INTRODUCTION
Since the first encounter with hydrothermal vents on the Galápagos Rift was published in 1977 (Lonsdale, Reference Lonsdale1977; Corliss & Ballard, Reference Corliss and Ballard1977), scientific interest and effort in exploring deep-sea chemosynthetic ecosystems have remained very high. Chemosynthetic ecosystems are now known to manifest in a variety of ways and settings other than vents such as cold seeps, and organic falls and are considered to be widespread (Baker et al., Reference Baker, Ramirez-Llodra, Tyler, German, Boetius, Cordes, Dubilier, Fisher, Levin, Metaxas and McIntyre2010). The discovery of the Lost City hydrothermal field illuminated the presence of serpentine-hosted chemosynthetic systems (Kelley et al., Reference Kelley, Karson, Blackman, Früh-Green, Butterfield, Lilley, Olson, Schrenk, Roe, Lebon and Rivizzigno2001, Reference Kelley, Karson, Früh-Green, Yoerger, Shank, Butterfield, Hayes, Schrenk, Olson, Proskurowski, Jakuba, Bradley, Larson, Ludwig, Glickson, Buckman, Bradley, Brazelton, Roe, Elend, Delacour, Bernasconi, Lilley, Baross, Summons and Sylva2005, Reference Kelley, Früh-Green, Karson and Ludwig2007). Serpentinization produces methane, CH4, the anaerobic oxidation of which results in the production of hydrogen sulphide, H2S, both of which are key to powering chemosynthetic ecosystems through chemosynthetic microbes (Kelley et al., Reference Kelley, Karson, Früh-Green, Yoerger, Shank, Butterfield, Hayes, Schrenk, Olson, Proskurowski, Jakuba, Bradley, Larson, Ludwig, Glickson, Buckman, Bradley, Brazelton, Roe, Elend, Delacour, Bernasconi, Lilley, Baross, Summons and Sylva2005; Ohara et al., Reference Ohara, Reagan, Fujikura, Watanabe, Michibayashi, Ishii, Stern, Pujana, Martinez, Girard, Ribeiro, Brounce, Komori and Kino2012). Unlike hydrothermal vents, the fluids of which are usually highly acidic (Van Dover, Reference Van Dover2000), the fluids from such serpentinite-hosted systems are often highly alkaline (Kelley et al., Reference Kelley, Karson, Früh-Green, Yoerger, Shank, Butterfield, Hayes, Schrenk, Olson, Proskurowski, Jakuba, Bradley, Larson, Ludwig, Glickson, Buckman, Bradley, Brazelton, Roe, Elend, Delacour, Bernasconi, Lilley, Baross, Summons and Sylva2005; Takai et al., Reference Takai, Moyer, Miyazaki, Nogi, Hirayama, Nealson and Horikoshi2005). Megafauna dependent on chemosynthesis have been reported to be common in such environments, although the details are little-known (Kelley et al., Reference Kelley, Früh-Green, Karson and Ludwig2007).
Gastropod molluscs comprise a major constituent of megafauna communities in various chemosynthetic ecosystems, and have received considerable taxonomic effort (reviewed in Warén & Bouchet, Reference Warén and Bouchet2001; Sasaki et al., Reference Sasaki, Warén, Kano, Okutani, Fujikura and Kiel2010). Gastropods in the superfamily Abyssochrysoidea currently assigned to the family Provannidae are restricted to chemosynthetic ecosystems (Johnson et al., Reference Johnson, Warén, Lee, Kano, Kaim, Davis, Strong and Vrijenhoek2010). The genus Provanna within it includes medium-sized snails (generally < 15 mm in shell height) and is a common endemic of such communities around the world (Sasaki et al., Reference Sasaki, Warén, Kano, Okutani, Fujikura and Kiel2010). They are deposit feeders grazing on bacteria and detritus, and produce lecithotrophic larvae (Warén & Bouchet, Reference Warén and Bouchet1986, Reference Warén and Bouchet1993; Levesque et al., Reference Levesque, Juniper and Limén2006; Sasaki et al., Reference Sasaki, Warén, Kano, Okutani, Fujikura and Kiel2010). It is the most species-rich genus of Abyssochrysoidea, currently with 22 described extant species (Johnson et al., Reference Johnson, Warén, Lee, Kano, Kaim, Davis, Strong and Vrijenhoek2010) and eight fossil species (Amano & Jenkins, Reference Amano and Jenkins2013; Amano & Little, Reference Amano and Little2014).
