Author Correction: Chloropicophyceae, a new class of picophytoplanktonic prasinophytes.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Ontogenetic analysis of siliceous cell wall formation in Triparma laevis f. inornata (Parmales, Stramenopiles).

Triparma laevis f. inornata is a unicellular alga belonging to the Bolidophyceae, which is most closely related to diatoms. Like diatoms, T. laevis f. inornata has a siliceous cell wall. The cell wall of T. laevis f. inornata consists of four round plates (three shields and one ventral plate) and one dorsal and three girdle plates. But, unlike diatoms, T. laevis f. inornata cells can grow when concentrations of silica are depleted. We took advantage of this ability, using TEM to study the ontogeny of the siliceous plate, pattern center formation, and development. Two types of pattern centers (annulus and sternum) were observed in the early and middle stage of plate formation. During their formation, the annuli were initially crescent-shaped but eventually their ends fused to make a ring. Only outward silica deposition of the branching ribs occurred on the growing annulus until it became a ring, resulting in an unfilled circle inside the annulus. The pattern center of the shield plate was always an annulus, but in ventral plates both annulus and sternum were observed. The annuli and sterna in T. laevis f. inornata round plates were very similar to the annuli and sterna in diatom valves. These results suggested that the round plates of Parmales are homologous to diatom valves. This information on the plate ontogeny of T. laevis f. inornata provides new insights into the evolution of the siliceous cell wall in the Parmales and diatoms.

Chloropicophyceae, a new class of picophytoplanktonic prasinophytes.

Prasinophytes are a paraphyletic group of nine lineages of green microalgae that are currently classified either at the class or order level or as clades without formal taxonomic description. Prasinophyte clade VII comprises picoplanktonic algae that are important components of marine phytoplankton communities, particularly in moderately oligotrophic waters. Despite first being cultured in the 1960s, this clade has yet to be formally described. Previous phylogenetic analyses using the 18S rRNA gene divided prasinophyte clade VII into three lineages, termed A, B and C, the latter formed by a single species, Picocystis salinarum, that to date has only been found in saline lakes. Strains from lineages A and B cannot be distinguished by light microscopy and have very similar photosynthetic pigment profiles corresponding to the prasino-2A pigment group. We obtained phenotypic and genetic data on a large set of prasinophyte clade VII culture strains that allowed us to clarify the taxonomy of this important marine group. We describe two novel classes, the Picocystophyceae and the Chloropicophyceae, the latter containing two novel genera, Chloropicon and Chloroparvula, and eight new species of marine picoplanktonic green algae.

Diversity and oceanic distribution of prasinophytes clade VII, the dominant group of green algae in oceanic waters.

Prasinophytes clade VII is a group of pico/nano-planktonic green algae (division Chlorophyta) for which numerous ribosomal RNA (rRNA) sequences have been retrieved from the marine environment in the last 15 years. A large number of strains have also been isolated but have not yet received a formal taxonomic description. A phylogenetic analysis of available strains using both the nuclear 18S and plastidial 16S rRNA genes demonstrates that this group composes at least 10 different clades: A1-A7 and B1-B3. Analysis of sequences from the variable V9 region of the 18S rRNA gene collected during the Tara Oceans expedition and in the frame of the Ocean Sampling Day consortium reveal that clade VII is the dominant Chlorophyta group in oceanic waters, replacing Mamiellophyceae, which have this role in coastal waters. At some location, prasinophytes clade VII can even be the dominant photosynthetic eukaryote representing up to 80% of photosynthetic metabarcodes overall. B1 and A4 are the overall dominant clades and different clades seem to occupy distinct niches, for example, A6 is dominant in surface Mediterranean Sea waters, whereas A4 extend to high temperate latitudes. Our work demonstrates that prasinophytes clade VII constitute a highly diversified group, which is a key component of phytoplankton in open oceanic waters but has been neglected in the conceptualization of marine microbial diversity and carbon cycle.

Diversity and oceanic distribution of the Parmales (Bolidophyceae), a picoplanktonic group closely related to diatoms.

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.

Application of cryopreservation to genetic analyses of a photosynthetic picoeukaryote community.

