Phylogenetic relationships within the class Spirotrichea (Ciliophora) inferred from small subunit rRNA gene sequences.

The small subunit rDNAs of five species belonging to the Euplotidae and eight species of the Oxytrichidae were sequenced to obtain a more detailed picture of the phylogenetic relationships within the Spirotrichea (Ciliophora). Various tree reconstruction algorhythms yielded nearly identical topologies. All Euplotidae were separated from the other Spirotrichea by a deep split. Further, a large genetic distance between the marine genus Moneuplotes and the freshwater species of Euplotoides was found. Differences between the methods used occurred only within the Oxytrichidae. Whereas the monophyly of the Stylonychinae was supported in all trees, the monophyly of the Oxytrichinae was not. However, the molecular data support the morphological and ontogenetic evidence that the pattern of 18 frontal-ventral-transversal cirri evolved in the stemline of the Oxytrichidae and was modified several times independently. Our results are also in agreement with taxonomic revisions: the separation of both Sterkiella nova from Oxytricha and Tetmemena pustulata from Stylonychia.

Phylogenetic relationships of orders within the class colpodea (Phylum ciliophora) inferred from small subunit rRNA gene sequences

Molecular analyses have been used recently to refine our knowledge of phylogenetic relationships within the ciliated protozoa (phylum Ciliophora). A current Hennigian phylogeny of the orders in the class Colpodea, based on light and electron microscopic analyses, makes three important assumptions with regard to apomorphic character states, namely, (1) that the kreyellid silver line evolved early in colpodean phylogeny, separating bryometopids, such as Bryometopus, from all other colpodeans; (2) that the macro-micronuclear complex is an autapomorphy of the cyrtolophosidids, such as Platyophrya; and (3) that merotelokinetal stomatogenesis is an apomorphic character of colpodids, such as Colpoda, Bresslaua, and Pseudoplatyophrya. These predictions of relationships within the class Colpodea were investigated by determining the complete small subunit rRNA gene sequences for the colpodid Bresslaua vorax, the grossglockneriid Pseudoplatyophrya nana, and the cyrtolophosidid Platyophrya vorax and a partial sequence for the bryometopid Bryometopus sphagni. These sequences were combined with the previously published complete SSrRNA sequences for the colpodid Colpoda inflata and the bursariomorphid Bursaria truncatella. The affiliations were assessed using both distance matrix and maximum-parsimony analyses. The tree topologies for the class Colpodea were identical in all analyses, with bootstrap support for bifurcations always exceeding 60%. The results suggest the following. (1) Since the clade including Bryometopus and its sister taxon, Bursaria, is never basal, the kreyellid silver-line system evolved later in colpodean phylogeny and does not separate bryometopids from all other colpodeans. (2) Since Platyophrya is always the sister taxon to the other five genera, there is a fundamental phylogenetic significance for its macro-micronuclear complex. (3) Since the colpodids, Colpoda, Bresslaua, and Pseudoplatyophrya, always group in one clade, merotelokinetal stomatogenesis appears to be a derived character state.

Ciliates in rapid gravity filters of waterworks exploiting deep groundwaters.

Potable water is increasingly produced from deep (>100 m) tertiary groundwaters which often are completely reduced and contain high amounts of ammonium, methane, and hydrogen sulphide. They thus require special treatment which includes oxygenation and removal of the reduced contaminants by the biofilm developing in rapid gravity filters. The biofilm is heavily colonized with ciliates and microinvertebrates. A total of 38 species of ciliates was found in 42 samples taken from 4 waterworks in Germany during a period of 2 years. Only six species occurred in high numbers and in more than half of the samples: Acineria uncinata, Aspidisca lynceus, Cinetochilum margaritaceum, Colpidium colpoda, Glaucoma scintillans, and Holosticha pullaster. Five to thirteen species occurred per sample, and up to 6,000 individuals ml-1 biofilm were counted. There was a considerable fluctuation in the number of species and individuals, which could not be related to specific process parameters. The number of species and individuals decreased markedly from the filter surface to its centre. Colonization of the filters very likely occurs randomly via impure surface waters. The ciliate communities found consist mainly of alphamesosaprobic to polysaprobic species and thus closely resemble those known from activated-sludge processes. This is explained by the specific conditions near and in the biofilm, which is probably microaerobic and highly productive, providing microaerobic bacterial feeders with copious food. Obviously, it is the microenvironment which determines the occurrence of certain species. Thus, future research on the autecology of the indicator species used in the saprobic system should concentrate on their microenvironments.