Serendipitously discovered in September 2010 in a submersible dive originally aimed to investigate the geology of the Southern Mariana Forearc, the ‘Shinkai Seep Field’ (SSF; Figure 1) is a serpentinite-hosted system located on a steep inner slope of the Mariana Trench, Southern Mariana Forearc (Ohara et al., Reference Ohara, Reagan, Fujikura, Watanabe, Michibayashi, Ishii, Stern, Pujana, Martinez, Girard, Ribeiro, Brounce, Komori and Kino2012). The dominant fauna is a large vesicomyid clam, Calyptogena (Abyssogena) mariana Okutani et al., 2013. In June 2015, further detailed investigations and sampling of the SSF was undertaken on the YK15-11 cruise of R/V Yokosuka. Biological samples were taken during the DSV Shinkai 6500 dive 1433, and numerous specimens of a rissoiform gastropod were collected. Taxonomic investigations which followed revealed that these specimens represented a highly characteristic Provanna species previously unknown to science. It is among the first Abyssochrysoid gastropod discovered from serpentinite-hosted ecosystems along with a Desbruyeresia species from South Chamorro Seamount (Chen et al., Reference Chen, Ogura, Hirayama, Watanabe, Miyazki and Okutani2016). Here this new species is formally described and named, as Provanna cingulata n. sp., and the bathymetric distribution range of the genus Provanna is discussed.
Location of the Shinkai Seep Field (SSF): (A) index map indicating the SSF by an asterisk; (B) detailed bathymetry of the SSF area (after Ohara et al., Reference Ohara, Reagan, Fujikura, Watanabe, Michibayashi, Ishii, Stern, Pujana, Martinez, Girard, Ribeiro, Brounce, Komori and Kino2012). Contours in 20 m intervals.
MATERIALS AND METHODS
Sample collection
All specimens of the new Provanna were collected from the SSF by the DSV Shinkai 6500 dive 1433 during cruise YK15-11. Upon recovery onto R/V Yokosuka, the specimens were immediately placed into 99% ethanol to dehydrate for preservation and storage.
Morphology
Morphological investigation and dissection were carried out under an Olympus SZX9 dissecting microscope. The radula was dissected and protoconch was illustrated from specimens preserved in 99% ethanol. Scanning electron microscopy (SEM) and shell morphometric measurements were carried out as in Chen et al. (Reference Chen, Ogura, Hirayama, Watanabe, Miyazki and Okutani2016).
Type specimens are deposited in the University Museum, the University of Tokyo (UMUT), the American Museum of Natural History, New York City (AMNH), the National Museum of Nature & Science, Tsukuba (NSMT) and Japan Agency for Marine-Earth Science and Technology (JAMSTEC).
