Assessment of ciliates in the sheep rumen by DGGE.

AIMS: This work was carried out to develop a rapid molecular profiling technique to screen ciliate populations in the rumen of sheep. METHODS AND RESULTS: DGGE was used to study the ciliate diversity in the rumen of sheep. There was considerable variation between sheep which were co-housed, and fed the same diet. However, no difference in the major banding patterns was detected, when samples were collected from a single sheep sampled at different points. Following dietary changes, use of a pair-wise comparison of lanes, demonstrated that although there was still diversity between the ciliate population of sheep, the effects as a result of dietary changes were greater. CONCLUSIONS: The technique generated molecular profiles which are sufficiently different to allow comparison between samples, and to permit molecular ecological studies on the rumen ciliate population. SIGNIFICANCE AND IMPACT OF THE STUDY: The outcome of this study means that ciliate diversity in the rumen may now be studied by those unfamiliar with morphological identification of these organisms.

Oceanic 18S rDNA sequences from picoplankton reveal unsuspected eukaryotic diversity.

Picoplankton--cells with a diameter of less than 3 microm--are the dominant contributors to both primary production and biomass in open oceanic regions. However, compared with the prokaryotes, the eukaryotic component of picoplankton is still poorly known. Recent discoveries of new eukaryotic algal taxa based on picoplankton cultures suggest the existence of many undiscovered taxa. Conventional approaches based on phenotypic criteria have limitations in depicting picoplankton composition due to their tiny size and lack of distinctive taxonomic characters. Here we analyse, using an approach that has been very successful for prokaryotes but has so far seldom been applied to eukaryotes, 35 full sequences of the small-subunit (18S) ribosomal RNA gene derived from a picoplanktonic assemblage collected at a depth of 75 m in the equatorial Pacific Ocean, and show that there is a high diversity of picoeukaryotes. Most of the sequences were previously unknown but could still be assigned to important marine phyla including prasinophytes, haptophytes, dinoflagellates, stramenopiles, choanoflagellates and acantharians. We also found a novel lineage, closely related to dinoflagellates and not previously described.

Hydrogenosomes: convergent adaptations of mitochondria to anaerobic environments.

Hydrogenosomes are membrane-bound organelles that compartmentalise the final steps of energy metabolism in a number of anaerobic eukaryotes. They produce hydrogen and ATP. Here we will review the data, which are relevant for the questions: how did the hydrogenosomes originate, and what was their ancestor? Notably, there is strong evidence that hydrogenosomes evolved several times as adaptations to anaerobic environments. Most likely, hydrogenosomes and mitochondria share a common ancestor, but an unequivocal proof for this hypothesis is difficult because hydrogenosomes lack an organelle genome - with one remarkable exception (Nyctotherus ovalis). In particular, the diversity of extant hydrogenosomes hampers a straightforward analysis of their origins. Nevertheless, it is conceivable to postulate that the common ancestor of mitochondria and hydrogenosomes was a facultative anaerobic organelle that participated in the early radiation of unicellular eukaryotes. Consequently, it is reasonable to assume that both, hydrogenosomes and mitochondria are evolutionary adaptations to anaerobic or aerobic environments, respectively.

Rapid evolutionary divergence of Photosystem I core subunits PsaA and PsaB in the marine prokaryote Prochlorococcus.

The nucleotide sequences of the genes coding for the subunits of the Photosystem I (PS I) core, PsaA and PsaB were determined for the marine prokaryotic oxyphototrophs Prochlorococcus sp. MED4 (CCMP1378), P. marinus SS120 (CCMP1375) and Synechococcus sp. WH7803. Divergence of these sequences from those of both freshwater cyanobacteria and higher plants was remarkably high, given the conserved nature of PsaA and PsaB proteins. In particular, the PsaA of marine prokaryotes showed several specific insertions and deletions with regard to known PsaA sequences. Even in between the two Prochlorococcus strains, which correspond to two genetically different ecotypes with shifted growth irradiance optima, the sequence identity was only 80.2% for PsaA and 88.9% for PsaB. Possible causes and implications of the fast evolution rates of these two PS I core subunits are discussed.

Diversity and abundance of Bolidophyceae (Heterokonta) in two oceanic regions.

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.

Symbiomonas scintillans gen. et sp. nov. and Picophagus flagellatus gen. et sp. nov. (Heterokonta): two new heterotrophic flagellates of picoplanktonic size.

Two new oceanic free-living heterotrophic Heterokonta species with picoplanktonic size (< 2 microm) are described. Symbiomonas scintillans Guillou et Chrétiennot-Dinet gen. et sp. nov. was isolated from samples collected both in the equatorial Pacific Ocean and the Mediterranean Sea. This new species possesses ultrastructural features of the bicosoecids, such as the absence of a helix in the flagellar transitional region (found in Cafeteria roenbergensis and in a few bicosoecids), and a flagellar root system very similar to that of C. roenbergensis, Acronema sippewissettensis, and Bicosoeca maris. This new species is characterized by a single flagellum with mastigonemes, the presence of endosymbiotic bacteria located close to the nucleus, the absence of a lorica and a R3 root composed of a 6+3+x microtubular structure. Phylogenetical analyses of nuclear-encoded SSU rDNA gene sequences indicate that this species is close to the bicosoecids C. roenbergensis and Siluania monomastiga. Picophagus flagellatus Guillou et Chrétiennot-Dinet gen. et sp. nov. was collected in the equatorial Pacific Ocean. Cells are naked and possess two flagella. This species is characterized by the lack of a transitional helix and lateral filaments on the flagellar tubular hairs, the absence of siliceous scales, two unequal flagella, R1 + R3 roots, and the absence of a rhizoplast. SSU rDNA analyses place this strain at the base of the Chrysophyceae/Synurophyceae lineages.