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Abstract: Protargol staining is a crucial method to reveal the infraciliature of ciliates, which is the most important morphological character for species identification. In the present study, Wilbert's protocol of protargol staining was emended mainly toward the highly happened improper bleaching. Through reciprocal treatments, both insufficient and excessive bleachings were much eliminated from the protargol protocol and the tests performed with four different species of ciliates established that the stainings were considerably improved and more reliable with optimized bleaching. Compared to the original protocol, the optimized method was proved to be more suitable for the groups difficult to stain, and it is also friendlier for the beginners and researchers in related fields. [ABSTRACT FROM AUTHOR]

Abstract: The mixotroph Yihiella yeosuensis is a small‐ and fast‐swimming dinoflagellate. To investigate its protistan predators, interactions between Y. yeosuensis and 11 heterotrophic protists were explored. No potential predators were able to feed on actively swimming Y. yeosuensis cells, which escaped via rapid jumps, whereas Aduncodinium glandula, Oxyrrhis marina, and Strombidinopsis sp. (approximately 150 μm in cell length) were able to feed on weakly swimming cells that could not jump. Furthermore, Gyrodinium dominans, Luciella masanensis, and Pfiesteria piscicida were able to feed on heat‐killed Yihiella cells, whereas Gyrodinium moestrupii, Noctiluca scintillans, Oblea rotunda, Polykrikos kofoidii, and Strombidium sp. (20 μm) did not feed on them. Thus, the jumping behavior of Y. yeosuensis might be primarily responsible for the observed lack of predation. With increasing Yihiella concentration, the growth rate of O. marina decreased, whereas that of Strombidinopsis did not change. However, with increasing Yihiella concentration (up to 530 ng C/ml), the ingestion rate of Strombidinopsis on Yihiella increased linearly. The highest ingestion rate was 24.1 ng C per predator per d. The low daily carbon acquisition from Yihiella relative to the body carbon content of Strombidinopsis might be responsible for its negligible growth. Thus, Y. yeosuensis might have an advantage over its competitors due to its low mortality rate. [ABSTRACT FROM AUTHOR]

Some of the most extreme genome wide rearrangements are found in ciliates, which are unique in possessing both germline micronucleus and somatic macronucleus in every cell/organism. A series of DNA rearrangement events, including DNA elimination, chromosomal fragmentation, gene unscrambling and alternative processing, happen during macronuclear development. To assess the molecular evolution of macronuclear and germline-limited sequences in different cryptic species of Chilodonella uncinata, we characterized the actin, α-tubulin and β-tubulin genes in the micronucleus and macronucleus genomes of USA-SC2 strain and compared them with other strains (i.e. cryptic species). Three main results are: (i) rearrangement patterns between germline and soma are conserved for each gene among the cryptic species of C. uncinata; (ii) in contrast, the germline-limited regions are highly divergent in sequence and length among the cryptic species; (iii) pointer shifting is frequent among the cryptic species. We speculate that pointer sequences may serve as the buffer between the conserved macronuclear destined sequences and rapidly-evolving internal eliminated sequences. The data combined with previous studies demonstrate the plasticity of gene rearrangement among different groups of ciliates and add to the growing data for the role of genome rearrangements in species differentiation. [ABSTRACT FROM AUTHOR]

Epibiosis is very common between crustaceans and ciliates where the calcified surface of the crustacean body provides a suitable substrate for ciliate colonization. The aim of this contribution is to provide data about a new record between the epistylid ciliate Epistylis hentscheli Kahl, 1935, and the crayfish Procambarus (Austrocambarus) sp. The distribution of the epistylid on the basibiont body and its cellular/ colonial characteristics were analyzed. Procambarus (Austrocambarus) sp. harbored colonies of E. hentscheli only on the pereiopods. This is the first record of this peritrich ciliate as an epibiont on Crustacea, having been previously found on algae and fish. [ABSTRACT FROM AUTHOR]

The exon junction complex (EJC) is a key element of the splicing machinery. The EJC core is composed of eIF4A3, MAGO, Y14 and MLN51. Few accessory proteins, such as CWC22 or UPF3, bind transiently to the EJC. The EJC has been implicated in the control of the splicing of long introns. To ascertain whether the EJC controls the splicing of short introns, we used Paramecium tetraurelia as a model organism, since it has thousands of very tiny introns. To elucidate whether EJC affects intron splicing in P. tetraurelia , we searched for EJC protein-coding genes, and silenced those genes coding for eIF4A3, MAGO and CWC22. We found that P. tetraurelia likely assembles an active EJC with only three of the core proteins, since MLN51 is lacking. Silencing of eIF4A3 or CWC22 genes, but not that of MAGO, caused lethality. Silencing of the MAGO gene caused either an increase, decrease, or no change in intron retention levels of some intron-containing mRNAs used as reporters. We suggest that a fine-tuning expression of EJC genes is required for steady intron removal in P. tetraurelia . Taking into consideration our results and those published by others, we conclude that the EJC controls splicing independently of the intron size. [ABSTRACT FROM AUTHOR]