Morphogenesis in some freshwater Tintinnids (Ciliophora, Oligotrichida).

The morphogenesis of four freshwater tintinnids was investigated using protargol silver impregnation and scanning electron microscopy. Division is rather similar in Tintinnopsis cylindrata, Tintinnidium pusillum and T. semiciliatum, e.g. the oral primordium develops apokinetally posterior of somatic kinety 10 and the somatic ciliature originates by two rounds of intrakinetal basal body proliferations before cytokinesis. The peculiar ventral organelles form without apparent contact with parental ciliary structures as do the adorai membranelles and the paroral membrane; thus, the ventral organelles might be part of the oral apparatus. The morphogenesis of Codonella cratera differs from that of the other species by the reorganization of some parental ciliary rows and by a second round of somatic basal body proliferation after cytokinesis. Based on morphologic and morphogenetic similarities, Tps. cylindrata is transferred from the Codonellidae to the Tintinnidiidae, and Tps. baltica and Tps. subacuta are newly combined with Codonella: C. baltica nov. comb., C. subacuta nov. comb. We could not discern a unique character defining oligotrichs as a monophyletic group because the enantiotropic cell division is possibly less pronounced in some tintinnids and occurs also in peritrichs and a few prostomatids. We suggest, however, that the enantiotropic cell division evolved convergently in these taxa, and thus adhere to our view that this special mode of cell division is the most reliable apomorphy for the oligotrichs. In addition, the character combination, polar oral apparatus and apokinetal origin of the oral primordium, occurs only in oligotrichs. Morphogenetic similarities suggest a sister group relationship between tintinnids and strobilidiids.

Stomatogenesis in the ditransversal ciliate Homalozoon vermiculare (Ciliophora, Rhabdophora).

H. vermiculare possesses about 10 somatic kineties on the right lateral side, 3 somatic kineties on the left lateral side and a single circumoral kinety. The somatic kineties are composed of monokinetids exc\~ept for the anterior ends of the brush kineties which are composed of dikinetids. The circumoral kinety consists of paired kinetosomes one of which is nonciliated and associated with a microtubular lamella and a nematodesma. Stomatogenesis commences when the anteriormost somatic kinetosomes in the opisthe are transformed into the nonciliated kinetosomes of the future oral dikinetid. They lose the somatic infraciliary fibers and the ciliary shaft and each gives rise to a nematodesma and a microtubular ribbon. Adjacent to each of the transformed somatic kinetosomes, a new kinetosome is assembled, thus producing an oral dikinetid anlage. The new anterior kinetosome bears a cilium and becomes the ciliferous kinetosome of the oral wreath of cilia. In protargol stained specimens, proliferation of kinetosomes can first be observed in the left lateral kineties but, eventually, each of the somatic kineties produces one kinetofragment. Thus, H. vermiculare has a holotelokinetal type of stomatogenesis. The cirumoral kinety arises from a counter-clockwise rotation (as viewed from outside the cell) of all 15 dikinetid kinetofragments and subsequent "head to tail" fusion of the fragments after cell division has been completed. The oral apparatus of the proter seems to be largely conserved during division. Some aspects of the evolution of the oral apparatus and the origin of the oral microtubular ribbons are discussed.

Divisional morphogenesis in Bakuella pampinaria nov. spec, and reevaluation of the classification of the urostylids (ciliophora, hypotrichida).

