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International audience; In the Arctic Ocean, the small green alga Micromonas polaris dominates picophytoplankton during the summer months but is also present in winter. It has been previously hypothesized to be phago-mixotrophic (capable of bacteria ingestion) based on laboratory and field experiments. Prey uptake was analyzed in several M. polaris strains isolated from different regions and depths of the Arctic Ocean and in Ochromonas triangulata, a known phago-mixotroph used as a control. Measuring ingestion of either fluorescent beads or fluorescently labeled bacteria by flow cytometry, we found no evidence of phago-mixotrophy in any M. polaris strain while O. triangulata was ingesting both beads and bacteria. In addition, in silico predictions revealed that members of the genus Micromonas lack a genetic signature of phagocytotic capacity.

Two-dimensional (2-D) protein analysis of the mastigoneme fraction of the chromophyte alga Ochromonas danica E. G. Pringsh. showed the presence of several component proteins of the tubular mastigoneme. Adding to the reported gene Ocm1, three new genes (Ocm2, Ocm3, and Ocm4) belonging to the Ocm gene family were isolated using degenerate primers designed from predicted Ocm1 amino acid sequences. The predicted polypeptides encoded by Ocm2, Ocm3, and Ocm4 were smaller in size than Ocm1. However, they shared four highly conserved, cysteine-rich, epithelial growth factor (EGF)-like motifs, potentially involved in protein-protein interaction. In addition, Ocm2, Ocm3, and Ocm4 showed homology to the SIG protein family in the centric diatom Thalassiosira weissflogii (Grunow) Fryxell et Hasle, which is up-regulated during early stages of sexual reproduction. Immunofluorescence analysis with a polyclonal antibody against the partial amino acid sequences of Ocm2, Ocm3, and Ocm4 showed that Ocm2 and Ocm3 were located in the basal segment region of mastigonemes attached on the surface of the anterior flagellum, and that Ocm4 was located within the tubular shaft portion similar to Ocm1.

The phylogenetic group stramenopiles refers to the systematic groups that possess tripartite tubular hairs (stramenopiles) on their flagella. There have been a number of studies describing the fine structure of these mastigonemes and a few studies isolating the component proteins; however, these proteins and their gene sequences have not yet been identified. In the present study, we identified a mastigoneme protein (Ocm1) of the chrysophycean alga Ochromonas danica Pringsh. (UTEX LB1298). Its corresponding gene, Ocm1, was identified by using degenerate primers that correspond to the partial amino acid sequences of a protein (85 kDa) obtained from a mastigoneme-rich fraction of isolated flagella. The polypeptide encoded by Ocm1 has four cysteine-rich, epithelial growth factor (EGF)-like motifs, potentially involved in protein-protein interactions. It lacks obvious hydrophobic regions characteristic of transmembrane domains, suggesting that this polypeptide is not likely a protein for anchoring the mastigoneme. In addition, a polyclonal antibody against Ocm1 labeled the area where the tubular shafts of the mastigonemes are located, but not the basal portion or the terminal filaments.

The growth and grazing characteristics of Poterioochromonas malhamensis (Pringsheim) Peterfi (= Ochromonas malhamensis Pringsheim) (ca. 8 μm) feeding on phytoplankton, including the cyanobacteria Synechococcus sp. (ca. 2 μm) and Microcystis viridis (A. Brown) Lemmermann (ca. 6 μm) and the green alga Chlorella pyrenoidosa Chick (ca. 13 μm), were investigated in laboratory experiments involving the following treatments: (1) light without added algal prey (autotrophy), (2) light with added algal prey (mixotrophy), and (3) dark with added algal prey (phagotrophy). There were significantly higher cell numbers under mixotrophic and phagotrophic growth than under autotrophic growth. With phytoplankton as food, growth rates under both mixotrophy and phagotrophy were about two or three times higher than those under autotrophy, indicating that the algal diets were readily able to support the population growth of P. malhamensis. There were no significant differences in growth rate between mixotrophic and phagotrophic cultures during exponential growth. The ingestion rate of P. malhamensis with algal prey was also similar under both continuous light and dark. Poterioochromonas malhamensis ingested on average 0.27 M. viridis cells·flagellate−1·h−1 and 0.18 C. pyrenoidosa cells·flagellate−1·h−1 in continuous light and 0.25 M. viridis cells·flagellate−1·h−1 and 0.18 C. pyrenoidosa cells·flagellate−1·h−1 in continuous dark during exponential growth. The results showed that light had no effect on the growth and ingestion rates of P. malhamensis for phagotrophy during exponential growth. However, phagotrophic populations of P. malhamensis were incapable of growth in continuous darkness for longer than 5 days. Populations of P. malhamensis showed no increase when prey was added again after 4 days in continuous darkness, indicating that light is necessary for sustained phagotrophic growth of P. malhamensis. The study suggests that P. malhamensis, which has strong tolerance for light, is light dependent for phagotrophy.