Abstract: Ciliated protozoa possess two morphologically and functionally distinct nuclei: a macronucleus (MAC) and a micronucleus (MIC). The MAC is transcriptionally active and functions in all cellular events. The MIC is transcriptionally inactive during cell growth, but functions in meiotic events to produce progeny nuclei. Thus, these two nuclei must be distinguished by the nuclear proteins required for their distinct functions during cellular events such as cell proliferation and meiosis. To understand the mechanism of the nuclear transport specific to either MAC or MIC, we identified specific nuclear localization signals (NLSs) in two MAC‐ and MIC‐specific nuclear proteins, macronuclear histone H1 and micronuclear linker histone‐like protein (Mlh1), respectively. By expressing GFP‐fused fragments of these proteins in Tetrahymena thermophila cells, two distinct regions in macronuclear histone H1 protein were assigned as independent MAC‐specific NLSs and two distinct regions in Mlh1 protein were assigned as independent MIC‐specific NLSs. These NLSs contain several essential lysine residues responsible for the MAC‐ and MIC‐specific nuclear transport, but neither contains any consensus sequence with known monopartite or bipartite NLSs in other model organisms. Our findings contribute to understanding how specific nuclear targeting is achieved to perform distinct nuclear functions in binucleated ciliates. [ABSTRACT FROM AUTHOR]

Holospora and related bacteria are a group of obligate Paramecium symbionts. Characteristic features are their infectivity, the presence of two distinct morphotypes, and usually a strict specialization for a single Paramecium species as host and for a nuclear compartment (either somatic or generative nucleus) for reproduction. Holospora caryophila steps out of line, naturally occurring in Paramecium biaurelia and Paramecium caudatum. This study addresses the phylogenetic relationship among H. caryophila and other Holospora species based on 16S rRNA gene sequence comparison analyzing the type strain and seven new macronuclear symbionts. Key aspects of Holospora physiology such as infectivity, symbiosis establishment and host range were determined by comprehensive infection assays. Detailed morphological investigations and sequence-based phylogeny confirmed a high similarity between the type strain of H. caryophila and the novel strains. Surprisingly, they are only distantly related to other Holospora species suggesting that they belong to a new genus within the family Holosporaceae, here described as Preeria caryophila comb. nov. Adding to this phylogenetic distance, we also observed a much broader host range, comprising at least eleven Paramecium species. As these potential host species exhibit substantial differences in frequency of sexual processes, P. caryophila demonstrates which adaptations are crucial for macronuclear symbionts facing regular destruction of their habitat. [ABSTRACT FROM AUTHOR]

Industrial wastewater is of particular importance. In most wastewater treatments, physicochemical parameters are used to determine the performances which are costly while protozoa can be used as less expensive biological indicators to determine the efficiency of the treatment plant. The purpose of this study was to investigate the relationship between protozoa and physic-chemical characteristics of the Abadan wastewater treatment industry. Samples were collected from wastewater stabilization ponds in the Abadan industrial park. A total of 66 samples for biochemical and chemical tests and protozoa counts were taken. The sites of sampling were influent of sewage and effluent from maturation pond. For the parameters from influent, first 300 mL of samples was collected in depths of 0.1, 0.35 and 1 m and then mixed to provide a composite sample. 300 mL sample was directly taken from the effluent. The results of this study indicate that there is a positive relationship between the number and population percentage of amoeba and mastigophora (flagellate) with BOD and COD, whereas is the negative relationship between sticking ciliates with BOD and COD. Furthermore there is a positive relation between DO, the percentage, number of ciliates and amoeba population and there is a correlation, with flagellate population. Increase of flagellate numbers is indicative of high effluent Kjeldahl nitrogen. There was also an increase of input TSS and amoeba population percentage and a reduction of free ciliates numbers. The type and abundance of protozoa in industrial wastewater treatment following biological methods is an indicator of the system performance and effluent quality. This study shows the use of biological indicators can be used for monitoring as a much cheaper alternative resulting in a better economy. It is also a good indicator for the detection of toxic materials entering the plant or refinery and also its organic load shocks. [ABSTRACT FROM AUTHOR]

