Ultrastructure of the marine predatory flagellate <Emphasis Type="Italic">Metromonas simplex</Emphasis> Larsen et Patterson, 1990 (Cercozoa)

The ultrastructure of the marine predatory flagellate Metromonas simplex Larsen et Patterson was studied. The cell is surrounded by a low-contrast fibrous layer composed of thin hairs covered by a thin bilayer membrane and an outer layer of thin short fibers. The plasmalemma lies under these layers. The predator captures whole cells of the prey, usually bodonids or chrysomonads. The cytostome as a cell pocket is undetectable. The long flagellum bears very thin mastigonemes (hairs) with lengths of 0.8–1.0 μm; the short flagellum is naked and reduced in length. The transitional zone lacks spirals or other additional elements. The transversal plate is elevated on the cell surface. The flagellar root system is very simple and has one microtubular band which originates near the kinetosomes. The latter are parallel to each other and interconnected by fibrous bridges. The vesicular nucleus, Golgi apparatus, and endoplasmic reticulum are of typical structures. The oval mitochondria of 0.6–2.5 μm contain lamellar cristae. The cylindrical extrusomes (trichocysts) found in the cytoplasm have lengths of 1.0–1.4 μm and diameters of 0.12–0.08 μm. The trichocysts have a wheel-shaped structure with 13 spokes visible in cross-sections. The contractile vacuole is absent. The similarity that M. simplex shares with Metopion fluens Larsen et Patterson, cryothecomonads, and other predatory flagellates is discussed.     

Food selection by bacterivorous protists: insight from the analysis of the food vacuole content by means of fluorescence in situ hybridization

A modified fluorescence in situ hybridization (FISH) method was used to analyze bacterial prey composition in protistan food vacuoles in both laboratory and natural populations. Under laboratory conditions, we exposed two bacterial strains (affiliated with beta- and gamma-Proteobacteria -- Aeromonas hydrophila and Pseudomonas fluorescens, respectively) to grazing by three protists: the flagellates Bodo saltans and Goniomonas sp., and the ciliate Cyclidium glaucoma. Both flagellate species preferably ingested A. hydrophila over P. fluorescens, while C. glaucoma showed no clear preferences. Differences were found in the digestion of bacterial prey with B. saltans digesting significantly faster P. fluorescens compared to two other protists. The field study was conducted in a reservoir as part of a larger experiment. We monitored changes in the bacterial prey composition available compared to the bacteria ingested in flagellate food vacuoles. Bacteria detected by probe HGC69a (Actinobacteria) and R-BT065 were negatively selected by flagellates. Bacteria detected by probe CF319a were initially positively selected but along with a temporal shift in bacterial cell size, this trend changed to negative selection during the experiment. Overall, our analysis of protistan food vacuole content indicated marked effects of flagellate prey selectivity on bacterioplankton community composition.     

Small but Manifold – Hidden Diversity in “Spumella‐like Flagellates”

Colourless, nonscaled chrysophytes comprise morphologically similar or even indistinguishable flagellates which are important bacterivors in water and soil crucial for ecosystem functioning. However, phylogenetic analyses indicate a multiple origin of such colourless, nonscaled flagellate lineages. These flagellates are often referred to as “Spumella‐like flagellates” in ecological and biogeographic studies. Although this denomination reflects an assumed polyphyly, it obscures the phylogenetic and taxonomic diversity of this important flagellate group and, thus, hinders progress in lineage‐ and taxon‐specific ecological surveys. The smallest representatives of colourless chrysophytes have been addressed in very few taxonomic studies although they are among the dominant flagellates in field communities. To overcome the blurred picture and set the field for further investigation in biogeography and ecology of the organisms in question, we studied a set of strains of specifically small, colourless, nonscaled chrysomonad flagellates by means of electron microscopy and molecular analyses. They were isolated by a filtration‐acclimatisation approach focusing on flagellates of around 5 μm. We present the phylogenetic position of eight different lineages on both the ordinal and the generic level. Accordingly, we describe the new genera Apoikiospumella, Chromulinospumella, Segregatospumella, Cornospumella and Acrispumella Boenigk et Grossmann n. g. and different species within them.     

