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The taxonomic position of Asian Holopedium (Crustacea: Cladocera) confirmed by morphological and genetic analyses

  • Ayaka Yamamoto
  • Wataru MakinoEmail author
  • Jotaro Urabe
Research paper


The cladoceran Holopedium gibberum Zaddach, 1855 (Ctenopoda: Holopediidae) was once thought to occur broadly in the northern hemisphere, but its cryptic sister species was recently separated from H. gibberum sensu stricto (s.s.) as a new species, Holopedium glacialis. In East Asia, although “H. gibberum” occurrence has been recorded in many water bodies, the identity of the surveyed populations has rarely been confirmed via molecular analyses. Thus, it is unclear whether it is actually H. gibberum s.s. or H. glacialis that is distributed in East Asia. We used DNA-barcoding techniques to check the taxonomic status of Holopedium samples collected in Japan. We sequenced mitochondrial cytochrome c oxidase subunit 1 (mtCOI) and nuclear 18S ribosomal DNA (nr18S) of Japanese Holopedium and compared the results with those of H. gibberum s.s. collected in Norway and H. glacialis collected in Canada. The mtCOI sequence divergences between Norwegian H. gibberum s.s. and Japanese Holopedium were at most 2.4%, which was within the degree of intraspecific differentiation in cladocerans. Norwegian H. gibberum s.s. and Japanese Holopedium shared identical nr18S haplotypes. Individuals of Canadian H. glacialis were genetically different from those of Japanese Holopedium. We therefore concluded that Japanese Holopedium can be identified as H. gibberum s.s.


DNA barcoding Freshwater biodiversity Integrative taxonomy Zooplankton 



Marcia Kyle (Norwegian University of Life Sciences) and Tom Andersen (University of Oslo) collected Norwegian Holopedium specimens, and Paul Frost, Andrea Conine, and Clay Prater (Trenton University) collected Canadian Holopedium specimens. We are truly grateful for their very kind support. The present study was also supported by grants from the Japan Society for the Promotion of Science (16770011, 19770010, 23570015, 15H02380, 15K07211, and 18K06407); by funds from the Ministry of the Environment, Japan (the Environment Research and Technology Development Fund, 4-1602); by funds from the Water Resources Environment Technology Center (2008-06, and 2018-03) to WM; and by grants from the Japan Society for the Promotion of Science (25291094 and 16H02522) and the Ministry of the Environment, Japan (the Environment Research and Technology Development Fund, D-1002) to JU.

Supplementary material

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Supplementary material 1 (PDF 1575 kb)


