Zootaxa 3682 (4): 594–598 ISSN 1175-5326 (print edition) www.mapress.com/zootaxa/ Article ZOOTAXA Copyright © 2013 Magnolia Press ISSN1175-5334(online edition) http://dx.doi.org/10.11646/zootaxa.3682.4.12 http://zoobank.org/urn:lsid:zoobank.org:pub:77C3BFBB-68D4-476C-9894-1C8651207963 DNA sequences and morphological variation in Lophiodes iwamotoi Ho, Serét & Shao, 2011 based on new material from New Caledonia HSUAN-CHING HO1,2,4& WEI-JEN CHEN3 1National Museum of Marine Biology & Aquarium, Pingtung, Taiwan, no. 2, Houwan Rd., Checheng, Pingtung, Taiwan. E-mail:[email protected] 2Institute of Marine Biodiversity & Evolutionary Biology, National Dong Hwa University; No. 2, Houwan Rd., Checheng, Pingtung, Taiwan 3Institute of Oceanography, National Taiwan University, No.1, Sec. 4 Roosevelt Rd. Taipei 10617, Taiwan 4Corresponding author Abstract Iwamoto’s anglerfish Lophiodes iwamotoi is recorded from New Caledonia for the first time. Study of molecular features further support the validity of the species. Moloecular sequence data from the cytochrome c oxidase subunit-I and Rho- dopsin loci, along with morphological variation are provided, as well as information on its fresh coloration. Key words: Pieces, Teleotsei, Lophiiformes, Lophiidae, New Caledonia Introduction Hoet al. (2011) documented the species of the family Lophiidae from the South Pacific Ocean, recognizing seven valid species and describing two new species from Polynesia. One of these, Lophiodes iwamotoi Ho, Serét & Shao, 2011 was described on the basis of five specimens collected from the Savannah Seamount of Polynesia at depths of 706-710 m in 1977. These specimens, despite being in good condition, were strongly faded due to long-term preservation. Recently, the authors examined fish specimens collected during the 39-day exploratory cruise (campaign: EXBODI) by the R/V Alis deployed by the French Oceanographic Fleet on seamounts off New Caledonia, in which nine lophiid specimens were recognized. Among them, four are identified as L. iwamotoi. Of the remaining five, three were identified as L. mutilus (Alcock 1894) and two as Lophiomus setigerus (Vahl 1797). The purpose of the present work is to record this recently described species and its congeners represented in the New Caledonian ichthyofauna. A molecular comparison was undertaken to obtain further data regarding the validity of the species. We discuss this DNA sequence data as well as morphological variation in L. iwamotoi, and provide fresh color information on this species for the first time (Fig. 1). Material and methods Nine lophiidspecimenswere included in the morphological and molecular examination. Whole genomic DNA was extracted from fin clips or muscle tissue of specimens using the Qiagen DNeasy extraction kit (Qiagen, Valencia, CA) according to manufacturer's instructions. Fragments of two protein-coding genes (cytochrome c oxidase subunit-I [COI] and rhodopsin [Rhod]) located in mitochondrial and nuclear genomes, respectively, were amplified and sequenced for this study. Protocols for collecting molecular data follow those outlined in Ward et al. (2005) for COI and Chen et al. (2003) for Rhod. Collected DNA sequences were deposited in NCBI GenBank (see Fig. 2 for the accession numbers of the corresponding gene sequences). Compiled sequences were manually aligned based on the inferred amino acid translation using Se-Al v2.0a11 (available at http://tree.bio.ed.ac.uk/software/seal/). 594 Accepted by M.R. de Carvalho: 12 Jun. 2013; published: 1 Jul. 2013 FIGURE 1. Freshly caught specimens of Lophiodes iwamotoi. A. ASIZP 73486, 248 mm SL. B. ASIZP 73489, 247 mm SL. C. ASIZP 73488, 154 mm SL. Not to scale. LOPHIODES IWAMOTOI FROM NEW CALEDONIA Zootaxa 3682 (4) © 2013 Magnolia Press · 595 Methods for taking counts and proportional measurements generally followed Caruso (1981, 1983). Specimens examined are deposited in the fish collection of the Academia Sinica (ASIZP). Meristic and morphometric values of the four newly collected specimens compared to those of the type-series are provided in Table 1. TABLE 1. Morphomeric data of the type-series and four additional specimens of Lophiodes iwamotoi. Types New Caledonian specimens Standard length (mm SL) 169–226 (n=4) 145–248 (n=3) 76.7 (n=1) Proportion as %SL Head length (HL) 36.2–37.2 36.6–38.3 43.5 Illicial length 23.0–26.7 19.8–23.5 25.4 