Nataliya Rybalka

Systematics of the Green Algae: A Brief Introduction to the Current Status

Green algae are a monophyletic lineage of Archaeplastida, a supergroup of photosynthetic eukaryotes exclusively originated from primary endosymbiosis. The green algae are divided into two clades, the Chlorophyta and Streptophyta. The Chlorophyta comprises the vast majority of green algae, with three major lineages (Trebouxiophyceae, Ulvophyceae, and Chlorophyceae) forming the “crown group” of Chlorophyta and, as their early offspring, an assemblage of various monophyletic lineages of unicellular prasinophyte algae. Chloroplast phylogenomics and multigene phylogenetic analyses not only largely confirmed previous assumptions based on single gene (18S or rbcL) analyses, but also revealed novel robust groupings. While a bifurcation of the Chlorophyceae and a clear distinction of four major lineages within the Ulvophyceae have been supported, the internal phylogenetic structure and circumscription of Trebouxiophyceae still remain ambiguous. The Streptophyta comprises relatively few algal lineages and embryophytes. The embryophyte land plants are a sister group of streptophyte green algae (Zygnematophyceae or a lineage comprising Zygnematophyceae and Coleochaetophyceae), but transitions of the green algae to the land have taken place several times, even outside the Streptophyta. Progress in green algal systematics at the species level revealed ITS2 rDNA as an appropriate candidate for DNA barcoding and species distinction. Approaches to integrate sequence analyses of several genes with morphology are now state of the art for species delimitations. An improved taxon sampling with consideration of traditional gathered expert knowledge combined with multigene phylogenetic analyses, and improved phylogeny inference methods will be required to clarify areas of ambiguity in the green algal phylogenies. A revisit of morphology is essential to establish synapomorphies for the novel clades in molecular groupings.

Testing for endemism, genotypic diversity and species concepts in Antarctic terrestrial microalgae of the Tribonemataceae (Stramenopiles, Xanthophyceae)

The genetic diversity of all available culture strains of the Tribonemataceae (Stramenopiles, Xanthophyceae) from Antarctica was assessed using the chloroplast-encoded psbA /rbcL spacer region sequences, a highly variable molecular marker, to test for endemism when compared with their closest temperate relatives. There was no species endemic for Antarctica, and no phylogenetic clade corresponded to a limited geographical region. However, species of the Tribonemataceae may have Antarctic populations that are distinct from those of other regions because the Antarctic strain spacer sequences were not identical to sequences from temperate regions. Spacer sequences from five new Antarctic isolates were identical to one or more previously available Antarctic strains, indicating that the Tribonemataceae diversity in Antarctic may be rather limited. Direct comparisons of the spacer sequences and phylogenetic analyses of the more conserved rbcL gene revealed that current morphospecies were inadequate to describe the actual biodiversity of the group. For example, the genus Xanthonema, as currently circumscribed, was paraphyletic. Fortunately, the presence of distinctive sequence regions within the psbA/rbcL spacer, together with differences in the rbcL phylogeny, provided significant autoapomorphic criteria to re-define the Tribonemataceae species.

Congruence of chloroplast- and nuclear-encoded DNA sequence variations used to assess species boundaries in the soil microalga Heterococcus (Stramenopiles, Xanthophyceae).

BackgroundHeterococcus is a microalgal genus of Xanthophyceae (Stramenopiles) that is common and widespread in soils, especially from cold regions. Species are characterized by extensively branched filaments produced when grown on agarized culture medium. Despite the large number of species described exclusively using light microscopic morphology, the assessment of species diversity is hampered by extensive morphological plasticity.ResultsTwo independent types of molecular data, the chloroplast-encoded psbA/rbcL spacer complemented by rbcL gene and the internal transcribed spacer 2 of the nuclear rDNA cistron (ITS2), congruently recovered a robust phylogenetic structure. With ITS2 considerable sequence and secondary structure divergence existed among the eight species, but a combined sequence and secondary structure phylogenetic analysis confined to helix II of ITS2 corroborated relationships as inferred from the rbcL gene phylogeny. Intra-genomic divergence of ITS2 sequences was revealed in many strains. The ‘monophyletic species concept’, appropriate for microalgae without known sexual reproduction, revealed eight different species. Species boundaries established using the molecular-based monophyletic species concept were more conservative than the traditional morphological species concept. Within a species, almost identical chloroplast marker sequences (genotypes) were repeatedly recovered from strains of different origins. At least two species had widespread geographical distributions; however, within a given species, genotypes recovered from Antarctic strains were distinct from those in temperate habitats. Furthermore, the sequence diversity may correspond to adaptation to different types of habitats or climates.ConclusionsWe established a method and a reference data base for the unambiguous identification of species of the common soil microalgal genus Heterococcus which uses DNA sequence variation in markers from plastid and nuclear genomes. The molecular data were more reliable and more conservative than morphological data.

