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New record of two marine ulvalean species (Chlorophyta) in Korea
New record of two marine ulvalean species (Chlorophyta) in Korea
Journal of Ecology and Environment. 2014. Nov, 37(4): 379-385
Copyright © 2014, The Ecological Society of Korea
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial Licens (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
  • Received : October 02, 2014
  • Accepted : November 04, 2014
  • Published : November 28, 2014
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About the Authors
Seung Hee Lee
Pil Joon Kang
Ki Wan Nam
kwnam@pknu.ac.kr

Abstract
Two marine ulvalean species (Chlorophyta) were collected from the southern and eastern coasts of Korea. One is morphologically characterized by usually unbranched or little branched ribbon-shaped thalli, fronds with spirally twisted basal portions and usually undulate margins and 1 (-2) pyrenoids per cell. The other has irregularly shaped thalli, undulate and dentate margins in the fronds, small macroscopic denticulations along the margin and (1-) 2-3 pyrenoids per cell. In phylogenetic tree based on molecular data, the two species nest in the same clade with Ulva flexuosa and U. rigida , respectively. These two Korean entities are identified as U. flexuosa and U. rigida , respectively, based on morphological and molecular analyses. This is the first record of Ulva flexuosa and U. rigida in Korea
Keywords
INTRODUCTION
Ulva Linnaeus, which is a large genus within Ulvaceae J.V. Lamouroux ex Dumortier, includes about 100 species worldwide ( Guiry and Guiry 2014 ). This genus is found in all aquatic habitats from freshwater to seawater environments ( Loughnane et al. 2008 ). In the past, the taxonomy of Ulva had been based mainly on morphology ( Bliding 1963 , 1968 , Koeman and van den Hoek 1981 , Hoeksema and van den Hoek 1983 ). However, this makes it difficult to identify Ulva species, because of the simple thallus structure ( Heesch et al. 2009 ). Recently, molecular analysis, which is an useful tool for differentiating taxa, has been adopted in the identification, taxonomy and phylogeny of species within this genus ( Malta et al. 1999 , Tan et al. 1999 , Hayden et al. 2003 , Loughnane et al. 2008 , Hofmann et al. 2010 ). Many synonyms reported in Ulva have been the result of previous molecular analyses ( Hayden et al. 2003 , Brodie et al. 2007 , Sfriso 2010 , Alongi et al. 2014 , Guiry and Guiry 2014 ).
A total of 15 species are currently recorded in marine algal flora of Korea ( Lee and Kang 1986 , 2002 , Lee 2008 , Bae 2010 , Kim et al. 2013 ). During survey of indigenous species, two marine ulvalean species (Chlorophyta) were collected from the southern and eastern coasts of Korea. These two Korean entities were identified based on morphological and molecular analyses and are newly recorded in marine algal flora of Korea in the present study.
MATERIALS AND METHODS
Samples for the present study were collected from Busan and Pohang, Korea, in 2014. All specimens were preserved in 5-10% formalin seawater, and pressed on herbarium sheets. A portion of the material was dried and preserved in silica gel for molecular analysis. Species identification was based on thallus morphology following the criteria of Bliding (1963 , 1968) and Koeman and van den Hoek (1981) . Sections of the thallus were mounted in 20% corn syrup for permanent preparation.
Total genomic DNA was extracted from silica-gel-preserved sample using the DNeasy Plant Mini Kit (Qiagen, Hilden, Germany) according to the manufacturer’s protocol. Before extraction, dried material was crushed with liquid nitrogen using a mortar and pestle. Concentrations of extracted DNA were assessed by using gel electrophoresis on a 1% agarose gel. Extracted DNA was used for amplification of the internal transcribed spacer (ITS) regions using published primers ( Blomster et al. 1998 ). PCR amplifications were performed in a TaKaRa PCR Thermal Cycler Dice (TaKaRa Bio Inc., Otsu, Japan) with an initial denaturation step at 94℃ for 5 min followed by 35 cycles at 94℃ for 1 min, 56℃ for 1 min, and 72℃ for 2 min and a final extension at 72℃ for 7 min. The reaction volume was 20 μL, consisting of 20 ng of genomic DNA, 2 μL of 10x PCR buffer, 2 μL of 200 μM dNTP, 1 μL of each forward and reverse primer, and 0.5 units of Taq polymerase (TaKaRa Bio Inc.). Amplifications were examined using gel electrophoresis in a 1% agarose gel and amplified ITS region products were purified using a QIAquick Gel Extraction Kit (Qiagen). The PCR products were moved to Macrogen Sequencing Service for sequencing (Macrogen, Seoul, Korea). The PCR primers were also used for sequencing.
