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Nearly complete (> 97%) chloroplast-encoded rbcL gene sequences were determined for Bolidomonas mediterranea from the eastern Mediterranean Sea and two strains of B. pacifica var. eleuthera from the equatorial Pacific Ocean. The three sequences were added to an rbcL data matrix comprising 45 taxa representing 11 of the 12 recognized classes of Hetero-kontophyta. Phylogenetic analyses that used maximum parsimony (MP), maximum likelihood (ML) and neighbour-joining (NJ) algorithms supported their sister group relationship to the Bacillariophyceae. Centric and pennate diatoms and the three novel Bolidomonas rbcL sequences share a two-amino-acid insertion at position 1336, relative to the homologous position in the complete sequence for the centric diatom Odontella sinensis. This synapomorphic character state further supports a common evolutionary history of bolidophyceans and diatoms. Chloroplast- and nuclear-encoded molecular markers thus favour the idea that the ancestor of the diatoms possessed tw...

International audience; Bolidomonas is a genus of picoplanktonic flagellated algae that is closely related to diatoms. Triparma laevis, a species belonging to the Parmales, which are small cells with a siliceous covering, has been shown to form a monophyletic group with Bolidomonas. We isolated several novel strains of Bolidophyceae that have permitted further exploration of the diversity of this group using nuclear, plastidial and mitochondrial genes. The resulting phylogenetic data led us to formally emend the taxonomy of this group to include the Parmales within the Bolidophyceae, to combine Bolidomonas within Triparma and to define a novel species, Triparma eleuthera sp. nov. The global distribution of Bolidophyceae was then assessed using environmental sequences available in public databases, as well as a large 18S rRNA V9 metabarcode data set from the Tara Oceans expedition. Bolidophyceans appear ubiquitous throughout the sampled oceans but always constitute a minor component of the phytoplankton community, corresponding to at most ~ 4% of the metabarcodes from photosynthetic groups (excluding dinoflagellates). They are ~ 10 times more abundant in the small size fraction (0.8– 5 μm) than in larger size fractions. T. eleuthera sp. nov. constitutes the most abundant and most widespread operational taxonomic unit (OTU) followed by T. pacifica, T. mediterranea and the T. laevis clade. The T. mediterranea OTU is characteristic of Mediterranean Sea surface waters and the T. laevis clade OTU is most prevalent in colder waters, in particular off Antarctica.

Diatoms are perhaps the most diverse lineage of eukaryotic algae, with their siliceous cell wall and diplontic life history often considered to have played important roles in their extraordinary diversification. The characteristic diminution of the diatom cell wall over the course of vegetative growth provides a reliable, intrinsic trigger for sexual reproduction, establishing a direct link between the evolution of their cell-wall and life-history features. It is unclear, however, whether the diplontic life cycle of diatoms represents an ancestral or derived trait. This uncertainty is based in part on our lack of understanding of the life cycle of the sister lineage to diatoms, which includes a mix of two free-living and separately classified forms: naked biflagellate unicells in the genus Bolidomonas and silicified forms in the order Parmales. These two forms might represent different life-history stages, although directly establishing such links can be difficult. We sequenced transcriptomes for Bolidomonas and two diatoms and found that ~0.1% of the coding regions in the two diploid diatoms are heterozygous, whereas Bolidomonas is virtually devoid of heterozygous alleles, consistent with expectations for a haploid genome. These results suggest that Bolidomonas is haploid and predict that parmaleans represent the diploid phase of a haplodiplontic life cycle. These data fill an important gap in our understanding of the origin of the diplontic life history of diatoms, which may represent an evolutionarily derived, adaptive feature.

The diversity and abundance of the Bolidophyceae (Heterokonta), a newly described picoplanktonic algal class which is a sister group to the diatoms, was assessed in the equatorial Pacific Ocean and in the Mediterranean Sea by culture isolation, molecular biology techniques, and pigment analyses. Eight strains of Bolidophyceae were isolated in culture from different mesotrophic and oligotrophic areas. The corresponding small subunit (SSU) rRNA gene sequences allowed us to design two probes specific for the Bolidophyceae. These probes have been used in natural samples (i) to selectively amplify and detect Bolidophyceae sequences and (ii) to quantify the relative abundance of Bolidophyceae within the picoeukaryote community. Sequences available to date indicate that the class Bolidophyceae comprises at least three different clades, two corresponding to the previously described species Bolidomonas pacifica and Bolidomonas mediterranea and the third one corresponding to a subspecies of B. pacifica . Amplification of the SSU rRNA gene from natural samples with universal primers and hybridization using a Bolidomonas -specific probe followed by a eukaryote-specific probe allowed us to estimate the contribution of the Bolidophyceae to the eukaryotic DNA in both Pacific and Mediterranean waters to be lower than 1%. Similarly, high-performance liquid chromatography analyses of fucoxanthin, the major carotenoid present in Bolidophyceae, indicated that less than 4% of the total chlorophyll a in the picoplanktonic fraction in the equatorial Pacific was due to Bolidophyceae. Consequently, although strains of Bolidophyceae have been isolated from samples collected at several stations, this new class seems to have been a minor component of the natural picoeukaryotic populations in the ecosystems investigated, at least during the periods sampled.

