Peritrich Ciliate Research Articles

Prevalent ciliate symbiosis on copepods: high genetic diversity and wide distribution detected using small subunit ribosomal RNA gene.

Toward understanding the genetic diversity and distribution of copepod-associated symbiotic ciliates and the evolutionary relationships with their hosts in the marine environment, we developed a small subunit ribosomal RNA gene (18S rDNA)-based molecular method and investigated the genetic diversity and genotype distribution of the symbiotic ciliates on copepods. Of the 10 copepod species representing six families collected from six locations of Pacific and Atlantic Oceans, 9 were found to harbor ciliate symbionts. Phylogenetic analysis of the 391 ciliate 18S rDNA sequences obtained revealed seven groups (ribogroups), six (containing 99% of all the sequences) belonging to subclass Apostomatida, the other clustered with peritrich ciliate Vorticella gracilis. Among the Apostomatida groups, Group III were essentially identical to Vampyrophrya pelagica, and the other five groups represented the undocumented ciliates that were close to Vampyrophrya/Gymnodinioides/Hyalophysa. Group VI ciliates were found in all copepod species but one (Calanus sinicus), and were most abundant among all ciliate sequences obtained, indicating that they are the dominant symbiotic ciliates universally associated with copepods. In contrast, some ciliate sequences were found only in some of the copepods examined, suggesting the host selectivity and geographic differentiation of ciliates, which requires further verification by more extensive sampling. Our results reveal the wide occurrence and high genetic diversity of symbiotic ciliates on marine copepods and highlight the need to systematically investigate the host- and geography-based genetic differentiation and ecological roles of these ciliates globally.

Evolution of Variations in the Common Pattern of Stomatogenesis in Peritrich Ciliates: Evidence from a Comparative Study Including a New Description of Stomatogenesis in Pseudepistylis songi Peng et al.,

The stomatogenesis of peritrich ciliates is an important developmental process but has been studied relatively little for such a large, diverse taxon. Complex oral structures and an inability of staining techniques to reveal them clearly have been the major factors hindering investigation of this process. In the present study, an improved method of staining with protargol was used to investigate the entire process of stomatogenesis in a large, colonial species of peritrich, Pseudepistylis songi, and to compare it to descriptions of stomatogenesis in several other species. We found that P. songi and other peritrichs have the same general type of ophryobuccokinetal stomatogenesis, with the parental oral complex being inherited by one daughter and the new oral complex by the other daughter; however, some differences between individual taxa appear to have arisen in the course of evolution. Reorganization of the entire germinal kinety (Gk) to form the germinal anlage may be plesiomorphic, with restriction of reorganization to its abstomal part being apomorphic. Development of the entire new haplokinety of one daughter cell (2Hk) from the germinal band or its homologue also appears to be plesiomorphic, with development of peristomial and infundibular parts of 2Hk from separate rudiments being apomorphic. Furthermore, development of the new Gk of the parental oral complex (1Gk) from the entire infundibular part of the parental haplokinety (1Hk) may be plesiomorphic, and development from just the abstomal part may be apomorphic. Finally, development of the Gk of the new oral complex (2Gk) from residual kinetosomes of the germinal band appears to be plesiomorphic.

Spatial genetic variation, phylogeography and barcoding of the peritrichous ciliate Carchesium polypinum

Most recent studies of geographic distribution of microbial eukaryotes have focused on marine rather than freshwater protists. Here, we used the freshwater peritrich ciliate Carchesium polypinum to quantify the degree of genetic diversity of four closely related and previously described lineages and to determine whether patterns of genetic differentiation showed geographic partitioning. Using an expanded dataset of 100 isolates and employing the mitochondrial marker cytochrome oxidase c subunit I (cox-1), we enriched the 6 previously identified clades of Carchesium polypinum. We found a large degree of geographic overlap among the different clades (e.g. to the level of range of sampling), but also a spatially restricted clade (e.g. to the level of one river basin). Furthermore, we present evidence of a clear geographic separation in one of the lineages with Canadian and North Carolinian isolates grouping in two distinct clusters.

