The effect of long-term low bacterial density on the growth kinetics of three marine heterotrophic nanoflagellates.

Three marine helerotrophic nanoflagellates were each maintained under two cultural conditions for one month. They were either subcultured every 5 days in batch culture, to familiarise the cells to high bacterial density, or they were maintained in continuous culture in the presence of low bacterial density to familiarise the flagellate cells to this biotic state. The kinetic behaviour of each of the species and each preconditioned state were compared in batch culture grazing experiments. The maximum specific growth rate (/gmmax) and half-saturation constant (Ks) were the parameters used for comparison. After low prey preconditioning, Paraphysmnonas imperforuta was found to have a μmax value less than half that obtained if the culture was maintained with high prey density and this species did not increase its affinity for the bacterium under chemostat conditions. Bodo designis however, increased its affinity for the bacterium but the μmax value was severely reduced. Stephanoeca diplocostata increased its affinity for the bacterium and the μamxvalue after low prey density conditions. It appeared that the maintenance of the cultures at <μmax in the chemoslats and the absence of bacterial aggregates may have reduced the performance of both P. imperforata, because it prefers being attached whilst feeding on bacteria in suspension, and B. designis, because it prefers to feed on attached bacteria. S. diplocostata on the other hand, appeared not to rely on aggregate formation for good performance and could therefore be a representative of pelagic flagellates known to occur as unattached cells in relatively oligotrophic waters

Differential thermal adaptation of clonal strains of a protist morphospecies originating from different climatic zones

Eco-physiological variation and local adaptation are key issues in microbial ecology. Here, we investigated the thermal adaptation of 19 strains of the same Spumella morphospecies (Chrysophyceae, Heterokonta). In order to test for local adaptation and the existence of specific ecotypes we analysed growth rates of these strains, which originated from different climate regions. We applied temperature-adaptation as an eco-physiological marker and analysed growth rates of the different Spumella strains at temperatures between 0°C and 35°C. The temperatures allowing for maximal growth of strains from temperate and warm climatic zones ranged between 19.9°C and 33.4°C. Phylogenetically, most of these 'warm'-adapted strains fall into two different previously defined 18S rDNA Spumella clusters, one of them consisting of mostly soil organisms and the other one being a freshwater cluster. As a rule, the 'warm'-adapted strains of the soil cluster grew slower than the 'warm'-adapted isolates within the freshwater cluster. This difference most probably reflect different strategies, i.e. the formation of cysts at the expense of lower growth rates in soil organisms. In contrast, as expected, all isolates from Antarctica were cold-adapted and grew already around melting point of freshwater. Surprisingly, optimum temperature for these strains was between 11.8°C and 17.7°C and maximum temperature tolerated was between 14.6°C and 23.5°C. Our data indicate that despite the relatively high optimal temperature of most Antarctic strains, they may have a relative advantage below 5–10°C only. Based on the thermal adaptation of the flagellate strains the Antarctic strains were clearly separated from the other investigated strains. This may indicate a limited dispersal of flagellates to and from Antarctica. Even if the latter assumption needs support from more data, we argue that the high levels of eco-physiological and molecular microdiversity indicate that the current species concepts do not sufficiently reflect protist eco-physiological differentiation

