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Figure 8 in Re-discovery and novel contributions to morphology and multigene phylogeny of Myxophyllum steenstrupi (Ciliophora: Pleuronematida), an obligate symbiont of terrestrial pulmonates

Figure 8. Phylogenetic tree based on the nuclear 18S rRNA gene as well as on the mitochondrial COI and 16S rRNA gene sequences, showing the systematic position of M. steenstrupi isolated from the plaited door snail Cochlodina laminata. The class Colpodea was used as the outgroup. Bootstrap values for maximum likelihood conducted in IQTrees as well as posterior probabilities for Bayesian inferences conducted in MrBayes and Phycas were mapped onto the best scoring IQ tree. Dash indicates mismatch between maximum likelihood and Bayesian tree topologies. Sequences marked in red were obtained during this study. Fully statistically supported nodes are marked with red solid circles. GenBank accession numbers can be found in Supporting Information (Table S6). The scale bar denotes six substitutions per one hundred nucleotide positions.

Figure 6 in Re-discovery and novel contributions to morphology and multigene phylogeny of Myxophyllum steenstrupi (Ciliophora: Pleuronematida), an obligate symbiont of terrestrial pulmonates

Figure 6. Phylogenetic tree based on the nuclear 18S, 5.8S, and 28S rRNA gene sequences, showing the systematic position of M. steenstrupi isolated from the plaited door snail Cochlodina laminata. The subclass Scuticociliatia is represented by the orders Philasterida and Pleuronematida, the subclass Astomatia was used as the outgroup. Bootstrap values for maximum likelihood conducted in IQTrees as well as posterior probabilities for Bayesian inferences conducted in MrBayes and Phycas were mapped onto the best scoring IQ tree. Dash indicates mismatch between maximum likelihood and Bayesian tree topologies. Sequences marked in red were obtained during this study. Fully statistically supported nodes are marked with red solid circles. GenBank accession numbers can be found in Supporting Information (Table S2). The scale bar denotes seven substitutions per one hundred nucleotide positions.

Figure 2. M. steenstrupi after protargol impregnation. A, B in Re-discovery and novel contributions to morphology and multigene phylogeny of Myxophyllum steenstrupi (Ciliophora: Pleuronematida), an obligate symbiont of terrestrial pulmonates

Figure 2. M. steenstrupi after protargol impregnation. A, B, top and bottom overviews of the same specimen, showing the ciliary pattern and nuclear apparatus. Black arrowheads mark the anterior suture, white double arrowheads denote the right lateral suture, white arrowheads denote the left lateral suture, and arrows mark the posterior suture. C, detail of the silverline system in the anterior body portion. D, semischematic diagram, showing the ciliary pattern within and around the mouth pocket. MA, macronuclear nodules; MI, micronucleus; PM, paroral membrane; SK, somatic kineties; SL, silverline; VK, vestibular kineties. Scale bars = 20 Μm (C), 55 Μm (A, B).

Figure 4. M in Re-discovery and novel contributions to morphology and multigene phylogeny of Myxophyllum steenstrupi (Ciliophora: Pleuronematida), an obligate symbiont of terrestrial pulmonates

Figure 4. M. steenstrupi after protargol impregnation (A–D, H) and after silver nitrate impregnation (E–G). A, detail of the anterolateral body region, showing the anterior suture (black arrowhead) made by abutting anterior ends of ventral kineties and thigmotactic kineties. B, detail of the posterolateral body region, showing the posterior suture. C, frontal view, showing the entrance to the mouth pocket (arrow). Note that somatic kineties enter the mouth pocket to become vestibular kineties. D, optical section through the mouth pocket, showing the vestibular kineties, adoral organelle and paroral membrane. E–G, details of the cell surface, showing the somatic ciliary pattern and the silverline system. Black arrowheads mark the anterior suture, opposed white arrowheads denote the blank stripe within the right lateral suture, red arrows show the dikinetids of the thigmotactic field, and red arrowheads mark the horizontal silverlines. H, detail of the anterior body region, showing the thigmotactic field and the blank stripe within the right lateral suture (opposed white arrowheads). AO, adoral organelle; F, fibres, MP, mouth pockets; PM, paroral membrane; VK, vestibular kineties; SK, somatic kineties. Scale bars = 5 Μm (G), 10 Μm (B), 15 Μm (H), 20 Μm (A, C, E).