A new hypotrichous ciliare, Bakuella pampinaria nov. spec, colonizing vineleaf and pear-tree litter, is described. Bakuella pampinaria differs from the other species of the genus by having distinct rows of yellowish cortical granules. Several morphogenetic differences separate Bakuella pampinaria from Bakuella edaphoni and other congeners, especially in that the transverse cirri do not participate in the formation of the oral primordium. The type population of Pseudourostyla cristata was reinvestigated. Two frontoterminal cirri are recognizable in dividing specimens indicating that this genus is valid, i.e. different from Urostyla, which very likely lacks such cirri. The urostyline hypotrichs are recognized as a monophyletic taxon by two apomorphies, viz. the midventral cirri and the partial or complete reorganization of the proter's adorai zone of membranelies during cell division. Phylogeny and evolution within the urostylids are much less clear since character states (apomorphies, plesiomorphies, convergencies) are uncertain and morphogenetic data are still too sparse or inaccurate. This is exemplified on a selected set of genera using Hennig's phylogenetic method.

Basic light and scanning electron microscopic methods for taxonomic studies of ciliated protozoa.

The following methods for taxonomic studies of ciliated protozoa are described in detail: live observation, supravital staining with methyl green-pyronin, dry silver nitrate impregnation, wet silver nitrate impregnation, silver carbonate impregnation, protargol impregnation (three procedures), and scanning electron microscopy. Familiarity with these methods (or modifications) is an absolute prerequisite for successful taxonomic work. No staining method is equally appropriate to all kinds of ciliates. A table is provided which indicates those procedures which work best for certain groups of ciliates. A second table relates to the structures revealed by the procedures. Good descriptions usually demand at least live observation, silver nitrate and protargol or silver carbonate impregnation. Some instructions are provided for distinguishing mono- and dikinetids as well as ciliated and non-ciliated basal bodies in silvered ciliates. The brilliancy of the silver preparations has unfortunately recently tempted some taxonomists to neglect live observation. However, many important species characters cannot be seen or are changed in silvered specimens. I thus consider all species descriptions based exclusively on silver slides as incomplete and of doubtful value for both α-taxonomists and ecologists. Especially the latter are usually not trained to correlate the silvered structures with the live appearance of the cell.

Ultrastructure of the mycophagous ciliate Grossglockneria acuta (Ciliophora, Colpodea) and phylogenetic affinities of colpodid ciliates.

Ultrastructural features of Grossglockneria acutaFoissner, 1980 are very similar to those known in the confamilial species, Pseudoplatyophrya nana. The somatic infraciliature corresponds to the typical colpodid dikinetid pattern. The peculiar oral apparatus is described in detail. The feeding tube contains about 17 concentric cytostomial microtubular lamellae and postciliary microtubules. The lamellae are interconnected by dense plaques and their innermost microtubules anchor in dense material at the distal end of the tube. The postciliary ribbons of the paroral kinetosomes extend subpellicularly into the proximal region of the tube. It is supposed that supernumerary postciliary microtubules are proliferated and assemble to the cytostomial microtubular lamellae during stomatogenesis. Few transverse microtubules are associated to some adorai and presumably all paroral kinetosomes; they do not extend into the tube. Most basal bodies of the single adorai organelle lack kinetosomal derivates. The Grossglocknerida and Colpodida are suggested to be sister groups because they undergo a telokinetal stomatogenesis in reproductive cysts. The colpodid kinetid is characterized by a compound microtubular structure of posterior transverse ribbons to the left of the kinety. Several hypotheses as to the origin of the typical colpodid kinetid pattern, especially the posteriorly extending transverse microtubules, are discussed. Available evidence suggests haptorid or nassulid ancestors of the colpodids. Some data indicate a paraphyletic composition of the class.

Morphogenesis and ultrastructure of the soil ciliate Engelmanniella mobilis (Ciliophora, Hypotrichida).

