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The biodiversity of terrestrial algae is still grossly understudied, and African deserts in particular are barely touched in this respect. Here, four coccoid green algae from oases in the Western Desert of Egypt were characterized using a combination of morphotaxonomic, ecological and 18S rDNA data, with additional carotenoid and lipid analyses for two of the strains. Three strains were identified as affiliated with known taxa: Mychonastes sp., Asterarcys sp. (first report of this genus from a desert soil), and Stichococcus cf. deasonii. The fourth strain is proposed to represent a new cryptic genus Pharao gen. nov., with the type species P. desertorum sp. nov. The new taxon is sister to the clade of uncharacterized North American desert strains of Radiococcaceae (Chlorophyceae, Chlorophyta). The pigment profile of P. desertorum gen. et sp. nov. revealed carotenoids and chlorophylls typical of green algae. Bioorganic analysis showed a complex lipidome based on phospho- (PC), galacto- (MGDG and DGDG), betaine- (DGTS), and sulfoquinovosyl- (SQDG) membrane lipids, besides significant amounts of storage neutral lipids such as diacyl- (DAG) and triacylglycerols (TAG). The presence of saturated alkyl chains within all the membrane lipid classes in P. desertorum and Asterarcys sp. appears to reflect the need to maintain membrane fluidity and viscosity. In summary, African deserts likely still harbor new taxa to be described, and lipidomic analyses of such taxa may provide clues about their ability to survive in the extremely harsh desert habitats.

As Scenedesmus komarekii Hegewald was cultured under high light intensity and nitrogen limitation, the color of cells progressed from green to brown, and finally through orange to brick red. The secondary carotenoids astaxanthin and canthaxanthin, as well as apolar carotenoids, were detected in the brown and orange cells. These carotenoids were contained in lipoidal globules that were first formed at the periphery of the cell and progressively propagated toward its inside, eventually filling most of it. The chloroplast was single and parietal in the green cells. As the cells turned brown, the chloroplast divided into several small lobes and was pushed toward the interior by the accumulating lipoidal globules. Sometimes the outer layer of the wall of the brown cell developed one or two diametrically opposed swellings. Once the cells became orange or red, neither lipoidal globules nor any major organelles were distinguishable. The cell wall in the orange cells became thick because of the formation of electron-dense granules between its outer and inner layers. The mode of the secondary carotenoid accumulation in S. komarekii differs from that of Haematococcus, an alga well known for its ability to accumulate secondary carotenoids, but resembles that of “Chlorella”zofingiensis (=Mychonastes zofingiensis (Donz) Kalina et Puncochařova).

Nuclear-encoded small subunit ribosomal RNA gene (18SrDNA) sequences were determined for Coelastrella multistriata (Trenkwalder) Kalina et Puncochařova, two species of Scotiellopsis (S. oocystiformis (Lund) Kalina et Puncochařova and S. terrestris (Reisigl) Kalina et Puncochařova) and two species of Muriella (M. aurantiaca Vischer and M. terrestris Boye- Petersen). Coelastrella and Scotiellopsis are members of the subfamily Scotiellocystoideae, and Muriella is a member of the subfamily Chlorelloideae in the family Chlorellaceae. Phylogenetic trees were constructed based on these sequence data and on previously known 18SrDNA sequences of 25 taxa. Coelastrella and Scotiellopsis were closely related to each other and formed a cluster with Scenedesmus vacuolatus (Shihira et Krauss) Kessler et al. This cluster shared a monophyletic ancestry with other Scenedesmus species. Muriella aurantiaca formed a sister relationship with the monophyletic lineage of Scenedesmus. However, another species, M. terrestris, was placed in the Trebouxiophyceae and was strongly related to Chlorella. The genus Mychonastes belonging to the Scotiellocystoideae was also not monophyletic. This study suggests that the subfamily Scotiellocystoideae should be removed from the Chlorellaceae.

Species of Mychonastes, Pseudodictyosphaerium, and Korshpalmella all have spherical (or nearly so) coccoid cells without pyrenoids. Mychonastes and Pseudodictyosphaerium are apparently frequent components of the picoplankton of lakes, although they had not been reported from North America prior to our studies. We have examined several isolates of Mychonastes, Pseudodictyosphaerium, and Korshpalmella from lakes and ponds in Itasca State Park, Minnesota (USA) and Arrowwood National Wildlife Refuge, North Dakota (USA). These isolates comprise 5 different 18S rDNA sequence types, none of which have a sequence identical to any published sequence for Mychonastes or Pseudodictyosphaerium. Four of these 18S types produce solitary cells, consistent with Mychonastes spp. However, other isolates all have identical 18S rDNA sequences, but are either unicellular, colonial with connecting stalks, or colonial in an amorphous matrix, consistent with Mychonastes spp., Pseudodictyosphaerium spp. or Korshpalmella spp, respectively. For some isolates, the colonial character is plastic, with different morphologies seen with different growth conditions. Ribosomal ITS and rbcL sequences also indicate little or no diversity among the isolates with various morphologies. These results suggest that the colonial habit is not a reliable generic character. Phylogenetic analyses of both rbcL and 18S rDNA sequences indicate that Mychonastes and Pseudodictyosphaerium spp. are a distinct lineage within the Chlorophyceae and should be placed in a new family. This material is based upon work supported by the National Science Foundation under Grant Nos. DEB-0128953, DEB-0129030, DBI-0070387 and MCB-0084188.

The coccoid algae (a.k.a ‘little green balls’) are important primary producers in freshwater communities, but very little is known about their diversity. This lack of knowledge results from the extremely small size and simple morphology of these organisms, which makes identification by light microscopy essentially impossible. However, molecular techniques allow differentiation of taxa at various levels and can ultimately lead to species identification. We have isolated several hundred cultures of coccoid algae from lakes and ponds in Itasca State Park in north-western Minnesota and Arrowwood National Wildlife Refuge in central North Dakota. Many of these isolates have been examined by light microscopy, 18S rDNA PCR–RFLP analysis, and 18S rDNA sequence analysis. In addition, some taxa with multiple isolates have been examined by rbcL and ribosomal RNA ITS sequence analysis. Our results indicate a high level of diversity, including possible new species of Chlorella, Muriella, Pseudomuriella and Mychonastes, among others. We will discuss the results of our analyses, including implications for identification of taxa and higher level systematics. Supported by National Science Foundation Grants DBI-00703867, MCB-0084188 and DEB-0128952.