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Doppler velocimetry measurements of phototactic response in flagellated algae.

A periodic shading mechanism (about 0.5 s period) is believed to be used by flagellated algae to detect light direction. Time changes in the orientation of a population of Haematococcus pluvialis under positive photoaxis conditions (cell samples were stimulated alternatively by two diametrically opposed beams of actinic light), have been investigated by analysing the Doppler shifts of laser light scattered by the cells by means of heterodyne detection techniques. This technique allows us to measure the mean value of the component of the cells' swimming velocities along the light stimulus axis. Preliminary data indicate that the time taken by a cell population to change orientation is about 1 s.

Cell Movement↗

Motility and gravitactic orientation of the flagellate, Euglena gracilis, impaired by artificial and solar UV-B radiation.

The effects of ultraviolet radiation on the gravitactic orientation of the freshwater flagellate, Euglena gracilis, were determined by a real time image analysis system. Both artificial UV radiation and solar radiation in a temperature-controlled growth chamber were employed. Histograms of gravitaxis showed that the degree of orientation decreased with increasing exposure time; this can be quantified using the Rayleigh test and upper quadrant summation. The effects of artificial UV radiation on the orientation are considerably stronger than those of solar radiation, probably because the radiation source emits higher fluence rates below 300 nm than found in solar radiation. The effects of monochromatic ultraviolet radiation on motility have been determined, and an action spectrum has been calculated.

Animals↗

Orientation of the photosynthetic flagellate, Peridinium gatunense, in hypergravity.

The photosynthetic freshwater flagellate, Peridinium gatunense, uses both positive phototaxis and negative gravitaxis to move upwards in the water column. At higher fluence rates approaching those at the surface of their habitat, the cells tend to become unoriented and thus stop their upward movement. Orientation and motility of Peridinium gatunense has been studied in the slow rotating centrifuge microscope (NIZEMI), which allows observation of swimming behavior during centrifugation acceleration between 1 g and 5g. The movement vectors were analyzed by real time image analysis capable of tracking many cells simultaneously. At 1 g the orientation was not very precise, but the degree of orientation increased significantly at higher acceleration forces up to about 3 g. Most cells were capable of swimming even against an acceleration vector of 3.8 g; at higher acceleration forces the cells were not able to cope with the centrifugal force. The linear velocity of cells swimming against 1 g was about 20% lower than that of cells moving in other directions. The velocity decreased even more in cells swimming against higher acceleration forces.

Animals↗

Effects of some chemical factors on flagellation and swarming of Vibrio alginolyticus.

Vibrio alginolyticus strains recently isolated from Dutch coastal seawater changed flagellar organization when cultivated in the presence of certain chemical agents. On agar media with more than 4.0% (w/v) NaCl the number of lateral flagella per cell decreased with increasing salt concentration. Both on agar media and in broth cultures with 6.0-9.0% (w/v) NaCl, cells with polar tufts of 2-4 sheathed or unsheathed flagella were frequently found. Cells grown on agar media with 7.3-9.8% (w/v) Na2SO4 had drastically reduced numbers of lateral flagella, but lacked polar tufts. EDTA suppressed growth, but did not affect flagellar arrangement. In the presence of 0.1-0.3% boric acid or 0.05-0.1% aluminum hydroxide, cells in liquid media tended to produce lateral, in addition to the polar flagella normally observed in broth cultures. Of a number of surface-active agents tested, Tween 80 and Na-taurocholate, even in high concentrations, did not affect flagellation. Bile salts (0.1%) and Na deoxycholate (0.05%) strongly reduced the number of both polar and lateral flagella. In agar cultures, Na-lauryl sulphate (0.01-0.1%) inhibited the formation of lateral, but increased the incidence of polar flagella. Teepol (0.05-0.2%) had a similar effect and also it had a deteriorating effect on the sheaths of the polar flagella. Concomitant with the reduction in the number of lateral flagella, induced by these agents, swarming on agar media was inhibited.

Agar↗

Plastid state- and light-dependent regulation of the expression of nucleus-encoded genes for chloroplast proteins in the flagellate Euglena gracilis.