Molecular and Morphological Characterization of a Poorly Known Marine Ciliate, Myoschiston duplicatum Precht, : Implications for Phylogenetic Relationships between Three Morphologically Similar Genera — Zoothamnium, Myoschiston, and Zoothamnopsis (Ciliophora, Peritrichia, Zoothamniidae)

We studied the morphology and molecular phylogeny of Myoschiston duplicatum, a peritrich ciliate that has been recorded as an epibiont of crustaceans, but which we also identified on marine algae from Korea. The important morphological characteristics revealed by silver staining of Myoschiston species have not been described because they are rarely collected. Using morphological methods, we redescribed the type species of the genus, Myoschiston duplicatum, and provided an improved diagnosis of Myoschiston. In addition, the coding regions for nuclear small subunit (SSU) rRNA and internal transcribed spacer 1-5.8S-internal transcribed spacer 2 sequences were sequenced. Phylogenetic analyses that included available SSU rDNA sequences of peritrichs from GenBank strongly supported a position of M. duplicatum within the family Zoothamniidae. In addition, phylogenetic analyses were performed with single datasets (ITS1-5.8S-ITS2) and combined datasets (SSU rDNA+ITS1-5.8S-ITS2) to explore further the phylogenetic relationship in the family Zoothamniidae between the three morphologically similar genera-Zoothamnium, Myoschiston, and Zoothamnopsis.

Track analysis and geographic distribution of some Lagenophrys Stein, 1852 (Protozoa: Ciliophora: Peritrichia) species

Records of species of the peritrich ciliate genus Lagenophrys, obligate epibionts of crustaceans, were analysed using panbiogeographical track analysis to detect its worldwide distribution patterns. Track analysis of 28 species of this genus found five generalized tracks: (1) Euroamerican; (2) American; (3) Caribbean; (4) Tasmanian; and (5) South American. Two panbiogeographic nodes were found at the intersection of generalized tracks 1 and 2, and 2 and 5. The distributions of the Lagenophrys species show complex patterns that could be related to trophic opportunities, habitat and distribution of hosts, and other factors. The distribution patterns suggest that a number of allopatric events have caused regionalization of the taxa.

Revealing the Diversity and Quantity of Peritrich Ciliates in Environmental Samples Using Specific Primer-based PCR and Quantitative PCR

Peritrichs are a diverse, ecologically important ciliate group usually with a complex life cycle. To date, the community of the peritrichs has been investigated by using morphology-based methods such as living observation and silver staining. Here we show a molecular approach for characterizing the diversity and quantity of free-living peritrichs in environmental samples. We newly designed four peritrich-specific primers targeting 18S rRNA genes that allow clone library construction, screening and analysis. A quantitative real-time PCR (qPCR) assay was developed to quantify peritrichs in environmental samples by using rDNA copy number as an indicator. DNA extracted from four water samples of contrasting environmental gradients was analysed. The results showed that the peritrich community was differentiated among these samples, and that the diversity decreased with the increase of water salinity. The qPCR results are consistent with the library sequence analysis in terms of quantity variations from sample to sample. The development of peritrich-specific primers, for the first time, for conventional PCR and qPCR assays, provides useful molecular tools for revealing the diversity and quantity of peritrich ciliates in environmental samples. Also, our study illustrates the potential of these molecular tools to ecological studies of other ciliate groups in diverse environments.

Ciliate Epibionts Associated with Crustacean Zooplankton in German Lakes: Distribution, Motility, and Bacterivory

Ciliate epibionts associated with crustacean zooplankton are widespread in aquatic systems, but their ecological roles are little known. We studied the occurrence of ciliate epibionts on crustacean zooplankton in nine German lakes with different limnological features during the summer of 2011. We also measured the detachment and re-attachment rates of the ciliates, changes in their motility, and the feeding rates of attached vs. detached ciliate epibionts. Epibionts were found in all lakes sampled except an acidic lake with large humic inputs. Epibiont prevalence was as high as 80.96% on the cladoceran Daphnia cucullata, 67.17% on the cladoceran Diaphanosoma brachyurum, and 46.67% on the calanoid copepod Eudiaptomus gracilis. Both cladoceran groups typically had less than 10 epibionts per individual, while the epibiont load on E. gracilis ranged from 1 to >30 epibionts per individual. After the death of the zooplankton host, the peritrich ciliate epibiont Epistylis sp. detached in an exponential fashion with a half-life of 5 min, and 98% detached within 30 min, leaving behind the stalks used for attachment. Immediately after detachment, the ciliates were immotile, but 62% became motile within 60 min. When a new host was present, only 27% reattached after 120 min. The average measured ingestion rate and clearance rate of Epistylis were 11,745 bacteria ciliate−1 h−1 and 24.33 μl ciliate−1 h−1, respectively. Despite their high feeding rates, relatively low epibiont abundances were observed in the field, which suggests either diversion of energy to stalk formation, high metabolic loss by the epibionts, or high mortality among the epibiont populations.

