitida Haswell 1878; Type locality: Cape Grenville, Queensland, Australia (Hartman 1959) ?Amphinome pulchra Horst 1912; Type locality: Banda Sea (Hartman 1959) Amphinome longosetosa Horst 1886; Type locality: unknown Amphinome praelonga Haswell 1878; Type locality: Papua New Guinea As with many amphinomid species, widespread distributions have been attributed to extensive synonymies based on morphological similarities, of which Eurythoe complanata and Archinome rosacea are prime examples (Hartman 1959; Kudenov 1974; Desbruyeres et al. 2006; Barroso et al. 2010; Borda et al., in preparation). The name Amphinome 9 Wood fall amphinomids and systematics d E. Borda et al. rostrata represents a morpho-species that exhibits a circumtropical distribution and the inclusion of representatives in our phylogenetic analyses from the west Pacific (eastern Australia and Mariana Islands) and the Mexican Caribbean (Yucatan), does not contradict this notion. We recognize, however, that a more thorough morphological evaluation, and inclusion of faster evolving nuclear gene data (e.g. ITS-1, Calmodulin; e.g. Pleijel et al. 2009; Audzijonyte & Vrijenhoek 2010) in phylogenetic analyses are highly recommended. Genus Cryptonome gen. n. Type species. Cryptonome conclava sp. n., here designated. Diagnosis. Body elongate, rectilinear, with indeterminant number of chaetigers. Prostomium reduced, with anterior and posterior lobes present. Five cephalic appendages present on prostomium: paired antennae and palps on anterior lobe; median antenna on posterior lobe. Caruncle highly reduced, low-lying ridge; confined and fused to chaetiger 2. Chaetiger 1 reduced, dorsally incomplete. Parapodia biramous: notopodia conical and encompassed by conspicuous collars; neuropodia mound-shaped, situated on large inflated lobes. Notochaetae include harpoons, delicate spurred to bifurcate capillaries, and bifurcate chaetae; notopodial hooks absent. Neurochaetae principally include bifurcate chaetae, all resilient internally solid, non-retractile, with long prongs basally inflated and delicate spurred capillary. Noto- and neuroaciculae hastate. Branchiae dichotomously branching tufts from chaetiger 3 and continuing almost to end of body. Dorsal cirri cirriform. Ventral cirri conical to subulate, distally pointed. Dorsal anus opening on terminal one or two chaetigers. Pygidial cirrus unpaired. Etymology. This genus is derived from the Greeks terms (i) kryptos, referring to the habitation of nooks and crannies hidden from view, and (ii) nomos, the word or law. Also refers to the long overlooked systematic complexity of Amphinomidae. Remarks. Cryptonome gen. n. is superficially similar to Linopherus Quatrefages 1866 and Paramphinome Sars, 1872 in having a sessile caruncle confined to chaetiger 2. However, in contrast to Cryptonome gen. n., branchiae in the latter two genera are restricted to anterior chaetigers and neurochaetae are both brittle and unmodified. In addition, the solid, unidentate notopodial hooks of chaetiger 1 in Paramphinome are altogether lacking in Cryptonome gen. n. and Linopherus. This new genus is established to distinguish from all other previously described rectilinear genera in 10 lacking notochaetal hooks, having a reduced caruncle, modified neurochaetae and branchiae on nearly all segments. Furthermore, the phylogenetic evidence supported Cryptonome gen. n. as belonging to a distinct lineage allied to species of Amphinome and Hipponoa and distantly related to Eurythoe, Hermodice, Paramphinome and Pareurythoe within Clade II (Fig. 2). Species Cryptonome conclava, sp. n. (Figs 1C,D and 3B– Q). Type material. HOLOTYPE (SIO-BIC A2388): in crevices of degraded wood falls near cold seeps in ‘‘Central Zone 2A’’, Nile Deep-sea Fan, off the coast of Egypt, 3237.078"N, 3021.386"E, 1694 m, 1 specimen measuring approximately 21 mm preserved length, 4.5 mm wide excluding chaetae for 47 chaetigers, dissected to expose caruncle, November 2007. PARATYPES: SIO-BIC A2387—1 specimen preserved in formalin (stored in 80% ethanol), measuring 23 mm preserved length, 5 mm wide excluding chaetae, collection date and locality as per HOLOTYPE and SIO-BIC A2383—2 specimens preserved in 95% ethanol, 46 chaetigers, oocytes present, collection date and locality as per