Epifluorescence microscopy reveals the presence of fluorescence in the living cells of at least three classes of flagellates. In Ochromonas cells, the fluorescence is blue-green in color and is found only in the short flagellum, both in the flagellar swelling and throughout the length of the flagellum. As recognized by the locale and color of the flagellar fluorescence, the same fluorescence is observed in only certain other heterokont algal groups but is also found in one of the two isokont flagella of the prymnesiophyte Prymnesium parvum.

The 16S-like ribosomal RNA genes from Mallomonas papillosa Harris et Bradley (Synurophyceae) and Tribonema aequale Pascher (Xanthophyceae) were sequenced and compared to those of other eukaryotes. Mallomonas is closely related to Ochromonas (Chrysophyceae) and supports the general hypothesis of a close phylogenetic relationship between the Synurophyceae and Chrysophyceae. Tribonema is specifically related to Costaria costata (C. A. Agardh) Saunders (Phaeophyceae) demonstrating an unexpected phylogenetic relationship between the Xanthophyceae and Phaeophyceae. Distance and parsimony analysis place these four chromophyte genera in a complex evolutionary assemblage that includes the Bacillariophyceae and Oomycetes but excludes the Dinophyceae. The close relationship between the chromophyte algae and the Oomycete fungi supports the hypothesis that protists with tripartite hairs form a natural assemblage.

Colorado and Wyoming lakes with high pH values generally ranging from 8 to 10 are dominated by phytoplankton belonging to the chrysophytes, diatoms and cryptomonads. Our studies on phytoplankton diversity in Dowdy, South Delaney Buttes and Cowdrey Lakes in Colorado and Diamond and Twin Buttes Lakes in Wyoming have yielded several isolates of rare or new chrysophytes. These isolates were examined with light microscopy, SEM, TEM, and novel information regarding their structure is presented. Proposed new species belong to the genera Saccochrysis, Prymnesium and Chrysochromulina. Chrysophytes that were examined for the first time include Monochrysis aphanaster, Uroglenopsis, and a mixotrophic species of Ochromonas, which ingests small diatoms. Probable new species are the following: Saccochrysis sp. nov. differs in chloroplast structure, cell shape and cell number/colony, and absence of a posterior vacuole; Prymnesium sp. nov. is the first freshwater form examined with EM and differs in scale morphology; and Chrysochromulina sp. nov. is similar to Chrysochromulina parva but lacks scales. The ultrastructure of M. aphanaster is unique and appears to be a member of the Chrysophyceae. It has two flagella, one of which is emergent, while the other is hidden within an invagination of the cytoplasm. Diacronema and Sarcinochrysis spp. from these lakes also were isolated and reinvestigated, and some unique cellular features will be discussed.

Ochromonas sensu lato is the largest genus in the Chrysophyceae, containing over 100 names. Ochromonas species are biflagellate, naked, plastid-bearing single cells, distinguished from loricate, scaled, colonial and colorless genera. Most, if not all, species of Ochromonas are mixotrophic, i.e., they photosynthesize but they also engulf bacteria and other small prey. Preliminary evidence from SSU rRNA sequences show that Ochromonas is a polyphyletic genus. Ochromonas tuberculata is the most distinct from all other Ochromonas species. The other Ochromonas species (examined thus far) are scattered in three clades. For example, O. danica and O. sphaerocystis are sister to Poterioochromonas stipitata and P. malhamensis. Four additional species (identified by light microscopy as O. elegans Doflein, O. globosa Skuja, O. ovalis Dolfein, O. sociabilis Pringheim) have SSU rRNA sequences identical to P. malhamensis. Of these, only O. sociabilis has been transferred to Poterioochromonas. Thus, at least some species may be synonymous with others. Two clades of marine species are also known, one containing coastal species and the other containing open ocean species. A number of genera (some also polyphyletic) are interspersed amongst the Ochromonas species (e.g., Chrysolepidomonas, Chrysonephele, Chrysoxys, Cyclonexis, Dinobryon, Epipyxis, Uroglena, Uroglenopsis). The goal of this research (just beginning) is to establish a monophyletic Ochromonas, probably by assigning some species to other genera (existing or new). One major problem is that the type species, O. triangulata Vysotskii, hasn't been observed in over 100 years, and it is unclear which of several clades of Ochromonas contains the type. Results will be discussed.

