Abstract
Chloromonas nivalis (Volvocales, Chlorophyceae) is considered a cosmopolitan species of a snow-inhabiting microalga because cysts morphologically identifiable as zygotes of the species are distributed worldwide. However, recent molecular data demonstrated that field-collected cysts identified as the zygotes consist of multiple species. Recently, we demonstrated that species identification of snow-inhabiting Chloromonas species is possible based on light and electron microscopy of asexual life cycles in strains and molecular phylogenetic analyses. Vegetative cells without eyespots and of inverted-teardrop shape have been reported once in North American material of C. nivalis; however, strains with such vegetative cells in snow-inhabiting species of Chloromonas have not been examined taxonomically in detail. Here, we used light and transmission electron microscopy together with molecular analyses of multiple DNA sequences to examine several C. nivalis strains. The morphological data demonstrated that one North American strain could be identified as C. nivalis, whereas three other strains should be re-classified as C. hoshawii sp. nov. and C. remiasii sp. nov. based on vegetative cell morphology, the number of zoospores within the parental cell wall during asexual reproduction, and whether cell aggregates (resulting from repeated divisions of daughter cells retained within a parental cell wall) were observed in the culture. This taxonomic treatment was supported by multigene phylogeny and comparative molecular analyses that included a rapidly evolving DNA region. Our molecular phylogenetic analyses also demonstrated that the North American strain of C. nivalis was phylogenetically separated from the Austrian and Japanese specimens previously identified as C. nivalis based on zygote morphology.
Highlights
During the snow melt season, snowfields in polar regions and snowpacks in mountainous areas are sometimes stained green, red, or other colors
Three strains assigned to C. nivalis in previous studies (CCCryo 005–99, UTEX SNO66 and UTEX SNO74) [19,23,24,25], one North American strain labeled as C. nivalis (UTEX SNO71) [21], and Chloromonas sp. strain CCCryo 047–99 were provided by the Culture Collection of Cryophilic Algae (CCCryo) at the Fraunhofer Institute for Cell Therapy and Immunology [22] and the Culture Collection of Algae at the University of Texas at Austin (UTEX) [21,27] (S1 Table)
Light and epifluorescence microscopy (Figs 1 and 2) demonstrated that the strains could be subdivided into three morphological species (C. hoshawii, C. nivalis, and C. remiasii) based on differences in cell shape and size, chloroplast morphology, presence of eyespots, number of zoospores formed within the parental cell wall during asexual reproduction, and presence of cell aggregates in culture (Table 1)
Summary
During the snow melt season, snowfields in polar regions and snowpacks in mountainous areas are sometimes stained green, red, or other colors. Among the snow-inhabiting Chloromonas species, C. nivalis (Chodat) Hoham et Mullet was considered cosmopolitan because of the world-wide distribution of cysts morphologically identified as zygotes of this species [formerly classified as Scotiella nivalis (Chodat) F.E. Fritsch] based on studies of North American material [1,11]. The species was generally identified solely on the basis of zygote morphology [1,14,15,16,17] according to the species concept of previous studies [11,18] This reliance on morphology possibly arises from the difficulty of inducing vegetative cell production from field-collected zygotes of snow-inhabiting Chloromonas. We document phylogenetic relationships between the North American strain of C. nivalis and previously examined specimens of C. nivalis zygotes
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