A DNA mini-barcoding system for endangered unionid mussels in the Lake Biwa system in Japan


A mini-barcoding system was developed to help identify endangered unionid mussels in the Lake Biwa system in Japan. A single primer set was designed to amplify a ca. 140-bp barcode fragment based on the published mitochondrial 16S rRNA gene sequences (ca. 350 bp) of Japanese unionids. The new primers successfully amplified genomic DNA from the foot tissues of 81 specimens of 12 target taxa from the Lake Biwa region. Sequencing, BLAST-searches and phylogenetic analyses of the amplified fragments confirmed that the mini-barcode had the same identification ability as the original sequences for Lake Biwa taxa. The mini-barcoding system also obtained the short barcodes from excreted materials, indicating its usefulness in nondestructive identification of these mussels.

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  1. Carella F, Villari G, Maio N, De Vico G (2016) Disease and disorders of freshwater unionid mussels: a brief overview of recent studies. Front Physiol 7:489. https://doi.org/10.3389/fphys.2016.00489

    Article  PubMed  PubMed Central  Google Scholar 

  2. Felsenstein J (1985) Confidence limits on phylogenies: an approach using the bootstrap. Evolution 39:783–791. https://doi.org/10.1111/j.1558-5646.1985.tb00420.x

    Article  Google Scholar 

  3. Henley WF, Grobler PJ, Neves RJ (2006) Non-invasive method to obtain DNA from freshwater mussels (Bivalvia: Unionidae). J Shellfish Res 25:975–977. https://doi.org/10.2983/0730-8000(2006)25[975:NMTODF]2.0.CO;2

    Article  Google Scholar 

  4. Katoh K, Rozewicki J, Yamada KD (2017) MAFFT online service: multiple sequence alignment, interactive sequence choice and visualization. Briefings Bioinf 20(4):1160–1166. https://doi.org/10.1093/bib/bbx108

    CAS  Article  Google Scholar 

  5. Kawabe T (1989) Stratigraphy of the lower part of the Kobiwako Group around the Ueno basin, Kinki district, Japan. J Geosci Osaka City Univ 32:39–90

    Google Scholar 

  6. Kihira H, Matsuda M, Uchiyama R (2003) Freshwater Mollusks of Japan 1. Freshwater Mollusks in Lake Biwa and Yodo River. Tokyo, Pisces

    Google Scholar 

  7. Kimura M (1980) A simple method for estimating evolutionary rate of base substitutions through comparative studies of nucleotide sequences. J Mol Evol 16:111–120. https://doi.org/10.1007/BF01731581

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  8. Kondo T (2008) Monograph of unionoida in Japan (Mollusca: Bivalvia). Special Publication of the Malacological Society of Japan, No. 3, 69 p.

  9. Kondo T, Tabe M, Fukuhara S (2011) Separateing Anodonta lauta and Anodonta japonica by shell shape. Chiribotan 41:84–88

    Google Scholar 

  10. Kumar S, Stecher G, Tamura K (2016) MEGA7: molecular evolutionary genetics analysis version 7.0 for bigger datasets. Mol Biol Evol 33:1870–1874. https://doi.org/10.1093/molbev/msw054

    CAS  Article  Google Scholar 

  11. Mabuchi K, Nishida K, Nakajima N (2018) Complete F mitochondrial genomes of the Biwa pearl mussel, Hyriopsis schlegelii: the first report from the species' native lake in Japan. Mitochondr DNA Part B 3:1294–1295. https://doi.org/10.1080/23802359.2018.1534562

    Article  Google Scholar 

  12. Nishino M (2017) Ecological structures of lakes and Biodiversity of Lake Biwa. In: Nishino M, Akiyama M, Nakajima T (eds) The messages from the shores of Lake Biwa: a viewpoint aiming at the conservation and restoration. Sunrise, Hikone, pp 37–43

    Google Scholar 

  13. Saitou N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4:406–425. https://doi.org/10.1093/oxfordjournals.molbev.a040454

    CAS  Article  Google Scholar 

  14. Sano I, Shirai A, Kondo T, Miyazaki JI (2017) Phylogenetic relationships of Japanese unionoida (Mollusca: Bivalvia) based on mitochondrial 16S rDNA sequences. J Water Res Prot 9:493–509. https://doi.org/10.4236/jwarp.2017.95032

    Article  Google Scholar 

  15. Scientific Committee for Research into the Wildlife in Shiga Prefecture (2016) Red data book of Shiga Prefecture 2015. Sunrise, Hikone

    Google Scholar 

  16. Vaughn CC, Spooner DE (2006) Unionid mussels influence macroinvertebrate assemblage structure in streams. J N Am Benthol Soc 25:691–700. https://doi.org/10.1899/0887-3593(2006)25[691:UMIMAS]2.0.CO;2

    Article  Google Scholar 

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We thank Masanari Matsuda (Lake Biwa Museum; LBM) for registering our specimens in the LBM shell collection. We also thank the following institutions and individuals for help collecting Hyriopsis specimens: Shiga Prefectural Fisheries Experiment Station, Fumio Mikami, and Toshiaki Ibuki. This work was supported by the Collaborative Research Fund of Shiga Prefecture “Studies on the conservation and ecosystem management of Lake Biwa” under a Japanese Grant for Regional Revitalization.

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Correspondence to Kohji Mabuchi.

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Mabuchi, K., Nishida, K. A DNA mini-barcoding system for endangered unionid mussels in the Lake Biwa system in Japan. Conservation Genet Resour 12, 581–584 (2020). https://doi.org/10.1007/s12686-020-01160-w

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  • DNA barcode
  • Unionidae
  • 16S rRNA
  • Lake biwa