Figure 3. M in Re-discovery and novel contributions to morphology and multigene phylogeny of Myxophyllum steenstrupi (Ciliophora: Pleuronematida), an obligate symbiont of terrestrial pulmonates

Figure 3. M. steenstrupi in vivo (A–D) and after protargol impregnation (E, F). A, B, ventral overviews of two representative individuals, showing the nuclear apparatus and the contractile vacuole. Arrow in (A) indicates the entrance to the mouth pocket. C, D, details of the posterior body region, showing the entrance to the mouth pocket (arrow) containing vestibular kineties, the nuclear and contractile vacuole apparatus as well as the posterior suture (opposed white arrowheads). E, top overview, showing the nuclear apparatus and the ciliary pattern. Black arrowheads mark the anterior suture, the white arrowhead denotes the left lateral suture. F, detail of nuclear apparatus of the specimen shown in (E). CV, contractile vacuole; MA, macronuclear nodules; MI, micronucleus; MP, mouth pocket; SK, somatic kineties; TF, thigmotactic field; VK, vestibular kineties. Scale bars = 15 Μm (C), 25 Μm (D), 30 Μm (F), 55 Μm (E), 60 Μm (A, B).

Figure 7 in Taxonomy and phylogeny of three heterotrich ciliates (Protozoa, Ciliophora), with description of a new Blepharisma species

Figure 7. Bayesian inference (BI) tree inferred from small subunit (SSU) rDNA sequences showing the positions of Anigsteinia clarissima, Blepharisma penardi sp. nov., and Blepharisma undulans (all three taxa in bold). Members of the class Karyorelictea are the outgroup taxa. Numbers near branches show the posterior probabilities of BI and bootstrap values from maximum likelihood (ML). Black circles indicate full support (1.00 BI, 100% ML) in both analyses. Disagreements between BI and ML are shown by asterisks. All branches are drawn to scale. The scale bar corresponds to five substitutions per 100 nucleotide positions.

Figure 3 in Taxonomy and phylogeny of three heterotrich ciliates (Protozoa, Ciliophora), with description of a new Blepharisma species

Figure 3. Photomicrographs of Anigsteinia clarissima after protargol staining. A, right view of a typical individual. B–D, anterior of cell (right view), arrows show paroral membrane. E, F, left (E) and right (F) views of anterior portion. G, portion of infraciliature, arrows mark postoral kineties. H, posterior end of oral field and the moniliform macronucleus. Scale bars = 140 μm (A), 30 μm (B–H).

Figure 6 in Taxonomy and phylogeny of three heterotrich ciliates (Protozoa, Ciliophora), with description of a new Blepharisma species

Figure 6. Nominal isolates of Blepharisma penardi sp. nov. (A, C, D), Blepharisma steini (B), and Blepharisma undulans (E–K). A, C, D, B. penardi sp. nov., originally described as an unnamed variety of Blepharisma lateritium, from Penard (1922). B, B. steini, from Kahl (1932). E, B. undulans, from Bhandary (1962). F, B. undulans, from Lee & Shin (2009). G, B. undulans, from Kahl (1932). H, B. undulans, from Cela (1972). I, B. undulans, from Suzuki (1954). J, K, B. undulans, from Dragesco & Dragesco-Kernéis (1991). Abbreviation: Ma, macronuclear nodules. Scale bars = 90 μm (B, I), 80 μm (G, F) 50 μm (C, E, H), 40 μm (J, K).

Figure 5 in Taxonomy and phylogeny of three heterotrich ciliates (Protozoa, Ciliophora), with description of a new Blepharisma species

Figure 5. Different isolates under the names of Anigsteinia clarissima (A–N, Q), Anigsteinia candida (O), and Anigsteinia oligonucleata (P). A, from Yagiu (1943). B, from Anigstein (1912). C, from Dragesco (1960). D, reported as Blepharisma clarissimum forma arenicola by Kahl (1932). E, from Kahl (1932). F, from Kahl (1932), after Anigstein (1912). G, from Ricci et al. (1982). H, from Kahl (1928). I, from Bock (1952). J, from Raikov (1960). K, L, from Agamaliev (1968). M, N, from Czapik & Jordan (1976). O, A. candida, reported as A. clarissima (misidentification) by Dragesco (2002). P, A. oligonucleata, reported as A. clarissima (misidentification) by Dragesco (1966). Q, from Dragesco & Dragesco-Kernéis (1986). Abbreviation: Ma, macronuclear nodules. Scale bars = 375 μm (O), 200 μm (E, I, M, N), 180 μm (H), 150 μm (D, G, J), 110 μm (F, Q), 80 μm (B), 60 μm (C, K, L, P), 40 μm (A).