Cyanobacteria and Algae of Biological Soil Crusts

Filamentous cyanobacteria are the key organisms in biological soil crust formation in all biomes of the world. However, especially in temperate, arctic, and high alpine regions, as well as in few dry Savannah ecosystems, filamentous green algae may act in a similar role. Here, we give an overview on the role, diversity, and biogeography of cyanobacteria and eukaryotic algae in biocrusts from all climatic regions and continents of the Earth. We refer to the species level wherever this is possible. Currently, there have been 320 species of cyanobacteria and more than 350 species of eukaryotic algae described from biocrusts. Despite this high diversity, only a minority of the cyanobacterial and algal species found is responsible for the bulk of biocrust formation. Others likely are opportunistic, utilizing the habitat created by biocrusts in the harsh regions of the Earth where habitable space is rare. We also discuss methods for the sampling and identification of biocrust algae and cyanobacteria.

Biotechnological screening of microalgal and cyanobacterial strains for biogas production and antibacterial and antifungal effects.

Microalgae and cyanobacteria represent a valuable natural resource for the generation of a large variety of chemical substances that are of interest for medical research, can be used as additives in cosmetics and food production, or as an energy source in biogas plants. The variety of potential agents and the use of microalgae and cyanobacteria biomass for the production of these substances are little investigated and not exploited for the market. Due to the enormous biodiversity of microalgae and cyanobacteria, they hold great promise for novel products. In this study, we investigated a large number of microalgal and cyanobacterial strains from the Culture Collection of Algae at Göttingen University (SAG) with regard to their biomass and biogas production, as well antibacterial and antifungal effects. Our results demonstrated that microalgae and cyanobacteria are able to generate a large number of economically-interesting substances in different quantities dependent on strain type. The distribution and quantity of some of these components were found to reflect phylogenetic relationships at the level of classes. In addition, between closely related species and even among multiple isolates of the same species, the productivity may be rather variable.

Influence of Different CO2 Concentrations on Microalgae Growth, α-Tocopherol Content and Fatty Acid Composition

A prerequisite for the growth of microalgae on an industrial scale is the optimization of cultivation conditions and the reduction of production costs. The nutrient elements, heat and carbon dioxide provided by flue gas can be used to increase biomass production. The aim of this study was to investigate the effects of different CO2 concentrations on algal strains with regard to biomass production, α-tocopherol content, fatty acid composition and fatty acid content. For this, we grew the three microalgae Coccomyxa sp., Desmodesmus sp. and Muriella terrestris that were isolated from two tufa-forming karstwater creeks characterized by high CO2 partial pressures. No significant differences in biomass production rate were observed when the cultures were gassed with air or 15% (v/v) CO2. However, cell growth increased substantially when 5% (v/v) CO2 was used. The content of α-tocopherol increased in the stationary phase compared to the logarithmic phase in all investigated strains. In contrast, it decreased significantly when the cultures were aerated with either 5% (v/v) CO2 or 15% (v/v) CO2. The saturation level of fatty acids was significantly higher in cultures aerated with CO2 in the stationary phase compared to the logarithmic phase. In contrast, it decreased in the stationary phase when the cultures were gassed with air. Fatty acid contents increased in the stationary phase compared to the logarithmic phase. It was also higher by gassing with 5% (v/v) CO2 and decreased when the cultures were aerated with air or 15% (v/v) CO2. When the biomass production rates were taken into consideration together with contents of α-tocopherol and total fatty acid, the productivity of both increased considerably when the cultures experienced gassing by a 5% (v/v) CO2.

