Phylogenetic Position of the Genus Cyrtostrombidium,with a Description of Cyrtostrombidium paralongisomum nov. spec. and a Redescription of Cyrtostrombidium longisomum Lynn & Gilron, 1993 (Protozoa,Ciliophora) Based on Live Observation,Protargol Impregnation,and 18S rDNA Sequences

We redescribe Cyrtostrombidium longisomum Lynn & Gilron, 1993, the type species of the genus Cyrtostrombidium, and describe the new species Cyrtostrombidium paralongisomum n. sp. using live observation, protargol staining and molecular data. The morphological characters of these two species are clearly distinct, i.e., dikinetid numbers in the girdle and ventral kineties; however, it is difficult to separate them by 18S rDNA sequences because they differ by only 8 bp, indicating that 18S rDNA sequences are insufficient for separating different species in the genus Cyrtostrombidium. We not only observed the position of the oral primordium in the genus Cyrtostrombidium but also observed a possibly homoplasious trait, a dorsal split in the girdle kinety, in (1) Apostrombidium, (2) Varistrombidium, and (3) Cyrtostrombidium/Williophrya. This partially supports the hypothesis of somatic ciliary pattern evolution recently put forth by Agatha and Strüder‐Kypke.     

Trimyema compressum Lackey, 1925: Morphology,Morphogenesis and Systematic Implications1

ABSTRACT Trimyema compressum is a species included in the family Trimyemidae with the single genus Trimyema. This species has 50–60 somatic kinetics and three rows of kinetosomes surrounding the oral cavity. Two isolated groups of kinetosomes can also be observed on the right side of the oral region. The morphogenesis of bipartition is telokinetal; all the new infraciliary structures of the opisthe come from the longitudinal and postero-anterior proliferation of the last kinetosome of all the somatic kinetics. In the proter there is a reorganization of the oral infraciliature. As a result of our observations, we suggest that the systematic position of the genus Trimyema be changed from the subclass Vestibulifera to the subclass Gymnostomata. We also consider that this genus must be included in the suborder Trimyemina Jankowski, 1980.     

Description of a New Freshwater Ciliate Epistylis wuhanensis n. sp. (Ciliophora,Peritrichia) from China,with a Focus on Phylogenetic Relationships within Family Epistylididae

Two populations of Epistylis wuhanensis n. sp., a new freshwater peritrich ciliate, were isolated from different freshwater ponds located in Hubei, China. Their morphological characteristics were investigated using live observation, protargol impregnation, and scanning electron microscopy (SEM). Specimens from the two populations showed identical arrangement of the infraciliature and identical small subunit ribosomal RNA (SSU rRNA) gene and ITS1‐5.8S‐ITS2 sequences. The zooids present bell‐shaped and 90–175 × 27–54 μm in vivo. Macronucleus is variable in shape and located in the middle of cell. Pellicle is usually smooth with 139–154 and 97–105 striations above and below the trochal band, respectively. SSU rRNA gene and ITS1‐5.8S‐ITS2 sequences of E. wuhanensis n. sp. did not match any available sequences in GenBank. Phylogenetically, E. wuhanensis n. sp. clusters with the other Epistylis within the family Epistylididae, but is distinct from the major clades of Epistylis. Above all, the morphological characteristics and molecular analyses support that the present Epistylis is a new species. Expanded phylogenetic analyses of sessilids based on both SSU rRNA gene sequences and ITS1‐5.8S‐ITS2 sequences reveal that the genus Epistylis consists of Epistylis morphospecies and taxonomic revision of the genus is needed.     