HOLOTYPE. COLL: Christina Bienhold and Antje Boetius. Description. Body rectilinear, exhibiting asexual reproduction (architomic scissiparity) in anteriormost 7 chaetigers; paired ventral and lateral nerve chords present, each connected by lateral nerves; subdermal pigment canals observed on ventrum of chaetiger 8. Adjacent or contiguous segments fused along lateral margins. Colour in life pink to blue-purple (Fig. 1C,E); preserved uniformly yellow-brown. Prostomium encompassed laterally by chaetigers 1–2 consisting of an anterior and posterior lobe (Fig. 3B,C). Posterior lobe with two pairs of inconspicuous eyes and subulate median antenna, lacking ceratophore extending to chaetiger 3 (Fig. 3B). Anterior lobe with paired lateral antennae; anterior margin bilobed in dorsal view. Paired palps similar in form to lateral antennae, stout, inserted ventrolaterally on anterior prostomial lobe (Fig. 3C). Caruncle reduced, inconspicuous and confined to chaetiger 2, resembling a low, smooth and bluntly triangular-shaped lobe completely fused to dorsum, about twice as long as wide, smaller than posterior prostomial lobe; ciliation pattern not observed; separated by a transverse groove from and not confluent with posterior prostomial lobe (Fig. 3C). Peristomium confluent with incised frontal margin of anterior prostomial lobe, deepening as a shallow Ushaped midventral ciliated groove leading to mouth through chaetiger 3 (Fig. 3E). Posterior lip of mouth formed by chaetiger 4 (Fig. 3E). Pharynx not observed. ª 2012 The Authors d Zoologica Scripta ª 2012 The Norwegian Academy of Science and Letters E. Borda et al. Parapodia biramous with dorsal and ventral cirri on all segments, appearing to alternate in ‘zig-zag’ pattern, particularly in non-regenerating segments of anterior half of body (Fig. 3D). Notopodia confined to dorsolaterum when segment is viewed in cross section; notopodial lobes conical in frontal view; collar present at base of notopodial fascicular lobes; latter conical. Neuropodia confined to ventrolaterum when segment viewed in cross section; neuropodial lobes tumid, enlarged and rounded in frontal view; collar circumscribes neuropodial fascicular lobes; latter small, mound-shaped. Neuropodia of non-regenerating segments conspicuously alternate relative positions along body in a zigzag pattern. Chaetiger 1 strongly reduced, incomplete dorsally and ventrally; notopodia projecting anterodorsally; neuropodia anteroventrally. Chaetiger 2–3 complete dorsally, incomplete ventrally; chaetiger 4 the first complete annular ring. Dorsal and ventral cirri each with conspicuous cirriphores, present on all segments; branchial cirri absent. Dorsal cirri with cirriform styles, arising from apex of notofascicular lobe. Ventral cirri subulate, tapering distally to points, arising from body wall, outside neurofascicular lobes and projecting ventrolaterally (Fig. 3B,D). All chaetae simple, each with a calcareous outer cortex and gelatinous to solid medulla. Notochaetae brittle, with gelatinous medulla. Notochaetae include: (i) spurred capillaries (Fig. 3H,H¢); (ii) bifurcate capillaries (Fig. 3I,I¢); (iii) bifurcate chaetae (Fig. 3L); (iv) harpoon chaetae (Fig. 3K); and (v) aciculae (Fig. 3M). Both spurred and bifurcate capillaries have long prongs delicately serrated; spurred capillaries twice as numerous but shorter than spurred capillaries (Fig. 3H,I); long prongs averaging 524 lm long in chaetiger 2, 613 lm in chaetiger 20 and 578 lm in chaetiger 43 from spur. Bifurcate notocapillaries present in chaetigers 2–44; long and short prongs averaging 880 and 32 lm long in chaetiger 20, and 676 and 45 lm long in chaetiger 44; an angle of 8 formed at junction of long and short prongs. Bifurcate notochaetae associated with regenerating anterior chaetigers 1–4, infrequently present, numbering 3–5 per notofascicle with long prongs smooth; ratio of long to short prongs ranging from 2.6 to 11. Harpoon chaetae arranged around the outer margin of notofascicle, appearing delicate, with single row of reverse barbs; those from far posterior chaetigers with more pronounced barbs (Fig. 3K). Aciculae (Fig. 3M) numbering 4–6 per notofascicle, hastate with distally enlarged blunt tips, and arranged in an arc directly in front of and immediately adjacent to