Sequences for the ITS region were aligned using BioEdit ( Hall 1999 ). Phylogenetic analyses were performed using the neighbor-joining (NJ) and maximum-likelihood (ML) methods. Bootstrap values were calculated with 1,000 replications. ITS sequences of other species (excluding U. flexuosa and U. rigida collected from Korea during the present study) were obtained from GenBank. Umbraulva japonica was used as an outgroup.
RESULTS AND DISCUSSION
- Ulva flexuosaWulfen 1803: xxii
Korean name: Yeon-gal-pa-rae nom. nov. (신칭: 연갈파래)
Specimens examined: NIBRAL0000143269, PKNU0000134878 (Cheongsapo: 17 May 2014).
Habitat: Epilithic near the lower intertidal.
Morphology: Thalli 15-25 cm high ( Fig. 1a and 1b ), erect, membranous, distromatic, usually unbranched or little branched conical to ligulate shape, light to dark green in color, soft in texture, attached by a small holdfast (<5 mm) on rocks near the lower intertidal; frond ribbon-shaped, with a spirally twisted basal portion, usually undulate at the margin, 40-60 μm thick in the upper portion ( Fig. 1d ), 80-120 μm thick in the basal portion ( Fig. 1f ); cells usually arranged in pairs, rectangular to polygonal near the middle to upper portion ( Fig. 1c ), oval to rectangular with round corners near the basal portion in the surface view ( Fig. 1e ), transformed into rhizoidal cells near the base, 15-25 μm × 10-15 μm, with a length to width ratio of 1.6-1.8 in the transverse section; chloroplasts cap-like, parietal, with 1 (-2) pyrenoids (5-6 μm).
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Ulva flexuosa Wulfen. (a, b) Habit of vegetative plant. (c, d) Surface view (c) and transverse section (d) of the rectangular to polygonal cells with round corner near upper portion. (e, f) Surface view (e) and transverse section (f) of the oval to rectangular cells near basal portion. Scale bars are presented on the figures.
- Ulva rigidaC. Agardh 1823: 410
Korean name: Gyeong-gal-pa-rae nom. nov. (신칭: 경갈파래).
Specimens examined: NIBRAL0000143268, PKNU 0000134875, PKNU 0000134879 (Duhodong: 3 July 2014).
Habitat: Epilithic near the subtidal.
Morphology: Thalli 20-30 cm high ( Fig. 2a ), membranous, distromatic, irregular in shape, light to dark green in color, stiff in texture, attached by small holdfast on rocks near subtidal; frond irregular in shape, usually undulate and dentate at the margin ( Fig. 2d ), with small macroscopic denticulations along the margin ( Fig. 2b ), 40-60 μm thick ( Fig. 2f ); denticulations usually branched ( Fig. 2c ); cells usually arranged in pairs, rectangular to polygonal with round corners in the surface view ( Fig. 2e ), 5-15 μm × 5-10 μm, with length to width ratio of 1.2-2.0 in the transverse section; chloroplasts cap-like, parietal, with (1-) 2-3 pyrenoids (2-3 μm).
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Ulva rigida C. Agardh. (a) Habit of vegetative plant. (b) Details of frond with many denticulations (arrows) along margin. (c) Denticulations with branching. (d) Dentate margin of frond. (e, f ) Surface view (e) and transverse section (f ) of rectangular to polygonal cells with round corner near basal portion. Scale bars are presented on the figures.
Ulva flexuosa , which occurs in various localities including China and Japan ( Guiry and Guiry 2014 ), was originally described from Duino, Adriatic Sea ( Womersley 1984 ). This species is very similar to U. linza and U. lactuca in habit. However, U. flexuosa is distinguished from the two species by thalli with a spirally twisted basal portion, undulate margin and ribbon-shaped frond. Specimens collected from Cheongsapo, Busan, during the present study share these features as described above ( Fig. 1a and 1b ).