Tunicothrix rostrata n. g., n. sp., a new urostylid ciliate (Ciliophora, Stichotrichia) from the Yellow Sea

The morphology and infraciliature of a new ciliate, Tunicothrix rostrata n. g., n. sp., isolated from the Yellow Sea, are investigated using live observation and protargol impregnation. Tunicothrix rostrata measures about 160 x 40 mum in vivo, and has a frontal beak-like protrusion, a conspicuous cortical alveolar layer, two right marginal rows, and usually three distinct midventral pairs. The discovery of T. rostrata enables us to reconsider the classification of Erniella wilberti, a curious ciliate with obscure midventral pairs. Both species are highly similar in overall appearance and nuclear and ciliary pattern. Thus, they are united in a new genus of the family Urostylidae, Tunicothrix, and E. wilberti is transferred to Tunicothrix: Tunicothrix wilbertiLin and Song, 2004 n. comb. By contrast, Erniella filiformis, type species of Erniella, has several ventral rows and does not belong to the urostylids. Tunicothrix rostrata is easily distinguished from T. wilberti by its beak-like anterior protrusion and by the distinctly elongated right marginal row 2, which curves anteriorly on the dorsal side of the cells. Tunicothrix is closely related to Parabirojimia, differing by the invariably two (vs. five-eight) right marginal rows and the conspicuous (vs. ordinary) alveolar layer, a unique feature in urostylid ciliates.     

Morphological Characterizations of Four Species of Parallelostrombidium (Ciliophora,Oligotrichia), with a Note on the Phylogeny of the Genus

The morphology and phylogeny of four oligotrichid ciliates, Parallelostrombidium paraellipticum sp. n., P. dragescoi sp. n., P. jankowskii (Xu et al. 2009) comb. n., and P. kahli (Xu et al. 2009) comb. n., are described or redescribed based on live observation, protargol stained material, and SSU rRNA gene sequences. The new species P. paraellipticum sp. n. is characterized by its obovoidal cell shape, adoral zone composed of 17–21 collar, 9–11 buccal, and two thigmotactic membranelles, and extrusomes attached in one row along the girdle kinety. The new species P. dragescoi sp. n. is distinguished from its congeners by its obovoidal cell shape and a lack of thigmotactic membranelles. Based on ciliary patterns recognizable in the original slides, Omegastrombidium jankowskii Xu et al. 2009 and O. kahli Xu et al. 2009 should be transferred to the genus Parallelostrombidium Agatha 2004. Phylogenetic analyses based on SSU rRNA gene sequence data demonstrate that all four new sequences cluster with previously described congeners. The genus Parallelostrombidium is separated into two clusters, suggesting its non‐monophyly and probably corresponding to the two subgenera proposed by Agatha and Strüder‐Kypke (2014), as well as their morphological difference (cell dorsoventrally flattened vs. unflattened).     

Pseudonotohymena antarctica n. g., n. sp. (Ciliophora,Hypotricha), a New Species from Antarctic Soil

A new soil ciliate, Pseudonotohymena antarctica n. g., n. sp., from King George Island, Antarctica, is described based on live observation, protargol impregnation, and its 18S rRNA gene. The new genus Pseudonotohymena is morphologically similar to the genus Notohymena Blatterer and Foissner 1988 in the following characteristics: 18 fronto‐ventral‐transverse cirri, a flexible body, undulating membranes, dorsomarginal kineties, and the number of cirri in the marginal rows. However, Pseudonotohymena differs from Notohymena particularly in the dorsal ciliature, that is, in possessing a nonfragmented dorsal kinety (vs. fragmented). In addition, the molecular phylogenetic relationship of the new species differs from that of Notohymena species. On the basis of the morphological features, the genetic data, and morphogenesis, we establish P. antarctica n. g., n. sp. In addition, the cyst morphology of this species is described.     

Morphology and Morphogenesis of Strobilidium caudatum (Fromentel), Meseres corlissi N. Sp., Halteria grandinella (Müller), and Strombidium rehwaldi N. Sp., and a Proposed Phylogenetic System for Oligotrich Ciliates (Protozoa,Ciliophora)1

