Molecular diversity of green corticolous microalgae from two sub-Mediterranean European localities

<div><p>Green algae in corticolous biofilms are simple coccoid cells or filamentous thalli with strikingly low morphological diversity. Consequently, microscopic identification of these organisms is difficult, and often possible only to higher taxonomic units. We investigated the taxonomic and phylogenetic composition of green microalgae isolated from biofilms growing on the bark of <i>Quercus pubescens</i> and <i>Pinus nigra</i>. The study was based on 122 partial sequences of the plastid-encoded <i>rbc</i>L gene. In total, 29 operational taxonomic units (OTUs), differing in their <i>rbc</i>L sequences, were encountered. Members of the Trebouxiophyceae formed 97.5% of the isolates; Streptophyta made up 2.5%. The most frequently occurring OTUs were in the genera <i>Coccomyxa, Parachloroidium</i> and <i>Stichococcus</i>. Within the <i>Watanabea</i> clade, we have probably discovered an as-yet undescribed generic lineage with chlorelloid morphology. OTUs belonging to the recently described trebouxiophycean genera <i>Kalinella, Leptochlorella</i> and <i>Xylochloris</i> were also encountered, which indicates that these genera are probably widely distributed in subaerial microhabitats, such as tree bark. The samples taken from oak trees were more diverse in their OTU composition than those taken from pine trees, but the average phylogenetic distances of OTUs in samples did not differ between the host tree taxa. Host tree species had a stronger effect on the community structure of algae than the sampling locality. This indicates that habitat filtering is important for the distribution of individual microalgal phylogenetic taxa.</p></div

Morphological delineation and distribution patterns of four newly described species within the <i>Synura petersenii</i> species complex (Chrysophyceae, Stramenopiles)

<div><p>The <i>Synura petersenii</i> species complex represents a common, cosmopolitan and highly diverse taxon of autotrophic freshwater flagellates. In this paper, we describe and characterize four new species (<i>S. borealis</i>, <i>S. heteropora</i>, <i>S. hibernica</i> and <i>S. laticarina</i>) that have been identified during our extensive sampling of freshwater habitats in 15 European countries. Morphometric analyses of siliceous scales led to the significant phenotypic differentiation of all four newly described species, and their separation from other related species of the <i>S. petersenii</i> complex. Two of these newly described species (<i>S. hibernica</i> and <i>S. borealis</i>) can be clearly distinguished by characteristic large colonies consisting of elongated, lanceolate-shaped cells. Development of strongly elongated, narrow cells in <i>S. hibernica</i> could be explained by the adaptation of this species to oligotrophic conditions. Though morphologically distinct, <i>S. borealis</i> possesses an exceptionally high degree of genetic diversity, possibly indicating recent speciation and evolutionary diversification within this taxon. Three of the four newly described species exhibit restricted biogeographic distribution. The evolutionarily related <i>S. borealis</i> and <i>S. laticarina</i> occur only in Northern Europe, and seem to be adapted to colder areas. The most remarkable distribution pattern was observed for <i>S. hibernica</i>, which has a geographic distribution that is restricted to western Ireland.</p></div

Physico-chemical parameters and metal concentration of water at the station in Rio Irvi where <i>Coccomyxa melkonianii</i> SCCA 048 was collected.

<p>Physico-chemical parameters and metal concentration of water at the station in Rio Irvi where <i>Coccomyxa melkonianii</i> SCCA 048 was collected.</p

Ecological differentiation of putative species clusters, based on the 98-taxa dataset.

<p>Ecological differentiation of putative species clusters, based on the 98-taxa dataset.</p

Phylogeny of the genus <i>Coccomyxa</i> obtained by Bayesian inference of the concatenated SSU and ITS rDNA dataset.

<p>Values at the nodes indicate statistical support estimated by three methods—MrBayes posterior-node probability (left), maximum-likelihood bootstrap (middle), and weighted maximum parsimony bootstrap (right). Full statistical support (1.00/100/100) is marked with an asterisk. Thick branches represent nodes receiving the highest PP support (1.00). The newly sequenced strain of <i>Coccomyxa</i> SCCA 048 is marked in bold. Sequences obtained from authentic strains are marked by arrowheads. Clades belonging to four species circumscribed by Darienko et al.[<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0151137#pone.0151137.ref004" target="_blank">4</a>], i.e., <i>C</i>. <i>simplex</i>, <i>C</i>. <i>subellipsoidea</i>, <i>C</i>. <i>polymorpha</i> and <i>C</i>. <i>viridis</i>, are marked in blue, green, yellow, and red, respectively. The clades recognized are either assigned to previously described <i>Coccomyxa</i> species or labelled alphabetically based on GMYC analyses (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0151137#pone.0151137.g005" target="_blank">Fig 5</a>). Scale bar represents the expected number of substitutions per site.</p

Linear regression of measured DNA content on the chromosome data in six <i>Micrasterias</i> strains.