The aim of the present article is to analyse the evolutionary links between protozoa and neuronal and neurosecretory cells. To this effect we employ functional and topological data available for ciliates, in particular for Paramecium . Of note, much less data are available for choanoflagellates, the progenitors of metazoans, which currently are in the focus of metazoan genomic data mining. Key molecular players are found from the base to the highest levels of eukaryote evolution, including neurones and neurosecretory cells. Several common fundamental mechanisms, such as SNARE proteins and assembly of exocytosis sites, GTPases, Ca 2+ -sensors, voltage-gated Ca 2+ -influx channels and their inhibition by the forming Ca 2+ /calmodulin complex are conserved, albeit with different subcellular channel localisation, from protozoans to man. Similarly, Ca 2+ -release channels represented by InsP 3 receptors and putative precursors of ryanodine receptors, which all emerged in protozoa, serve for focal intracellular Ca 2+ signalling from ciliates to mammalian neuronal cells, eventually in conjunction with store-operated Ca 2+ -influx. Restriction of Ca 2+ signals by high capacity/low affinity Ca 2+ -binding proteins is maintained throughout the evolutionary tree although the proteins involved differ between the taxa. Phosphatase 2B/calcineurin appears to be involved in signalling and in membrane recycling throughout evolution. Most impressive example of evolutionary conservation is the sub-second dynamics of exocytosis-endocytosis coupling in Paramecium cells, with similar kinetics in neuronal and neurosecretory systems. Numerous cell surface receptors and channels that emerge in protozoa operate in the human nervous system, whereas a variety of cell adhesion molecules are newly “invented” during evolution, enabled by an increase in gene numbers, alternative splice forms and transcription factors. Thereby, important regulatory and signalling molecules are retained as a protozoan heritage. [ABSTRACT FROM AUTHOR]

The superoxide dismutase (SOD) family is a first line antioxidant enzyme group involved in transformation of the superoxide anion (O 2 − ) into hydrogen peroxide (H 2 O 2 ) and O 2 . SOD gene expression patterns and enzyme activities therefore have a role as molecular biomarkers in evaluating the oxidative stress status of aquatic organisms. However, antioxidant enzyme systems are yet to be fully explored in the marine ciliates. In this study, we identified and characterized two types of Cu/Zn SODs (Ec-Cu/ZnSOD1 and Ec-Cu/ZnSOD2) and Ec-Mn SOD in the marine ciliate Euplotes crassus . Subsequently, SOD activity and transcriptional modulation of the relevant genes were investigated after the exposure to Cd and Cu for 8 h. All Ec-SODs showed conserved domains and metal binding sites on their active sites. Total SOD activity was induced at 1 h after exposure to Cd (125 and 1000 μg/L), and showed a marginal increase at 1-h exposure to Cu (10 and 100 μg/L). However, SOD activity was maintained at a steady level under Cd and decreased under Cu exposure conditions at 3 h and 8 h. mRNA expression of both the Ec-Cu/Zn-SOD s and Mn-SOD were remarkably elevated after the exposure to Cd (250–1000 μg/L, maximum 4-fold, p  < 0.05) and, in particular, Cu (25–100 μg/L, maximum > 20-fold, p <  0.05), in a concentration – dependent manner. These findings suggest that Ec-SODs may be actively involved in cellular protection against metal – mediated oxidative stress. This study is therefore helpful in understanding the molecular responses for metal toxicity in the ciliates. [ABSTRACT FROM AUTHOR]

Chlorpyrifos and dimethoate are overused agricultural pesticides that can trigger trophic cascades, resulting in toxicity to both terrestrial and aquatic organisms as well as altered ecosystems. In previous studies, substantial attention has been given to the effects of pesticides on vertebrate species and, to a lesser extent, species of zooplankton. The present study was designed to show that the fission time effective concentration in ciliates is a potential aquatic detection index for environmental monitoring. The ciliate Urostyla grandis was treated with doses of chlorpyrifos and dimethoate. After exposed to the pesticides, the LC50 (i.e., concentration that killed 50% of the ciliate cells within 24 h) values were 0.029 mg L−1 for chlorpyrifos and 0.0685 mg L−1 for dimethoate. The fission time effective concentrations after 168 h of exposure were 0.0075-0.0093 mg L−1 for chlorpyrifos and 0.2640-0.2750 mg L−1 for dimethoate. These results show that the fission time effective concentration is lower than the LC50 value in ciliates, indicating that fission time effective concentration is more suitable than the LC50 value for environmental monitoring using ciliates. The effects of chlorpyrifos and dimethoate on ciliate cell ultrastructures included agglutination of chromatin in the macronucleus, protruded and discontinuous macronuclear and micronuclear membranes, loss of integrity of mitochondrial membranes and contents, and abscission and deformation of the adoral zone of membranelles. [ABSTRACT FROM AUTHOR]