The cysts (statospores) of Ochromonas tuberculata Hibberd are produced within a cytoplasmic silica deposition vesicle (SDV) whose membrane (silicalemma) appears to be formed by the coalescence of golgi vesicles. Silica is first deposited as small nodules and the collar and spines develop by centrifugal growth only after a complete but still thin wall has been laid down. Small vesicles appear to be attached to the SDV only in the region overlying the developing collar; a cap of radially arranged, moderately electron-dense material occurs at the tip of the growing spines. The cyst pore is formed at the anterior end of the flagellate cell, by lack of silica deposition over a small region of the SDV and rupture of the SDV and other membranes crossing this region. When the cyst wall is complete, an extracystic plug is formed in the pore, resulting in the loss of some extracystic cytoplasm and the plasmalemma, and the inner SDV membrane becomes the functional plasmalemma. The plug develops first by coalescence with the cell membrane of golgi-derived vesicles containing dense but apparently nonsiliceous spicules surrounded by amorphous material. During later stages of plug formation only fibrous material is deposited, some of which may be extruded through the pore forcing out some of the spiculate component. Scanning electron micrographs of the mature wall show it is smooth except for the concentrically wrinkled inner face of the flared collar and that the real pore diameter is only ca. half that of the collar. At germination the plug completely disappears in an unknown way and a single cell, similar to a normal vegetative cell emerges through the pore. Chrysophycean cyst formation generally resembles cell wall formation in diatoms, but differs in some details.

SUMMARY Ochromonas danica Prings., a chrysomonad alga which demonstrates a high degree of nutritional versatility, is capable of feeding on the toxic blue-green alga Microcystis aeruginosa Kuetz. In this paper light microscopic, electron microscopic, and cytochemical examinations of endocytosis in O. danica are reported, with particular emphasis on the vicissitudes of endocytic and lysosomal activities during intra-cellular digestion. An interpretation of the function of organelles associated with endocytosis is presented.

SUMMARY The following marine chrysomonads, originally described by Carter, are redescribed: Pseudopedinella pyriforme, Ochromonas oblonga, and Pyrmnesium parvum. Parachrysiclalis n.g. is set up for P. estuariale, with a flattened body and 2 equal flagella inserted in the middle of one side. Redescribed Xanthophy-cean species include Nephrochloris salina and Olis-thocliscus luteus. Olisthodiscus carterae and O. magnus are new species, the first distinguished by the position of the flagella and large number of chromatophores, O. magnus by its size (30-18 μ X 15-25 μ) and anteriorly inserted but posteriorly trailing flagellum. Among Cryptophyceae, Chroomonas vectensis Carter, Rhodomonas amphioxeia Conrad, and Cryptomonas salina and C. stigmatica Wislouch are redescribed along with Rhodomonas minuta v. apyrenoidosa var. nov. Redescribed Chlorophyta are Bipedinomonas pyriformis Carter, Pyramimonas obovata Carter, P. amylifera Conrad, and what may be Tetra-selmis suecica (Kylin) Butcher. The loricate, probably colorless Calycomonas gracilis (Lohmann) Wulff is redescribed.

SUMMARY Ochromonas danica grown on a chemically defined medium under controlled conditions in the light synthesized the following vitamins: ascorbate, B6, N5-methyltetrahydrofolate, tetrahydrofolate polyglutamates, oxidized folate monoglutamates, nicotinate, pantothenate, riboflavin, vitamin A, β-carotene, and vitamin E but no vitamin. B12. The cells also secreted molecules into their growth medium including the vitamins ascorbate, B6, the above folates, nicotinate, pantothenate, riboflavin, vitamin E, and the amino acids alanine, aspartic acid, leucine, and valine. The role of such secretions in nature is discussed.

SUMMARY Ochromonas danica grown in a chemically defined medium under controlled conditions at different incubation temperatures showed marked changes in chemical composition and secretory pattern. As the incubation temperature rose from 15 to 30 C, the cell number per unit volume of medium increased; the cell mass reached a maximum and then declined slightly, and the weight per single cell decreased. The chemical composition of the cells changed with increasing temperature: carbohydrates decreased dramatically; proteins quadrupled at 20 C and then declined 25%; lipids increased; and RNA increased to 25 C and then decreased to about 40% of maximum at 30 C. The extracellular secretion by the cells of carbohydrates seemed to increase and then decline; RNA decreased; proteins increased; and lipids remained approximately constant, as the temperature increased. The implications of these findings for thermal pollution are discussed.