dorsal cirrus. Notopodial hooks absent. Neurochaetae less numerous than notochaetae and include: (i) spurred capillaries (Fig. 3J); (ii) bifurcate chaetae (Fig. 3N,P); and (iii) aciculae (Fig. 3O). Spurred capillaries (Fig. 3J) particularly associated with regenerating anterior chaetigers 1–5, num- ª 2012 The Authors d Zoologica Scripta ª 2012 The Norwegian Academy of Science and Letters d Wood fall amphinomids and systematics bering c. 15 per neurofascicle in chaetigers 1–2, decreasing to 1–2 in chaetigers 4–5; similar in form and size to those of notopodia (e.g. Fig. 3H). Bifurcate neurochaetae resilient, flexible, considerably heavier and less numerous than notochaetae; inner medulla solid; those from non-regenerating and regenerating chaetigers differ. Bifurcate neurochaetae from non-regenerating chaetigers numbering 4–5 per neurofascicle in chaetiger 20 (Fig. 3Q), 5–6 in chaetiger 40, 10–11 in chaetiger 43, and decreasing to 7–8 in each of last segments. Prongs of bifurcate neurochaetae from non-regenerating segments conspicuously stout; short prongs reduced to smooth spurs; long prongs with girths enlarged basally and smooth cutting margins, tapering distally to blunt tips (Fig. 3Q); ratio of long to short prongs ranging from 15 to 23:1 (averaging 19:1) in chaetiger 20, decreasing to 3.9 to 25:1 (averaging 12.4:1) in chaetiger 44. Bifurcate neurochaetae from regenerating chaetigers 1– 8 larger, numbering 4–5 per neurofascicle in chaetigers 1– 4, and 15–18 per neurofascicle by chaetigers 7–8 (Fig. 3N,P). Most prongs of these neurochaetae from regenerating segments and last 1–2 segments generally not stout, in that short prongs well-developed, smooth; most long prongs gradually tapering to blunt tips. Long prongs of one to two neurochaetae from chaetigers 7 to 8 with enlarged basal girths (Fig. 3N¢) as described earlier for chaetae from non-regenerating segments (Fig. 3Q). Ratio of long to short prongs ranging from 3.6 to 5.9:1 (averaging 4.5) for bifurcate neurochaetae from chaetigers 7 to 8. Neuroaciculae hastate with distally enlarged blunt tips, numbering around 6 per neurofascicle and arranged in superior arc along distal apex of neurofascicular lobe (Fig. 3O); best developed along anterior margin, least so along posterior margin. Branchiae first present from chaetiger 3 (Fig. 3B), maximally developed from chaetigers 9–10 to 24–25 (Figs 1D and 3D), continuing to end of body save the last 1–2 chaetigers (Fig. 3F). Terminal gill filaments slender and cirriform, numbering 17–20 per branchia in chaetiger 3, increasing to around 60–70 where maximally developed (Fig. 3D,G) and gradually decreasing to 1–3 in last branchiferous chaetigers; lacking in last two chaetigers (Fig. 3F). Branchiae emerging from single, basally enlarged trunk on body wall behind notopodia (Fig. 3G); trunk branching dichotomously around three times, forming two primary and around eight secondary limbs; each of the latter terminating in 4–8 terminal cirriform filaments; margins of branchial filament conspicuously ciliated. Pygidium with dorsal anus opening on last 2 chaetigers (Fig. 3F); pygidial cirrus an inconspicuous median unpaired, bilobed papilla, latter with paired bubble-shaped capsules each containing a central spherical inclusion (Fig. 3F¢). 11 Wood fall amphinomids and systematics d E. Borda et al. Ecology. A xylophile that colonizes galleries and tunnels excavated by Xylophaga and other invertebrates in wood falls (Fig. 1C) near cold seeps. Reproduction. Sexual and asexual reproduction. All observed specimens exhibited evidence of architomic scissiparity (see Kudenov 1974) in anteriormost and ⁄ or posteriormost chaetigers. Paratype SIO-BIC A2387 was regenerating the anteriormost 6 and posteriormost 9 chaetigers, and left chaetigers 1–2 not fully formed; PARATYPES SIO-BIC A2383 showed regeneration in anteriormost chaetigers and oocytes could be observed through the body wall. Distribution. >1694 m. Eastern Mediterranean, Nile Delta, Etymology. The specific epithet is derived from the Latin conclava, meaning an apartment or parlour, and refers to the network of excavated chambers connected by corridors in timber samples from which this species was collected. Remarks. The type locality and depth of Cryptonome conclava sp. n. does not coincide