The ecophysiology of the photoprotective xanthophyll cycle (XC) was compared in six chlorophyll c-containing pico- and nano- phytoplankton species. Different accessory pigment combinations and ecological properties characterize the six studied species, Bolidomonas mediterranea, Pelagomonas calceolata, Phaeocystis cordata, Phaeocystis sp.(strain RCC186), Mesopedinella arctica and Ochromonas sp. The first experimental set consisted in the study of the activity of the xanthophyll cycle and non-photochemical quenching (NPQ) during two gradual light increases, with photon flux density (PFD) ranging from 40 to 400 mu mol photons m(-2) s(-1). Pigments, absorption spectra, flow cytometry measurements for cell counts and chlorophyll a autofluorescence and Electron Transport Rate (ETR) vs. light curves were determined at different times during the experiment. The second set of experiments consisted in using two inhibitors: carotenogenesis inhibitor (norflurazon) and deepoxidation step (occurring in the xanthophyll cycle) inhibitor (dithiotreitol) during high-light shift, to compare the functioning of the xanthophyll cycle among the different species. Results highlighted a large diversity in the xanthophyll cycle functioning, possibly related to the ecological traits of the species. In view of the results, three groups of species have been identified as (i) highlight-adapted, (ii) low light-adapted and (iii) variable light-adapted species. (C) 2009 Elsevier GmbH. All rights reserved.

In late 2004, the BIOSOPE cruise sailed between the equatorial influenced waters off the Marquesas Islands and the nutrient enriched waters of the Chilean upwelling. Along the way, it explored the Southeast Pacific gyre centred around Easter Island, which is probably the most oligotrophic oceanic region on earth. During this cruise, we undertook a vigorous effort to isolate novel photosynthetic picoplanktonic eukaryotes. Two strategies were attempted on board: enrichment of filtered samples with culture medium and sorting of specific populations by flow cytometry based on size and chlorophyll fluorescence. Over 1900 pre-cultures were started and then further purified by flow cytometry, serial dilution or pipette isolation to yield a total of 212 strains. These strains were characterized morphologically and for more than 50% of them, genetically, through partial sequencing of the 18 S rRNA gene. Among the characterized strains, the largest number belongs to stramenopiles (Heterokontophyta) with a record of 38 strains belonging to the species Pelagomonas calceolata (Pelagophyceae). Strains from the recently described genera Bolidomonas and Florenciella have been re-isolated for the first time since their description. Two other abundant groups are the Chlorophyta, especially Prasinophyceae, and the Haptophyta, especially the genera Phaeocystis and Emiliania. A limited number of heterotrophic flagellates have also been isolated, all of them belonging to groups containing known species. Finally, over a dozen of unicellular cyanobacterial Synechococcus strains have been obtained, some forming unusual short chains. Overall our strategy was quite successful since it allowed us to isolate a large number of picoplankton strains. Still it failed in two respects. First, apparently very few novel taxa have been obtained. One set of strains is related to Prasinoderma coloniale (Prasinococcales, Prasinophyceae) but their sequences are sufficiently different from the latter to probably belong to a new genus or species. The sequences of two other strains, unfortunately later lost, were phylogenetically affiliated to stramenopile environmental sequences, probably corresponding to a new algal class. Second, very few strains have been obtained from the very oligotrophic central gyre itself. In order to be successful, future work in similar waters should probably combine flow cytometry sorting with culture media and cultivation approaches specifically developed for oligotrophic water species.

International audience; A new algal class, the Bolidophyceae (Heterokonta), is described from one genus, Bolidomonas, gen, nov., and two species, Bolidomonas pacifica, sp, nov and Bolidomonas mediterranea, sp, nov., isolated from the equatorial Pacific Ocean and the Mediterranean Sea, respectively. Both species are approximately 1.2 mu m in diameter and have two unequal flagella; the longer flagellum bears tubular hairs, whereas the shorter is smooth. The flagellar basal apparatus is restricted to two basal bodies, and there is no transitional helix. Cells are naked, devoid of walls or siliceous structures. The internal cellular organization is simple with a single plastid containing a ring genophore and a girdle lamella, one mitochondrion with tubular cristae, and one Golgi apparatus close to the basal bodies. The Mediterranean and the Pacific species differ in the insertion angle between their flagella and their pattern of swimming, these differences possibly being linked to each other. Analyses of the SSU rDNA gene place the two strains as a sister group to the diatoms, Moreover, pigment analyses confirm this position, as fucoxanthin is found as the major carotenoid in both lineages. These data strongly suggest that the ancestral heterokont that gave rise to the diatom lineage was probably a biflagellated unicell.