Members of the genus Thuricola are a highly specialized group of peritrichous ciliates possessing a protective barrel-shaped lorica. The genus presents many difficulties in terms of species separation and definition, and in this context the present study investigates three species by protargol staining and analyses of SSU rDNA sequences. Based on their morphologic characteristics and biotope, they were identified as three poorly known forms in Thuricola, namely T. obconica Kahl, 1933, T. kellicottiana (Stokes, 1887) Kahl 1935 and T. folliculata Kent, 1881, respectively. T. obconica is characterized by possessing curved lorica and a single valve in vivo. T. kellicottiana is distinguished by two valves with a spine on the main valve, and a generally long internal stalk upon which the zooids sit. T. folliculata also has two valves but lacks a spine. The ciliature of the three species are basically similar. The main features are characterized as follows: infundibular polykineties 1-3 (P1-3) are relatively long and composed of three rows each; P1 bends twice and comprises three equally long rows; P2 ends near the second bend of P1 with all rows exhibiting a staggered arrangement; P3 converges with P1 at adstomal end, its row 1 at least twice as long as the other two rows; epistomial membranes 1 and 2 are present. Silverline system in Vorticella-pattern. Phylogenetic analyses indicate that the three ciliates in this study cluster together within one of the two major sub-clades within the Vaginicolidae. [ABSTRACT FROM AUTHOR]

Ciliates have been reported as pathogens of many species of economically important bivalves. Mussel protozoan X (MPX), is an uncharacterised intracellular ciliate of mussels and has been widely reported in Mytilus spp . around the world. In order to characterise this ciliate, Mytilus edulis samples were collected from a site on the West coast of Scotland, and four different fixatives for histological examination were tested. Fresh preparations of mussel digestive glands were also examined by laser scanning confocal microscopy. Intracellular ciliates were prepared by laser capture microdissection and partial sequences of small subunit ribosomal RNA gene and of large subunit ribosomal RNA gene were generated, using Phyllopharyngea primers. Methacarn solution proved to be the best fixative for both histological and molecular characterisation. The morphological and molecular investigations confirmed that this ciliate belongs to the class Phyllopharyngea, order Rhynchodida. However, this organism does not belong to any known family, genus or species, therefore, a new description is necessary, following further morphological analyses. Most mussel samples containing MPX displayed mild to moderate infections, with no signs of necrosis or haemocytic response, although a single sample displayed a severe infection (∼10 3 ciliates per section). The localisation of this ciliate in tissues other than the digestive gland, the presence of necrosis in infected tissue of the most severely infected mussel and the binary fission of this ciliate have been observed here for the first time. We also report the first observation of the live ciliate isolated from tissue. Although MPX remains of unknown significance to the mussel industry, tools and protocols described here will be useful in further characterising these and other ciliates (subclass Rhynchodia) known as pathogens for bivalves. [ABSTRACT FROM AUTHOR]

Understanding the behaviours of swimming microorganisms in various environments is important for understanding cell distribution and growth in nature and industry. However, cell behaviour in complex geometries is largely unknown. In this study, we used Tetrahymena thermophila as a model microorganism and experimentally investigated cell behaviour between two flat plates with a small angle. In this configuration, the geometry provided a 'dead end' line where the two flat plates made contact. The results showed that cells tended to escape from the dead end line more by hydrodynamics than by a biological reaction. In the case of hydrodynamic escape, the cell trajectories were symmetric as they swam to and from the dead end line. Near the dead end line, T. thermophila cells were compressed between the two flat plates while cilia kept beating with reduced frequency; those cells again showed symmetric trajectories, although the swimming velocity decreased. These behaviours were well reproduced by our computational model based on biomechanics. The mechanism of hydrodynamic escape can be understood in terms of the torque balance induced by lubrication flow. We therefore conclude that a cell's escape from the dead end was assisted by hydrodynamics. These findings pave the way for understanding cell behaviour and distribution in complex geometries. [ABSTRACT FROM AUTHOR]