ABSTRACT. The morphology and morphogenesis of some oligotrichs were investigated using protargol impregnation, silver carbonate impregnation and scanning electron microscopy. The somatic kineties of Strobilidium caudatum form a spiral at the posterior pole. Strobilidiids without such a spiral are transferred to the genus Rimostrombidium. Fourteen new combinations and a nomen novum, Strobilidium kahli, are necessary, Meseres corlissi n. sp. is characterized by eight somatic kineties composed of long cilia which are not fused to “bristles” as they are in Halteria. Strombidium oblongum shows similar characteristics and is thus combined with Meseres. Strombidium rehwaldi n. sp. has an anterior and an equatorial girdle of extrusomes. The morphogenesis of Meseres and Halteria is very similar, i.e. the entire somatic ciliature and the oral primordium originate apokinetally on the cell surface; the parental somatic ciliature is resorbed. In strobilidiids and tintinnids, the oral anlagen develop in a subsurface pouch and the parental somatic kineties, which are not resorbed, elongate by intrakinetal proliferation of basal bodies. In strombidiids, the oral primordium develops in an intracellular sac or tube. These morphogenetic peculiarities and distinct morphologic characters (e.g. arrangement of adoral membranelles) were applied in constructing a phylogenetic system for oligotrichs using hypotrichs as outgroup. This shows that halteriids are more closely related to hypotrichs than they are to other oligotrichs. The Halteriidae are thus raised to ordinal and subclass ranks, Halteriida n. ord., Halteriia n. subcl.     

Phylogenetic Analysis and Fluorescence <Emphasis Type="Italic">In Situ</Emphasis> Hybridization Detection of Archaeal and Bacterial Endosymbionts in the Anaerobic Ciliate <Emphasis Type="Italic">Trimyema Compressum</Emphasis>

The anaerobic free-living ciliate, Trimyema compressum, is known to harbor both methanogenic archaeal and bacterial symbionts in the cytoplasm. To clarify their phylogenetic belongings, a full-cycle rRNA approach was applied to this symbiosis. Phylogenetic analysis showed that the methanogenic symbiont was related to Methanobrevibacter arboriphilicus, which was distantly related to symbionts found in other Trimyema species. This result suggested that Trimyema species do not require very specific methanogenic symbionts, and symbiont replacement could have occurred in the history of Trimyema species. On the other hand, the bacterial symbiont was located near the lineage of the family Syntrophomonadaceae in the phylum Firmicutes. The sequence similarity between the bacterial symbiont and the nearest species was 85%, indicating that bacterial symbionts may be specific to the Trimyema species. The elimination of bacterial symbionts from the ciliate cell by antibiotic treatment resulted in considerably decreased host growth. However, it was not restored by stigmasterol addition (<2 μg ml⁻¹), which was different from the previous report that showed that the symbiont-free strain required exogenous sterols for growth. In addition, the decline of host growth was not accompanied by host metabolism shift toward the formation of more reduced products, which suggested that the contribution of bacterial symbionts to the host ciliate was not a dispose of excessive reducing equivalent arising from the host’s fermentative metabolism as methanogenic symbionts do. This study showed that bacterial symbionts make a significant contribution to the host ciliate by an unknown function and suggested that interactions between bacterial symbionts and T. compressum are more complicated than hitherto proposed.     

Systematic and morphological diversity of endosymbiotic methanogens in anaerobic ciliates

The identities and taxonomic diversity of the endosymbiotic methanogens from the anaerobic protozoaMetopus contortus, Metopus striatus, Metopus palaeformis, Trimyema sp. andPelomyxa palustris were determined by comparative analysis of their 16S ribosomal RNA sequences. Fluorescent oligonucleotide probes were designed to bind to the symbiont rRNA sequences and to provide direct visual evidence of their origins from methanogenic archaea contained within the host cells. Confocal microscopy was used to analyze the morphology of the endosymbionts in whole cells ofMetopus palaeformis, Metopus contortus, Trimyema sp. andCyclidium porcatum. The endosymbionts are taxonomically diverse and are drawn from three different genera;Methanobacterium, Methanocorpusculum andMethanoplanus. In every case the symbionts are closely related to, but different from, free-living methanogens for which sequences are available. It is thus apparent that symbioses have been formed repeatedly and independently. Ciliates which are unrelated to each other (Trimyema sp. andMetopus contortus) may contain symbionts which are closely related, and congeneric ciliates (Metopus palaeformis andM. contortus) may contain symbionts which are distantly related to each other. This suggests that some of the symbiotic associations must be relatively recent. For example, at least one of the symbioses inMetopus must postdate the speciation ofM. palaeformis andM. contortus. Despite this,Metopus contortus, Trimyema sp., Cyclidium porcatum and their respective endosymbionts show sophisticated morphological interactions which probably facilitate the exchange of materials between the partners.     

