Next-Generation Sequencing of an 88-Year-Old Specimen of the Poorly Known Species Liagora japonica (Nemaliales, Rhodophyta) Supports the Recognition of Otohimella gen. nov.

Takahiro Segawa,Hiroshi Kawai,Ayumi Akiyoshi,Hiroshi Mori,Taiju Kitayama,Ryo Ootsuki,Tsuyoshi Abe,Kazuhiro Kogame,Hisayoshi Nozaki,Masahiro Suzuki,Akira Kurihara,Hidetoshi Sakayama

doi:10.1371/journal.pone.0158944

Abstract

Liagora japonica is a red algal species distributed in temperate regions of Japan. This species has not been collected from its type locality on the Pacific coast of Japan since 1927 and seems to have become extinct in this area. For molecular characterization of L. japonica, we extracted DNA from the topotype material of L. japonica collected in 1927, analyzed seven genes using Illumina next-generation sequencing, and compared these data with sequences from modern samples of similar red algae collected from the Japan Sea coast of Japan. Both morphological and molecular data from modern samples and historical specimens (including the lectotype and topotype) suggest that the specimens from the Pacific and Japan Sea coasts of Japan should be treated as a single species, and that L. japonica is phylogenetically separated from the genus Liagora. Based on the phylogenetic results and examination of reproductive structures, we propose Otohimella japonica gen. et comb. nov., characterized morphologically by diffuse carposporophytes, undivided carposporangia, and involucral filaments initiated only from the cortical cell on the supporting cell.

Highlights

Taxonomic studies are based on type specimens, which are permanently attached to taxonomic names
Based on limited publicly available sequence data from the Nemaliales, including Liagoraceae (Table C in S2 File), reliable sequences from Liagora japonica were determined for four genes: 28S rRNA, psaA, rbcL, and cytochrome oxidase 1 (COI) (Fig 1, Figures B-E in S1 File)
Liagora japonica showed < 0.3% intraspecific divergence of rbcL, which was within the range of that of Liagoraceae [31], while the intraspecific divergence of COI was < 1.5%, which was higher than typical intraspecific divergence of the Nemaliales [34]

Summary

Introduction

Taxonomic studies are based on type specimens, which are permanently attached to taxonomic names. Morphological information from type specimens is not sufficient for taxonomic studies, because many type materials are fragmented and lack reproductive structures required for comparison with modern collections. Hughey et al [7] performed NGS to determine the complete plastid and mitochondrial genomes from 140-year-old type specimens of the bangiophycean species of Pyropia using the published complete organelle genome data from Pyropia spp. for reference mapping and sequence assembly. In the case of Florideophyceae, the largest group of red algae, limited genomic data are available as references in NGS, compared with bangiophycean red algae, and old DNA sequences have not been determined previously by NGS for molecular phylogenetic analyses

Methods

Results

Discussion

Conclusion