  1. Amaral-Zettler LA, McCliment EA, Ducklow HW, Huse SM (2009) A method for studying protistan diversity using massively parallel sequencing of V9 hypervariable regions of small-subunit ribosomal RNA Genes. PLoS One 4:e6372CrossRefPubMedGoogle Scholar
  2. Baloch WA, Maeda H, Saisho T (1998) Seasonal abundance and vertical distribution of zooplankton in Lake Ikeda, southern Japan. Microbes Environ 13:1–8CrossRefGoogle Scholar
  3. Ban S, Makino W, Sakano H, Haruna H, Ueda H (2013) Annual variation in biomass and the community structure of crustacean zooplankton over 5 years in Lake Toya, Japan. Limnology 14:59–70CrossRefGoogle Scholar
  4. Belyaeva M, Taylor DJ (2009) Cryptic species within the Chydorus sphaericus species complex (Crustacea: Cladocera) revealed by molecular markers and sexual stage morphology. Mol Phylogenet Evol 50:534–546CrossRefPubMedGoogle Scholar
  5. Costa FO, deWaard JR, Boutillier F, Ratnasingham S, Dooh ST, Hajibabaei M, Hebert PDN (2007) Biological identifications through DNA barcodes: the case of the Crustacea. Can J Fish Aquat Sci 64:272–295CrossRefGoogle Scholar
  6. Cox JA, Hebert PDN (2001) Colonization, extinction, and phylogeographic patterning in a freshwater crustacean. Mol Ecol 10:371–386CrossRefPubMedGoogle Scholar
  7. Crease TJ, Colbourne JK (1998) The unusually long small-subunit ribosomal RNA of the Crustacean, Daphnia pulex: sequence and predicted secondary structure. J Mol Evol 46:307–313CrossRefPubMedGoogle Scholar
  8. Crease TJ, Taylor DJ (1998) The origin and evolution of variable-region helices in V4 and V7 of the small-subunit ribosomal RNA of Branchiopod crustaceans. Mol Biol Evol 15:1430–1446CrossRefPubMedGoogle Scholar
  9. Cristescu ME (2014) From barcoding single individuals to metabarcoding biological communities: towards an integrative approach to the study of global biodiversity. Trends Ecol Evol 29:566–571CrossRefPubMedGoogle Scholar
  10. De Melo R, Hebert PDN (1994) A taxonomic reevaluation of North American Bosminidae. Can J Zool 72:1808–1825CrossRefGoogle Scholar
  11. Folmer O, Black M, Hoeh W, Lutz R, Vrijenhoek R (1994) DNA primers for amplification of mitochondrial cytochrome c oxidase subunit I from diverse metazoan invertebrates. Mol Mar Biol Biotechnol 3:294–299PubMedGoogle Scholar
  12. Frey DG (1982) Questions concerning cosmopolitanism in Cladocera. Arch Hydrobiol 93:484–502Google Scholar
  13. Ha J-Y, Hanazato T, Chang K-H, Jeong K-S, Kim D-K (2015) Assessment of the lake biomanipulation mediated by piscivorous rainbow trout and herbivorous daphnids using a self-organizing map: a case study in Lake Shirakaba, Japan. Ecol Inform 29:182–191CrossRefGoogle Scholar
  14. Hanazato T, Nohara S (1992) Seasonal succession and vertical distribution of zooplankton in Lake Ozenuma. Jpn J Limnol 53:55–63CrossRefGoogle Scholar
  15. Hasegawa M, Kishino H, Yano T (1985) Dating of the human-ape splitting by a molecular clock of mitochondrial DNA. J Mol Evol 22:160–174CrossRefPubMedGoogle Scholar
  16. Hebert PDN, Ratnasingham S, de Waard JR (2003a) Barcoding animal life: cytochrome c oxidase subunit 1 divergences among closely related species. Proc R Soc Lond B 270:S96–S99Google Scholar
  17. Hebert PDN, Cywinska A, Ball SF, de Waard JR (2003b) Biological identifications through DNA barcodes. Proc R Soc Lond B 270:313–321CrossRefGoogle Scholar
  18. Ishida S, Taylor DJ (2007) Quaternary diversification in a sexual Holarctic zooplankter, Daphnia galeata. Mol Ecol 16:569–582CrossRefPubMedGoogle Scholar
  19. Ishida S, Kotov AA, Taylor DJ (2006) A new divergent lineage of Daphnia (Cladocera: Anomopoda) and its morphological and genetical differentiation from Daphnia curvirostris Eylmann, 1887. Zool J Linn Soc 146:385–405CrossRefGoogle Scholar
