Eberhard Hegewald

Phylogenetic relationship of Chlorella and Parachlorella nov. gen. (Chlorophyta, Trebouxiophyceae)

Abstract Chlorella is one of the archetypical forms of coccoid green algae and one of the best-studied phototrophic eukaryotes. However, its systematics remains enigmatic due to conflicts between morphological and molecular phylogenetic approaches. The sequences of the 18S ribosomal RNA gene of nine strains of Chlorella and related taxa, and the ITS2 region of 17 strains of Chlorellaceae were determined and included in phylogenetic analyses. The secondary structure of the ITS2 region of C. vulgaris was compared to that of Parachlorella beijerinckii. All phylogenetic analyses showed that the Chlorellaceae form a clade within the Trebouxiophyceae. The Chlorellaceae studied here are divided into two sister groups: (1) the Parachlorella-clade with Parachlorella beijerinckii gen. et sp. nov. and P. kessleri comb. nov. as well as Diclosler acuatus and Closteriopsis acicularis; and (2) the Chlorella-clade including the ‘true’ spherical Chlorella species C. vulgaris, C. lobophora and C.sorokiniana. The latter are intermixed with taxa that differ in morphology and were formerly classified in other families of coccoid green algae: Actinastrum hantzschii (formerly Coelastraceae), Diacanthos belenophorus (formerly Oocystaceae), Dictyosphaerium pulchellum (formerly Botryococcaceae), Didymogenes anomala and D. palatina (formerly Scenedesmaceae) and Micractinium pusilium (formerly Micractiniaceae). It was shown that morphological criteria traditionally used for classification (e.g. spines, mucilaginous envelopes or the formation of colonies or coenobia) are burdened with a high degree of phenotypic plasticity. These plastic traits represent adaptations to environmental factors in order to optimize floating in the water column and to resist grazing pressure. A genus and species concept in Chlorellaceae remains provisional, because, in most cases, there is still a lack of classical and molecular approaches to identify clear phylogenetic lineages.

PHYLOGENY OF THE HYDRODICTYACEAE (CHLOROPHYCEAE): INFERENCES FROM rDNA DATA1

The hydrodictyacean green algal lineage has been the focus of much research due to the fossil record of at least some members, their ornamented cell walls, and their distinctive reproductive strategies. The phylogeny of the family was, until recently, exclusively morphology based. This investigation examines hydrodictyacean isolates from several culture collections, focusing on sequences from ribosomal data: 18S rDNA, 26S rDNA (partial), and internal transcribed spacer (ITS)‐2 data. Results from phylogenetic analyses of independent and combined data matrices support the Hydrodictyaceae as a monophyletic lineage that includes isolates of Chlorotetraedron, Hydrodictyon, Pediastrum, Sorastrum, and Tetraedron. Phylogenetic analyses of rDNA data indicate that the three‐dimensional coenobium of Hydrodictyon is evolutionarily distinct from the three‐dimensional coenobium of Sorastrum. The more robust aspects of the ITS‐2 data corroborate the 18S+26S rDNA topology and provide a structural autapomorphy for the Hydrodictyaceae and Neochloridaceae, that is, an abridgment of helix IV in the secondary structure. The rDNA data do not support monophyly of Pediastrum but rather suggest the existence of four additional hydrodictyacean genera: Monactinus, Parapediastrum, Pseudopediastrum, and Stauridium.

ITS2 sequence-structure phylogeny in the Scenedesmaceae with special reference to Coelastrum (Chlorophyta, Chlorophyceae), including the new genera Comasiella and Pectinodesmus

Hegewald E., Wolf M., Keller A., Friedl T. and Krienitz L. 2010. ITS2 sequence-structure phylogeny in the Scenedesmaceae with special reference on Coelastrum (Chlorophyta, Chlorophyceae). Phycologia 49: 325–335. DOI: 10.2216/09-61.1 Sequences and secondary structures of the nuclear-encoded internal transcribed spacer 2 (ITS2) ribosomal RNA of nine Coelastrum taxa, Asterarcys quadricellulare (Coelastraceae), Westella botryoides (hitherto Oocystaceae) and Dimorphococcus lunatus (Scenedesmaceae) were determined and compared with existing GenBank entries of scenedesmacean taxa (Desmodesmus, Enallax, Neodesmus, Scenedesmus). Phylogenetic analyses showed that the studied Coelastrum taxa belong to several different lineages within the Scenedesmaceae: five Coelastrum taxa (Coelastrum microporum, Coelastrum astroideum, C. astroideum var. rugosum  =  Coelastrum rugosum, Coelastrum pseudomicroporum and Coelastrum sphaericum incl. Coelastrum proboscideum) form monophyletic clades, whereas two strains labeled Coelastrum morum belong to different genera. The African strain of C. morum clusters with Coelastrum cambricum. The Finnish strain labeled C. morum clusters with Asterarcys, Dimorphococcus and Hariotina. According to its morphology this strain belongs to Coelastrella, related to Coelastrella saiponensis. Westella botryoides belongs to a separate clade within the Scenedesmaceae. Coelastrum reticulatum is positioned in the clade with Asterarcys, Dimorphococcus and Coelastrella; hence its separation in a separate genus, as originally described (Hariotina), is justified. In general, the phylogenetic analysis of ITS2 data shows that the Coelastraceae are included in the monophyletic Scenedesmaceae, and thus the splitting into two families is not justified, but they belong to the monophyletic subfamily Coelastroidea. The genera Comasiella and Pectinodesmus are newly erected, and several new combinations are proposed.