自由生活的异养鞭毛虫多样性及生态功能

随着对水生态系统结构与功能的深入研究,自由生活的异养鞭毛虫在水生态系统中的作用成为当前生态学领域的热点之一.已有的研究结果表明,异养鞭毛虫物种丰富,摄食方式多样,周转迅速,是微食物环的重要成分,在水生态系统中起着十分重要的作用.异养鞭毛虫物种多样性及生态学方面的系统研究将有助于对水生态系统结构、功能和过程深入了解.综述了异养鞭毛虫多样性、群落结构、摄食生态学以及在氮、磷循环中的作用,分析了其在生态系统中的功能.     

Heterotrophic flagellate biodiversity and community structure in freshwater streams

The species diversity and community structure of heterotrophic flagellates in watercourses from different geographical regions were investigated. Forty-one species have been identified. The species diversity of cercomonads, kinetoplastids, choanoflagellates, and flagellates is the highest. Bodo saltans, B. designis and Goniomonas truncata are the most common species. Planktonic communities in different streams are most similar in terms of species composition. The highest similarity within benthic cenoses was noted between geographically distant communities. Communities of heterotrophic flagellates seem to be highly heterogenous systems; species distribution depends mainly on the type of habitat and not on geographical factors.     

Easy Visualization of the Protist <Emphasis Type="Italic">Oxyrrhis</Emphasis><Emphasis Type="Italic">marina</Emphasis> Grazing on a Live Fluorescently Labelled Heterotrophic Nanoflagellate

Planktonic heterotrophic flagellates are ubiquitous eukaryotic microorganisms that play a crucial role in carbon and nutrient fluxes through pelagic food webs. Here we illustrate for the first time a grazing model of planktonic dinoflagellate, Oxyrrhis marina, on the heterotrophic nanoflagellate Goniomonas amphinema, using the DNA-binding fluorescent dye Hoechst 33342. A solution of 1 microg/mL of the fluorochrome allowed viability of the prey for at least 48 hours, provided low fluorescence quenching, and labelled the flagellate without masking the cytoplasm. After 2 hours of contact between the fluorescent prey and the predator, O. marina population had preyed on live G. amphinema at an ingestion rate of 2.2 prey Oxyrrhis (-1) h(-1). Results show that this model is a time-effective and inexpensive approach for the direct observation of heterotrophic flagellate grazing. The fact that prey remain alive while predation occurs, as well as the low rate of quenching, could be of help in studying the fate of real-time trophic interactions between protists in microbial webs.     

Feeding spectra and pseudoconoid structure in predatory alveolate flagellates

It is revealed experimentally that predatory alveolate flagellates Colpodella angusta Simpson et Patterson, C. edax Simpson et Patterson, and Voromonas pontica (Mylnikov, 2000) Cavalier-Smith, 2004 eat its prey by sucking out the insides through the front of its own body (rostrum). Feeding spectra of these predators include bodonids, percolomonads and colorless chrysomonads, while colorless euglenoids, cryptomonads, ciliates and naked amoebas are unsuitable food for them. Predators’ rostrum contains a microtubular structure (pseudoconoid) complemented by microtubular bands, micronemas or rhoptries. It is shown that the pseudoconoid begins near the kinetosomes of flagella and passes along the flagellate pocket into the rostrum. The pseudoconoid forms an unlocked cone in Colpodella angusta and Voromonas pontica, while, in Colpodella edax, it coils significantly without forming a cone. The similarity between the pseudoconoid and associated structures of predatory flagellates with analogous structures in other colpodellids, percinzoids, dinoflagellates and sporozoans is discussed.     

High diversity of the 'Spumella-like' flagellates: an investigation based on the SSU rRNA gene sequences of isolates from habitats located in six different geographic regions

We isolated 28 strains of 'Spumella-like' flagellates from different freshwater and soil habitats in Austria, People's Republic of China, Nepal, New Zealand, Uganda, Kenya, Tanzania and Hawaii by use of a modified filtration-acclimatization method. 'Spumella-like' flagellates were found in all of the samples and were often among the dominant bacterivorous flagellates in the respective environments. The small subunit ribosomal RNA (SSU rRNA) gene sequence of the isolates was determined and aligned with previously published sequences of members belonging to the Chrysophyceae sensu stricto. Phylogenetic analysis of the 28 new sequences confirmed their position within the Chrysophyceae sensu stricto and positioned them within different clades. Most of the sequences grouped within clade C and formed several subclusters separated from each other by green taxa including flagellates belonging to Ochromonas, Dinobryon, Poterioochromonas and others. All soil isolates clustered together (subcluster C1) with the soil strain Spumella elongata and the undescribed soil strain 'Spumella danica'. Aquatic isolates were affiliated with at least two branches (C2 and C3). Sequence similarity to the closest related member of the Chrysophyceae ranged between 92% and 99.6%, sequence divergence among the 'Spumella-like' flagellates was as high as 10%. We conclude that (i) the 'Spumella-like' flagellates are a diverse group both in terms of sequence dissimilarity between isolates and in terms of the number of genotypes, (ii) Spumella and Ochromonas are polyphyletic, and (iii) based on the SSU rRNA gene no biogeographical restriction of certain branches could be observed even though different ecotypes may be represented by the same genotype.     