  20. Kimura M (1980) A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences. J Mol Evol 16:111–120CrossRefPubMedGoogle Scholar
  21. Korovchinsky NM (1992) Sididae and Holopedidae (Crustacea: Daphniiformes). SPB Academic Publishing, AmsterdamGoogle Scholar
  22. Korovchinsky NM (2005) New species of Holopedium Zaddach, 1855 (Crustacea: Claocera: Ctenopoda) from Greenland. J Limnol 64:103–112CrossRefGoogle Scholar
  23. Korovchinsky NM (2006) The Cladocera (Crustacea: Branchiopoda) as a relict group. Zool J Linnean Soc 147:109–124CrossRefGoogle Scholar
  24. Korovchinsky NM (2009) The genus Leptodora Lillijeborg (Crustacea: Branchiopoida: Cladocera) is not monotypic: description of a new species from the Amur River basin (Far East of Russia). Zootaxa 2120:39–52Google Scholar
  25. Kotov AA, Taylor DJ (2019) Contrasting endemism in pond-dwelling cyclic parthenogenesis: the Daphnia curvirostris species group (Crustacea: Cladocera). Sci Rep 9:6812CrossRefPubMedGoogle Scholar
  26. Kotov AA, Ishida S, Taylor DJ (2006) An assessment of species diversity in the Daphnia curvirostris (Crustacea: Cladocera) complex with phylogenetic evidence for the independent origin of neckteeth. J Plankton Res 28:1067–1079CrossRefGoogle Scholar
  27. Kotov AA, Ishida S, Taylor DJ (2009) Revision of the genus Bosmina Baird, 1845 (Cladocera: Bosminidae), based on evidence from male morphological characters and molecular phylogenies. Zool J Linn Soc 156:1–51CrossRefGoogle Scholar
  28. Kotov AA, Karabanov DP, Bekker EI, Neterina TV, Taylor DJ (2016) Phylogeography of the Chydorus sphaericus group (Cladocera: Chydoridae) in the Northern Palearctic. PLoS One 11:e0168711CrossRefPubMedGoogle Scholar
  29. Lakatos C, Urabe J, Makino W (2015) Cryptic diversity of Japanese Diaphanosoma (Crustacea: Cladocera) revealed by morphological and molecular assessments. Inland Waters 5:253–262CrossRefGoogle Scholar
  30. Makino W, Tanabe AS (2009) Extreme population genetic differentiation and secondary contact in the freshwater copepod Acanthodiaptomus pacificus in the Japanese Archipelago. Mol Ecol 18:3699–3713CrossRefPubMedGoogle Scholar
  31. Makino W, Ohtsuki H, Urabe J (2013) Finding copepod footprints: a protocol for molecular identification of diapausing eggs in lake sediments. Limnology 14:269–282CrossRefGoogle Scholar
  32. Makino W, Maruoka N, Nakagawa M, Takamura N (2017) DNA barcoding of freshwater zooplankton in Lake Kasumigaura, Japan. Ecol Res 32:481–493CrossRefGoogle Scholar
  33. Makino W, Tanabe AS, Urabe J (2018) The fauna of freshwater calanoid copepods in Japan in the early decades of the 21st Century: implications for the assessment and conservation of biodiversity. Limnol Oceanogr 63:758–772CrossRefGoogle Scholar
  34. Mills S, Alcántara-Rodríguez JA, Ciros-Pérez J, Gómez A, Hagiwara A, Galindo KH, Walsh EJ et al (2017) Fifteen species in one: deciphering the Brachionus plicatilis species complex (Rotifera, Monogononta) trough DNA taxonomy. Hydrobiologia 796:39–58CrossRefGoogle Scholar
  35. Mizuno T, Takahashi E (eds) (2000) An illustrated guide to freshwater zooplankton in Japan (in Japanese). Tokai University Press, TokyoGoogle Scholar
  36. Pace ML (1986) An empirical analysis of zooplankton community size structure across lake trophic gradients. Limnol Oceanogr 31:45–55CrossRefGoogle Scholar
  37. Petrusek A, Černý M, Audenaert E (2004) Large intercontinental differentiation of Moina micrura (Crustacea: Anomopoda): one less cosmopolitan cladoceran? Hydrobiologia 526:73–81CrossRefGoogle Scholar
  38. Porter TM, Hajibabaei M (2018) Automated high throughput animal CO1 metabarcode classification. Sci Rep 8:4226CrossRefPubMedGoogle Scholar
  39. R Core Team (2013) R: a language and environment for statistical computing. R Foundation for Statistical Computing, Vienna. ISBN 3-900051-07-0.