Ulva rigida resembles U. laetevirens Areschoug in having macroscopic denticulations along the thallus margin ( Sfriso 2010 ). However, the denticulations in U. rigida are very numerous and branched, while those in U. laetevirens are occasionally isolated and rarely branched ( Sfriso 2010 ). This type of denticulation is also found in Korean specimens collected during the present study.
Many studies have used the ITS region to analyze the phylogenetic relationships among Ulva species ( Malta et al. 1999 , Hayden et al. 2003 , Hayden and Waaland 2004 , Hofmann et al. 2010 , O’Kelly et al. 2010 ). In phylogenetic tree obtained from the present study based on the molecular data ( Figs. 3 and 4 ), the former Korean entity forms the same clade with U. californica as a subclade of the group containing U. flexuosa from Greece (near type locality) and China and U. californica from USA (type locality) and Japan with relatively high bootstrap value of 92% in NJ tree and 86% in ML tree. In this group supported by the undulate margin of thalli, genetic distance between the Korean entity and U. flexuosa ranged from 1.1% to 1.3%, while that between U. californica and U. flexuosa was 0.8 - 1.3%. According to Mareš et al. (2011) , the genetic distance for ITS sequences among U. flexuosa subspecies range from 3.8% to 5.9%. Based on this report, the Korean entity and U. californica are categorized into a specific variation within U. flexuosa . The present analyses, which show interspecific genetic distance of 2.1-13.3% in ulvalean species, also support strongly the intraspecific variation between those species. Accordingly, Ulva californica Wille in Collins et al. (1899) , which is currently accepted ( Guiry and Guiry 2014 ), is synonymized with U. flexuosa , here.
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Phylogenetic tree of selected taxa obtained from neighbor-joining analysis based on ITS sequences. Bootstrap percentages (1000 replicates samples) are shown above branches. Scale bar = 0.02 substitutions/site.
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Phylogenetic tree of selected taxa obtained from maximum-likelihood analysis based on ITS sequences. Bootstrap percentages (1000 replicates samples) are shown above branches. Scale bar = 0.02 substitutions/site.
The second Korean entity nests in the same clade with U. armoricana and U. scandinavica . Recently, however, U. armoricana and U. scandinavica were reduced to synonymy with U. rigida based on molecular studies ( Brodie et al. 2007 ). Although this Korean entity forms a sister clade to other U. rigida from Spain (type locality) and Scotland in NJ tree, genetic distance between those species is 0.5% within intraspecific variation range.
Based on these morphology and molecular data, our Korean entities are identified as U. flexuosa and U. rigida , respectively. This is the first record of these two ulvalean species in Korea.
Acknowledgements
This work was supported by a grant from the National Institute of Biological Resources (NIBR) funded by the Ministry of Environment (MOE) of the Republic of Korea (NIBR201401204), and by a grant from Marine Biotechnology Program funded by the Ministry of Oceans and Fisheries of the Korean Government.
References
Agardh CA 1823 Species Algarum Rite Cognitae, Cum Synonymis, Differentiis Specificis et Descriptionibus Succinctis. Volumen Primum Pars Posterior Ex Officina Berlingiana Lundae
Alongi G , Cormaci M , Furnari G 2014 A nomenclatural reassessment of some of Bliding's Ulvaceae Webbia: J Plant Taxon Geogr 69 89 - 96
Bae HB , Bae EH , Kim HS , Kwon CJ , Hwang IK , Kim GH , Klochkova TA 2010 Algal Flora of Korea. Volume 1, Number 1. Chlorophyta: Ulvophyceae: Ulotrichales, Ulvales, Cladophorales, Bryopsidales. Marine Green Algae National Institute of Biological Resources Incheon Ulotrichales, Ulvales 7 - 52
Bliding C 1963 A critical survey of European taxa in Ulvales. Part I. Capsosiphon, Percursaria, Blidingia, Enteromorpha Op Bot Soc Bot Lund 8 1 - 160