Parasites and pathogens of the endosymbiotic pea crab ( Pinnotheres pisum) from blue mussels ( Mytilus edulis) in England

A histopathological survey of the commensal pea crab ( Pinnotheres pisum) from the mantle cavities of blue mussels ( Mytilus edulis) has been conducted. A total of 266 pea crabs from eight sites around the English coastline were examined. Of these, 82 were negative for any visible infections by histology. The remaining pea crabs were infected with an intranuclear bacilliform virus designated as P. pisum bacilliform virus ( PpBV) in the hepatopancreatic epithelial cells, peritrichous ciliates on the gills, an intracytoplasmic microsporidian infection of the hepatopancreatocytes, a myophilic microsporidian infection, the gregarine Cephaloidophora fossor in the hepatopancreas, the entoniscid isopod Pinnotherion vermiforme, a low level nematode infection and an acanthocephalan cystacanth. Host reactions to infections were generally subdued. Results are discussed in relation to the endocommensal habitat of the pea crabs.

Redescriptions of five species of marine peritrichs, Zoothamnium plumula, Zoothamnium nii, Zoothamnium wang, Pseudovorticella bidulphiae, and Pseudovorticella marina (Protista, Ciliophora)

The significant similarity of congeners and an accumulation of inaccurate descriptions or misinterpretations in previous studies can make it difficult to separate species of peritrich ciliates from one another. Isolates collected from Chinese coastal waters allowed investigation and redescription of three marine species of peritrichs, Zoothamnium plumula Kahl, 1933, Z. nii Ji et al., 2005 and Z. wangi Ji et al., 2005, emphasizing in particular details of characters observed in vivo, infraciliature and comparisons with morphologically similar congeners. Brief redescriptions based on new material also are provided for two other marine peritrichs, Pseudovorticella bidulphiae (Stiller, 1939) Ji et al., 2009 and P. marina (Greeff, 1870) Ji et al., 2009.

A new interpretation of stomatogenesis in a peritrich ciliate: Using Campanella umbellaria as a model system

The process of stomatogenesis in peritrich ciliates is still incompletely understood. Previous studies on the stomatogenesis of four species of peritrichs, Telotrochidium sp., Carchesium polypinum, Opercularia coarctata, and Astylozoon pyriforme conflict with one another in some cases and omit details of events in others. We described the entire process of stomatogenesis in the peritrich ciliate Campanella umbellaria (C. umbellaria) using an improved method of staining with protargol. Our results disagree with some previous studies with regard to the formation of some rudimentary structures, reorganization of the parental haplokinety, formation of new germinal rows, and separation of daughter oral complexes. The pattern of stomatogenesis characteristic of peritrichs is compared to the stomatogenetic patterns of three other oligohymenophorean subclasses and a hypothesis about the evolution of stomatogenesis in the class Oligohymenophorea is offered. Details of stomatogenesis need to be described and verified in a greater variety of peritrichs to clarify possible differences between taxa and make it possible to relate stomatogenesis to evolution within the subclass Peritrichia. Ultrastructural studies are the next step in description of morphogenetic processes in peritrichs, and characteristics of C. umbellaria make it a useful model for this work.