Interorganellar regulatory interactions in the flagellate Euglena gracilis were shown to be more complicated than in green algae and higher plants. Euglena plastids have a much more complex influence on nuclear gene expression than was previously thought. The petJ gene for cytochrome c6 represents a group of nucleus-encoded genes for chloroplast proteins, the expression of which is influenced by the state of plastids at the transcriptional level. Moroever, the regulation of these genes might be light-dependent. In contrast, for nucleus-encoded small subunit of ribulose-bisphosphate carboxylase, chlorophyll a/b binding protein, and porphobilinogen deaminase transcript levels are unchanged in wild-type cells relative to white mutants. In these cases there is no plastid-derived signal operative during transcription. Porphobilinogen deaminase appeared to be regulated even at the post-translational level.

Animals↗

Actin phylogeny identifies Mesostigma viride as a flagellate ancestor of the land plants.

Green algae and land plants trace their evolutionary history to a unique common ancestor. This "green lineage" is phylogenetically subdivided into two distinct assemblages, the Chlorophyta and the Streptophyta. The Chlorophyta includes the Chlorophyceae, Trebouxiophyceae, Ulvophyceae, and Prasinopohyceae, whereas the Streptophyta includes the Charophyceae plus the bryophytes, ferns, and all other multicellular land plants (Embryophyta). The Prasinophyceae is believed to contain the earliest divergences within the green lineage. Phylogenetic analyses using rDNA sequences identify the prasinophytes as a paraphyletic taxon that diverges at the base of the Chlorophyta. rDNA analyses, however, provide ambiguous results regarding the identity of the flagellate ancestor of the Streptophyta. We have sequenced the actin-encoding cDNAs from Scherffelia dubia (Prasinophyceae), Coleochaete scutata, Spirogyra sp. (Charophyceae), and the single-copy actin gene from Mesostigma viride (Prasinophyceae). Phylogenetic analyses show Mesostigma to be the earliest divergence within the Streptophyta and provide direct evidence for a scaly, biflagellate, unicellular ancestor for this lineage. This result is supported by the existence of two conserved actin-coding region introns (positions 20-3, 152-1), and one intron in the 5'-untranslated region of the actin gene shared by Mesostigma and the embryophytes.

Actins↗

Effects of temperature and turbulence on the predator-prey interactions between a heterotrophic flagellate and a marine bacterium.

Biotic and abiotic factors can influence interactions between microbial grazers and their prey, thus impacting both the cycling of biogenic carbon within the surface layer of the ocean and the export of carbon to the deep ocean and higher trophic levels. In this study, microcosm experiments were used to evaluate the combined effect of temperature and turbulence on the growth rate of a marine bacterium (Vibrio splendidus), a protistan predator (Paraphysomonas sp.), and the community grazing impact of Paraphysomonas sp. on V. splendidus. It was found that the artificial turbulence generated (1.35 x 10-1 cm2 s-3) significantly increased the rates of growth of Paraphysomonas sp. at high (>10 degrees C), but not low (<5 degrees C) temperatures, and that turbulence had no effect on the growth of V. splendidus. Both flagellate and bacterial growth were temperature dependent and decreased 4- to 6-fold as temperatures decreased from 15 to 0 degrees C. Bacterial grazing mortality by Paraphysomonas sp. was 1.3- to 2.5-fold greater in the turbulent than static treatments among all four temperatures, and the rates of cell-specific ingestion of bacteria by Paraphysomonas sp. was 2-fold greater at 15 and 10 degrees C in the turbulent than in the static treatment. Hence, this study shows that turbulence can influence nanoflagellate grazing at temperatures >5 degrees C and suggests that at low temperatures, increased viscosity may limit the size of organisms that can be affected by small-scale turbulence.

Animals↗

Unexplained complexity of the mitochondrial genome and transcriptome in kinetoplastid flagellates.