Genetics
Two specimens of the provannid from the SSF were sequenced for the barcoding gene cytochrome oxidase c subunit I (COI). In addition, COI of one specimen of Provanna shinkaiae Okutani & Fujikura, Reference Okutani and Fujikura2002 from the JAMSTEC collection (No. 051346–051358, 70% ethanol, from methane seep, Japan Trench, 39°6.432′N 143°53.478′E, 5352 m, ROV KAIKO Dive #258, R/V Kairei cruise KR02-09) was also sequenced for comparison. DNA extraction and quality checks were performed as in Chen et al. (Reference Chen, Ogura, Hirayama, Watanabe, Miyazki and Okutani2016). The COI region was amplified with the primer pairs LCO1490 and HCO2198 (Folmer et al., Reference Folmer, Black, Hoeh, Lutz and Vrijenhoek1994) as well as Pg501L (5′-TATACGATGACGGGGAATGC-3′) and Pg1253R (5′-TGTTGAGGAAAGAAAGTAATATTAA-3′) (Ogura et al., unpublished). The polymerase chain reaction was carried out in 20 µl reactions, including 1 µl DNA template (15–30 ng μl−1), 1 µl each of forward and reverse primers (10 µM), 1.6 µl dNTP mixture (TaKaRa Bio, Japan), 2 µl 10 × buffer, TaKaRa Ex Taq DNA polymerase solution (TaKaRa Bio, Japan), and 13.25 µl double-distilled water. A Veriti 200 Thermal Cycler (Applied Biosystems) was used for thermo cycling. The protocol used was: 95°C for 1 min followed by 35 cycles of [95°C for 15 s, 40°C for 15 s, 72°C for 30 s], ending with 72°C for 7 min. Amplification was confirmed with 1.4% agarose gel electrophoresis using ethidium bromide. ExoSAP-IT (Affymetrix) was used following standard protocols to purify successful PCR products. Details of cycle sequencing reaction and purification are as listed in Chen et al. (Reference Chen, Ogura, Hirayama, Watanabe, Miyazki and Okutani2016). Sequences were resolved from precipitated products using Applied Biosystems 3130xl DNA sequencer.
The complementary sequences from the forward and reverse primers were aligned to check the sequencing accuracy using a software Alignment Explorer mounted on the software package MEGA 6 (Tamura et al., Reference Tamura, Stecher, Peterson, Filipski and Kumar2013). Phylogenetic analyses were carried out using the same package using the resulting sequences plus the abyssochrysoid COI sequences available on GenBank. Sequences of the whelk Neptunea amianta (Dall, 1890), N. antiqua (Linnaeus, 1758), and the periwinkle Littorina littorea (Linnaeus, 1758) from distantly related gastropod groups were included as outgroup taxa (after Johnson et al., Reference Johnson, Warén, Lee, Kano, Kaim, Davis, Strong and Vrijenhoek2010).
The Model Selection (ML) program in MEGA 6 was applied to the dataset to select the most suitable nucleotide substitution model, which was HKY + G + I. Then the maximum-likelihood (ML) tree was generated using this model also in MEGA 6. The ML tree was bootstrapped 2000 times. Restricted by the length of some shorter sequences on GenBank, the sequence length used in the final phylogenetic analyses was 411 bp. New sequences generated from this study are deposited in DNA Data Bank of Japan (DDBJ) under the accession numbers LC094443, LC094444 and LC095875.
SYSTEMATICS
Clade CAENOGASTROPODA Cox, 1960
Superfamily ABYSSOCHRYSOIDEA Tomlin, Reference Tomlin1927
Family PROVANNIDAE Warén & Ponder, Reference Warén and Ponder1991
Genus Provanna Dall, 1918
Provanna cingulata n. sp.
(Figures 2–4)
Provanna cingulata n. sp.: (A–D) holotype, shell height 11.0 mm (UMUT RM32589); (E–F) paratype #1, shell height 11.4 mm (AMNH_IZC 250202); (G–H) paratype #3, shell height 14.3 mm (NSMT-Mo 78975); (I–L) paratype #4, shell height 16.5 mm (UMUT RM32590). Scale bars: A–L = 2 mm.
Provanna cingulata n. sp.: (A) early whorls of a juvenile specimen (paratype #4, UMUT RM32591); (B–C) protoconch, white arrow in (B) indicates boundary between the protoconch and the teleoconch; (D) operculum. Scale bars: A, D = 500 µm; B = 200 µm; C = 50 µm.
Provanna cingulata n. sp., radula: (A) middle section of an adult radula ribbon; (B) Most anterior section of the same adult radula ribbon, showing obvious dental wear; (C) a single marginal teeth. Scale bars: A–B = 50 µm; C = 20 µm.