Morphology and Morphogenesis of a Freshwater Ciliate, Epistylis chlorelligerum Shen, 1980 (Ciliophora, Peritrichia)

An oligohalobic peritrichous ciliate, Epistylis chlorelligerum Shen, 1980, was collected from a ditch in Hangzhou, China. The morphology, oral infraciliature, and morphogenesis of the species were studied using living and protargol-impregnated specimens. Zooids of E. chlorelligerum are 160-230 × 50-60 μm in vivo, and characterized by green-colored endoplasm containing symbiotic algae. The oral infraciliature presents a well-developed filamentous reticulum linked to the circular fiber of the cytostome; the outer two rows of P3 extend adstomally over P1 and usually enfold it. During binary fission, one daughter cell inherits most part of the old buccal apparatus and the reorganized haplokinety and germinal kinety (Hk' and G'), and new buccal apparatus of the other daughter cell is mostly developed from the original germinal kinety (G) and haplokinety (Hk): new peniculi 2, 3 (2P2, 2P3), new haplokinety (2Hk), and new germinal kinety (2G) are formed from G, while the new peniculus 1 (2P1) and its peristomial extention (2Pk) originate from Hk. The epistomial membrane can be observed until the two sets of buccal apparatus begin to separate from each other.

Long‐Term Dynamics of Microbial Biofilm Communities of the River Rhine with Special References to Ciliates

AbstractThe aim of this study was to quantify and qualify seasonal changes of all important components of a microbial biofilm community. We explored the development of the biofilm community structure on submerged glass slides for 15 months including all organisms from bacteria to macro‐invertebrates. Besides bacteria, heterotrophic flagellates were the most abundant biofilm component followed by ciliates, meiofauna organisms and algae. Most important were sessile choanoflagellates, peritrichous ciliates and some crustaceans. Ciliates and macrofauna were the most important components with regard to the total biovolume. The biofilm architecture was strongly influenced by extracellular structures produced by protozoans and macro‐invertebrates. Alterations within the biofilm community were mainly due to changes in abundances rather than in the composition except for heterotrophic flagellates and macro‐invertebrates. Biofilm organisms were dominated by planktivorous organisms exerting a strong grazing impact on the plankton organisms in this large river. (© 2011 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)

Evidence for Cryptic Speciation in Carchesium polypinum Linnaeus, 1758 (Ciliophora: Peritrichia) Inferred from Mitochondrial, Nuclear, and Morphological Markers

Protist diversity is currently a much debated issue in eukaryotic microbiology. Recent evidence suggests that morphological and genetic diversity might be decoupled in some groups of protists, including ciliates, and that these organisms might be much more diverse than their morphology implies. We sought to assess the genetic and morphological diversity of Carchesium polypinum, a widely distributed peritrich ciliate. The mitochondrial marker cytochrome c oxidase subunit I and the nuclear small subunit ribosomal RNA were used to examine genetic diversity. For the morphological assessment, live microscopy and Protargol staining were used. The mitochondrial marker revealed six robust, deeply diverging, and strongly supported clades, while the nuclear gene was congruent for three of these clades. There were no major differences among individuals from the different clades in any of the morphological features examined. Thus, the underlying genetic diversity in C. polypinum is greater than what its morphology suggests, indicating that morphology and genetics are not congruent in this organism. Furthermore, because the clades identified by the mitochondrial marker are so genetically diverse and are confirmed by a conserved nuclear marker in at least three cases, we propose that C. polypinum be designated as a "cryptic species complex." Our results provide another example where species diversity can be underestimated in microbial eukaryotes when using only morphological criteria to estimate species richness.

Morphometric study of a Brazilian strain of Carchesium polypinum (Ciliophora: Peritrichia) attached to Pomacea figulina (Mollusca: Gastropoda), with notes on a high infestation

During an ecological study of the epibiotic relationship between ciliate protists and Pomacea figulina (Spix, 1827) (Gastropoda, Ampullariidae), originating from an urban stream in southeast Brazil, a high infestation by the peritrich ciliate Carchesium polypinum (Linnaeus, 1758) Ehrenberg, 1830 (Ciliophora, Peritrichia) associated to the shell of one mollusc among 23 was observed. We provided a morphological and morphometric study of C. polypinum using observations of specimens in vivo, after protargol staining, and examined using scanning electron microscopy. The Brazilian-population of C. polypinum is characterized by: size of zooid in vivo 89 µm x 57 µm on average; colony regularly dichotomously branched with usually up to 40 zooids; macronucleus usually J-shaped; single contractile vacuole located in the upper third of body; myoneme not continuous throughout the colony; stalks contract despite the discontinuity of their individual myonemes; polykinety comprises three peniculi, each consisting of three kineties. The high infestation showed here could be related to the preference for eutrophic environments showed by C. polypinum and suggested that ciliate epibionts may be ecologically important in aquatic habitats.