Kinetoplastids are flagellated protozoans, whose members include the pathogens Trypanosoma brucei, T. cruzi and Leishmania species, that are considered among the earliest diverging eukaryotes with a mitochondrion. This organelle has become famous because of its many unusual properties, which are unique to the order Kinetoplastida, including an extensive kinetoplast DNA network and U-insertion/deletion type RNA editing of its mitochondrial transcripts. In the last decade, considerable progress has been made in elucidating the complex machinery of RNA editing. Moreover, our understanding of the structure and replication of kinetoplast DNA has also dramatically improved. Much less however, is known, about the developmental regulation of RNA editing, its integration with other RNA maturation processes, stability of mitochondrial mRNAs, or evolution of the editing process itself. Yet the profusion of genomic data recently made available by sequencing consortia, in combination with methods of reverse genetics, hold promise in understanding the complexity of this exciting organelle, knowledge of which may enable us to fight these often medically important protozoans.

Animals↗

Indications for acceleration-dependent changes of membrane potential in the flagellate Euglena gracilis.

The effects of the calcium sequester EGTA on gravitactic orientation and membrane potential changes in the unicellular flagellate Euglena gracilis were investigated during a recent parabolic-flight experiment aboard of an Airbus A300. In the course of a flight parabola, an acceleration profile is achieved which yields subsequently about 20 s of hypergravity (1.8 g(n)), about 20 s of microgravity, and another 20 s of hypergravity phases. The movement behavior of the cells was investigated with real-time, computer-based image analysis. Membrane potential changes were detected with a newly developed photometer which measures absorption changes of the membrane potential-sensitive probe oxonol VI. To test whether the data obtained by the oxonol device were reliable, the signal of non-oxonol-labelled cells was recorded. In these samples, no absorption shift was detected. Changes of the oxonol VI signals indicate that the cells depolarize during acceleration (very obvious in the step from microgravity to hypergravity) and slightly hyperpolarize in microgravity, which can possibly be explained with the action of Ca-ATPases. These signals (mainly the depolarization) were significantly suppressed in the presence of EGTA (5 mM). Gravitaxis in parallel was also inhibited after addition of EGTA. Initially, negative gravitaxis was inverted into a positive one. Later, gravitaxis was almost undetectable.

Acceleration↗

Chemotaxis in the green flagellate alga Chlamydomonas.

Behavior of the green flagellate alga Chlamydomonas changes in response to a number of chemical stimuli. Specific sensitivity of the cells to different substances might appear only at certain stages of the life cycle. The heterogamous species C. allensworthii demonstrates chemotaxis of male gametes towards pheromones excreted by female gametes. In C. reinhardtii chemotaxis towards tryptone occurs only in gametes, whereas chemotaxis towards ammonium, on the contrary, only in vegetative cells. Chemotaxis to different chemical stimuli might involve different mechanisms of reception and signal transduction, elucidation of which has only recently begun. Indirect evidences show that the cells likely respond to tryptone with changes in the membrane electrical conductance. The recently completed project of sequencing the whole nuclear genome of C. reinhardtii provides the basis for future identification of molecular elements of the chemosensory cascade in this alga.

Animals↗

Degradation of collagen types I, III, IV and V by extracellular proteinases of an oral flagellate Trichomonas tenax.

Proteinases secreted by an axenic strain of Trichomonas tenax were active against native types I, III, IV and V collagens when evaluated by polyacrylamide gel electrophoresis. Degradation of all four collagen types was temperature dependent. Basement membrane type IV collagen was digested most effectively. An inhibition of all collagenolytic activities by a specific inhibitor of cysteine proteinases, E-64, and activation by a reducing agent, dithiothreitol, indicated the involvement of cysteine proteinases of the oral flagellate in the cleavage of collagen.

Animals↗

Effects of low temperature and calcium on microfilament structure in flagellates of Physarum polycephalum.