Confusing Selective Feeding with Differential Digestion in Bacterivorous Nanoflagellates

Food selectivity and the mechanisms of food selection were analyzed by video microscopy for three species (Spumella, Ochromonas, Cafeteria) of interception-feeding heterotrophic nanoflagellates. The fate of individual prey particles, either live bacteria and/or inert particles, was recorded during the different stages of the particle-flagellate-interaction, which included capture, ingestion, digestion, and egestion. The experiments revealed species-specific differences and new insights into the underlying mechanisms of particle selection by bacterivorous flagellates. When beads and bacteria were offered simultaneously, both particles were ingested unselectively at similar rates. However, the chrysomonads Spumella and Ochromonas egested the inert beads after a vacuole passage time of only 2-3 min, which resulted in an increasing proportion of bacteria in the food vacuoles. Vacuole passage time for starved flagellates was significantly longer compared to that of exponential-phase flagellates for Spumella and Ochromonas. The bicosoecid Cafeteria stored all ingested particles, beads as well as bacteria, in food vacuoles for more then 30 min. Therefore "selective digestion" is one main mechanism responsible for differential processing of prey particles. This selection mechanism may explain some discrepancies of former experiments using inert particles as bacterial surrogates for measuring bacterivory.     

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 degrees C and 35 degrees C. The temperatures allowing for maximal growth of strains from temperate and warm climatic zones ranged between 19.9 degrees C and 33.4 degrees 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 degrees C and 17.7 degrees C and maximum temperature tolerated was between 14.6 degrees C and 23.5 degrees C. Our data indicate that despite the relatively high optimal temperature of most Antarctic strains, they may have a relative advantage below 5-10 degrees 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.     

Leptomonas ciliatorum n. sp. (Kinetoplastida,Trypanosomatidae) in the Macronucleus of a Hypotrichous Ciliate1

A leptomonad flagellate found in the macronucleus of the hypotrichous ciliate Paraholosticha sterkii carries out its whole life cycle there. The ciliates can divide, encyst, and excyst even when parasitized, and the flagellates are maintained throughout. Parasite-free ciliates may be rapidly infected. The light- and electron-microscopic structure of the flagellate resembles that of other leptomonads.     

Food capture and adhesion by the heliozoonActinophrys sol

Summary The heliozoonActinophrys sol is characterized by needle-like axopodia radiating from the spherical cell body. When helio-zoons capture food organisms, the prey is caught by adhesion to the surface of axopodia where numerous extrusomes are present close to the plasma membrane. To understand the molecular mechanism by which the heliozoons capture prey organisms, crude isolation and characterization of the adhesive substance was carried out. Prey flagellates (Chlorogonium elongatum) adhered and aggregated to remnants of heliozoon cells which had been killed by freezing or treatment at high temperature (80 °C for 10 min). Isolated extrusomes, which were prepared as the supernatant of cells homogenized and centrifuged after freezing and thawing, showed strong adhesion to the prey flagellates which responded to the supernatant by adhering their flagella and cell bodies to each other to form bouquet-like cell clusters. The adhesive substance was further extracted from heat-treatedA. sol. This fraction contained filamentous material similar to the secreted contents of the extrusomes observed during feeding. Its adhesive activity was not inhibited by trypsin treatment.     

The quest for the ancestor of the Ciliophora: a brief review of the continuing problem

Although the assumption has long been made that the Ciliophora arose from flagellates, limited progress has been made in determining exactly what extant group - flagellate or other - may best resemble the ancestral "preciliate" assemblage. The distinctiveness of the ciliates and of the many protist phyla containing flagellated stages in their life cycles makes the recognition of potentially homologous features difficult. The suggestion that there may be a phylogenetic relationship between dinoflagellates and ciliates, while seeming unlikely from consideration of a number of specialized characters, is attracting increased attention. This is because some basic similarities (possibly shared derived characters) are characteristic of many species from both groups: cortical alveoli, tubular mitochondrial cristae, kinetidal systems, acentric mitoses with persisting nuclear envelope, functional cytostome, extrusomes (mucocytsts and explosive trichocysts), locomotory organelles, and small subunit rRNAs (close structural similarity values). But many questions remain unanalyzed or unanswered, and caution is still advisable in drawing any firm conclusions about the evolutionary closeness of ciliates and dinoflagellates.     