Figure 4 in Taxonomy and phylogeny of three heterotrich ciliates (Protozoa, Ciliophora), with description of a new Blepharisma species

Figure 4. Photomicrographs of Blepharisma penardi sp. nov. from life (A–E, I) and after protargol staining (F–H), and Blepharisma undulans from life (J, M) and after protargol staining (K, L). A, C, D, left view, to show the shape variants, arrows in C, D mark the contractile vacuole. Note the cell colour: colourless (A) and dark-brownish (C). B, cytoplasmic granules. E, portion of cortex, showing the tiny, densely arranged cortical granules. F, G, general infraciliature. H, portion of buccal field, arrows point to paroral membrane. I, macronucleus. J, left view of a typical individual. K, right view, to show the two macronuclei. L, posterior portion of oral field, arrows indicate paroral membrane. M, to show cortical granules, note there are some white granules (arrows) amongst the coloured ones. Abbreviations: CV, contractile vacuole; Ma, macronucleus. Scale bars = 70 μm (A, C, D, F, G, J, K), 30 μm (H), 15 μm (B, E, I, L, M).

Figure 2 in Taxonomy and phylogeny of three heterotrich ciliates (Protozoa, Ciliophora), with description of a new Blepharisma species

Figure 2. Photomicrographs of Anigsteinia clarissima from life. A–D, general views of typical individuals, arrows in B show lacunar contractile vacuole system, arrow in C indicates the paroral membrane. E, anterior region of cell, arrows mark the colourless cortical granules. F, posterior portion of buccal field, arrows show paroral membrane. G, macronuclear nodules. H, portion of cortex (right view), showing cortical granules between somatic kineties. Abbreviation: Ma, macronucleus. Scale bars = 120 μm (A–D), 25 μm (E–H).

Figure 1 in Taxonomy and phylogeny of three heterotrich ciliates (Protozoa, Ciliophora), with description of a new Blepharisma species

Figure 1. Morphology and infraciliature of Anigsteinia clarissima (A–D), Blepharisma penardi sp. nov. (E, F, K, L), and Blepharisma undulans (G–J). A, left view of a typical individual of A. clarissima. B, paroral membrane of A. clarissima, anterior (double arrowheads) and posterior (arrow) parts are formed of dikinetids with only left basal body ciliated, middle part (arrowhead) formed by trikinetid fragments with all basal bodies ciliated. C, D, general infraciliature of A. clarissima, arrows show paroral membrane. E, left view of a typical individual of B. penardi sp. nov. F, L, dorsal (F) and ventral (L) views of infraciliature of B. penardi sp. nov. from the same specimen, arrow marks paroral membrane. G, left view of a typical individual of B. undulans. H, paroral membrane of B. undulans, anterior part with only right basal bodies ciliated and posterior part with both basal bodies ciliated. I, J, right (I) and left (J) views of B. undulans showing the infraciliature, arrow points to paroral membrane. K, paroral membrane of B. penardi sp. nov., anterior part consists of ciliated monokinetids, posterior part consists of dikinetids with only left basal body ciliated. Abbreviations: AZM, adoral zone of membranelles; CV, contractile vacuole; LCV, lacunar contractile vacuole system; SK, somatic kineties. Scale bars = 100 μm (A, C, D), 70 μm (E, G, I, J, L), 50 μm (B, H, K).

Figure 7 in Re-discovery and novel contributions to morphology and multigene phylogeny of Myxophyllum steenstrupi (Ciliophora: Pleuronematida), an obligate symbiont of terrestrial pulmonates

Figure 7. Phylogenetic tree based on the mitochondrial COI nucleotide sequences, showing the systematic position of M. steenstrupi isolated from the plaited door snail Cochlodina laminata. The class Colpodea was used as the outgroup. Bootstrap values for maximum likelihood conducted in IQTrees as well as posterior probabilities for Bayesian inferences conducted in MrBayes and Phycas were mapped onto the best scoring IQ tree. Dash indicates mismatch between maximum likelihood and Bayesian tree topologies. Sequences marked in red were obtained during this study. Fully statistically supported nodes are marked with red solid circles. GenBank accession numbers can be found in Supporting Information (Table S3). The scale bar denotes twenty substitutions per one hundred nucleotide positions.