ZOOBANK REGISTRATION
urn:lsid:zoobank.org:act:F7E7D074-0924-4F4B-9980-3CE3A1E144EE
TYPE LOCALITY
Shinkai Seep Field, 11°39.3652′N, 143°02.8786′E, 5687 m in depth. Collected during the R/V Yokosuka cruise YK15-11, DSV Shinkai 6500 Dive 1433, 2015/7/15, by Tomoyo Okumura.
TYPE MATERIAL
Holotype: Shell height (SH) 11.0 mm, shell width (SW) 6.8 mm, live collected, 99% ethanol, Figure 2A–D (UMUT RM32589).
Paratypes: #1 (Figure 2E–F): SH 11.4 mm, SW 7.3 mm, live collected, 99% ethanol (AMNH_IZC 250202). #2: SH 8.8 mm, SW 5.3 mm, live collected, 99% ethanol (NSMT-Mo 78974). #3 (Figure 2G, H): SH 14.3 mm, SW 10.1 mm, dead collected shell only, 99% ethanol (NSMT-Mo 78975). #4 (Figure 2I–L): SH 16.5 mm, SW 11.0 mm, dead collected shell only, 99% ethanol (UMUT RM32590). #5: Juvenile specimen, dead collected with broken aperture, SH 3.6 mm, SW 2.3 mm, dried and mounted for protoconch SEM (UMUT RM32591). #6: Two live collected specimens, dissected for radula and a section of foot taken for DNA, 99% ethanol (UMUT RM32592). #7: Five intact specimens in 99% ethanol (JAMSTEC 1150051773).
All type materials originate from the type locality with identical collection data (original accession number for the lot is 12541-15002-6 K#1433-B06).
FURTHER MATERIAL EXAMINED
There were 18 further specimens collected together from the type locality, which were also examined. Of these, eight were live collected, two were dead collected, and eight specimens were only shell fragments (Table 1).
Collection conditions and shell measurements of all available specimens (both type series and non-types).
DIAGNOSIS
A large Provanna up to 16.5 mm in shell height with rounded whorls and surface sculpture consisting of numerous spiral keels, lacking axial sculpture except for part of protoconch.
DESCRIPTION
Shell (Figure 2). Rissoiform, about 5.5 whorls. Large sized for the genus, up to 16.5 mm in height. Protoconch (Figure 3B, C) preserved in juveniles, slightly corroded in adults. Only protoconch I present, protoconch II lacking. White, about 1.5 whorls and 0.6 mm in height. Earlier part generally smooth with evenly dispersed fine granulation. Granules gradually becoming to disappear, continually replaced by axial ribbing. Teleoconch whorls rather inflated, rounded, convex. Suture moderately deep, impressed. Surface sculpture consists of spiral cords, increasing in number with growth, cross section always triangular. Very earliest teleoconch smooth but two prominent spiral cords emerging after 0.5 whorls and a third emerging from interspace after another whorl (Figure 3A). Lower of two initial cords especially sharp and strong throughout all growth stages. Penultimate whorl with eight to nine spiral cords above suture, weak and strong cords alternating. Strength of spiral cords increasing with growth. On body whorl of adults most cords of equal strength, only most peripheral one significantly stronger. Spiral cords present in close proximity on base, decreasing in strength anteriorly. Cord numbers increasing with growth (8–12 in specimens investigated). No axial ribbing or clearly visible growth lines present on teleoconch. Aperture rounded, slightly taller than wide. Outer-lip sharp, not thickened. Anterior siphonal notch shallow but distinct. Inner lip simple and smooth with no plicae or extension of parietal callous. Columella straight. No umbilical opening present. Ostracum thin, white, partly or completely corroded in body whorl leaving only periostracum layer. Periostracum golden brown in colouration, decreasing in intensity towards apex.
Operculum (Figure 3D). Paucispiral, nucleus eccentric, a little more than 3.5 volutions. Thin, semi-transparent, corneous. Yellowish-brown in colouration. Oval shaped, bluntly pointed at top.
