TAXONOMIC REEXAMINATION OF CHARA GLOBULARIS (CHARALES, CHAROPHYCEAE) FROM JAPAN BASED ON OOSPORE MORPHOLOGY AND rbcL GENE SEQUENCES, AND THE DESCRIPTION OF C. LEPTOSPORA SP. NOV.¹

Abstract

Chara globularis Thuillier (=f. globularis sensu R. D. Wood) is a widespread species of the genus and inhabits fresh- and brackish-water environments. In an attempt to reexamine the taxonomic status of C. globularis collected from Japan, we reassessed vegetative and oospore morphology of Japanese material and herbarium specimens originating from Europe (including the type specimen) and conducted molecular phylogenetic analyses based on rbcL gene sequences. Although the other vegetative morphologies were consistent with the description of C. globularis f. globularis sensu R. D. Wood, we identified two types of branchlets within the Japanese materials: one has elongate end segments (EL type), and the other has short end segments (SH type) corresponding to the type material. Moreover, the oospore wall of the EL type was different from that in the SH type. The oospores of the EL type were dark brown to reddish brown and had a spongy pattern with the pusticular elevations on the fossa wall, whereas the fossa wall of the SH type was black with a granulate to papillate or fine pusticular pattern. In addition, our sequence data demonstrated that these two types are separated phylogenetically from each other. Therefore, we describe the EL type as a new species, C. leptospora sp. nov.

Taxonomic status of Chara tenuspina A. Br. (Streptophyta: Charales) based on LM morphology, matK, atpB and rbcL of cpDNA sequences

“…The quality and quantity of the DNA were determined using a fluorometer (Eppendorf, Hamburg, Germany), but the integrity of the extracted DNA was estimated on 1% TBE-agarose gel. The PCR amplification and sequencing of the atpB gene were accomplished using the primers described by Sakayama et al (2004), while that of the rbcL and matK made use of primers described by Sakayama et al (2009) andsanders et al (2003), respectively. Analyses were performed in a Veriti gradient Thermal Cycler (Applied Carlsbad, CA, USA Each 20 μl reaction contained 10 mM each of dATP, dCTP, dGTP, and dTTP; 0.5 μM of each primer, 4.0 μl 5´ reaction buffer, 0.2 μl Phusion Hot Start II HF DNA Polymerase (Thermo Scientific, Waltham, MA, USA.)…”

“…12). Representatives of genera Nitellopsis, Nitella, Lychothamnus and Tolypella were selected as an outgroup according to previous study (mCCoUrT et al 1996(mCCoUrT et al , 1999karol et al 2001;sakayama et al 2002sanders et al 2003 andSakayama et al 2009). The rbcL DNA sequences analysed in this study were 1,244 bp (excluding gaps) and correspond to position 5 -1,249 of the Chara vulgaris rbcL gene (TUrmel et al 2006).…”

“…Species level taxonomy in the genera Chara L. and Nitella agardh is controversial, mostly due to overlapping morphological variations, apparent intermediate forms between taxa, and the unknown extent to which a phenotypic plasticity or developmental differences contribute to for morphological variation (O'Reilly et al 2007;Sakayama et al 2002Sakayama et al , 2009Urbaniak 2009Urbaniak , 2010Urbaniak , 2011aUrbaniak , 2011bUrbaniak & Combik 2013).…”

Taxonomical analysis of closely related species of Chara L. section Hartmania (Streptophyta: Charales) based on morphological and molecular data

“…Detailed statistical analyses of oospore dimensions have also been studied by mandaL et al (2002) and Urbaniak (2009). Sakayama et al (2009Sakayama et al ( , 2015 and Perez et al 2014) showed that the combination of different types of data (SEM oospore morphology and molecular data) can be more informative than when taken separately, and can be used for taxonomic distinction, especially in closely related species of the genus Nitella or Tolypella.…”

“…For the genus Chara, molecular phylogenetic analyses, based on chloroplast and nuclear DNA gene sequences, together with analyses of morphological characteristics, have also been carried out to resolve the relationships at species or genic level (mccoUrt et al 1996;meierS et al 1999;SanderS et al 2003;Sakayama et al 2009;Schneider et al 2015;noWak et al 2016). By contrast, in the same way using combined morphological characteristics of the species and analysis of about 6214 bp, noWak et al (2016) coudl not separate Chara individuals and all of the studied species.…”