or overlap with those of other described ‘Amphinome’ species (designated incertae sedis above). Cryptonome conclava sp. n. is similar to ‘Amphinome’ djiboutiensis Gravier 1902 in both having a sessile caruncle confined to chaetiger 2 [not chaetiger 1 as described by Çinar (2008)], but differs from ‘Amphinome’ djiboutiensis by having distally smooth bifurcate neurochaetae instead of coarsely serrated, non-bifurcate neurochaetae with distally bent tips. Cryptonome conclava sp. n. is most similar morphologically and in habit (in part) to Eurythoe turcica and Eurythoe parvecaunculata. Both Eurythoe species have reduced caruncles confined to chaetiger 2, modified bifurcate neurochaetae and highly dichotomous branchiae on most segments from chaetiger 3. Therefore, not only have these species been incorrectly assigned to Eurythoe by Horst (1912) and Çinar (2008), they are congeneric with Cryptonome gen. n. (see proposed new combinations and remarks below). Both Horst (1912) and Çinar (2008) associated the caruncle of their taxa with chaetiger 1. However, the first segment, which is chaetigerous in amphinomids, is always reduced and dorsally incomplete in all genera (Gustafson 1930; Kudenov 1993), except in Amphinome sensu stricto, Hipponoa and Paramphinome. The caruncle is a direct extension of the posterior brain (Gustafson 1930), emerging through the middorsal body wall in such a manner that it appears attached to a variable number of chaetigers, depending on the genus. The caruncle of Cryptonome gen. n., therefore, occupies a middorsal position on chaetiger 2. 12 Cryptonome conclava sp. n. can be distinguished from Eurythoe turcica by having bifurcate capillary notochaetae and from Eurythoe parvecarunculata by lacking bifurcate capillary neurochaetae. Cryptonome conclava sp. n. differs from both species in having a dorsal anus opening through two terminal chaetigers, rather than one, and having a bilobed (incised) midventral pygidial papilla, instead of a unilobed one. While Cryptonome conclava sp. n. and Eurythoe parvecarunculata both have bifurcate capillary notochaetae, Eurythoe turcica lacks them (Çinar 2008). Cryptonome conclava sp. n. differs from Eurythoe parvecarunculata by having finely denticulate bifurcate capillary notochaetae present in most body segments (chaetigers 2–44), in contrast to coarsely denticulate bifurcate capillary notochaetae restricted to the anterior half of the body (chaetigers 1–23). Moreover, the angle formed at the junction of long and short prongs in bifurcate notocapillaries is 8 in Cryptonome conclava sp. n., in contrast to 12 in chaetigers 1–2, and 24 in chaetigers 3– 23 of Eurythoe parvecarunculata. The large prongs of bifurcate neurochaetae from nonregenerating segments of Cryptonome conclava sp. n. are stout with larger girths compared with those of both Eurythoe parvecarunculata and Eurythoe turcica; small prongs tend to be most reduced and spur-like (Çinar 2008). Prong ratios for such bifurcate neurochaetae of Cryptonome conclava sp. n. are greater than those present in Eurythoe parvecarunculata and Eurythoe turcica. For example, Cryptonome conclava ratios average 19:1 in chaetiger 20, and decrease to 12.4 in chaetiger 43; those for Eurythoe parvecarunculata range from 5.5:1 in chaetiger 7, and decrease to 3.3:1 in chaetiger 31; and for Eurythoe turcica range from 6.2:1 in chaetiger 2; 3.4:1 in chaetiger 7; 11.9:1 in chaetiger 31; and 5.5:1 in chaetiger 63. Prong ratios cited for the latter two species apparently reflect single observations of largest chaetae (Çinar 2008), and those calculated for Eurythoe turcica mirror the pattern we observed in Cryptonome conclava sp. n. when chaetae from regenerating and non-regenerating segments were compared. Species Cryptonome turcica (Çinar 2008), comb. n., emended. Eurythoe turcica Çinar 2008:1976–1983, figs 2–5. Type locality. Levantine Sea (coast), Turkey (40–60 m). Diagnosis. Body elongate, rectilinear, with 80 chaetigers. Anterior prostomial lobe marginally rounded, with paired conical antennae and palps; posterior lobe with two pairs of conspicuous eyes and cirriform median antenna extending to anterior margin of chaetiger 3. Caruncle small, round; confined and fused to chaetiger 2. Chaetiger 1 reduced, dorsally incomplete. Parapodia