2nd dorsal-spine length 27.2–31.7 29.1–30.5 27.4 3rd dorsal-spine length 56.9–70.8 66.5–68.8 44.3 5th dorsal-spine length 10.5–13.1 13.0–17.5 10.7 Tail length 26.3–29.0 25.3–29.8 32.7 Proportion as % HL Head width 52.1–53.8 51.8–53.0 56.3 Head depth 65.8–67.9 62.4–66.7 65.0 Snout length 55.4–57.5 57.6–59.6 58.1 Snout width 19.2–22.5 19.3–23.3 19.5 Distance between inner sphenotic spines 41.1–42.5 40.4–41.7 45.2 Distance between posterior frontal spines 41.0–43.2 43.2–44.6 45.2 Distance between pterotic and sphenotic spines 19.2–20.5 18.4–20.3 19.5 Distance between quadrate and anterior palatine 67.1–72.6 64.5–68.8 50.9 spines Distance between opercular and subopercular spines 41.6–47.4 41.5–43.9 46.4 Results and discussion Among the species in the Lophiodes mutilus-species group (sensu Caruso, 1981), L. iwamotoi is closest to L. mutilus in having a long third dorsal-fin spine, a simple small escal bulb, and many tendrils on the third dorsal-fin spine. Several characters have been used to readily separate it from L. mutilus: 19–20 pectoral-fin rays (vs. 15–18; mostly 16), a much longer third dorsal-fin spine (56.9–70.8% SL vs. 36.6–63.9% SL) that reaches the caudal peduncle, but a relatively short fifth dorsal-fin spine (10.5–13.1% SL vs. 16.9–29.3% SL) that reaches the origin of the soft dorsal fin. The New Caledonian specimens agree well with the original description, except for the following variations. The inner frontal spine is present in all type-specimens and in the two smaller New Caledonian specimens, but is strongly reduced in two larger specimens. The same change of the inner frontal spine morphology was also observed in other members of the L. mutilus species group (Ho, pers. obs.), implying that an ontogenic change of this particular character might be common in Lophiodes species. The inner sphenotic spines are strongly reduced in the 248 mm specimen. The illicium is much shorter in the 248 mm specimen, only 19.8% SL (vs. 23.0–26.7% SL in type series), whereas that of the 226 mm holotype is 26.7% SL. The length of the illicium in the 154 mm specimen is also below the average value for the type-series. No trend in growth in the spine is observed. The 248 mm specimen has a relatively long fifth dorsal-fin spine at 17.5% SL (vs. 10.5–13.1% SL in the type-series). However, the spine reaches the origin of the dorsal fin when fully laid back, identically to the type-series. Several proportional values are different in the 76.7 mm specimen: it has a longer head (43.5% SL), a wider head (56.5% HL), a shorter 3rd dorsal-fin spine (44.3% SL) that reaches only the middle portion of dorsal-fin base, a longer tail (32.7% SL), a greater distance between inner sphenotic spines (45.2% HL), a greater distance between 596 · Zootaxa 3682 (4) © 2013 Magnolia Press HO & CHEN frontal spine (45.2% HL), and a smaller distance between quadrate and anterior palatine spines (50.9% HL). However, it has 19 pectoral-fin rays and similar coloration with the other three specimens; its molecular features also support our identification. All these differences may be attributed to changes in growth. FIGURE 2. Polymorphic nucleotide sites at the cytochrome c oxidase subunit-I locus (A) and Rhodopsin locus (B) in two Lophiodes species collected from seamounts off New Caledonia. Diagnostic nucleotides of L. iwamotoi to L. mutilus are highlighted. Numeration of nucleotide sites starts from first nucleotide of the corresponding genes. The sequences used for the examination were deposited in NCBI Genbank. Their accession numbers are given. Few knobs are found on the dorsal surface of the humeral spine in the type-series, but three blunt spines are present on the two smaller specimens and absent in two larger specimens. Ho et al. (2011) mentioned that “although the humeral spines are quite variable in most species of Lophiodes, ranging from simple to multifid, all specimens in the type series of L. iwamotoi have a simple humeral spine with few low knobs on it.” We confirm that L. iwamotoi has a simple humeral spine, especially in larger specimens. Arrangement and form of other spines on the head bones in the New Caledonian material are as recorded in the originally described specimens of the species. Figure 1 shows the coloration of the freshly caught New Caledonian specimens. The 248 mm specimen (Figs. 1A–B) is uniformly light brown dorsally