Wenche Eikrem

Green evolution and dynamic adaptations revealed by genomes of the marine picoeukaryotes Micromonas.

Picoeukaryotes are a taxonomically diverse group of organisms less than 2 micrometers in diameter. Photosynthetic marine picoeukaryotes in the genus Micromonas thrive in ecosystems ranging from tropical to polar and could serve as sentinel organisms for biogeochemical fluxes of modern oceans during climate change. These broadly distributed primary producers belong to an anciently diverged sister clade to land plants. Although Micromonas isolates have high 18S ribosomal RNA gene identity, we found that genomes from two isolates shared only 90% of their predicted genes. Their independent evolutionary paths were emphasized by distinct riboswitch arrangements as well as the discovery of intronic repeat elements in one isolate, and in metagenomic data, but not in other genomes. Divergence appears to have been facilitated by selection and acquisition processes that actively shape the repertoire of genes that are mutually exclusive between the two isolates differently than the core genes. Analyses of the Micromonas genomes offer valuable insights into ecological differentiation and the dynamic nature of early plant evolution.

The diversity of small eukaryotic phytoplankton (≤3 μm) in marine ecosystems

Small cells dominate photosynthetic biomass and primary production in many marine ecosystems. Traditionally, picoplankton refers to cells < or =2 microm. Here we extend the size range of the organisms considered to 3 microm, a threshold often used operationally in field studies. While the prokaryotic component of picophytoplankton is dominated by two genera, Prochlorococcus and Synechococcus, the eukaryotic fraction is much more diverse. Since the discovery of the ubiquitous Micromonas pusilla in the early 1950s, just over 70 species that can be <3 microm have been described. In fact, most algal classes contain such species. Less than a decade ago, culture-independent approaches (in particular, cloning and sequencing, denaturing gradient gel electrophoresis, FISH) have demonstrated that the diversity of eukaryotic picoplankton is much more extensive than could be assumed from described taxa alone. These approaches revealed the importance of certain classes such as the Prasinophyceae but also unearthed novel divisions such as the recently described picobiliphytes. In the last couple of years, the first genomes of photosynthetic picoplankton have become available, providing key information on their physiological capabilities. In this paper, we discuss the range of methods that can be used to assess small phytoplankton diversity, present the species described to date, review the existing molecular data obtained on field populations, and end up by looking at the promises offered by genomics.

Diversity of picoplanktonic prasinophytes assessed by direct nuclear SSU rDNA sequencing of environmental samples and novel isolates retrieved from oceanic and coastal marine ecosystems.

Picoplanktonic prasinophytes are well represented in culture collections and marine samples. In order to better characterize this ecologically important group, we compared the phylogenetic diversity of picoplanktonic prasinophyte strains available at the Roscoff Culture Collection (RCC) and that of nuclear SSU rDNA sequences from environmental clone libraries obtained from oceanic and coastal ecosystems. Among the 570 strains avalaible, 91 belonged to prasinophytes, 65 were partially sequenced, and we obtained the entire SSU rDNA sequence for a selection of 14 strains. Within the 18 available environmental clone libraries, the prasinophytes accounted for 12% of the total number of clones retrieved (142 partial sequences in total), and we selected 9 clones to obtain entire SSU rDNA sequence. Using this approach, we obtained a subsequent genetic database that revealed the presence of seven independent lineages among prasinophytes, including a novel clade (clade VII). This new clade groups the genus Picocystis, two unidentified coccoid strains, and 4 environmental sequences. For each of these seven lineages, at least one representative is available in culture. The three picoplanktonic genera Ostreococcus, Micromonas, and Bathycoccus (order Mamiellales), were the best represented prasinophytes both in cultures and genetic libraries. SSU rDNA phylogenetic analyses suggest that the genus Bathycoccus forms a very homogeneous group. In contrast, the genera Micromonas and Ostreococcus turned out to be quite complex, consisting of three and four independent lineages, respectively. This report of the overall diversity of picoeukaryotic prasinophytes reveals a group of ecologically important and diverse marine microorganims that are well represented by isolated cultures.