Xanthophyte, Eustigmatophyte, and Raphidophyte Algae

The Xanthophyceae, Eustigmatophyceae, and Raphidophyceae are three independent classes of stramenopile algae (Heterokontophyta or Ochrophyta); they are not closer related with each other. Most Xanthophytes are unicellular or colonial coccoid algae, others from multicellular filaments and or exhibit thalli composed of multinucleate siphons. The Eustigmatophyceae comprises only coccoid members which are very difficult to distinguish from the coccoid Xanthophytes. Freshwater Raphidophytes are rather distinct, because they form flagellated vegetative stages. The color of Xanthophytes and Eustigmatophytes is yellowish green due to the absence of the brown fucoxanthin, present in Raphidophytes and other stramenopile algae. Only the Eustigmatophytes lack chlorophyll c. Many Xanthophytes and Eustigmatophytes share that they predominately occur in terrestrial habitats, e.g. soil, representing a small group of terrestrial algae with their plastids obtained from an ancestral red alga by secondary symbiosis. For Raphidophytes only three genera are recognized in freshwater yet and they are observed within the plankton.

Screening of microalgae and cyanobacteria strains for α-tocopherol content at different growth phases and the influence of nitrate reduction on α-tocopherol production

Tocopherols (vitamin E) are only synthesized by photosynthetic organisms and have wide applications in cosmetics and as dietary supplements in human nutrition and aquaculture. Tocopherols from microalgae and cyanobacteria are rarely investigated, and little is known about their contents. Therefore, 130 strains of cultured microalgae and cyanobacteria were analyzed for α-tocopherol content under various culture conditions. The growth phase had a significant effect on content of α-tocopherol. Maximal amounts were observed at the stationary growth phase. Reduction of nitrate concentration in media caused an increased production of α-tocopherol. The contents were significantly enhanced when the nitrate concentration was reduced to one fourth in culture media used. The content of α-tocopherol was found to reflect phylogenetic relationships at the level of classes, with classes of Rhodophyta and Cyanobacteria accumulating the lowest contents. Within each class, contents varied widely at the species level emphasizing the importance of extensive screening procedures for the identification of strains with high α-tocopherol contents.

Species distinctions among closely related strains of Eustigmatophyceae (Stramenopiles) emphasizing ITS2 sequence-structure data: Eustigmatos and Vischeria

ABSTRACT There is an increasing interest in the Eustigmatophyceae, a class of stramenopile microalgae, because they offer a variety of high-value health-beneficial compounds, e.g. polyunsaturated fatty acids (PUFAs), while concomitantly producing high biomass. Clarification of the taxonomy of these organisms at the species level is important in order to achieve reproducible results and constant yields of valuable compounds in their exploitation. Here the distinction of the, so far exclusively, morphologically defined species of the genera Eustigmatos and Vischeria was tested. Distinctions inferred from almost full 18S and ITS2 rRNA as well as plastid-encoded rbcL gene sequences were evaluated following a morphological investigation. The ITS2 secondary-structure-based phylogenies separated independent lineages (species) with long internal branches. This recommends ITS2 as a promising marker for a DNA metabarcoding approach (culture-independent biodiversity assessment). In contrast, the 18S V4 region which is commonly used in metabarcoding was almost invariant, whereas the almost full length sequences distinguished eight groups/types of strains. Monophyly of the species was supported by shared ITS2 secondary structure features, making them distinct from other eustigmatophyte lineages in concordance with phylogenetic analyses. No groups of strains were congruently supported by all three markers. Consequently, the previous distinction of two genera on the basis of morphology cannot be retained and the species should be accommodated in a single genus, Vischeria. Taxonomic changes among the species with the definition of epitypes, on the basis of cryopreserved strains, are recommended. Two findings point to a more complex evolutionary history of the species. The rbcL and nuclear markers resulted in disparate groupings of strains. In three species divergent intragenomic ITS2 paralogues were revealed. Therefore, a still broader taxon sampling, in conjunction with a deep sequencing approach, is needed for a more comprehensive understanding of the complex evolution of eustigmatophyte species.