Radula (Figure 4A). Taenioglossate, formula 2 + 1 + 1 + 1 + 2. Teeth solid. Central tooth triangular with one single strong cusp having a triangular cutting edge, laterally supported on both sides and weakly in the centre. Lateral teeth laterally thickened, with two to three moderately strong inner cusps increasing in size outwards, one strong central cusp, and two to three very weak outer cusps. A weakly raised ridge present under outer lateral cusps. Variation in cusp numbers seen between rows, even within single radular membrane. Marginal teeth (Figure 4C) moderate in length, marginals truncated apically. Distal end evenly serrated into well-separated denticles, numbering ~15 in inner marginals and ~18 in outer. Shafts carry very fine, shallow serrations on outer side near distal end.
Soft parts. Head-foot simple with two cephalic tentacles of equal length. No penis or neck furrow observed. Position of eyes indicated by an unpigmented bulge near base of cephalic tentacles. Gill monopectinate, not hypertrophied. Epipodial tentacles lacking, both anterior and posterior pedal glands present. All available material were fixed and preserved in 99% ethanol, tissues are thus brittle and bleached, preventing further detailed anatomical investigations.
DIMENSIONS
Largest known specimen 16.5 mm in shell height (Paratype #4) but with much corroded apex; maximum size is estimated to be up to 20 mm. Shell measurements for all available specimens are shown on Table 1.
COMPARATIVE REMARKS
Provanna cingulata n. sp. is most similar to Provanna macleani Warén & Bouchet, Reference Warén and Bouchet1989 from wood falls of the East Pacific (Johnson et al., Reference Johnson, Warén, Lee, Kano, Kaim, Davis, Strong and Vrijenhoek2010), which has very strong, numerous spiral cords like P. cingulata n. sp. and although axial sculpture is present it is very weak and indistinct (Warén & Bouchet, Reference Warén and Bouchet1989). The periostracum colouration of both species is yellowish brown. There are, however, more numerous spiral cords which are more closely spaced in P. cingulata n. sp. compared with P. macleani (7–8 above suture vs 6 above suture, 8–12 basal ribs vs 5 basal ribs). Furthermore, the radula of P. macleani is very different from that of P. cingulata n. sp. in having a reduced and membranaceous central tooth (Warén & Bouchet, Reference Warén and Bouchet1989), as the central tooth of P. cingulata n. sp. is prominent and solid. Provanna macleani is also much smaller in size with specimens only reaching 7.1 mm (Warén & Bouchet, Reference Warén and Bouchet1989).
Provanna cingulata n. sp. may also confused with young specimens of Provanna reticulata Warén & Bouchet, Reference Warén and Bouchet2009 from seeps off West Africa. Although the adult shells of P. reticulata have strong axial ribbing lacking in P. cingulata n. sp., in addition to spiral ribbing, and therefore easily separable, the young shells only have spiral ribbing (Warén & Bouchet, Reference Warén and Bouchet2009, figure 10I) and are very similar to those of P. cingulata n. sp. The radula differs slightly between the two species, however, with regards to the inner cusps of laterals. In P. reticulata there are always two inner cusps, whereas in P. cingulata n. sp. there are two or three (variable among rows within same specimen). Otherwise the juveniles of the two species are difficult to separate.
Except these two, Provanna cingulata n. sp. differs from the other described species of Provanna by having the sculpture consisting entirely of spiral cords and completely lacking in axial ribbing. Furthermore, its size when fully grown is unusually large for the genus.
The shell of Provanna cingulata n. sp. bears a certain resemblance to Cordesia provannoides Warén & Bouchet, Reference Warén and Bouchet2009 but it lacks the characteristic penis. Provanna cingulata n. sp. also has radula and protoconch characteristic of genus Provanna, which significantly differs from those of Cordesia (Warén & Bouchet, Reference Warén and Bouchet2009). Therefore we can safely exclude the possibility of the new species being a member of Cordesia and place it in the genus Provanna with good confidence.
Only specimens up to 11.4 mm shell height (Paratype #2) were collected alive, all large specimens were dead collected. In such dead collected specimens the apex was always corroded with only about four whorls remaining, although they were also clearly corroded from the aperture inwards indicating they may have been dead for quite a while. If the protoconch is intact when alive, the largest specimens could perhaps reach 20 mm in shell length. The vast majority of the live collected specimens had a well-preserved apex.