and pale gray ventrally, with a pale posterior pectoral-fin margin. The LOPHIODES IWAMOTOI FROM NEW CALEDONIA Zootaxa 3682 (4) © 2013 Magnolia Press · 597 remaining specimens have many small irregular black patches on a light brown background; these patches faded after preservation. Distribution of L. iwamotoi is now extended westward from the Society Islands to southeastern New Caledonia, and the depth range is extended to 460–780 m. From the sample collection data and molecular evidence from both mitochondrial and nuclear gene markers (see below) (Fig. 2), the two Lophiodes species are suggested to be genetically and reproductively isolated, living sympatrically at least on seamounts of the South Durand Bank. It is notably that one of our Lophiomus setigerus specimens was collected at a depth of 815–970 m, which represents the deepest record for the species. No nucleotide insertions or deletions were found in our aligned sequences for the samples of the two Lophiodes species. Fifty-one and 12 variable sites were observed along the 630 bp and 828 bp sequenced fragments for the COI and Rhod loci, respectively. This represents 7.82% and 0.84 % of interspecific nucleotide divergence (evaluated by uncorrected pairwise p-distance) from COI and Rhodopsin genes. A few single nucleotide polymorphism sites (SNPs) were detected among the samples of the same species (Fig. 2). In addition to morphological characters, the two Lophiodes species co-occuring in New Caledonian waters can be unambiguously distinguished from each other by four and more diagnostic nucleotides along the fragment of the two genes sequenced (Fig. 2). Specimens examined. Lophiodes iwamotoi: ASIZP 73486 (248 mm), stn. EXB 097, CP3853, Banc Sud Durand, 22°18'S, 168°46'E, 692 m, 14 Sep. 2011. ASIZP 73487 (76.7 mm), stn. EXB 111, CP3854, Banc Sud Durand, 22°18'S, 168°45'E, 570 m, 14 Sep. 2011. ASIZP 73488 (154 mm), EXB 158, stn. CP3864, Banc de L'Orne/Walpole, 22°22'S, 168°57'E, 460–708 m, 15 Sep. 2011. ASIZP 73489 (247 mm), stn. EXB 182, CP3871, Banc Ellet, 22°53'S, 169°25'E, 580–780 m, 16 Sep. 2011. Lophiodes mutilus: ASIZP 73483 (107 mm SL) and ASIZP 73484 (56.3 mm), stn. EXB 088, CP3852, Banc Sud Durand, 22°17'S, 168°43'E, 582 m, 14 Sep. 2011. ASIZP 73485 (177 mm), stn. EXB 110, CP3854, Banc Sud Durand, 22°18'S, 168°45'E, 570 m, 14 Sep. 2011. Lophiomus setigerus: ASIZP 73490 (121 mm), stn. EXB532, CP3834, Au large passe Yaté, 22°06'S, 167°04'E, 257–258 m, 9 Sep. 2011. ASIZP 73491 (235 mm), stn. EXB538, CP3844, Au large passe de la sarcelle, 22°20'S, 167°22'E, 815–970 m, 10 Sep. 2011. Acknowledgements Our gratitude goes to the crews of the R/V Alis and participants of the oceanographic cruise (campaign: EXBODI; PIs, Sarah Samadi and Laure Corbari) involved in organizing the survey and the capture of the samples. We wish to thank J.-N. Chen, C.-J. Chen, C.-H. Huang, and H. Lee for their efforts in sequencing. The EXBODI is supported by UMS Flotte Océanographique Française. This study is partly supported by the National Science Council to HCC (NSC 101-2621-B291-001), and to WJC (NSC 102-2923-B-002 -001 -MY3). References Caruso, J.H. (1981) The systematics and distribution of the lophiid anglerfishes: I. A revision of the genus Lophiodes, with the description of two new species. Copeia,1981, 522–549. http://dx.doi.org/10.2307/1444556 Caruso, J.H. (1983) The systematics and distribution of the lophiid anglerfishes: II. Revisions of the genera Lophiomus and Lophius.Copeia, 1983, 11–30. http://dx.doi.org/10.2307/1444694 Chen, W.-J., Bonillo, C. & Lecointre, G. (2003) Repeatability of clades as criterion of reliability: a case study for molecular phylogeny of Acanthomorpha (Teleostei) with larger number of taxa. Molecular Phylogenetics and Evolution,26, 262– 288. http://dx.doi.org/10.1016/s1055-7903(02)00371-8 Ho, H.-C., Séret, B. & Shao, K.-T. (2011) Records of anglerfishes (Lophiiformes: Lophiidae) from the western South Pacific Ocean, with descriptions of two new species. Journal of Fish Biology, 79, 1722–1745. http://dx.doi.org/10.1111/j.1095-8649.2011.03106.x Ward, R.D., Zemlak, T.S., Innes, B.H., Last, P.R. & Hebert, P.D. (2005) DNA barcoding Australia's fish species. Philosophical Transactions of the Royal Society B-Biological Sciences, 360, 1847–1857. http://dx.doi.org/10.1098/rstb.2005.1716 598 · Zootaxa 3682 (4) © 2013 Magnolia Press HO & CHEN