Many anaerobic ciliated protozoa contain organelles of mitochondrial ancestry called hydrogenosomes. These organelles generate molecular hydrogen that is consumed by methanogenic Archaea, living in endosymbiosis within many of these ciliates. Here we describe a new species of anaerobic ciliate, Trimyema finlayi n. sp., by using silver impregnation and microscopy to conduct a detailed morphometric analysis. Comparisons with previously published morphological data for this species, as well as the closely related species, Trimyema compressum, demonstrated that despite them being similar, both the mean cell size and the mean number of somatic kineties are lower for T. finlayi than for T. compressum, which suggests that they are distinct species. This was also supported by analysis of the 18S rRNA genes from these ciliates, the sequences of which are 97.5% identical (6 substitutions, 1479 compared bases), and in phylogenetic analyses these sequences grouped with other 18S rRNA genes sequenced from previous isolates of the same respective species. Together these data provide strong evidence that T. finlayi is a novel species of Trimyema, within the class Plagiopylea. Various microscopic techniques demonstrated that T. finlayi n. sp. contains polymorphic endosymbiotic methanogens, and analysis of the endosymbionts' 16S rRNA gene showed that they belong to the genus Methanocorpusculum, which was confirmed using fluorescence in situ hybridization with specific probes. Despite the degree of similarity and close relationship between these ciliates, T. compressum contains endosymbiotic methanogens from a different genus, Methanobrevibacter. In phylogenetic analyses of 16S rRNA genes, the Methanocorpusculum endosymbiont of T. finlayi n. sp. grouped with sequences from Methanomicrobia, including the endosymbiont of an earlier isolate of the same species, 'Trimyema sp.,' which was sampled approximately 22 years earlier, at a distant (~400 km) geographical location. Identification of the same endosymbiont species in the two separate isolates of T. finlayi n. sp. provides evidence for spatial and temporal stability of the Methanocorpusculum- T. finlayi n. sp. endosymbiosis. T. finlayi n. sp. and T. compressum provide an example of two closely related anaerobic ciliates that have endosymbionts from different methanogen genera, suggesting that the endosymbionts have not co-speciated with their hosts. [ABSTRACT FROM AUTHOR]

Endosymbiosis is a widespread phenomenon and hosts of bacterial endosymbionts can be found all-over the eukaryotic tree of life. Likely, this evolutionary success is connected to the altered phenotype arising from a symbiotic association. The potential variety of symbiont's contributions to new characteristics or abilities of host organisms are largely unstudied. Addressing this aspect, we focused on an obligate bacterial endosymbiont that confers an intraspecific killer phenotype to its host. The symbiosis between Paramecium tetraurelia and Caedibacter taeniospiralis, living in the host's cytoplasm, enables the infected paramecia to release Caedibacter symbionts, which can simultaneously produce a peculiar protein structure and a toxin. The ingestion of bacteria that harbor both components leads to the death of symbiont-free congeners. Thus, the symbiosis provides Caedibacter-infected cells a competitive advantage, the "killer trait." We characterized the adaptive gene expression patterns in symbiont-harboring Paramecium as a second symbiosis-derived aspect next to the killer phenotype. Comparative transcriptomics of infected P. tetraurelia and genetically identical symbiont-free cells confirmed altered gene expression in the symbiont-bearing line. Our results show up-regulation of specific metabolic and heat shock genes whereas down-regulated genes were involved in signaling pathways and cell cycle regulation. Functional analyses to validate the transcriptomics results demonstrated that the symbiont increases host density hence providing a fitness advantage. Comparative transcriptomics shows gene expression modulation of a ciliate caused by its bacterial endosymbiont thus revealing new adaptive advantages of the symbiosis. Caedibacter taeniospiralis apparently increases its host fitness via manipulation of metabolic pathways and cell cycle control. [ABSTRACT FROM AUTHOR]

Ciliated protists are unicellular eukaryotes that play important roles in aquatic ecosystems. One of the major tasks of ciliate taxonomy is to re-evaluate the systematic confusing taxa using modern methods. In the present study, two peniculid ciliates, Marituja cf. caudata and Disematostoma minor collected from east China, were studied using a multi-method approach. New morphological observations supplied additional information for species identification and systematic revision of the order Peniculida. The small subunit ribosomal RNA gene sequences of M. cf. caudata, D. minor, and Frontonia terricola were characterized for the first time and provided new insights into the phylogeny of Peniculida. The family Stokesiidae Roque, 1961, was expanded to include the genera Disematostoma and Marituja in addition to its type genus Stokesia, since the three genera formed a well-supported clade in the phylogenetic analyses. The diagnosis of Stokesiidae was improved to include the newly recognized synapomorphies, i.e., barren kinetosomes on the dorsal side, a ciliated dorsal suture, and the somatic ciliature that can be recognized as transversely oriented circles. Additionally, the systematic relationships of the genera and families of Peniculida were hypothesized. We argue that more diversified morphological features should be considered when assessing diagnostic traits for ciliate taxa during systematic revisions. [ABSTRACT FROM AUTHOR]