Microfilament structures of the ridge and the backbone in Physarum flagellates disintegrated selectively within a few minutes upon cooling by ice-water. The elongated cells concurrently rounded up to spherical or irregular and amoeboid shape. When such rounded cells were warmed to 25 degrees C, the microfilament structures were reconstructed within 1 min and cells subsequently returned to an elongated shape. Disruption of microfilaments by cytochalasin A also caused the rounding up of cells, indicating that the rounding up resulted from disintegration of microfilament structures. This transformation induced by the cold treatment was retarded by preincubation of the cells with EGTA for 15 min, but addition of EGTA immediately before the onset of the cold treatment was less effective. The effect of EGTA was cancelled by simultaneous addition of excess Ca2+. Addition of procaine also inhibited the transformation induced by the cold treatment, while caffeine inhibited the recovery of the elongated shape when returned to 25 degrees C. Furthermore, addition of A23187 at 25 degrees C in the presence of Ca2+ mimicked the effect of the cold treatment. Thus, intracellular release of Ca2+ was suggested to be involved in the transformation induced by the cold treatment. Lability of the microfilament structures at a high concentration of Ca2+ was directly proved using Triton-permeabilized cells. Therefore we concluded that low temperature disrupts microfilament structures that are necessary for the maintenance of the elongated cell shape by inducing intracellular Ca2+ release. However, microfilament structures in Physarum amoeba cells were affected neither by the cold treatment nor by high Ca2+ concentration.

Actin Cytoskeleton↗

Identification of 11-cis and all-trans-retinal in the photoreceptive organelle of a flagellate green alga.

Isolation of intact photoreceptive organelles (eyespot apparatuses) involved in blue-light mediated photoresponses in a flagellate green alga (Spermatozopsis similis) allowed for the first time the identification of both 11-cis- and all-trans-retinal in a plant cell. Both isomers were identified by HPLC analysis in conjunction with UV spectra. Additionally, reconstitution of a distinct absorption band, centered around 540 nm, was achieved by addition of exogenous 9-cis-retinal to bleached, isolated eyespot apparatuses.

Carrier Proteins↗

The evolutionary expansion of the trypanosomatid flagellates.

The trypanosomatids combine a relatively uniform morphology with ability to parasitise a very diverse range of hosts including animals, plants and other protists. Along with their sister family, the biflagellate bodonids, they are set apart from other eukaryotes by distinctive organisational features, such as the kinetoplast-mitochondrion and RNA editing, isolation of glycolysis enzymes in the glycosome, use of the flagellar pocket for molecular traffic into and out of the cell, a unique method of generating cortical microtubules, and bizarre nuclear organisation. These features testify to the antiquity and isolation of the kinetoplast-bearing flagellates (Kinetoplastida). Molecular sequencing techniques (especially small subunit ribosomal RNA gene sequencing) are now radically reshaping previous ideas on the phylogeny of these organisms. The idea that the monogenetic (MG) trypanosomatids gave rise to the digenetic (DG) genera is losing ground to a view that, after the bodonids, the African trypanosomes (DG) represent the most ancient lineage, followed by Trypanosoma cruzi (DG), then Blastocrithidia (MG), Herpetomonas (MG) and Phytomonas (DG), with Leptomonas (MG), Crithidia (MG), Leishmania (DG) and Endotrypanum (DG) forming the crown of the evolutionary tree. Vast genetic distances (12% divergence) separate T. brucei and T. cruzi, while the Leishmania species are separated by very short distances (less than 1% divergence). These phylogenetic conclusions are supported by studies on RNA editing and on the nature of the parasite surface. The trypanosomatids seem to be able to adapt with ease their energy metabolism to the availability of substrates and oxygen, and this may give them the ability to institute new life cycles if host behaviour patterns allow. Sexual processes, though present in at least some trypanosomatids, may have played only a minor part in generating diversity during trypanosomatid evolution. On the other hand, the development of altruistic behaviour on the part of some life cycle stages may be a hitherto unconsidered way of maximising fitness in this group. It is concluded that, owing to organisational constraints, the trypanosomatids can undergo substantial molecular variation while registering very little in the way of morphological change.

Animal Population Groups↗

Trypanosoma cruzi strain-specific monoclonal antibodies: identification of Colombian strain flagellates in the insect vector.