Cryopreservation is useful for long-term maintenance of living strains in microbial culture collections. We applied this technique to environmental specimens from two monitoring sites at Sendai Bay, Japan and compared the microbial diversity of photosynthetic picoeukaryotes in samples before and after cryopreservation. Flow cytometry (FCM) showed no considerable differences between specimens. We used 2500 cells sorted with FCM for next-generation sequencing of 18S rRNA gene amplicons and after removing low-quality sequences obtained 10,088-37,454 reads. Cluster analysis and comparative correlation analysis of observed high-level operational taxonomic units indicated similarity between specimens before and after cryopreservation. The effects of cryopreservation on cells were assessed with representative culture strains, including fragile cryptophyte cells. We confirmed the usefulness of cryopreservation for genetic studies on environmental specimens, and found that small changes in FCM cytograms after cryopreservation may affect biodiversity estimation.

Origins of plastids and glyceraldehyde-3-phosphate dehydrogenase genes in the green-colored dinoflagellate Lepidodinium chlorophorum.

The dinoflagellate Lepidodinium chlorophorum possesses "green" plastids containing chlorophylls a and b (Chl a+b), unlike most dinoflagellate plastids with Chl a+c plus a carotenoid peridinin (peridinin-containing plastids). In the present study we determined 8 plastid-encoded genes from Lepidodinium to investigate the origin of the Chl a+b-containing dinoflagellate plastids. The plastid-encoded gene phylogeny clearly showed that Lepidodinium plastids were derived from a member of Chlorophyta, consistent with pigment composition. We also isolated three different glyceraldehyde-3-phosphate dehydrogenase (GAPDH) genes from Lepidodinium-one encoding the putative cytosolic "GapC" enzyme and the remaining two showing affinities to the "plastid-targeted GapC" genes. In a GAPDH phylogeny, one of the plastid-targeted GapC-like sequences robustly grouped with those of dinoflagellates bearing peridinin-containing plastids, while the other was nested in a clade of the homologues of haptophytes and dinoflagellate genera Karenia and Karlodinium bearing "haptophyte-derived" plastids. Since neither host nor plastid phylogeny suggested an evolutionary connection between Lepidodinium and Karenia/Karlodinium, a lateral transfer of a plastid-targeted GapC gene most likely took place from a haptophyte or a dinoflagellate with haptophyte-derived plastids to Lepidodinium. The plastid-targeted GapC data can be considered as an evidence for the single origin of plastids in haptophytes, cryptophytes, stramenopiles, and alveolates. However, in the light of Lepidodinium GAPDH data, we need to closely examine whether the monophyly of the plastids in the above lineages inferred from plastid-targeted GapC genes truly reflects that of the host lineages.

A haptophyte bearing siliceous scales: ultrastructure and phylogenetic position of Hyalolithus neolepis gen. et sp. nov. (Prymnesiophyceae, Haptophyta).

A haptophyte alga bearing hyaline but conspicuous scales was discovered in surface water samples of Shiribeshi Seamount, Japan. X-ray elemental analysis confirmed that silica was the major element in these scales. These scales were hat-shaped, ellipsoidal in top view, 4-6mum wide and 5-7mum long, perforated by several small pores, and were deposited on cells in several layers. Beneath the siliceous scale layers, organic scales were present, which are typical of haptophytes. The cells were non-motile despite having two short flagella hidden in the scale case. The haptonema, extended over the scale case, and one and a half times the cell length. The intracellular features were typical of haptophytes, including the peripheral endoplasmic reticulum (PER), Golgi cisternae with peculiar dilations, and the flagellar apparatus. The siliceous scales were produced in vesicles in the posterior region of the cell. Motile cells lacking silica scales were observed occasionally when cultures were maintained at lower temperatures and under oligotrophic conditions. This alga was described as Hyalolithus neolepis gen. et sp. nov. Phylogenetic analysis using the SSU rDNA and rbcL gene sequences indicated that Hyalolithus is a member of the Prymnesiales and falls in a clade including Prymnesium, Platychrysis, and Chrysochromulina polylepis. Based on these results, the evolutionary implications of the presence of silicified scales in haptophytes is discussed.