DISTRIBUTION
So far only known from the type locality, the Shinkai Seep Field.
ETYMOLOGY
‘Cingulatus’ (Latin) meaning girdled or belted, referring to the numerous spiral cords prominently girdling the whorls. Used as an adjective.
MOLECULAR PHYLOGENY
Figure 5 shows the phylogenetic tree produced by ML (HKY + G + I) method, clearly showing that P. cingulata n. sp. is nested within a monophyletic clade with other species currently classified in the genus Provanna. All other currently recognized abyssochrysoid genera are also grouped together as monophyletic clades. This result supports the morphological characteristics in placing P. cingulata n. sp. in the genus Provanna. According to the resulting tree, of those species whose COI sequences are available on GenBank, Provanna cingulata n. sp. is closest related to an undescribed Provanna species from Beebe vent field on the Mid-Cayman Spreading Centre represented by a single COI sequence on GenBank (‘Provanna sp. SP-2014’ from Plouviez et al., Reference Plouviez, Jacobson, Wu and Van Dover2015). The two specimens of P. cingulata n. sp. sequenced differed only by 0.16% pairwise distance in their COI sequence (1197 bp), while their pairwise distance from the undescribed Mid-Cayman Provanna was 5.4–5.8%. Provanna cingulata n. sp. was genetically distinct from P. macleani, the morphologically most similar described species, with the two separated by the undescribed Provanna from Mid-Cayman.
Phylogenetic tree of Abyssochrysoidea based on 411 bp of the COI gene, showing the systematic position of Provanna cingulata n. sp. within the genus Provanna. Node values are ML (HKY + G + I) bootstrap values. GenBank accession numbers of the COI sequences used are shown in parentheses.
DISCUSSION
Provanna cingulata n. sp. co-occurs with several other gastropod species in the SSF, including Bayerius arnoldi (Lus, 1981), a trochoid and a xylodisculid. However, this is the only gastropod collected from the SSF that belongs to an obligatory chemosynthetically associated group (Sasaki et al., Reference Sasaki, Warén, Kano, Okutani, Fujikura and Kiel2010). Provanna cingulata n. sp. is thus likely to require chemosynthetically influenced habitats for survival; like Calyptogena (Abyssogena) mariana from the same seep field. It is also among the first definitive records of the superfamily Abyssochrysoidea from serpentinite-hosted chemosynthetic ecosystems, in addition to a recently described species of Desbruyeresia (Chen et al., Reference Chen, Ogura, Hirayama, Watanabe, Miyazki and Okutani2016). As the gill of P. cingulata n. sp. is not hypertrophied, it is unlikely to house endosymbiotic chemosymbionts. The radula ribbon shows significant and obvious wear in the anterior section (Figure 4B) compared with a more posterior, pristine, section (Figure 4A), strongly suggesting a deposit feeder which uses the radula to graze bacteria and perhaps also other particles. The same has been suggested for other Provanna species, for example P. variabilis Warén & Bouchet, Reference Warén and Bouchet1986 (Warén & Bouchet, Reference Warén and Bouchet1993); and all Provanna species are presumed to have similar feeding habits (Sasaki et al., Reference Sasaki, Warén, Kano, Okutani, Fujikura and Kiel2010). The well-preserved protoconch in P. cingulata n. sp. is virtually identical to that of the Provanna spp. protoconch illustrated in Warén & Ponder, Reference Warén and Ponder1991 (Figure 1), and is indicative of lecithotrophic development without a planktotrophic stage (Sasaki et al., Reference Sasaki, Warén, Kano, Okutani, Fujikura and Kiel2010).