Peritrichs are a major group of ciliates with worldwide distribution, and they play important roles in many habitats. Intrafamilial phylogeny of some peritrichs was investigated using information from three genes, which provided more robust interpretations than single-gene analyses. Sixty-seven new sequences including SSU rDNA, ITS1-5.8S-ITS2 and LSU rDNA were aligned with available sequences in GenBank to infer phylogenetic relationships within the families Zoothamniidae and Epistylididae. Results reveal the following relationships: (1) Epistylididae is polyphyletic, consisting of two clades that nest within the Zoothamniidae as part of the crown clade of peritrichs (order Vorticellida) and a third one that is part of the basal clade of peritrichs (order Opercularida); (2) Epistylis elongata falls within one of the clades of Zoothamnium rather than with congeners; (3) Zoothamnium is probably paraphyletic, consisting of three divergent clades, with the genera Myoschiston and Zoothamnopsis intermingled with species of Zoothamnium . The following evolutionary hypotheses can be inferred from these results: (1) the contractile stalk of Zoothamnium is plesiomorphic. (2) Myoschiston, Zoothamnopsis and clade II of Epistylididae are derived from the Zoothamnium morphotype by partial or incomplete development of the spasmoneme that forms the contractile center of the stalk around which the rigid cortex is secreted. (3) Clade I of the Epistylididae, which are primarily colonial forms that appear never to have evolved a spasmoneme of any sort, may represent the ancestral morphotype of peritrichs. [ABSTRACT FROM AUTHOR]

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To understand the transcriptional response of antioxidant defense system and heat shock protein ( Hsp) families of the marine ciliate Euplotes crassus, we analyzed the transcriptome profile using RNA-seq technology after exposure to cadmium (Cd) and copper (Cu). De novo sequence assembly produced 61,240 unigenes with 21,330 BLAST hits and showed high sequence orthology with transcriptomes of other ciliates. Gene annotation and gene ontology (GO) comparison revealed that E. crassus expressed highly diversified but conserved stress-responsive gene families of the antioxidant defense system and Hsps. After waterborne exposure to 250 μg/L of Cd and 25 μg/L of Cu, transcriptional responses of the gene families were significantly modulated, suggesting that even the unicellular E. crassus has a conserved molecular defense mechanism, such as modulating mRNA expression, for homeostasis. These transcriptional responses make E. crassus a potential model for understanding the molecular response of single cell ciliates to heavy metal contamination. [ABSTRACT FROM AUTHOR]

Recently, the heterotrophic nanoflagellate Katablepharis japonica has been reported to feed on diverse red-tide species and contribute to the decline of red tides. However, if there are effective predators feeding on K. japonica, its effect on red tide dynamics may be reduced. To investigate potential effective protist predators of K. japonica, feeding by the engulfment-feeding heterotrophic dinoflagellates (HTDs) Oxyrrhis marina, Gyrodinium dominans, Gyrodinium moestrupii, Polykrikos kofoidii, and Noctiluca scintillans, the peduncle-feeding HTDs Luciella masanensis and Pfiesteria piscicida, the pallium-feeding HTD Oblea rotunda, and the naked ciliates Strombidium sp. (approximately 20 μm in cell length), Pelagostrobilidium sp., and Miamiensis sp. on K. japonica was explored. We found that none of these heterotrophic protists fed on actively swimming cells of K. japonica. However, O. marina, G. dominans, L. masanensis, and P. piscicida were able to feed on heat-killed K. japonica. Thus, actively swimming behavior of K. japonica may affect feeding by these heterotrophic protists on K. japonica. To the contrary, K. japonica was able to feed on O. marina, P. kofoidii, O. rotunda, Miamiensis sp., Pelagostrobilidium sp., and Strombidium sp. However, the specific growth rates of O. marina did not differ significantly among nine different K. japonica concentrations. Thus, K. japonica may not affect growth of O. marina. Our findings suggest that the effect of predation by heterotrophic protists on K. japonica might be negligible, and thus, the effect of grazing by K. japonica on populations of red-tide species may not be reduced by mortality due to predation by protists. [ABSTRACT FROM AUTHOR]