biramous: anterior ª 2012 The Authors d Zoologica Scripta ª 2012 The Norwegian Academy of Science and Letters E. Borda et al. notopodia conical becoming mound-shaped in middle and posterior chaetigers; neuropodia mound-shaped. Notochaetae include harpoons, finely denticulate spurred capillaries and bifurcate chaetae with long prongs denticulate. Bifurcate notocapillaries absent. Bifurcate notochaetae restricted to chaetigers 1–4. Neurochaetae principally include bifurcate chaetae and finely denticulate spurred capillaries, the latter similar to those in notopodia, limited to chaetigers 1–39. Noto- and neuroaciculae hastate, numbering around 3 per ramus. Dorsal cirri long, cirriform. Ventral cirri short, conical, distally pointed. Branchiae from chaetiger 3, continuing almost to the end of the body, dichotomously branching tufts ending in 95 terminal filaments in anterior segments, best developed anteriorly. Dorsal anus opening on last chaetiger. Pygidial cirrus unpaired, reduced to rounded papilla. Ecology. A shallow water xylophile that occupies the galleries and tunnels excavated by other invertebrates in shallow wood falls. Distribution. Eastern Mediterranean, Levantine Sea. Remarks. The species diagnosis is here emended to clarify the position of the caruncle on chaetiger 2, rather than chaetiger 1 (sensu Çinar 2008). Eurythoe turcica is formally transferred to Cryptonome as new combination, Cryptonome turcica comb. n., because it has a small, caruncle confined to chaetiger 2 and lacks the sinuous bilobed caruncle characteristic of Eurythoe species (e.g. Eurythoe complanata; Gustafson 1930; Storch & Welsch 1970; Fauchald 1977). Species Cryptonome parvecarunculata (Horst 1912), comb. n. emended. Eurythoe parvecarunculata Horst 1912: 37; pl. X, figs 1–5; Augener 1918: 90; pl. II, fig. 3, pl. 3, figs 37–38; Bindra 1927: 4; Fauvel 1919: 472; 1923a: 9, 1923b: 9, 1927: 525, fig. 1, 1932: 46, 1939: 272, 1953: 85, fig. 38e–I; Day 1951: 6; 1967: 128, fig. 3.2i–l; Tebble 1955: 84; Uschakov & Wu 1962: 77, pl II A–E; 1965: 208–209; fig. 20A–E; Tampi & Rangarajan 1964: 103; James et al. 1969: 32; Riser 1970: 553; Intés & Loeuff 1975: 294; Perkins & Savage 1975: 25; Day & Hutchings 1979: 94; Rullier & Amoureux 1979: 156; Wu et al. 1980: 115; Salazar-Vallejo 1996: 23; Paxton & Chou 2000: 212; Çinar 2008: 1983– 1985, fig. 6; Amaral et al. 2006: 64; not Hartman 1959: 134; not Hartman 1974: 612. Not Eurythoe djiboutiensis Kirkegaard 1968: 170; not Bellan 2010. Not Amphinome incarunculata Day 1951: 6; 1967: 120. Not Amphinome maldivensis Fauvel 1953: 85; Hartman 1959: 129; 1974: 4. ª 2012 The Authors d Zoologica Scripta ª 2012 The Norwegian Academy of Science and Letters d Wood fall amphinomids and systematics Not Amphinome djiboutiensis Fauvel 1953: 85; Fauvel & Rullier 1957: 58; fig. 3, 1959: 509; Hartman 1959: 128; Wehe & Fiege 2002:18. Not Eurythoe heterotricha Fauvel 1953: 85. Type Locality. Saleh Bay, north coast of Sumbawa, Indonesia (274 m). Diagnosis. Body elongate, rectilinear, with 50 chaetigers. Anterior prostomial lobe marginally rounded, with paired conical antennae and longer digitiform palps; posterior lobe with 2 pairs of small eyes and cirriform median antenna extending to chaetiger 4. Caruncle small, round, confined and fused to chaetiger 2. Chaetiger 1 reduced, dorsally incomplete. Parapodia biramous: notopodia conical; neuropodia mound-shaped. Notochaetae include harpoons, coarsely denticulate spurred and bifurcate capillaries, and distally smooth bifurcate chaeta. Spurred notopodial capillaries absent. Bifurcate notocapillaries present in chaetigers 1–23, with short prongs measuring 40 lm long; a 12 angle formed at junction of long and short prongs of bifurcate notocapillaries in chaetigers 1–2, abruptly increasing to 24 in chaetigers 3–23. Bifurcate notochaetae restricted to chaetigers 1–3. Neurochaetae predominantly include bifurcate chaetae and coarsely denticulate spurred capillaries, the latter limited to chaetigers 1–23. Noto- and neuroaciculae hastate, numbering 3–4 per ramus. Dorsal cirri long, cirriform. Ventral cirri short, digitiform, distally pointed. Branchiae from chaetiger 3, continuing to the end of the body, dichotomously branching tufts ending in 50 terminal filaments in anterior segments, best