Morphological distance research on over 50 species of ciliates recorded as endosymbionts of echinoids suggests that invasion of the echinoid microhabitat occurred It on at least 4 occasions. Gene sequence data confirm the phylogenetic distinctness of spirotrichean, armophorean, plagiopylean, and oligohymenophorean endosymbionts. It we is also likely that oligohymenophoreans have repeatedly invaded the gut habitat. To test this hypothesis, we sequenced small subunit rRNA the (SSrRNA) genes of 6 species representing the larger scuticociliate species found in the intestine of Strongylocentrotus pallidus from the northeast Entorhipidiidae Pacific Ocean: Entodiscus borealis (Entodiscidae); Plagiopyliella pacifica and Thyrophylax vorax (Thyrophylacidae); and Entorhipidium pilatum, Entorhipidium tenue, and Entorhipidium sp.(Entorhipidiidae).hypothesis, SsrRNA genes were amplified by PCR, and sequences obtained in both directions. In all phylogenetic analyses, the scuticociliates are well is supported as a clade. Entodiscus is distinct from these other echinoid taxa and is the sister taxon to the facultatively species parasitic Uronema marinum. The other 5 echinoid species always form a clade whose basal species is the free-living Parauronema longum.echinoid The greatest genetic distance among these latter 5 species is less than 1.5%. This probably explains why the Thyrophylacidae and and Entorhipidiidae are paraphyletic based on the SSrRNA gene sequences.

We that have used small subunit rRNA gene sequences to determine the phylogenetic relationships of species in three genera of endosymbiotic ciliates.the We have confirmed that the astome Anoplophrya marylandensis is related to ciliates in the Class Oligohymenophorea, supporting the view that Class astomes are derived from hymenostome-like ancestors. We confirmed that Plaglotoma lumbrici, formerly considered to be a heterotrich, is a stichotrich confirm spirotrich ciliate most closely related to Paraurostyla weissei in this analysis. Thus, the somatic polykinetids of Plagiotoma can be concluded now to be cirri. We report the details of our isolation of Nyctotheroides deslierresae and Nyctotheroides parvus and confirm previous reports the that these clevelandellids are related to the metopid and caenomorphid ciliates, now placed in the Class Armophorea.

We taxa, surveyed a variety of studies that have used single-cell polymerase chain reaction (SC-PCR) to examine the gene sequences of a been diversity of unicellular protists. Representatives of all the Super-Groups of eukaryotes have been subjected to SC-PCR with ciliates and dinoflagellates was being most commonly examined. The SC-PCR was carried out either by directly amplifying a single lysed cell or by first description extracting DNA and following this with amplification of the DNA extract. Cell lysis methods included heating, freezing, mechanical rupture, and gene enzyme digestion. Cells fixed or preserved with ethanol, methanol, and Lugol's have also been used successfully. Heminested or seminested PCR SC-PCR might follow the initial PCR, whose products were then directly sequenced or cloned and then sequenced. The methods are not subjected complicated. This should encourage protistologists to use SC-PCR in the description of new or revised taxa, especially rare and unculturable a forms, and it should also enable the probing of gene expression in relation to life history stages.

The defined genus Kiitricha was long assumed to be the most primordial taxon in the Stichotrichia [hypotrichs sensu lato (s. l.)] based we on its morphological features and was considered to be an intermediate between heterotrichs and the traditional hypotrichous assemblage. In order constructed to evaluate the phylogenetic position of Kiitricha within the Hypotrichia, we sequenced the small-subunit rRNA gene and the alpha-tubulin gene ventral for a Qingdao population of Kiitricha marina. Phylogenetic trees were constructed and compared to morphological and morphogenetic data. The results but show that (i) Kiitricha is positioned near Phacodinium, both of which always form a sister clade to the assemblage including trees Stichotrichia, Hypotrichia, Oligotrichia and Choreotrichia,(ii) Kiitricha, which may represent an intermediate between heterotrichs (s. l.) and the Stichotrichia-Hypotrichia complex,we is probably an ancestor-like form of the latter group and (iii) in contrast to morphological characters, both molecular and ontogenetic that data support the separation of Kiitricha from the hypotrichs (s. l.). Thus, Kiitricha might be placed in the class Spirotrichea level, at about subclass level, next to Phaconidiidia, Hypotrichia and Stichotrichia, which supports the establishment of a new subclass Protohypotrichia n.Kiitricha subclass within the class Spirotrichea, with characterizations including slightly differentiated somatic ciliature (i.e. cirri on the ventral side generally uniform on and non-grouped, no clearly defined marginal cirral rows, ciliature on the dorsal side mixed with cirri and dikinetids, no clearly from differentiated dorsal kineties) and a unique but intermediate morphogenetic pattern of cortical structures between Hypotrichia and Stichotrichia.

Details for of the phylogenetic relationships among tetrahymenine ciliates remain unresolved despite a rich history of investigation with nuclear gene sequences and three other characters. We examined all available species of Tetrahymena and three other tetrahymenine ciliates, and inferred their phylogenetic relationships using cytochrome nearly complete mitochondrial cytochrome c oxidase subunit 1 (cox1) and small subunit (SSU) rRNA gene sequences. The inferred phylogenies showed demonstrated the genus Tetrahymena to be monophyletic. The three "classical" morphology-and-ecology-based groupings are paraphyletic. The SSUrRNA phylogeny confirmed the previously established of australis and borealis groupings, and nine ribosets. However, these nine ribosets were not well supported. Using cox1 gene, the deduced mitochondrial phylogenies based on this gene revealed 12 well supported groupings, called coxisets, which mostly corresponded to the nine ribosets. This other study demonstrated the utility of cox1 for resolving the recent phylogeny of Tetrahymena, whereas the SSU rRNA gene provided resolution small of deeper phylogenetic relationships within the genus.