Alpha‐Tubulin and Small Subunit rRNA Phylogenies of Peritrichs Are Congruent and Do Not Support the Clustering of Mobilids and Sessilids (Ciliophora, Oligohymenophorea)

Peritrich ciliates have been traditionally subdivided into two orders, Sessilida and Mobilida within the subclass Peritrichia. However, all the existing small subunit (SSU) rRNA phylogenetic trees showed that the sessilids and mobilids did not branch together. To shed some light on this disagreement, we tested whether or not the classic Peritrichia is a monophyletic group by assessing the reliability of the SSU rRNA phylogeny in terms of congruency with alpha-tubulin phylogeny. For this purpose, we obtained 10 partial alpha-tubulin sequences from peritrichs and built phylogenetic trees based on alpha-tubulin nucleotide and amino acid data. A phylogenetic tree from the alpha-tubulin and SSU rRNA genes in combination was also constructed and compared with that from the SSU rRNA gene using a similar species sampling. Our results show that the mobilids and sessilids are consistently separated in all trees, which reinforces the idea that the peritrichs do not constitute a monophyletic group. However, in all alpha-tubulin gene trees, the urceolariids and trichodiniids do not group together, suggested mobilids may not be a monophyletic group.

A New Flagship Peritrich (Ciliophora, Peritrichida) from the River Rhine, Germany:Apocarchesium arndtin. sp.

We discovered a free-living peritrich ciliate with outstanding features in the River Rhine. Its morphology and 18S rRNA gene sequence were studied with standard methods. Apocarchesium arndti n. sp. has several peculiarities. (i) There are ordinary zooids, macrozooids, and microzooids, which form a hemispherical rosette on a discoidal base, the stalk dish, locking the approximately 18 microm wide and up to 2 mm long, spirally contracting colony stalk. (ii) The stalk myoneme is connected only to the microzooids. (iii) A rosette contains up to 50 zooids not connected to each other but individually attached to the stalk dish with the scopula. (iv) The ordinary zooids are epistylidid, trumpet-shaped (approximately 6:1 length:width), about 180 x 30 microm in size, and have an ellipsoidal macronucleus subapically between oral cavity and dorsal side. (v) The myoneme system of the zooids, which can contract individually, forms a tube-like structure in the narrow posterior half of the cell. (vi) The silverline pattern belongs to the transverse-striate type. (vii) The oral apparatus is of usual structure, with kinety 1 of peniculus 3 distinctly shortened proximally. (viii) The 18S rRNA places A. arndti n. sp. as a distinct lineage near Vorticella and Carchesium. These data are used to provide an improved diagnosis of the genus Apocarchesium. Features (i)-(iii) and the molecular data indicate that Apocarchesium could be the type genus of a new peritrich family.

Analysis of the secondary structure of ITS transcripts in peritrich ciliates (Ciliophora, Oligohymenophorea): Implications for structural evolution and phylogenetic reconstruction

Despite extensive previous morphological work, little agreement has been reached about phylogenetic relationships among peritrich ciliates, making it difficult to study the evolution of the group in a phylogenetic framework. In this study, the nucleotide characteristics and secondary structures of internal transcribed spacers 1 and 2 (ITS1 and ITS2) of 26 peritrich ciliates in 12 genera were analyzed. Information from secondary structures of ITS1 and ITS2 then was used to perform the first systematic study of ITS regions in peritrich ciliates, including one species of Rhabdostyla for which no sequence has been reported previously. Lengths of ITS1 and ITS2 sequences varied relatively little among taxa studied, but their G+C content was highly variable. General secondary structure models of ITS1 and ITS2 were proposed for peritrich ciliates and their reliability was assessed by compensatory base changes. The secondary structure of ITS1 contains three major helices in peritrich ciliates and deviations from this basic structure were found in all taxa examined. The core structure of peritrich ITS2 includes four helices, with helix III as the longest and containing a motif 5'-MAC versus GUK-3' at its apex as well as a YU-UY mismatch in helix II. In addition, the structural motifs of both ITS secondary structures were identified. Phylogenetic analyses using ITS data were performed by means of Bayesian inference, maximum likelihood and neighbor joining methods. Trees had a consistent branching pattern that included the following features: (1) Rhabdostyla always clustered with members of the family Vorticellidae, instead of members of the family Epistylididae, in which it is now classified on the basis of morphology. (2) The systematically questionable genus Ophrydium closely associated with Carchesium, forming a clearly defined, monophyletic group within the Vorticellidae. This supported the hypothesis derived from previous study based on small subunit rRNA gene sequences that Ophrydium and its few relatives are morphologically anomalous vorticellids, not sufficiently distinct to be given familial status and should be placed within a more broadly defined family Vorticellidae. This study validated for the first time a secondary structure of ITS1 and ITS2 from peritrich ciliates and demonstrated its potential in helping to resolve deep phylogenetic relationships.