Morphological and taxonomical studies on two soil haptorid ciliates (Ciliophora, Litostomatea): Clavoplites haranti sp. n. and Enchelys terrenum (Foissner, 1984) comb. n., and taxonomy of the family Enchelyidae Ehrenberg, 1838

The morphology and infraciliature of two soil haptorid ciliates, Clavoplites haranti sp. n. and Enchelys terrenum (Foissner, 1984) comb. n., collected from Malé Karpaty Mts. and Biele Karpaty Mts. (Slovakia), were investigated using live observation and protargol impregnation. Clavoplites haranti is distinguished from its congeners by a combination of characters including: theronts spatulate to slenderly fusiform, trophonts bursiform, size about 60 x 25 microm in vivo; massive oral bulge with broadly clavate to lemon-shaped, 1.5-2 microm long extrusomes arranged in a ring in oral bulge; ellipsoidal macronucleus and one globular micronucleus; on average 13 ciliary rows, 3 anteriorly differentiated to an inconspicuous dorsal brush. Enchelys terrenum is about 85 x 30 microm in size and differs from its congeners by a combination of characters including spatulate shape; massive oral bulge with 4-6 microm long, fusiform extrusomes arranged in a ring in oral bulge; ellipsoidal to reniform macronucleus and one globular micronucleus; usually 19 ciliary rows, 3 anteriorly differentiated to an ordinary dorsal brush. The diagnoses of the family Enchelyidae and the genus Enchelys are improved. The new genus Armatoenchelys differs from the genera Enchelys and Apoenchelys in having both body and oral bulge extrusomes. Enchelys geleii, E. longitricha, and E. vermiformis are transferred to the new genus.     

Descriptions of two new marine scuticociliates,Pleuronema sinica n. sp. and P. wilberti n. sp. (Ciliophora: Scuticociliatida), from the Yellow Sea,China

Two new marine scuticociliates, Pleuronema sinica n. sp. and P. wilberti n. sp., collected from the sand beach of Qingdao, China, were investigated in vivo and following protargol impregnation. Ciliates of the genus Pleuronema are normally recognizable by their large sail-like paroral membrane although one species, P. grolierei, has shorter cilia in the paroral membrane. Neither of the new forms has the conspicuous paroral membrane in vivo so in this respect they are not typical members of this genus. Pleuronema sinica is characterized by its large, conspicuously flattened body, the possession of only one preoral kinety, the irregular-shaped macronucleus and the rather unusual structure of the oral apparatus. By contrast P. wilberti has a medium-size broad-oval body, six to eight preoral kineties and a highly differentiated membranelle 3 that is five- or six-rowed. An identification key is supplied for the 15 species of Pleuronema for which the infraciliature is known.     

Morphology and Phylogeny of Three New Loxophyllum Species (Ciliophora,Pleurostomatida) from Mangrove Wetlands of Southern China

Three new pleurostomatid ciliates, Loxophyllum meridionale sp. n., L. salinum sp. n., and L. planum sp. n., isolated from brackish waters of mangrove wetlands in southern China, were investigated using live observation and protargol impregnation. The main characteristic feature of L. meridionale sp. n. is the presence of three contractile vacuoles (CVs) along its dorsal margin. Loxophyllum salinum sp. n. can be identified by the presence of three closely spaced CVs along the ventral margin and 12–15 right and four to six left kineties. Loxophyllum planum sp. n. is characterized by the combination of two macronuclei, a single, subterminal CV, and the absence of cortical granules. Phylogenetic trees based on the small subunit rDNA sequence data recovered Loxophyllum as a monophyletic group within the pleurostomatids. Loxophyllum meridionale sp. n., L. salinum sp. n., and L. planum sp. n. were all nested within the Loxophyllum clade. Based on its ciliary pattern and general morphological features, Loxophyllum asetosum is transferred to the genus Amphileptus and a new combination is suggested: Amphileptus asetosus (Burkovsky 1970) comb. n. [basionym L. asetosum Burkovsky 1970].     