  40. Ronquist F, Huelsenbeck JP (2003) MrBayes 3: Bayesian phylogenetic inference under mixed models. Bioinformatics 19:1572–1574CrossRefPubMedGoogle Scholar
  41. Rowe CD, Adamovicz SJ, Hebert PDN (2007) Three new cryptic species of the freshwater zooplankton genus Holopedium (Crustacea: Branchiopoda: Ctenopoda), revealed by genetic methods. Zootaxa 1656:1–49CrossRefGoogle Scholar
  42. Somervo P, Yu DW, Xu CCY, Ji Y, Hultman J, Wirta H, Ovaskainen O (2017) Quantifying uncertainty of taxonomic placement in DNA barcoding and metabarcoding. Methods Ecol Evol 8:397–408Google Scholar
  43. Stefanni S, Stankovic D, Borme D, de Olazabal A, Juretic T, Pallavicini A, Tirelli V (2018) Multi-marker metabarcording approach to study mesozooplankton at basin scale. Sci Rep 8:12085CrossRefPubMedGoogle Scholar
  44. Tamura K, Stecher G, Peterson D, Filipski A, Kumar S (2013) MEGA6: molecular evolutionary genetics analysis version 6.0. Mol Biol Evol 30:2725–2729CrossRefPubMedGoogle Scholar
  45. Tanabe AS (2007) KAKUSAN: a computer program to automate the selection of a nucleotide substitution model and the configuration of a mixed model on multilocus data. Mol Ecol Notes 7:962–964CrossRefGoogle Scholar
  46. Tanabe AS, Nagai S, Hida K, Yasuike M, Fujiwara A, Nakamura Y, Katakura S et al (2016) Comparative study of the validity of three regions of the18S-rRNA gene for massively parallel sequencing-based monitoring of the planktonic eukaryote community. Mol Ecol Res 16:402–414CrossRefGoogle Scholar
  47. Tanaka M (1992) Nihon kosho-shi (Records of Japanese lakes). Nagoya University Press, Nagoya (in Japanese) Google Scholar
  48. Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG (1997) The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res 25:4876–4882CrossRefPubMedGoogle Scholar
  49. Urabe J, Ishida S, Nishimoto M, Weider LW (2003) Daphnia pulicaria, a zooplankton species that suddenly appeared in 1999 in the offshore zone of Lake Biwa. Limnology 4:35–41CrossRefGoogle Scholar
  50. Weider LJ, Hobaek A, Colbourne J, Crease TJ, Dufresne F, Hebert PDN (1999) Holarctic phylogeography of an asexual species complex. I. Mitochondrial DNA variation in Arctic Daphnia. Evolution 53:777–792CrossRefPubMedGoogle Scholar
  51. Xu S, Hebert PDN, Kotov AA, Cristescu ME (2009) The noncosmopolitanism paradigm of freshwater zooplankton: insights from the global phylogeography of the predatory cladoceran Polyphemus pediculus (Linnaeus, 1761) (Crustacea, Onychopoda). Mol Ecol 18:5161–5179CrossRefPubMedGoogle Scholar
  52. Xu L, Han B-P, Van Damme K, Vierstaete A, Vanfleteren JR, Dumont HJ (2011) Biogeography and evolution of the Holarctic zooplankton genus Leptodora (Crustacea: Branchiopoda: Haplopoda). J Biogeogr 38:359–370CrossRefGoogle Scholar
  53. Yang J, Zhang X, Xie Y, Song C, Zhang Y, Yu H, Burton GA (2017) Zooplankton community profiling in a eutrophic freshwater ecosystem-Lake Tai basin by DNA metabarcoding. Sci Rep 7:1773CrossRefPubMedGoogle Scholar
  54. Zuykova EI, Simonov EP, Bochkarev NA, Taylor DJ, Kotov AA (2018a) Resolution of the Daphnia umbra problems (Crustacea: Cladocera) using an integrated taxonomic approach. Zool J Linn Soc 184:969–998Google Scholar
  55. Zuykova EI, Simonov EP, Bochkarev NA, Abramov SA, Sheveleva NG, Kotov AA (2018b) Contrasting phylogeographic patterns and demographic history in closely related species of Daphnia longispina group (Crustacea: Cladocera) with focus on North-Eastern Eurasia. PLoS One 13:e0207347CrossRefPubMedGoogle Scholar

Copyright information

© The Japanese Society of Limnology 2019

Authors and Affiliations

  1. 1.Graduate School of Life SciencesTohoku UniversitySendaiJapan

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