PHYLOGENETIC POSITION OF THE OOCYSTACEAE (CHLOROPHYTA)

The complete 18S rRNA gene sequences of three Oocystis A. Braun species (Oocystaceae) and three other chlorococcal algae, Tetrachlorella alternans (G. M. Smith) Korš. (Scenedesmaceae), Makinoella tosaensis Okada (Scenedesmaceae), and Amphikrikos cf. nanus (Fott & Heynig) Hind. (Chlorellaceae) were determined and subjected to four different phylogenetic analysis algorithms. Independent of the reconstruction method, these taxa clustered together as a monophyletic group (Oocystaceae) within the Trebouxiophyceae. This result was supported by high bootstrap values. A comparison of morphological data with the phylogenetic reconstructions indicated that the evolution of Oocystaceae was accompanied by a reduction in the number of plastids. This study fully supports the taxonomic assignment of the Oocystaceae as a distinct family. The diacritic criterion that the cell walls are composed of several cellulose layers with perpendicular fibril orientations is in accordance with the molecular data.

Taxonomy of Desmodesmus serratus (Chlorophyceae, Chlorophyta) and related taxa on the basis of morphological and DNA sequence data

Fawley M.W., Fawley K.P. and Hegewald E. 2011. Taxonomy of Desmodesmus serratus (Chlorophyceae, Chlorophyta) and related taxa on the basis of morphological and DNA sequence data. Phycologia 50: 23–56. DOI: 10.2216/10-16.1 The green alga Desmodesmus serratus is a common small Desmodesmus species. The highly variable morphology among strains and natural populations suggested that this taxon was in need of revision using a combination of molecular and morphological characters. Thirty-six isolates that were assigned to D. serratus on the basis of morphology or that were known to be closely related to D. serratus on the basis of the results of DNA sequence analysis were examined. These isolates were evaluated by light microscopy, electron microscopy, and phylogenetic analysis of DNA sequences from the nuclear ribosomal internal transcribed spacer (ITS) regions, nuclear 5.8S ribosomal DNA, and the plastid rbcL gene. Results of the analyses of ITS and rbcL sequence data were congruent. On the basis of the results of these analyses, we propose the new species Desmodesmus itascaensis, Desmodesmus perdix, Desmodesmus serratoides, Desmodesmus pseudoserratus, and Desmodesmus santosii. Two of these species, D. perdix and D. pseudoserratus, possess morphologies identical to that of D. serratus. However, these species were all clearly separated on the basis of the results of DNA sequence analysis. The new species as well as D. serratus form a monophyletic lineage that is allied with the small, spineless species, Desmodesmus costato-granulatus, Desmodesmus lunatus, Desmodesmus elegans, Desmodesmus ultrasquamatus, and Desmodesmus regularis.

Classification of crucigenoid algae: phylogenetic position of the reinstated genus Lemmermannia, Tetrastrum spp. Crucigenia tetrapedia, and C. lauterbornii (Trebouxiophyceae, Chlorophyta)1

The subfamily Crucigenioideae was traditionally classified within the well‐characterized family Scenedesmaceae (Chlorophyceae). Several morpho‐logical revisions and questionable taxonomic changes hampered the correct classification of crucigenoid species resulting in a high number of synonymous genera. We used a molecular approach to determine the phylogenetic position of several Tetrastrum and Crucigenia species. The molecular results were correlated with morphological and ontogenetic characters. Phylogenetic analyses of the SSU rDNA gene resolved the position of Tetrastrum heteracanthum and T. staurogeniaeforme as a new lineage within the Oocystis clade of the Trebouxiophyceae. Crucigenia tetrapedia, T. triangulare, T. punctatum, and T. komarekii were shown to be closely related to Botryococcus (Trebouxiophyceae) and were transferred to Lemmer‐mannia. Crucigenia lauterbornii was not closely related to the other Crucigenia strains, but was recovered within the Chlorella clade of the Trebouxiophyceae.