With a maximum shell height of 16.5 mm and estimated to reach 20 mm if the apex is intact, P. cingulata n. sp. is the largest Provanna species so far known. Of the 25 described recent and fossil Provanna species, the largest is P. reticulata Warén & Bouchet, Reference Warén and Bouchet2009 from West Africa seeps which reaches a shell height of 14.0 mm (see Table 1 in Amano & Little, Reference Amano and Little2014). Another large Provanna species from the Antarctic hydrothermal vents of East Scotia Ridge (Rogers et al., Reference Rogers, Tyler, Connelly, Copley, James, Larter, Linse, Mills, Garabato, Pancost, Pearce, Pulunin, German, Shank, Boersch-Supan, Aker, Aquilina, Bennett, Clarke, Dinley, Graham, Green, Hawkes, Hepburn, Hilário, Huvenne, Marsh, Ramirez-Llodra, Reid, Roterman, Sweeting, Thatje and Zwirglmaier2012) is currently under description, but that species only reaches 15.0 mm in shell height (Katrin Linse, pers. comm.).
Furthermore, Provanna cingulata n. sp. was collected from 5687 m deep, which is so far the deepest occurrence record of not only Provanna but also the whole superfamily Abyssochrysoidea. Prior to the discovery of P. cingulata n. sp., the deepest abyssochrysoids known were P. abyssalis Okutani & Fujikura, Reference Okutani and Fujikura2002 and P. shinkaiae from methane seeps on the landward slope of Japan Trench (Okutani & Fujikura, Reference Okutani and Fujikura2002; Fujikura et al., Reference Fujikura, Okutani and Maruyama2012). Provanna abyssalis was collected from a depth of 5379 m and P. shinkaiae from 5343 m. These are followed by a yet-undescribed species of Provanna from the Beebe hydrothermal vent field of the Mid-Cayman Spreading Centre, the deepest hydrothermal vent field associated with a spreading centre on earth (German et al., Reference German, Bowen, Coleman, Honig, Huber, Jakuba, Kinsey, Kurz, Leroy, McDermott, deLepinay, Nakamura, Seewald, Smith, Sylva, Van Dover, Whitcomb and Yoerger2010), reaching the depth of 4966 m (Connelly et al., Reference Connelly, Copley, Murton, Stansfield, Tyler, German, Van Dover, Amon, Furlong, Grindlay, Hayman, Huhnerbach, Judge, Le Bas, McPhail, Meier, Nakamura, Nye, Pebody, Pedersen, Plouviez, Sands, Searle, Stevenson, Taws and Wilcox2012; Plouviez et al., Reference Plouviez, Jacobson, Wu and Van Dover2015). All other abyssochrysoids have bathymetric ranges reaching less than 4000 m. The discovery of P. cingulata n. sp. thus extends the bathymetric range of Abyssochrysoidea by more than 300 m.
Interestingly, although P. cingulata n. sp. is closely related to the Mid-Cayman species, these two are not closely related to P. shinkaiae which was also included in the phylogenetic analyses. This rejects the possibility of a single deep clade within Provanna, and the adaptation to living in very deep habitats probably evolved more than once in the genus. To further elucidate and consolidate the evolutionary history of Provanna species, multi-gene phylogenetic analyses with divergence time constraints including these species should be carried out in the future to be added to the current knowledge as presented by Johnson et al. (Reference Johnson, Warén, Lee, Kano, Kaim, Davis, Strong and Vrijenhoek2010).
ACKNOWLEDGEMENTS
The authors would like to thank the Captain and crews of the JAMSTEC RV ‘Yokosuka’, the DSV ‘Shinkai 6500’ team, as well as the scientific party on-board the expedition YK15-11 for their tireless support of the scientific activity during the cruise. Sincere gratitude is directed to Ms Ryoko Yamazaki (JAMSTEC) for her great help in obtaining genetic sequences of the new Provanna species. The Shinkai Seep Field is within the Mariana Trench Marine National Monument of the USA and our study was done under the special use permit #12541-15002. We thank the US Fish and Wildlife Service for approval of our study in the monument.
FINANCIAL SUPPORT
The genetic work in the present study was funded by a Japan Society for the Promotion of Science (JSPS) KAKENHI grant (No. 15K18602) awarded to Hiromi Kayama Watanabe.