Phylogenetic reconstructions of the Haptophyta inferred from 18s ribosomal DNA sequences and available morphological data

Abstract Most haptophytes are unicellular, photosynthetic flagellates, although some have coccoid, colonial, amoeboid, or filamentous stages. Nearly all have a characteristic filamentous appendage, the haptonema, arising between the two flagella. The small subunit ribosomal RNA gene (l8S ribosomal DNA) from 18 haptophyte species has been sequenced, and the sequences aligned with those of more than 300 published and unpublished chlorophyll a + c algae. Phylogenies were constructed using maximum likelihood, neighbor-joining, and weighted maximum parsimony analyses. The high divergence (6%) between members of Pavlova and the remaining haptophytes supports the division of Haptophyta into two classes: Prymnesiophyceae and Pavlovophyceae. Three major clades that correspond to known taxa within the Prymnesiophyceae were identified: one clade embraces Phaeocystis spp.; the second includes members of the genera Chrysochromulina, Prymnesium, and Imantonia; and the third includes coccolithophorid genera and the genus Isochrysis. Two other clades contain taxa whose sequences were derived from a gene clone library. These taxa are not strongly related to any of the cultured taxa included in this study. Based on 18S ribosomal DNA sequence data and available information on morphological structure and ultrastructure, we propose that the class Prymnesiophyceae be divided into four orders: Phaeocystales ord. nov., Prymnesiales, Isochrysidales, and Coccolithales. A total of 1–2% divergence at this level in the 18S ribosomal RNA gene analysis warrants a separation above the level of family. Within the Pavlovophyceae, a new genus is established, Rebecca J.C. Green gen. nov., into which Pavlova salina and Pavlova helicata are moved.

Telonemia, a new protist phylum with affinity to chromist lineages.

Recent molecular investigations of marine samples taken from different environments, including tropical, temperate and polar areas, as well as deep thermal vents, have revealed an unexpectedly high diversity of protists, some of them forming deep-branching clades within important lineages, such as the alveolates and heterokonts. Using the same approach on coastal samples, we have identified a novel group of protist small subunit (SSU) rDNA sequences that do not correspond to any phylogenetic group previously identified. Comparison with other sequences obtained from cultures of heterotrophic protists showed that the environmental sequences grouped together with Telonema, a genus known since 1913 but of uncertain taxonomic affinity. Phylogenetic analyses using four genes (SSU, Hsp90, alpha-tubulin and beta-tubulin), and accounting for gamma- and covarion-distributed substitution rates, revealed Telonema as a distinct group of species branching off close to chromist lineages. Consistent with these gene trees, Telonema possesses ultrastructures revealing both the distinctness of the group and the evolutionary affinity to chromist groups. Altogether, the data suggest that Telonema constitutes a new eukaryotic phylum, here defined as Telonemia, possibly representing a key clade for the understanding of the early evolution of bikont protist groups, such as the proposed chromalveolate supergroup.

Ribosomal DNA phylogenies and a morphological revision provide the basis for a revised taxonomy of the Prymnesiales (Haptophyta)

Nucleotide sequences of the nuclear-encoded small subunit (18S rDNA) and partial large subunit (28S rDNA) ribosomal DNA were determined in 30 different species of the haptophyte genera Prymnesium, Chrysocampanula, Chrysochromulina, Imantonia and Platychrysis, all belonging to the order Prymnesiales. Phylogenies based on these and other available haptophyte 18S, 28S and plastid 16S rDNA sequences were reconstructed, and compared with available morphological and ultrastructural data. The rDNA phylogenies indicate that the genus Chrysochromulina is paraphyletic and is divided into two major clades. This is supported by ultrastructural and morphological data. There is a major split between Chrysochromulina species with a saddle-shaped cell form (clade B2) and the remaining species in the genus (clade B1). Clade B2 includes the type species C. parva and taxa belonging to this clade thus retain the name Chrysochromulina. The non-saddle-shaped Chrysochromulina species analysed are closely related to Hyalolithus, Prymnesium and Platychrysis species. Imantonia species are sister taxa to these species within clade B1. An amendment to the classification of the order Prymnesiales and the genera Prymnesium, Platychrysis and Chrysochromulina is proposed with one new and one emended family (Chrysochromulinaceae and Prymnesiaceae, respectively), two new genera (Haptolina and Pseudohaptolina), and one new species (Pseudohaptolina arctica). We suggest a revision of the taxonomy of the Prymnesiales that is in accordance with available molecular evidence and supported by morphological data.

Nanoplankton of the equatorial Pacific with emphasis on the heterotrophic protists

Abstract In the equatorial Pacific during the boreal spring of 1992, nanoplankton, represented by chrysophytes, dinoflagellates, amoebae, choanoflagellates, naked flagellates and ciliates, dominated the heterotrophic protistan biomass. This component contributed 11–60% of the combined phytoplankton and heterotrophic protistan biomass. The heterotrophs, other than the dinoflagellates and ciliates, were represented by amoebae and flagellates at a mean (±SD) density and biomass of 5.85 ± 2.53 x 105 cells I −1 and 1.76 ± 1.37 μgC l −1 . Fifty-two species of lobose amoebae, apusomonads, bicosoecids, cercomonads, choanoflagellates, chrysomonads, euglenids, jakobids, kathablepharids, kinetoplastids, pedinellids and a number of taxa of uncertain position (incertaesedis taxa) were identified. The heterotrophs represent five different trophic types of organisms, defined by habitat and prey. Fifty per cent of the species identified in this study are principally associated with detritus (marine snow). The majority of free-living suspension feeders we identify in this study are choanoflagellates. Other suspension feeding planktonic taxa may not be detected using the protocols we employ. Only 42% of the species identified are obligately bacterivorous and 12% are not bacterivores at all but graze principally on algae. The remainder of heterotrophic species prey on an array of DOM, bacteria, other protists and detritus.