Morphology of Clapsiella magnifica gen. n., sp. n., a new hypotrichous ciliate with a curious dorsal ciliary pattern

The present work describes the morphology and infraciliature of a new hypotrichous ciliate, Clapsiella magnifica gen. n., sp. n., found in rewetted soil from a temporal pond in Argentina. It was studied by means of live observation and protargol impregnation. Its main diagnostic features are: Flexible hypotrich measuring 250–320 μm × 70–140 μm in vivo; two macronuclear nodules and 4–6 micronuclei. Single contractile vacuole. Cytoplasm transparent, cortical granules absent. Somatic ciliature composed of a tricorona of cirri, three buccal(?) cirri, 6–9 ventral rows, 3–5 right marginal(?) rows, one left marginal row, and 12–17 transverse cirri. Dorsal pattern rather complicated, with about 14 kineties and kinety fragments, with scattered kinetids among them; 17–28 caudal cirri arranged in three rows on dorsal kineties 1, 3, and 7. Remarkably, dorsal kinetids have two or four basal bodies, bearing a stiff bristle arising from left anterior basal body. Adoral zone composed of 70–92 membranelles, occupying about 40% of body length in protargol preparations; paroral and endoral curved, resembling a cyrtohymenid pattern. The peculiar dorsal ciliary arrangement and the unique combination of other characters require the establishment of a new genus for this new species, which is considered incertae sedis in the Hypotricha but possibly related to the oxytrichids.     

Redescription of Antetintinnidium mucicola (Claparède and Lachmann, 1858) nov. gen., nov. comb. (Alveolata,Ciliophora, Tintinnina)

Tintinnid ciliates have traditionally been described and classified exclusively based on their lorica features. Although information on the cell characters is urgently needed for a natural classification, more molecular than cytological data has been accumulated over recent years. Apparently, the tintinnids developed in the marine environment and entered freshwater several times independently. Typical freshwater tintinnids belong to the genera Tintinnidium and Membranicola. The species are comparatively well‐known regarding their morphology and characterised by two unusual de novo originating ciliary rows, the ventral organelles. In contrast, the cell features in the marine/brackish Tintinnidium species, specifically their somatic ciliary patterns, are insufficiently known or not known at all. Therefore, the morphology of a common marine/brackish representative, Tintinnidium mucicola, is redescribed based on live observation and protargol‐stained material. Furthermore, biogeographical and autecological data of the species are compiled from literature and own records. The phylogenetic relationships of T. mucicola are inferred and the diversity of the family Tintinnidiidae is assessed from 18S rDNA sequences. The study shows that T. mucicola is not only molecularly distinct, but also characterised by many plesiomorphic features, for instance, it does not possess a verifiable homologue to the ventral organelles. Hence, a new genus, Antetintinnidium nov. gen., is established for T. mucicola. The new insights into the diversity of Tintinnidiidae shed light on the early evolution of tintinnids and might provide clues on their adaptions to freshwater.     

Molecular Phylogeny and Taxonomy of Two Novel Brackish Water Hypotrich Ciliates,with the Establishment of a New Genus,Antiokeronopsis gen. n. (Ciliophora,Hypotrichia)