Grazing rates and functional diversity of uncultured heterotrophic flagellates

9 pages, 4 figuresAquatic assemblages of heterotrophic protists are very diverse and formed primarily by organisms that remain uncultured. Thus, a critical issue is assigning a functional role to this unknown biota. Here we measured grazing rates of uncultured protists in natural assemblages (detected by fluorescent in situ hybridization (FISH)), and investigated their prey preference over several bacterial tracers in short-term ingestion experiments. These included fluorescently labeled bacteria (FLB) and two strains of the Roseobacter lineage and the family Flavobacteriaceae, of various cell sizes, which were offered alive and detected by catalyzed reporter deposition-FISH after the ingestion. We obtained grazing rates of the globally distributed and uncultured marine stramenopiles groups 4 and 1 (MAST-4 and MAST-1C) flagellates. Using FLB, the grazing rate of MAST-4 was somewhat lower than whole community rates, consistent with its small size. MAST-4 preferred live bacteria, and clearance rates with these tracers were up to 2 nl per predator per h. On the other hand, grazing rates of MAST-1C differed strongly depending on the tracer prey used, and these differences could not be explained by cell viability. Highest rates were obtained using FLB whereas the flavobacteria strain was hardly ingested. Possible explanations would be that the small flavobacteria cells were outside the effective size range of edible prey, or that MAST-1C selects against this particular strain. Our original dual FISH protocol applied to grazing experiments reveals important functional differences between distinct uncultured protists and offers the possibility to disentangle the complexity of microbial food websThis study was supported by the project ESTRAMAR (CTM2004-12631/MAR, MEC) to RM. FN was supported by the Marie-Curie fellowship ESUMAST (MEIF CT-2005-025000) and TL by the project METAOCEANS (MEST-CT-2005-019678)Peer reviewe

Magyar Pedagógia

International audienceProtists (unicellular eukaryotes) arguably account for most eukaryotic diversity and are central players of the biosphere. Known protist diversity and biology is largely based on cultured strains. Yet, environmental molecular surveys have unveiled entirely novel lineages that, as their prokaryotic counterparts, are essentially uncultured. Culture bias is an important drawback for any microbe-related science and is particularly severe for heterotrophic protists, which depend on organic food sources for growth. Here, we show how ecologically significant bacterivorous protists have been brought into culture by mimicking in situ conditions. Single cells sorted by serial dilution or flow cytometry were inoculated into seawater amended with natural bacterial assemblage at nearly in situ abundances. Strains belonging to lineages only known so far from environmental sequencing were isolated. Among them, Minorisa minuta gen. nov. sp. nov. forms a novel branch within Rhizaria, holding a key evolutionary position, and with an average size of 1.4 mu m represents one of the smallest bacterial grazers known to date. It has a worldwide planktonic distribution and can account for 5% of heterotrophic protists communities in coastal waters. Physiological features of this strain can partly explain its success in the environment. Culturing ecologically relevant but elusive protists provide invaluable material for ecophysiology, genomics, ecosystem modeling and evolutionary issues. The ISME Journal (2013) 7, 351-358; doi:10.1038/ismej.2012.85; published online 19 July 201

Hidden diversity among aquatic heterotrophic flagellates: ecological potentials of zoosporic fungi

International audienceSince the emergence of the ‘microbial loop' concept, heterotrophic flagellates have received particular attention as grazers in aquatic ecosystems. These microbes have historically been regarded incorrectly as a homogeneous group of bacterivorous protists in aquatic systems. More recently, environmental rDNA surveys of small heterotrophic flagellates in the pelagic zone of freshwater ecosystems have provided new insights. (i) The dominant phyla found by molecular studies differed significantly from those known from morphological studies with the light microscope, (ii) the retrieved phylotypes generally belong to well-established eukaryotic clades, but there is a very large diversity within these clades and (iii) a substantial part of the retrieved sequences cannot be assigned to bacterivorous but can be assigned instead to parasitic and saprophytic organisms, such as zoosporic true fungi (chytrids), fungus-like organisms (stramenopiles), or virulent alveolate parasites (Perkinsozoa and Amoebophrya sp.). All these microorganisms are able to produce small zoospores to assure dispersal in water during their life-cycles. Based on the existing literature on true fungi and fungus-like organisms, and on the more recently published eukaryotic rDNA environmental studies and morphological observations, we conclude that previously overlooked microbial diversity and related ecological potentials require intensive investigation (i) for an improved understanding of the roles of heterotrophic flagellates in pelagic ecosystems and (ii) to properly integrate the concept of ‘the microbial loop' into modern pelagic microbial ecology