Spleen cells from mice immunized with insect-derived Trypanosoma cruzi metacyclic trypomastigotes were used to obtain Colombian strain-specific monoclonal antibodies. At least 4 different strain-specific antigens were recognized by the monoclonal antibodies on epimastigotes or metacyclic trypomastigotes. There was no reactivity with other stages of Colombian strain T. cruzi, nor with any stage of 15 other T. cruzi strains or isolates, nor with 22 other Trypanosomatidae. One of the monoclonal antibodies was used to identify, by indirect immunofluorescence, Colombian strain flagellates in cryostat sections or glass-slide smears of the insect vector's intestine.

Animals↗

Characterization of dolichol monophosphate- and dolichol diphosphate-linked saccharides in trypanosomatid flagellates.

Dolichol-P- and dolichol-P-P-linked saccharides were isolated from several trypanosomatid flagellates incubated with [U-14C]glucose. Formation of Glc-P-dolichol and Man-P-dolichol was observed in Herpetomonas muscarum and Leishmania adleri, whereas only the latter derivative was synthesized in Trypanosoma dionisii and Leptomonas samueli. The main and largest dolichol-P-P-linked oligosaccharide formed in Trypanosoma conorhini, T. dionisii, L. samueli, Herpetomonas samuelpessoai and H. muscarum appeared to be Man9GlcNAc2, whereas in Blastocrithidia culicis it was Man6GlcNAc2. In L. adleri there were two main oligosaccharides linked to dolichol-P-P, Man6GlcNAc2 and Man5GlcNAc2. The The structures of the oligosaccharides were identical with those of the intermediates in the formation of Glc3Man9GlcNAc2-P-P-dolichol in higher eucaryotes. It was concluded that similarly to Trypanosoma cruzi, Crithidia fasciculata and Leishmania mexicana, no glucosylated derivatives of dolichol-P-P were formed in the additional seven trypanosomatids studied here. The results obtained suggest that the defective step could be in some cases the formation of Glc-P-dolichol and in others, the transfer of glucose residues from the latter compound to dolichol-P-P-linked oligosaccharides.

Acetates↗

Graviperception in the flagellate Euglena gracilis during a shuttle space flight.

During a recent space flight, gravitaxis of the unicellular photosynthetic flagellate, Euglena gracilis, was studied on board of the American shuttle Columbia. Accelerations were varied between 0 and 1.5 x g using a slow rotating centrifuge microscope (NIZEMI). The cells showed a sigmoidal response curve for the dependence of the precision of gravitaxis on acceleration which is indicative of the involvement of an active, physiological gravireceptor with a threshold at g-values < or = 0.16 x g and a saturation at g-values > or = 1 x g. No adaptation to microgravity was found during the prolonged space mission. After return the cells showed a normal gravitactic behavior at 1 x g. Since the cells are heavier than water, their swimming velocity is affected by sedimentation. The velocity distribution at different accelerations closely follows Stokes' law for sedimentation indicating that, in contrast to the ciliate Paramecium, E. gracilis, does not show any gravikinesis.

Acceleration↗

Flagellar systems in the euglenoid flagellates.

The flagellar apparatus of euglenoids consists of two functional basal bodies, three unequal microtubular roots subtending the reservoir, and a fourth band of microtubules nucleated from one of the flagellar roots and subtending the reservoir membrane. The flagellar apparatus of some euglenoids may contain additional basal bodies, striated roots ("rhizoplasts"), fibrous roots, striated connecting fibers between basal bodies, layered structures, or various electron-dense connective substances. With the possible exception of Petalomonas cantuscygni, nearly all euglenoids are biflagellate although the length of one flagellum may be highly reduced. The flagellar transition zone and number of basal bodies are highly variable among species. In recent years a cytoplasmic pocket that branches off from the reservoir has been discovered. The microtubules of the ventral flagellar root are continuous with the microtubules which line this pocket. Based on positional and structural similarities, this structure is believed to be homologous with the MTR/cytostome of bodonids. Coupled with other ultrastructural and biochemical data, the fine structure of the flagellar apparatus supports the belief that the euglenoid flagellates are descendant from bodonid ancestors.

Animals↗