developed anteriorly. Dorsal anus opening on last chaetiger. Pygidial cirrus unpaired, reduced to rounded papilla. Ecology. A possible xylophile that occupies the burrows of the foliaceous shipworm Nototeredo knoxi (Mollusca: Teredinidae; Maddocks 1979; as reported from specimens in Panama). Distribution. Circumtropical (?); Indonesia and also recorded from: Moreton Bay, Australia (Day & Hutchings 1979), Mozambique (Augener 1918; Day 1967); Brazil (Rullier & Amoureux 1979; Amaral et al. 2006), Xisha Island, South China Sea (Wu et al. 1980; Paxton & Chou 2000); Sanibel Island, Boynton Beach, Florida (Riser 1970); Gold Coast, west Africa (Tebble 1955; Intés & Loeuff 1975); Yellow Sea, east China Sea (Uschakov & Wu 1962); French Guyana (Fauvel 1919, 1923b; Perkins & Savage 1975; Salazar-Vallejo 1996), St. Francis Bay and Point. St. John, South Africa (Day 1951); Galeta Island, 13 Wood fall amphinomids and systematics d E. Borda et al. Panama (Maddocks 1979), Bay of Bengal (Fauvel 1923a; Tampi & Rangarajan 1964; James et al. 1969). Remarks. The species diagnosis is here emended to clarify the position of the caruncle on chaetiger 2, rather than chaetiger 1 (sensu Horst 1912) and add data concerning the angles formed by the long and short prongs of notocapillaries. Eurythoe parvecarunculata is formally transferred to Cryptonome gen. n. as new combination, Cryptonome parvecarunculata comb. n., because it has a small caruncle confined to chaetiger 2 and lacks the sinuous bilobed caruncle characteristic of Eurythoe species. Horst (1912) did not justify the confusing placement of his species in Eurythoe. In retrospect, such confusion followed that Eurythoe parvecarunculata was considered a junior synonym of ‘Amphinome’ djiboutiensis for six decades (Fauvel & Rullier 1957; Hartman 1959; Kirkegaard 1968; Wehe & Fiege 2002), a notion deftly dispelled by Çinar (2008) and supported here. Both Fauvel (1953) and Hartman’s (1959) tentative attribution of Amphinome maldivensis to Amphinome djiboutiensis lacks support because the former species has, in part, bifurcate capillaries in contrast to the latter. Finally, Fauvel’s (1953) referral of Eurythoe heterotricha to Amphinome parvecarunculata fails to account for significant differences in the caruncle. Çinar (2008) illustrated bifurcate neurochaetae for both Cryptonome parvecarunculata comb. n. (and Cryptonome turcica comb. n.) with what he interpreted as damaged or broken short prongs. While such malformations can be wear induced, we suggest that architomy as observed in Cryptonome conclava sp. n. is a more likely explanation for such chaetal forms observed by Çinar (2008). Cryptonome parvecarunculata comb. n. has been recorded as having a widespread tropical distribution; however, we question the likelihood of this and warrants further study derived from both morphological and molecular evidence. Discussion The relationships within Amphinomida and its relationship to other Annelida are poorly understood, though their monophyly is not questioned (Rouse & Pleijel 2001; Roussset et al. 2007; Wiklund et al. 2008). Although this study provides a limited view of amphinomid relationships, our intention was to establish a working framework for continued elucidation of Amphinomidae systematics that explored the utility of both nuclear and mitochondrial gene data. The phylogenetic evidence supports the presence of a paraphyletic Amphinomidae (Wiklund et al. 2008; this study). The latter being strongly divided into a ‘fusiform’ clade, including Archinome (Archinomidae) and Chloeia (Amphinomidae) and a ‘rectilinear’ clade (Amphinomidae). At this time, the higher-level status of Amphi14 nomidae and Archinomidae can only be reconciled under an expanded taxonomic and phylogenetic scope and will be address elsewhere (Borda et al., in preparation). Thus, for the purposes of this study, we distinguished ‘fusiform’ versus ‘rectilinear’ amphinomids as well-supported Clades I and II, respectively. Phylogenetic affinities of Cryptonome The phylogenetic hypotheses supported Cryptonome conclava as being allied to Amphinome and ⁄ or Hipponoa and that this new species should be recognized as a new genus of Clade II. This designation was also reflected in analyses with an expanded taxonomic sampling (Borda et