<p>Chromosome data published by Kasprik <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0086247#pone.0086247-Kasprik1" target="_blank">[44]</a>, (r = 0.987, R²  = 0.974, P-value<0.001).</p

DNA-Based Taxonomy in Ecologically Versatile Microalgae: A Re-Evaluation of the Species Concept within the Coccoid Green Algal Genus <i>Coccomyxa</i> (Trebouxiophyceae, Chlorophyta)

<div><p><i>Coccomyxa</i> is a genus of unicellular green algae of the class Trebouxiophyceae, well known for its cosmopolitan distribution and great ecological amplitude. The taxonomy of this genus has long been problematic, due to reliance on badly-defined and environmentally variable morphological characters. In this study, based on the discovery of a new species from an extreme habitat, we reassess species circumscription in <i>Coccomyxa</i>, a unicellular genus of the class Trebouxiophyceae, using a combination of ecological and DNA sequence data (analyzed with three different methods of algorithmic species delineation). Our results are compared with those of a recent integrative study of Darienko and colleagues that reassessed the taxonomy of <i>Coccomyxa</i>, recognizing 7 species in the genus. Expanding the dataset from 43 to 61 sequences (SSU + ITS rDNA) resulted in a different delimitation, supporting the recognition of a higher number of species (24 to 27 depending on the analysis used, with the 27-species scenario receiving the strongest support). Among these, <i>C</i>. <i>melkonianii</i> sp. nov. is described from material isolated from a river highly polluted by heavy metals (Rio Irvi, Sardinia, Italy). Analyses performed on ecological characters detected a significant phylogenetic signal in six different characters. We conclude that the 27-species scenario is presently the most realistic for <i>Coccomyxa</i> and we suggest that well-supported lineages distinguishable by ecological preferences should be recognized as different species in this genus. We also recommend that for microbial lineages in which the overall diversity is unknown and taxon sampling is sparse, as is often the case for green microalgae, the results of analyses for algorithmic DNA-based species delimitation should be interpreted with extreme caution.</p></div

Correlation of 1C DNA content and morphometric parameters.

<p>(A) average cell length, (B) number of terminal lobes, (C) cell complexity, (D) centroid size in <i>Micrasterias</i> strains. Significant correlations are given by solid lines.</p

Multigene phylogeny of <i>Synura</i> (Synurophyceae) and descriptions of four new species based on morphological and DNA evidence

<p>We used phylogenetic analyses based on multiple gene sequences (partial nr SSU and LSU rDNA, partial pt LSU rDNA, <i>psa</i>A and <i>rbc</i>L) from 148 strains (including three outgroups) and scale ultrastructure to examine phylogenetic relationships among species of the colonial genera <i>Synura</i> and <i>Tessellaria</i>. The phylogenetic tree based on the combined dataset was congruent with ultrastructural characteristics of the scales. <i>Synura</i> was divided into three major clades, two including species in section <i>Synura</i>, and one representing section <i>Peterseniae</i>. One clade, consisting of seven strains of <i>S. uvella</i> (section <i>Synura</i>), diverged at the base of the genus. The second clade consisted of the remaining species belonging to the section <i>Synura</i>. The third clade, containing organisms in the section <i>Peterseniae</i> and characterized by scales possessing a keel, was monophyletic with strong support values. Based on our findings, <i>S. uvella</i> needs to be in a separate section from other spine-bearing species, and we therefore propose new sectional ranks; <i>Synura, Peterseniae, Curtispinae</i> (presence of body scales with slender spines, tubular scales and caudal scales). We further propose four new species based on phylogenetic analyses and unique scale characters: <i>S. longitubularis</i> sp. nov., <i>S. sungminbooi</i> sp. nov., <i>S. soroconopea</i> sp. nov. and <i>S. lanceolata</i> sp. nov. Lastly, we propose a new genus name, <i>Neotessella</i>, to replace the invalid use of the name <i>Tessellaria</i>.</p

Ecological differentiation of putative species clusters, based on the 62-taxa dataset.

<p>Ecological differentiation of putative species clusters, based on the 62-taxa dataset.</p