A revision of the species Desmodesmus perforatus and D. tropicus (Scenedesmaceae, Chlorophyceae, Chlorophyta)

S.L. Jeon and E. Hegewald. 2006. A revision of the species Desmodesmus perforatus and D. tropicus (Scenedesmaceae, Chlorophyceae, Chlorophyta). Phycologia 45: 567–584. DOI: 10.2216/05-63.1 We have made a synopsis of the morphology, ultrastructure, taxonomy, distribution and genetics of two widely distributed but little known species because this is important for the general understanding of species delineation in green algae. The taxa Desmodesmus perforatus and D. tropicus have in common central gaps between coenobial cells, but otherwise according to the type descriptions and type illustrations, they are morphologically well defined and easy to distinguish. However, in screening the literature, the situation becomes confusing and a clear delineation seems to be impossible. To solve the question of whether the two species are in fact different, or extremes of a single morphologically variable taxon, we isolated strains from different localities worldwide as well as strains of the related taxon D. maximus. The strains were studied under both optical and electron microscopes (EM) and the ITS-2r DNA of available strains was compared. With the results of the studied strains, and a comparison with natural populations, we could delimit the taxa D. perforatus var. perforatus, D. perforatus var. iberaënsis and D. tropicus and revise the literature data. Under the optical microscope, populations are clearly distinguishable, although single specimens may not be. According to ITS-2 rDNA all strains of D. perforatus (except Hegewald 1998–18) and D. tropicus differ by 10 bases and D. perforatus var. perforatus and D. perforatus var. iberaënsis by 2 bases. Regarding the compensatory base changes (CBC), there are zero exchanges between D. perforatus and D. tropicus (except strain Hegewald 1998–18) but D. tropicus and D. perforatus var. iberaënsis have one CBC. A tepwise xchange of bases was shown for CBCs. Other accepted taxa are D. perforatus f. bicaudatus (Compère) E. Hegewald, D. tropicus var. longiclathratus (Tell) S.L. Jeon & E. Hegewald and D. perforatus var. mirabilis (Massjuk) E. Hegewald. A sister group to D. perforatus and D. tropicus is D. maximus. This taxon is similar in EM cell wall structure to D. perforatus and D. tropicus, but has no holes between the cells.

Phylogenetic placement of Chlorotetraedron incus, C. polymorphum and Polyedriopsis spinulosa (Neochloridaceae, Chlorophyta)

Abstract We investigated unialgal strains of the type species of the genera Chlorotetraedron and Polyedriopsis, C. polymorphum and P. spinulosa, and also of C. incus, by means of 18S rRNA analyses. According to the phylogenetic analyses, C. incus, C. polymorphum and P. spinulosa are closely related and belong to the Chlorophyceae, family Neochloridaceae; this contrasts with the previous classification of Polyedriopsis in the Golenkiniaceae.

Desmodesmus baconii (Chlorophyta), a new species with double rows of arcuate spines

Fawley M.W, Fawley K.P. and Hegewald E. 2013. Desmodesmus baconii (Chlorophyta), a new species with double rows of arcuate spines. Phycologia 52: 565–572. DOI: 10.2216/12-116.1 A new species of the chlorophycean genus Desmodesmus, D. baconii, was described based upon analyses of the morphology of the coenobia and DNA sequences from the nuclear ribosomal 18S RNA gene and the internal transcribed spacer region. Desmodesmus baconii was unusual in that it possessed two rows of spines only on the terminal cells. This species was easily distinguished from other Desmodesmus taxa with two rows of spines by the point symmetry of the coenobia and the lack of rows of spines on median cells of four-celled coenobia. Spines tapered from prominent incurved spines near one pole to small spines at the opposite pole. Median cells and the pole of terminal cells opposite the large spines had two or three short spines. Cell sizes were slightly larger for two-celled (4.2–6.7 μm × 1.6–2.9 μm) than for four-celled coenobia (3.2–6.5 μm × 1.2–4.5 μm). Results of analyses of DNA sequence data indicated that D. baconii, although a member of the monophyletic genus Desmodesmus, was not closely allied with any other Desmodesmus taxon. Desmodesmus baconii was isolated from hypereutrophic Lake Chicot in Arkansas, USA, and is not known from any other location.