Two novel brackish water urostyloid ciliates, Anteholosticha paramanca sp. n. and Antiokeronopsis flava gen. n., sp. n., isolated from the Shenzhen Mangrove Nature Protection Area on the coast of the South China Sea, were investigated using live observation and protargol impregnation techniques. Anteholosticha paramanca sp. n. is characterized by its spherical yellowish cortical granules arranged in lines, shortened midventral complex and three transverse cirri. Morphogenesis is similar to that in Anteholosticha manca. The new genus Antiokeronopsis is diagnosed by having a continuous adoral zone of membranelles, frontal cirri arranged in a bicorona, midventral complex composed of midventral pairs only, one marginal cirral row on each side, the presence of frontoterminal and transverse cirri, and the lack of buccal and caudal cirri. The type species A. flava sp. n. is characterized by its elongated body shape, brown to yellowish body color and two types of cortical granules. Small subunit ribosomal RNA gene sequence data justify the classification of both species. Phylogenetic analyses indicate that A. paramanca clusters with Bakuella subtropica within a clade that includes two other Anteholosticha species, while Antiokeronopsis groups within the core urostylids and is most closely related to the well‐known genera Pseudokeronopsis and Uroleptopsis.     

Morphology and Small Subunit rDNA Gene Sequence of Pseudoamphisiella quadrinucleata n. sp. (Ciliophora,Urostylida) from the South China Sea

ABSTRACT. The urosylid genus Pseudoamphisiella was established by Song (1996) with hitherto only two known congeners. In the present work, the morphology and infraciliature of a new member, Pseudoamphisiella quadrinucleata n. sp., a form with conspicuous alveolar layer and four macronuclear nodules isolated from the coastal waters both near Hong Kong and near Guangzhou, South China were investigated using living observation and protargol silver impregnation methods. Pseudoamphisiella quadrinucleata differs from other two known forms mainly by the number of macronuclear nodules: constantly four vs. two in Pseudoamphisiella alveolata and 24–57 in Pseudoamphisiella lacazei. To support this, the sequence of the small subunit rDNA of P. quadrinucleata n. sp. showed 14 and 74 nucleotides in comparison with that of the two known congeners, respectively, which hence firmly supports the validity of the new species.     

Three new entocommensal ciliates from digestive tract of sea urchins of the Weddell Sea,Antarctica (Protozoa,Ciliophora)

Three new entocommensal ciliates (Spirostrombidium echini sp. nov., Entodiscus pseudoechini sp. nov. and Cryptochilum antarcticum sp. nov.) from the digestive tract of sea urchins in Antarctica (Weddell Sea) were studied using protargol and silver nitrate impregnation methods. Based on our reinvestigations, improved diagnoses for the genera Entodiscus and Cryptochilum are provided. According to the general morphology, E. pseudoechini is recognized by a higher number of ciliary rows, the absence of a caudal cilium which is typically present for other congeners, and shortened somatic kineties direct to the ventral surface, of which the last somatic kinety does not pass through the caudal pole as in most other scuticociliates. C. antarcticum differs from the known congeners in its particularly high number of somatic kineties. S. echini is distinguished by the combination of habitat, body shape, structure of buccal apparatus and arrangement of somatic kineties. Accepted: 29 March 1999     

Morphology and morphometry of Bresslauides pratensis n. sp., a new soil ciliate (Ciliophora, Colpodea) from Turkey

A new colpodid ciliate, Bresslauides pratensis n. sp., was discovered in soil from a meadow field in Turkey. Its morphology was investigated using live observation, protargol impregnation, silver nitrate impregnation, and scanning electron microscopy. The new species differs from the congeners by the following combination of features: the cell size in vivo is about 135-180 μm × 130-160 μm; the body has an almost circular outline with a conspicuous bulge on the ventral side; the somatic cilia are arranged in about 105 densely spaced, sigmoidal kineties; a diagonal groove extends to the left cell side, terminating in a pronounced postoral sack; one almost globular macronucleus and usually two globular micronuclei; the oral structures occupy almost the anterior half of the cell and comprise a distally elongated right polykinetid and a crescentic left oral polykinetid, both restricted to the vestibulum; on average 12 vestibular, 31 postoral, and 55 left oral kineties. Based on the morphological data, it was concluded that B. pratensis is a well-outlined and distinctive member of the genus Bresslauides. Additionally, the most recent molecular data on the order Colpodida and genera Colpoda, Bresslaua, and Bresslauides are briefly discussed.