Bliding C 1968 A critical survey of European taxa in Ulvales. II. Ulva, Ulvaria, Monostroma, Kornmannia Bot Notiser 121 535 - 629
Blomster J , Maggs CA , Stanhope MJ 1998 Molecular and morphological analysis of Enteromorpha intestinalis and E. compressa (Chlorophyta) in the British Isles J Phycol 34 319 - 340    DOI : 10.1046/j.1529-8817.1998.340319.x
Brodie J , Maggs CA , John DM 2007 Green Seaweeds of Britain and Ireland British Phycological Society London
Collins FS , Holden I , Setchell WA 1899 Phycotheca Boreali-Americana A Collection of Dried Specimens of the Algae of North America Malden, MA
Guiry MD , Guiry GM 2014 AlgaeBase World-wide electronic publication, National University of Ireland Galway http://www.algaebase.org
Hall TA 1999 BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT Nucleic Acids Symp Ser 41 95 - 98
Hayden HS , Blomster J , Maggs CA , Silva PC , Stanhope MJ , Waaland JR 2003 Linnaeus was right all along: Ulva and Enteromorpha are not distinct genera Eur J Phycol 38 277 - 294    DOI : 10.1080/1364253031000136321
Hayden HS , Waaland JR 2004 A molecular systematic study of Ulva (Ulvaceae, Ulvales) from the northeast Pacific Phycologia 43 364 - 382    DOI : 10.2216/i0031-8884-43-4-364.1
Heesch S , Broom JES , Neill KF , Farr TJ , Dalen JL , Nelson WA 2009 Ulva, Umbraulva and Gemina: genetic survey of New Zealand taxa reveals diversity and introduced species Eur J Phycol 44 143 - 154    DOI : 10.1080/09670260802422477
Hoeksema BW , van den Hoek C 1983 The taxonomy of Ulva (Chlorophyta) from the coastal region of Roscoff (Brittany, France) Bot Mar 26 65 - 86
Hofmann LC , Nettleton JC , Neefus CD , Mathieson AC 2010 Cryptic diversity of Ulva (Ulvales, Chlorophyta) in the Great Bay Estuarine System (Atlantic USA): introduced and indigenous distromatic species Eur J Phycol 45 230 - 239    DOI : 10.1080/09670261003746201
Kim HS , Boo SM , Lee IK , Sohn CH 2013 National List of Species of Korea, Marine Algae Jeonghaengsa Seoul (in Korean)
Koeman RPT , van den Hoek C 1981 The taxonomy of Ulva (Chlorophyceae) in the Netherlands Brit Phycol J 16 9 - 53    DOI : 10.1080/00071618100650031
Lee IK , Kang JW 1986 A check list of marine algae in Korea Kor J Phycol 1 311 - 325
Lee YP 2008 Marine Algae of Jeju Academy Publishing Co Seoul (in Korean)
Lee YP , Kang SY 2002 A Catalogue of the Seaweeds in Korea Jeju National University Press Jeju
Loughnane CJ , McIvor LM , Rindi F , Stengel DB , Guiry MD 2008 Morphology, rbcL phylogeny and distribution of distromatic Ulva (Ulvophyceae, Chlorophyta) in Ireland and southern Britain Phycologia 47 416 - 429    DOI : 10.2216/PH07-61.1
Malta EJ , Draisma SGA , Kamermans P 1999 Free-floating Ulva in the southwest Netherlands: species or morphotypes? A morphological, molecular and ecological comparison Eur J Phycol 34 443 - 454    DOI : 10.1080/09541449910001718801
Mareš J , Leskinen E , Sitkowska M , Skácelova O , Blomster J 2011 True identity of the European freshwater Ulva (Chlorophyta, Ulvophyceae) revealed by a combined molecular and morphological approach J Phycol 47 1177 - 1192    DOI : 10.1111/j.1529-8817.2011.01048.x
O’Kelly CJ , Kurihara A , Shipley TC , Sherweed AR 2010 Molecular assessment of Ulva spp. (Ulvophyceae, Chlorophyta) in the Hawaiian Islands J Phycol 46 728 - 735    DOI : 10.1111/j.1529-8817.2010.00860.x
Sfriso A 2010 Coexistence of Ulva rigida and Ulva laetevirens (Ulvales, Chlorophyta) in Venice Lagoon and other Italian transitional and marine environments Bot Mar 53 9 - 18
Tan IH , Blomster J , Hansen G , Leskinen E , Maggs CA , Mann DG , Sluiman HJ , Stanhope MJ 1999 Molecular phylogenetic evidence for a reversible morphogenetic switch controlling the gross morphology of two common genera of green seaweeds, Ulva and Enteromorpha Mol Biol Evol 16 1011 - 1018    DOI : 10.1093/oxfordjournals.molbev.a026190
Womersley HBS 1984 The Marine Benthic Flora of Southern Australia Government Printer Adelaide Part I