Verrucophora farcimen gen. et sp. nov. (Dictyochophyceae, Heterokonta) - a bloom-forming ichthyotoxic flagellate from the Skagerrak, Norway.

Since 1998, a heterokont flagellate initially named Chattonella aff. verruculosa has formed recurrent extensive blooms in the North Sea and the Skagerrak, causing fish mortalities. Cells were isolated from the 2001 bloom off the south coast of Norway, and monoalgal cultures were established and compared with the Chattonella verruculosa Y. Hara et Chihara reference strain NIES 670 from Japan. The cells in Norwegian cultured isolates were very variable in size and form, being large oblong (up to 34 μm long) to small rounded (5–9 μm in diameter) with two unequal flagella, numerous chloroplasts, and mucocysts. The SSU and partial LSU rDNA sequences of strains from Norway and Japan were compared and differed by 0.4% (SSU) and 1.3% (LSU), respectively. Five strains from Norway were identical in the LSU rDNA region. Phylogenetic analyses based on heterokont SSU and concatenated SSU + LSU rDNA sequences placed C. aff. verruculosa and the Japanese C. verruculosa within the clade of Dictyochophyceae, with the picoflagellate Florenciella parvula Eikrem as the closest relative. Ultrastructure, morphology, and pigment composition supported this affinity. We propose the name Verrucophora farcimen sp. et gen. nov. for this flagellate and systematically place it within the class Dictyochophyceae. Our studies also show that C. verruculosa from Japan is genetically and morphologically different but closely related to V. farcimen. The species is transferred from the class Raphidophyceae to the class Dictyochophyceae and renamed Verrucophora verruculosa. We propose a new order, Florenciellales, to accommodate V. farcimen, V. verruculosa, and F. parvula.

A review of the phylogeny of the Haptophyta

Most haptophytes are unicellular, photosynthetic flagellates, although some have coccoid, colonial, amoeboid or filamentous stages. Nearly all have a characteristic filamentous appendage, the haptonema, arising between the two flagella. We have amassed small subunit rRNA gene sequences (18S rDNA) from 125 haptophytes and aligned the sequences with those of over 300 published and unpublished chlorophyll a+c algae. Phylogenies were constructed using Bayesian, minimum evolution and weighted maximum parsimony analyses. The high divergence (6%) between members of Pavlova and the remaining haptophytes supports the division of the Haptophyta into two classes: the Prymnesiophyceae and the Pavlovophyceae (Edvardsen et al. 2000). Four major clades within the Prymnesiophyceae were identified that correspond to known taxa: one clade embraces Phaeocystales; the second includes members of the Prymnesiales; the third represents the Isochrysidales; and the fourth the Coccolithales. Two other minor clades contain taxa whose sequences were derived from a gene clone library. In the absence of information on cell morphology associated with these sequences we are unable to determine whether they belong to existing orders or if new orders should be erected. These taxa are not strongly related to any of the known cultured taxa. One to two per cent divergence in the 18S rRNA gene analysis warrants a separation above the level of family.

Carotenoids from further prasinophytes

Abstract The qualitative and quantitative carotenoid composition of seven prasinophytes (eight clones) have been examined by chromatographic (TLC and HPLC) and spectroscopic methods (VIS, CD and mass spectra). The prasinophytes studied fall into two pigment types: (A) those producing common green algal carotenoids (β,β-carotene, β,ϵ-carotene, lutein, zeaxanthin and the epoxides violaxanthin and neoxanthin) and (B) prasinophytes synthesising carotenoids peculiar to this algal class (prasinoxanthin, anhydroprasinoxanthin, uriolide, anhydrouriolide, micromonal, anhydromicromonal, micromonol, anhydromicromonol and dihydrolutein), where prasinoxanthin is a major carotenoid. Mantoniella squamata (clone 2) was grown under both low and high light intensity, revealing differences in carotenoid composition. Lutein together with lesser amounts of zeaxanthin and its epoxides were only detected at high light intensity. Three previously unidentified carotenoids were identified as prasinoxanthin (xanthophyll K), micromonal and dihydrolutein.