Figure 6 in Morphology, morphogenesis, and molecular phylogeny of a new marine urostylid ciliate (Ciliophora, Stichotrichia) from the South China Sea, and a brief overview of the convergent evolution of the midventral pattern within the Spirotrichea

Figure 6. The maximum-likelihood (ML) tree inferred from the small subunit ribosomal RNA (SSU rRNA) gene sequences of 63 spirotrichous taxa, showing the position of Bergeriella ovata gen. et sp. nov. (boxed), and the phylogenetic relationships among the taxa possessing midventral cirral rows (i.e. urostylids s.l.; branches are depicted by thick lines, and species names are highlighted in bold text). Nodal support for branches in the ML, Bayesian inference (BI), and neighbour-joining (NJ) trees are marked in order. Bootstrap values lower than 50% and Bayesian posterior probabilities lower than 0.70 are replaced with hyphens. Clades with different topologies in the NJ tree relative to the ML and BI trees are indicated with asterisks. All branches are drawn to scale. The scale bar corresponds to five substitutions per 100 nucleotide positions. Phacodinium and Protocruzia were taken as out-group taxa.

Figure 3 in Morphology, morphogenesis, and molecular phylogeny of a new marine urostylid ciliate (Ciliophora, Stichotrichia) from the South China Sea, and a brief overview of the convergent evolution of the midventral pattern within the Spirotrichea

Figure 3. Ventral (A, C, E, G) and dorsal (B, D, F, H) views of Bergeriella ovata gen. et sp. nov. in morphogenesis (A–D) and regeneration (E–H), after impregnation with protargol. A, an early divider, showing the oral primordium (arrow) and fronto-ventral-transverse (FVT) anlagen (arrowhead) of the proter. The double arrowheads mark the anlagen for the nonmigratory row, which comes from the posteriormost FVT streak. B, the same specimen as shown in (A), showing the enlarged macronuclear nodules and the formation of the dorsal kinety anlagen (arrows). C, an individual at a late stage of division, with all cirri developed; note the structures that will respectively form the enlarged postoral ventral cirri (arrowheads) and the delicate left ventral cirri (arrows). The double arrowheads indicate the anlagen for the nonmigratory row. D, the differentiating marginal row (arrows) and dorsal kineties; note that the macronuclear nodules are separating. E, F, an early reorganizer, showing the oral primordium (arrow in E), FVT streak (arrowhead), nonmigratory row (double arrowheads) and dorsal kinety anlagen; the arrows in (F) indicate the anlagen for the right and left marginal rows, which are derived within the parental structure. G, H, a middle-stage reorganizer, with a further proliferation of kinetosomes, showing the first frontal cirrus (arrow in G) generated from the undulating membrane anlagen, the basal bodies developed from FVT streaks (arrowhead), the anlagen for the nonmigratory row (double arrowheads), and the anlagen for the right and left marginal rows (arrows in H). Abbreviations: DK, dorsal kineties; DKA, dorsal kinety anlagen. Scale bars: 40 Mm.

Figure 2 in Morphology, morphogenesis, and molecular phylogeny of a new marine urostylid ciliate (Ciliophora, Stichotrichia) from the South China Sea, and a brief overview of the convergent evolution of the midventral pattern within the Spirotrichea

Figure 2. Photomicrographs of Bergeriella ovata gen. et sp. nov. from life. A, ventral view of a specimen. B, C, ventral view of slender and fat forms; the arrow indicates the wide and bright oral field. D, lateral view. E, F, lateral (E) and dorsal (F) views, showing the distribution of the granules; the arrows mark the granule rows near the base of each marginal cirrus, the arrowheads point to the granule rows along with midventral rows, and the double arrowheads mark the granule bands in the gap between the somatic kineties. G, H, showing the cortical granules (arrows), the fibres associated with cirri (arrowheads), and a dorsal cilium (double arrowheads). I, focusing on the oral field; note the paroral membrane (arrowhead) and the endoral membrane (arrow). J, the cortical granules (arrow) near the base of marginal cirri. K, lateral view of the posterior portion; arrows point to the enlarged postoral ventral cirri. L, globular lipid droplets in the cytoplasm. Scale bars: 50 Mm.