The morphogenetic pattern, freeze-fracture, transmission, and scanning electron-microscopy of interphasic cells were used to elucidate the enigmatic systematic position of Engelmanniella mobilis. Frontal, buccal, parabuccal, and marginal cirri are distinguishable during cortical development; transverse, frontoterminal (migratory), and caudal cirri are absent. Ontogenetic peculiarities include the conservation of parental and grandparental marginal cirri and the apokinetal origin of a dorsal kinety in the opisthe. Thus, interphasic cells of E. mobilis possess three generations of cirri. The pellicle is multilamellated. Prominent, undischargeable subpellicular granules consisting of a homogeneous osmiophilic mass develop from bundles of long fibers. Rhomboid crystal-like structures projecting from the lithosomes and regularly patterned mitoribosomes are additional remarkable ultrastructural characteristics. The somatic and frontal cirri comprise 2-10 kinetosomes. Very likely, all cirri are composed of closely adjacent "dikinetids" because their kinetosomes form pairs each possessing a transverse and a postciliary microtubular ribbon. The fourth row of kinetosomes of the adorai membranelies shows the usual transverse microtubular ribbons, while one or two transverse microtubules are closely adjacent to the third and the second row of its basal bodies. Our data suggest that Engelmanniella has descended from a kahliellid lineage. However, its placement in a well-founded (!) stichotrichid family is impossible at present.

The hemimastigophora (Hemimastix amphikineta nov. gen., nov. spec.), a new protistan phylum from gondwanian soils.

The morphology, morphogenesis and ultrastructure of Hemimastix amphikineta nov. gen., nov. spec, are described. This species occurred in some Australian and in 1 Chilean soil, but was absent from more than 1000 soil samples from Laurasian localities. Thus, it has probably a restricted Gondwanian distribution. Hemimastix amphikineta is a small (14-20 × 7-10 µn), colourless organism that looks distinctly Ciliophora-like because of its posteriorly located contractile vacuole and its 2 longitudinal somatic kineties each composed of about 12 cilia-like flagella. These 2 kineties are interposed between 2 large plicated and microtubule-bearing pellicular plates which are arranged inversely mirror-image like ("diagonal symmetry"). Hemimastix amphikineta has saccular to tubular mitochondrial cristae and complex extrusomes. It has 2 microtubular systems and a membranous sac associated with each kinetid. The nucleolus persists throughout nuclear division. A permanent cytostome-cytopharyngeal complex, pharyngeal rods, striated fibres, mastigonemes, and a paraflagellar rod are absent. This unique combination of characters dictates a very separate position for H. amphikineta within the known protists. Thus, the phylum Hemimastigophora nov. phylum (Hemimastigea nov. cl. and Hemimastigida nov. ord.), is established to include H. amphikineta and possibly Spironema multiciliatum Klebs, 1892. The structure of the pellicle and the nuclear apparatus of H. amphikineta indicate some relationship with the Euglenophyta. However, clear evidence for a certain affinity is lacking. Thus, the Hemimastigophora are placed in an incertae sedis position within the kingdom Protista Haeckel, 1866.

r-and K-selection in soil ciliates: a field and experimental approach.

Previously published field studies have suggested that the concept of r/K selection may be applied to terrestrial ciliates. A census was taken of the dominant groups, the Colpodea and Polyhymenophora, to determine species composition and absolute abundances in climatically unpredictable alpine and predictable lowland sites. In addition, two typical representatives of each of the two taxa (Colpoda aspera and Grossglockneria acuta, and Blepharisma undulans and Gonostomum affine, respectively) were selected for examination under laboratory conditions. Variables investigated were: the C/P index (ratio of Colpodea to Polyhymenophora), fertility, tolerance of altered environmental conditions, incidence in predictable or unpredictable biotopes, opportunism, competitive ability, and body size. Our own observations were supplemented by reference to the literature.The field census produced a higher C/P index in the unpredictable sites than in the predictable ones. The experiments show that C. aspera and G. acuta are less sensitive to changes in temperature and exhibit a more rapid rise in individual density as well as a higher number of individuals than do B. undulans and G. affine, although the former species have longer generation times at low temperatures. The explosive increase in number of individuals can be ascribed to the special reproductive strategy of the Colpodea (quadripartition in cysts). These results imply that the Colpodea are more r-selected as compared with the Polyhymenophora. That might be an explanation for their wide distribution and high density in unfavourable terrestrial habitats.