al., in preparation). Although Cryptonome is divergent in caruncular morphology from both Amphinome and Hipponoa (caruncle absent), this clade was supported by molecular evidence and is corroborated by the shared possession of chaetae that are internally solid and resilient. The latter feature contrasts the externally brittle chaetae and variably developed medullae of most other amphinomid taxa (Gustafson 1930). While members of all three genera possess solid neurochaetae, they are non-retractile in both Cryptonome and Amphinome and retractile in Hipponoa (Day 1967; Kudenov 1994a). The reproductive biology and larval stages of amphinomids have not been studied extensively and records are scarce (Allen 1957; Kudenov 1974, 1977; Pernet et al. 2001). The few studies that are available indicate that amphinomids exhibit a diversity of reproductive strategies and behaviours (Kudenov 1974; Schroeder & Hermans 1975; Wilson 1991; Yáñez-Rivera & Salazar-Vallejo 2011). However, the reproductive biology of the Euphrosinidae and genera of Clade I is essentially unknown. An interesting feature of Clade II, and manifested in Cryptonome conclava, is evidence of architomy, or asexual reproduction whereby worms fragment following megasepta formation, and fragments regenerate heads and ⁄ or tails (Kudenov 1974). Such an alternative strategy of asexual reproduction is characteristic of species belonging to several Clade II genera including: Eurythoe (Kudenov 1974), Hermodice (Yáñez-Rivera & Salazar-Vallejo 2011), Pareurythoe (Kudenov 1974), Amphinome (Kudenov, personal observation) and now Cryptonome. It is unknown if Hipponoa exhibits architomy. Though, it has been described as a protandric hermaphrodite (Kudenov 1977). Both Amphinome and Hipponoa have been observed to exhibit parental care by brooding behaviour of juveniles, which has not been observed or reported in Cryptonome. All Cryptonome conclava specimens examined in this study were clearly undergoing regeneration of anterior and ⁄ or posterior chaetigers and one simultaneously exhibited coelomic oocytes. Given this species’ apparent opportunistic ability to exploit temporally restricted resources (e.g. food, deep-sea habitats), ª 2012 The Authors d Zoologica Scripta ª 2012 The Norwegian Academy of Science and Letters E. Borda et al. a reproductive strategy that enables both sexual and asexual reproduction seems highly appropriate. Key to Cryptonome species and genera of Clade II 1 Caruncle not present; median ciliated nuchal organ present; neuropodia arising from ventral body surfaces; neurochaetae retractile.........................................Hipponoa Caruncle present, variably developed; median ciliated nuchal organ absent; neuropodia arising from lateral body surfaces; neurochaetae non-retractile ................2 2 Chaetiger 1 dorsally continuous, not complete ............. 3 Chaetiger 1 dorsally discontinuous, not incomplete ...4 3 Stout, distally curved hooks present in notopodia of chaetiger 1; caruncle round, sessile, without free lateral wings; neurochaetae not unidentate; harpoon notochaetae with 1 row of barb ..................... Paramphinome Stout, distally curved hooks not present in notopodia of chaetiger 1; caruncle stalked, broadly triangular to chordate with free lateral wings; neurochaetae unidentate; harpoon notochaetae with up to 5 rows of barbs .................................................. Amphinome 4 Caruncle large and conspicuous, extending beyond one external chaetiger posteriorly..........................................7 Caruncle small and inconspicuous, not extending beyond one external chaetiger posteriorly ............ Cryptonome................................................................. 5 5 Bifurcate capillary notochaetae present....................... 6 Bifurcate capillary notochaetae not present ..........................................................Cryptonome turcica comb. n. 6 Bifurcate capillary notochaetae finely denticulate, present in chaetigers 2–44; junction angle between long and short prongs 8...........................Cryptonome conclava sp. n. Bifurcate capillary notochaetae coarsely denticulate, present in chaetigers 1–23; junction angle between long and short prongs 12 in chaetigers 1–2, and 24 in chaetigers 3–23................Cryptonome parvecarunculata comb. n. 7 Caruncle without a median lobe, with paired lateral lobes forming a complex monopodial-like pattern of bipinnate chevrons opening anteriorly…...............................Hermodice Caruncle with a smooth median lobe, with paired lateral lobes not forming a complex monopodial-like pattern of bipinnate chevrons opening anteriorly.... 