Mycobacteria are widespread microorganisms that live in various environments, including man-made water systems where they cohabit with protozoa. Environmental mycobacterial species give rise to many opportunistic human infections and can infect phagocytic protozoa. Protozoa such as amoebae and ciliates feeding on bacteria can sometimes get rid of non-digestible or pathogenic material by packaging it into secreted fecal pellets. Usually, packaged bacteria are still viable and are protected against chemical and physical stresses. We report here that mycobacteria can be packaged into pellets by ciliates. The model bacterium Mycobacterium smegmatis survived digestion in food vacuoles of the ciliate Tetrahymena pyriformis and was included in expelled fecal pellets. LIVE/DEAD® staining confirmed that packaged M. smegmatis cells preserved their viability through the process. Scanning and transmission electron microscopy revealed that bacteria are packaged in undefined filamentous and/or laminar substances and that just a thin layer of material seemed to keep the pellet contents in a spherical shape. These results imply that packaging of bacteria is more common than expected, and merits further study to understand its role in persistence and dissemination of pathogens in the environment. [ABSTRACT FROM AUTHOR]

Mating pheromones (gamone 1 and gamone 2) in the ciliate Blepharisma are biologically active substances that trigger sexual reproduction (conjugation) under starvation conditions. Gamone 1 is a glycoprotein secreted by type I cells, and gamone 2 is a tryptophan (Trp)-derivative compound secreted by type II cells. Both gamones stimulate complementary mating type cells to promote each gamone production and induce pair formation. To elucidate the biosynthetic pathway of gamone 2, we investigated the enzymes involved in the pathway and the specificity of the enzymes. An RNA-seq analysis revealed that Blepharisma stoltei (Heterotrichea) possesses four indoleamine 2,3-dioxygenase ( IDO ) genes showing distinct expression patterns. Along with results from real-time PCR, these findings demonstrated that each IDO gene has different expression patterns that depend on the cellular conditions. Expression of IDO-I was correlated with the intensity of gamone 2 expression, and the recombinant IDO-I protein showed catalytic activity for 5-hydroxy-L-Trp (5-HTP) but very weak activity for L-Trp. Our results indicate that IDO-I is an enzyme evolutionary specialized to gamone 2 production in Blepharisma , and that the biosynthetic pathway for gamone 2 uses 5-HTP as an intermediate. [ABSTRACT FROM AUTHOR]

ATP-binding cassette (ABC) transporters participate in transporting various substances, including xenobiotics, in or out of cells. However, their genetic information and function in ciliates remain still unclear. In this study, we sequenced and characterized two ABC transporter genes ( EcABCB and EcABCC ), and investigated the effect of cadmium (Cd) and benzo[ a ]pyrene (B[ a ]P) on their function and gene expression, using efflux assay and real-time reverse transcription-polymerase chain reaction (qRT-PCR), respectively, in the marine ciliate, Euplotes crassus . Sequencing analysis and efflux assay showed that EcABCB and EcABCC are typical ABC transporters, possessing conserved function. Exposure to Cd (≥ 5 mg/L) and B[ a ]P (≥ 50.5 μg/L) enhanced accumulation of a substrate. A significant increase in the expression of EcABCB and EcABC mRNA was observed at lower concentration in response to Cd and B[ a ]P. Our findings indicate that Cd and B[ a ]P could inhibit the efflux function of ABC transporters, leading to cellular toxicity in the ciliate. [ABSTRACT FROM AUTHOR]

Despite their important role of linking microbial and classic marine food webs, data on biogeographical patterns of microbial eukaryotic grazers are limited, and even fewer studies have used molecular tools to assess active (i.e., those expressing genes) community members. Marine ciliate diversity is believed to be greatest at the chlorophyll maximum, where there is an abundance of autotrophic prey, and is often assumed to decline with depth. Here, we assess the abundant (DNA) and active (RNA) marine ciliate communities throughout the water column at two stations off the New England coast (Northwest Atlantic)--a coastal station 43 kmfromshore (40mdepth) and a slope station 135 km off shore (1,000m). We analyze ciliate communities using a DNA fingerprinting technique, Denaturing Gradient Gel Electrophoresis (DGGE), which captures patterns of abundant community members. We compare estimates of ciliate communities from SSU-rDNA (abundant) and SSU-rRNA (active) and find complex patterns throughout the water column, including many active lineages below the photic zone. Our analyses reveal (1) a number of widely-distributed taxa that are both abundant and active; (2) considerable heterogeneity in patterns of presence/absence of taxa in offshore samples taken 50m apart throughout the water column; and (3) three distinct ciliate assemblages based on position from shore and depth. Analysis of active (RNA) taxa uncovers biodiversity hidden to traditional DNA-based approaches (e.g., clone library, rDNA amplicon studies). [ABSTRACT FROM AUTHOR]