Figure 1 in Morphology, morphogenesis, and molecular phylogeny of a new marine urostylid ciliate (Ciliophora, Stichotrichia) from the South China Sea, and a brief overview of the convergent evolution of the midventral pattern within the Spirotrichea

Figure 1. Bergeriella ovata gen. et sp. nov. drawn from life (A–C, E–F) and after impregnation with protargol (D, G–J). A, ventral view of a specimen. B, different body shapes. C, section of the ventral infraciliature, showing the fibres associated with the postoral ventral cirri (double arrowheads) and the obliquely arranged left ventral cirri (arrow). D, distribution of cortical granules (arrow) near the marginal cirri. E, F, distribution of the cortical granules on the ventral (E) and dorsal (F) sides; the arrow indicates the granules along the nonmigratory row, and the arrowheads point to the granular rows along the dorsal kineties. G, left lateral side view of the infraciliature. H–I, ventral (H) and dorsal (I) views of the infraciliature; note the enlarged postoral ventral cirri (dashed lines), the frontal cirri (dashed lines), and the three dorsal kineties (arrows). J, ventral view of an early divider; the arrow indicates the oral primordium of the proter, the arrowheads mark the old endoral membranes in dedifferentiation, and the double arrowhead points to the oral primordium of the opisthe. Abbreviations: AZM, adoral zone of membranelles; BC, buccal cirri; DK, dorsal kineties; EM, endoral membrane; FC, frontal cirri; LMR, left marginal row; LVR, left ventral rows; MVR, midventral rows; NMR, nonmigratory row; PM, paroral membrane; PVR, postoral ventral rows; RMR, right marginal row. Scale bars: 40 Mm (A–C, F–J); 15 Mm (E).

Figure 5 in Morphology, morphogenesis, and molecular phylogeny of a new marine urostylid ciliate (Ciliophora, Stichotrichia) from the South China Sea, and a brief overview of the convergent evolution of the midventral pattern within the Spirotrichea

Figure 5. Photomicrographs of regeneration in Bergeriella ovata gen. et sp. nov. after impregnation with protargol. A, B, ventral views of middle reorganizers, showing the oral primordium (arrow in A), fronto-ventral-transverse (FVT) anlagen (arrowheads), and the anlagen for the nonmigratory row (double arrowhead); the arrow in (B) points to the undulating membrane anlagen. C, a middle-stage reorganizer; the arrow marks the first frontal cirrus generated from the undulating membrane anlagen, and the arrowheads indicate the anlagen for the nonmigratory row. D, dorsal view of the same specimen, showing the anlagen for the left marginal row (arrow) and the dorsal kinety anlagen (arrowheads).

Figure 4 in Morphology, morphogenesis, and molecular phylogeny of a new marine urostylid ciliate (Ciliophora, Stichotrichia) from the South China Sea, and a brief overview of the convergent evolution of the midventral pattern within the Spirotrichea

Figure 4. Photomicrographs of Bergeriella ovata gen. et sp. nov. after impregnation with protargol. A, infraciliature of the ventral posterior portion, showing the nonmigratory row (arrow), the postoral (arrowheads), and the left ventral cirri (double arrowheads). B, infraciliature of an anterior portion, showing the frontal cirri (arrows), buccal cirri (arrowhead), and the undulating membranes (double arrowheads). C, left lateral view, showing the left ventral rows, and the left marginal row (arrow). D, dorsal kineties, in which kinetosomes become more densely spaced from left to right (arrows). E, dorsal view, showing the anterior portion of the nonmigratory row (arrowhead) and the right marginal row (arrow). F, nuclear apparatus, some with replication bands can be seen. G, H, ventral and dorsal views of an early stage divider, showing the undulating membrane anlagen (arrow in G), FVT anlagen (arrowheads), and the anlagen for the nonmigratory row, which come from the posteriormost FVT streak (double arrowheads) of the opisthe. The arrows in (H) indicate the dorsal kineties anlagen. I, early divider showing the appearance of the oral primordium (arrow) and the enlarged macronuclear nodules of the proter. J, separating ellipsoid macronuclear nodules. K, L, ventral views of a late-stage divider (same specimen), showing the developed cirri of the proter (K) and the opisthe (L); the arrows indicate the migrating postoral and left ventral cirri, and the arrowheads indicate the anlagen for the nonmigratory row. M, dorsal view of the same specimen shown in (K) and (L), showing the anlagen of the marginal rows (arrows) and the new dorsal kineties.