Phylogenetic position of Cryothecomonas inferred from nuclear-encoded small subunit ribosomal RNA

The systematic position of the genus Cryothecomonas has been determined from an analysis of the nuclear-encoded small subunit ribosomal RNA gene of Cryothecomonas longipes and two strains of Cryothecomonas aestivalis. Our phylogenetic trees inferred from maximum likelihood, distance and maximum parsimony methods robustly show that the genus Cryothecomonas clusters within the phylum Cercozoa, and is related to the sarcomonad flagellate Heteromita globosa. Morphological data supporting the taxonomic placement of Cryothecomonas near the sarcomonad flagellates has been compiled from the literature. The high number of nucleotide substitutions found between two morphologically indistinguishable strains of Cryothecomonas aestivalis suggests the possibility of cryptic species within Cryothecomonas aestivalis.     

The morphology of predatory flagellate Colpodella pseudoedax Mylnikov et Mylnikov, 2007 (Colpodellida, Alveolata)

The structure of the predatory freshwater flagellate Colpodella pseudoedax was studied. The cell was found to contain two heterodynamic flagella, three-membrane pellicle, micropores, subpellicular microtubules, microtubular open-side conoid, roptries, micronemes, extrusive organelles (trichocysts), and mitochondria with vesicular and tubular cristae. Upon discharge, trichocysts form cross-striated bands. A thin-walled cylinder lies in the transitional zone of the flagella. Cells reproduce by means of longitudinal binary fission. This species differs from similar C. edax by their smaller cell size and lack of reproduction cyst. Similarities between C. pseudoedax and other colpodelids, as well as between colpodellids and perkinseids and sporozoans, are discussed.     

Concanavalin A-induced discharge of glycocalyx of raphidophycean flagellates, Chattonella marina and Heterosigma akashiwo

Chattonella marina and Heterosigma akashiwo, known as red tide phytoplankton, are naturally wall-less and have quite fragile cell structures. In this study, we found that an equilibrium dialysis technique allowed the study of lectin binding to these flagellates. The results suggested that concanavalin A (Con A) binds to these flagellate cells through the specific carbohydrate moieties on the cell surface. Interestingly, the binding of an excess of Con A on the cell surface caused morphological changes concomitant with discharge of glycocalyx, a polysaccharide-containing common structure on the external cell surface of these flagellates. Fluorescent microscopic observation using FITC-labeled Con A (F-Con A) confirmed that F-Con A molecules are localized on the discharged glycocalyx.     

The role of actin, actomyosin and microtubules in defining cell shape during the differentiation of Naegleria amebae into flagellates

Differentiation of Naegleria amebae into flagellates was used to examine the interaction between actin, actomyosin and microtubules in defining cell shape. Amebae, which lack microtubules except during mitosis, differentiate into flagellates with a fixed shape and a complex microtubule cytoskeleton in 120 min. Based on earlier models of ameboid motility it has been suggested that actomyosin is quiescent in flagellates. This hypothesis was tested by following changes in the cytoskeleton using three-dimensional reconstructions prepared by confocal microscopy of individual cells stained with antibodies against actin and tubulin as well as with phalloidin and DNase I. F-actin as defined by phalloidin staining was concentrated in expanding pseudopods. Most phalloidin staining was lost as cells rounded up before the onset of flagellum formation. Actin staining with a Naegleria-specific antibody that recognizes both F- and G-actin was confined to the cell cortex of both amebae and flagellates. DNase I demonstrated G-actin throughout all stages. Most of the actin in the cortex was not bound by phalloidin yet was resistant to detergent extraction suggesting that it was polymerized. The microtubule cytoskeleton of flagellates was intimately associated with this actin cortex. Treatment of flagellates with cytochalasin D produced a rapid loss of flagellate shape and the appearance of phalloidin staining while latrunculin A stabilized the flagellate shape. These results suggest that tension produced by an actomyosin network is required to maintain the flagellate shape. The rapid loss of the flagellate shape induced by drugs, which specifically block myosin light chain kinase, supports this hypothesis.