8 8 Caruncle dorsoventrally bilobed, not sinusoidal; median lobe of caruncle thickened, pronounced, overlapping lateral lobes; lateral lobes recessed and convoluted; paired ciliated ridges running obliquely to longitudinal body axis............................................................... Eurythoe Caruncle not dorsoventrally bilobed, sinusoidal; median lobe of caruncle not thickened, not pronounced, not ª 2012 The Authors d Zoologica Scripta ª 2012 The Norwegian Academy of Science and Letters d Wood fall amphinomids and systematics overlapping lateral lobes; paired ciliated ridges running parallel to longitudinal body axis................. Pareurythoe Conclusions This study highlights the poor state of Amphinomida systematics. The lack of a sound phylogenetic framework for revisionary activities has left confused morphological terminologies, vague generic definitions and underestimates in amphinomid diversity, particularly among species found in the deep sea. Our goals were to provide a classification scheme that better reflects phylogenetic history, to clarify the taxonomic status of a genus that had not been revised in nearly 150 years (Amphinome) and discuss the historical confusion associated with the genera Amphinome, Eurythoe and Hipponoa (in part). This reasonably allowed for establishing Cryptonome as a new genus to represent some poorly known amphinomids. Although the diversity and monophyly of Cryptonome species was not evaluated here, we hope that continued exploration and experimentation of wood falls and other organic strata will provide additional specimens and insights into the distribution of Cryptonome species at depth. Lastly, this contribution also aimed to draw attention to the discoveries that continue to be made across chemoautotrophic ecosystems. Acknowledgements The authors would like to thank Antje Boetius for the deployment and help during the recovery of the wood experiments. We thank the captains and crews of the R ⁄ Vs Meteor and Pourquois Pas? and the teams operating ROVs Quest 4000 (MARUM, Bremen, Germany) and Victor 6000 (IFREMER, Toulon, France) for their technical support. We would also like to thank the following for generously contributing specimens and ⁄ or literature for this study: Gustav Paulay (University of Florida) for Amphinome rostrata (Mariana Is.) and Chloeia viridis, Fredrik Pleijel (University of Gothenburg) for Archinome storchi, Euphrosine foliosa and Chloeia flava and Beatriz Yáñez-Rivera (Universidad Nacional Autónoma de Mexico, Mazatlan) for Amphinome rostrata (Mexico), Kristian Fauchald and Linda Ward (Smithsonian National Museum of Natural History), Danny Eibye-Jacobsen (Danish Museum of Natural History), and Leslie Harris (Los Angeles County Museum of Natural History). Thanks to Harim Cha (Scripps Institution of Oceanography) for accessioning specimens into the SIO Benthic Invertebrate Collections. This study was funded by the projects GDRE DIWOOD (European Research Group CNRS-MPG), CHEMECO (ESF EURODEEP), HERMIONE (EC 7th PF No. 226354) and the Max Planck Institute for Marine Microbiology, with additional funding sources to EB (NSF DBI: 0706856, Encyclopedia of Life Rubenstein Fellowship, Society of Systematic Biologists Mini15 Wood fall amphinomids and systematics d E. Borda et al. PEET Award and Census of Marine Life Chemosynthetic Ecosystem Science Training AWards for New Investigators) and GWR (NSF OCE-0826254 and OCE-0939557). References Allen, M. J. (1957). Histochemical studies on developmental stages of polychaetous annelids. Anatomical Record, 128, 515– 516. Amaral, A. C. Z., Lana, P. C., Fernandes, F. C. & Coimbra, J. C. (2006). Catálogo das Espécies de Annelida Polychaeta do Brasil. Available at: http://www.ib.unicamp.br/projbiota/bentos_marinho/ prod_cien/texto_poli.pdf. Audzijonyte, A. & Vrijenhoek, R. C. (2010). 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