The phytoplankton and ciliate biomasses coupled with environmental factors were investigated in 15 transects in north coasts of Sfax (Tunisia, Eastern Mediterranean Sea) in July 2007. The phytoplankton biomass was dominated by Bacillariophyceae (89.66%), followed by Dinophyceae (10.07%), Coccolithophorideae (0.96%), Cyanobacteriae (0.21%), Chlorophyceae (0.03%) and Euglenophyceae (0.01%). Coscinodiscus sp. (93.26%) was the most abundant species of Bacillariophyceae group and associated with a high nutrient availability. Ciliate biomass was highly variable, with a large dominance of Spirotrichea, up to 96.2%. Biomass followed distinct patterns because of differences in the observed organism biovolumes. The spatial distribution of the ciliates biomass seems to be dependent on environmental factors and probably on their capacity to exploit a wide range of food resources including phytoplankton. The pollution generated by the phosphate-treating manufactory influenced the spatial phytoplankton and ciliate community's distribution and their diversity along the north coast of Sfax. [ABSTRACT FROM AUTHOR]

[Display omitted] The transition from growth to division during the cell cycle encompasses numerous conserved processes such as large-scale DNA replication and protein synthesis. In ciliate cells, asexual cell division is accompanied by additional cellular changes including amitotic nuclear division, extensive ciliogenesis, and trichocyst replication. However, the molecular mechanisms underlying these processes remain elusive. In this study, we present single-cell gene expression profiles of Paramecium cf. multimicronucleatum cells undergoing cell division. Our results reveal that the most up-regulated genes in dividing cells compared to growing cells are associated with 1) cell cycle signaling pathways including transcription, DNA replication, chromosome segregation and protein degradation; 2) microtubule proteins and tubulin glycylases which are essential for ciliogenesis, nuclei separation and structural differentiation signaling; and 3) trichocyst matrix proteins involved in trichocyst synthesis and reproduction. Furthermore, weighted gene co-expression network analysis identified hub genes that may play crucial roles during cell division. Our findings provide insights into cell cycle regulators, microtubules and trichocyst matrix proteins that may exert influence on this process in ciliates. [ABSTRACT FROM AUTHOR]

Protists are nonfungal eukaryotes that represent a vastly diverse component of plant microbial communities. Predatory protists provide plant microbiome ecosystems with nutrient recycling, culling and dispersal of the bacterial community, and selection and activation of microbial biocontrol or growth promoting traits, all functions important for plant health. Here, we briefly review protist interactions with bacteria in the rhizosphere, following topics discussed at the 12th Japan-US Seminar in Plant Pathology. We discuss how understanding of the specificity and signaling mechanisms of predator-prey interactions at the plant interface could be used to develop novel products and strategies for sustainable crop production. • Protist predators are a diverse and ubiquitous part of the plant microbiome, but are functionally poorly understood. • Predators shape the rhizosphere by cycling nutrients, selecting for or activating bacterial defense, or protecting bacteria. • Understanding of protist-bacterial associations could improve discovery, delivery, and efficacy of microbial inoculants. [ABSTRACT FROM AUTHOR]

One of the most dramatic examples of nuclear morphogenesis occurs during conjugation in Tetrahymena when the micronucleus elongates to a size longer than the cell itself. After contraction to a spherical shape, the nucleus moves directly to chromosome separation in the first meiotic division. Here we investigate the consequences of interrupting the elongation process. Colchicine, a microtubule inhibitor, caused retraction of elongated structures. With time, cells began to lose their micronuclei, and by five hours more than half of the paired cells had at least one cell missing a micronucleus. After reversing the colchicine block, existing micronuclei did not undergo elongation again, nor did meiosis occur. These observations indicate that micronuclear elongation is critical to subsequent meiotic division. Further, nuclear elimination occurs, which could be due to meiotic failure or possibly a problem downstream from meiosis. An analysis of the process of colchicine-induced micronuclear degeneration indicated that it was regulated by a caspase-dependent mechanism, characteristic of apoptosis, and then resorbed by a lysosome-dependent autophagic mechanism. Amicronucleate cells failed to grow when returned to nutrient medium, likely because of a lesion in the post-conjugation reconstruction of a functioning oral apparatus. The ease by which a large number of nuclei are induced to “self-destruct” may make this system useful in investigating the link between colchicine treatment and nuclear death in Tetrahymena, and in investigating how nuclear death could be regulated in living cells more generally. Finally, we note that this phenomenon might relate to the evolution of amicronucleate species of Tetrahymena.