Prevalence and seasonality of Zoothamnium duplicatum (Protozoa: Ciliophora) epibiont on an estuarine mysid (Crustacea: Mysida) in tropical mangrove brackish water

The estuarine mysid Mesopodopsis tenuipes (Crustacea: Mysida) was often found carrying the peritrich ciliate Zootham- nium duplicatum (Protozoa: Ciliophora) on its body. The prevalence and seasonality of this association were studied on the basis of a year-round survey conducted in the Merbok mangrove brackish water in north-western Peninsular Malaysia from March 2005 to Feb- ruary 2006. The frequency of mysids hosting ciliates varied from 0 to 57.3%, with an annual mean of 17.0%. Juveniles showed a sig- nificantly lower prevalence of ciliates than adult mysids. There was no difference in infestation prevalence between the sexes. An epibi- otic renewal process could be assessed through observations in breeding females. The infestation prevalence and loads were apprecia- bly lower in the females carrying earlier stages of embryos than in those with developed ones, indicating that zoothamnid ciliates may require two or more days to reach a saturation phase in terms of prevalence when ciliate reproduction is high. This, in turn, suggests that the high moulting frequency of the tropical mysid may inhibit high colonisation pressure by the epibiont. A periodic seasonal trend was not determined, although mysids in lower salinity conditions were more likely to carry epibionts than those at higher salini- ties.

Reconsideration of Phylogenetic Relationships of the Subclass Peritrichia (Ciliophora, Oligohymenophorea) Based on Small Subunit Ribosomal RNA Gene Sequences, with the Establishment of a New Subclass Mobilia Kahl, 1933

ABSTRACT. Based on its characteristic oral apparatus, the ciliate subclass Peritrichia has long been recognized as a monophyletic assemblage composed of the orders Mobilida and Sessilida. Following the application of molecular methods, the monophyly of Peritrichia has recently been questioned. We investigated the phylogenetic relationships of the peritrichous ciliates based on four further complete small subunit ribosomal RNA sequences of mobilids, namely Urceolaria urechi, Trichodina meretricis, Trichodina sinonovaculae, and Trichodina ruditapicis. In all phylogenetic trees, the mobilids never clustered with the sessilids, but instead formed a monophyletic assemblage related to the peniculines. By contrast, the sessilids formed a sister clade with the hymenostomes at a terminal position within the Oligohymenophorea. We therefore formally separate the mobilids from the sessilids (Peritrichia sensu stricto) and establish a new subclass, Mobilia Kahl, 1933, which contains the order Mobilida Kahl, 1933. We argue that the oral apparatus in the mobilians and sessilid peritrichs is a homoplasy, probably due to convergent evolution driven by their similar life-styles and feeding strategies. Morphologically, the mobilians are distinguished from all other oligohymenophoreans by the presence of the adhesive disc, this character being a synapomorphy for the Mobilia.

A new species of Pachytrocha Kent, 1882 (Ciliophora, Peritrichia: Vaginicolidae)

The peritrich ciliate Pachytrocha zhytomirensis n. sp. is described from the sewage treatment system of Zhytomir (Ukraine). The new species is characterized by an elongate, subgeniculate lorica sheltering a single zooid with an annular ridge and a peristomial lip thickened noticeably near the entrance to the infundibulum. When contracting, the upper part of the cell bends toward the wall of the lorica and the inflated edge of the peristomial lip covers the body. The macronucleus is ribbon-like and oriented parallel to the longitudinal axis of the cell. Cothurnia asymmetrica Sommer, 1951 is recognized as a synonym of Pachytrocha cothurnoides Kent, 1882.