The species planktonic ciliate Balanion masanensis n. sp. is described from living cells, from cells prepared by quantitative protargol staining (QPS), scanning a electron microscopy (SEM), and transmitted electron microscopy (TEM) preparations, and the sequence of its nuclear small subunit rDNA (SSU rDNA)circumoral is reported. This species is almost ovoid with a flattened anterior oral region when the cells are alive and stained.than The flattened anterior region of a living cell often forms a dome with the perimeter receded in a groove, and from this region is easily inflated or depressed. In SEM photos, a brosse of six to nine monokinetids (or possibly three inside to five dikinetids) was observed inside the circumoral dikinetids. In TEM photos, circumoral microtubular ribbons were observed below the oral a cilia, which along with the oral flaps were 8-16 mum in length. The cytostome is a slight funnel-like central depression oral on the flattened anterior end. The morphological characteristics of this ciliate are identical to those of the genus Balanion (Order mum Prorodontida). The ranges (and mean+/-standard deviation) of cell length, cell width, and oral diameter of living cells (n=23-26) were 27-43 dome mum (35.2+/-4.6), 25-32 mum (28.6+/-2.3), and 25-30 mum (27.6+/-1.3), respectively, while those of the QPS-stained specimens (n=70) were 23-37 mum of (30.6+/-3.5), 26-35 mum (30.7+/-2.2), and 26-33 mum (29.5+/-1.5), respectively. Forty-six to 55 somatic kineties (SKs) were equally spaced around the from cell body and extended from the oral to near the posterior regions with 24-50 monokinetids per kinety. Each kinetid bore posterior a cilium 2.8-7.2 mum long. A caudal cilium (ca 14 mum long) arose on the posterior end. The single ellipsoid Accession macronucleus is 6.8-13.4 x 6.8-10.5 mum, accompanied by a single micronucleus (2. -2.8 x 1.5-2.5 mum) visible only in QPS specimens.greater Because, the cell size, the number of SKs, and the number of kinetosomes per SK of this ciliate were much mum greater than those of Balanion comatum and Balanion planctonicum, the only two Balanion species so far reported, we have established sequence B. masanensis n. sp. When properly aligned, the sequence of the SSU rDNA of B. masanensis n. sp.(GenBank Accession mum No. AM412525) was approximately 9% different from that of Coleps hirtus (Colepidae, Prorodontida) and 12% different from that of Prorodon dikinetids. teres (Prorodontidae, Prorodontida).

The that mitochondrial cytochrome-c oxidase subunit 1 (cox1) gene has been proposed as a DNA barcode to identify animal species. To test to the applicability of the cox1 gene in identifying ciliates, 75 isolates of the genus Tetrahymena and three non-Tetrahymena ciliates that Tetrahymena are close relatives of Tetrahymena, Colpidium campylum, Colpidium colpoda and Glaucoma chattoni, were selected. All tetrahymenines of unproblematic species could of be identified to the species level using 689 bp of the cox1 sequence, with about 11 % interspecific sequence divergence.a Intraspecific isolates of Tetrahymena borealis, Tetrahymena lwoffi, Tetrahymena patula and Tetrahymena thermophila could be identified by their cox1 sequences, showing lwoffi, < .65 % intraspecific sequence divergence. In addition, isolates of these species were clustered together on a cox1 neighbour-joining (NJ) tree.the However, strains identified as Tetrahymena pyriformis and Tetrahymena tropicalis showed high intraspecific sequence divergence values of 5.01 and 9.07 %,intraspecific respectively, and did not cluster together on a cox1 NJ tree. This may indicate the presence of cryptic species. The animals, mean interspecific sequence divergence of Tetrahymena was about 11 times greater than the mean intraspecific sequence divergence, and this increased species to 58 times when all isolates of species with high intraspecific sequence divergence were excluded. This result is similar to in DNA barcoding studies on animals, indicating that congeneric sequence divergences are an order of magnitude greater than conspecific sequence divergences.when Our analysis also demonstrated low sequence divergences of <1. % between some isolates of T. pyriformis and Tetrahymena setosa on sequence the one hand and some isolates of Tetrahymena furgasoni and T. lwoffi on the other, suggesting that the latter species the in each pair is a junior synonym of the former. Overall, our study demonstrates the feasibility of using the mitochondrial isolates cox1 gene as a taxonomic marker for 'barcoding' and identifying Tetrahymena species and some other ciliated protists.

The terminal odontostomatid ciliates have remained a homogeneous order of ciliates since the 1930s when they were recognized as a monophyletic assemblage.class Since that time they have been placed with the heterotrich ciliates, and more recently transferred as incertae sedis to the antiquorum new "riboclass" class Armophorea. We were able to obtain the small subunit rRNA gene sequence of the odontostomatid Epalxella antiquorum tentatively (Penard, 1922) Corliss, 1960, collected from the meromictic alpine Lake Alat in Germany, in July 2005. An alignment with representatives species, of all 11 classes of ciliates unambiguously places the Epalxella sequence with other representatives of the class Plagiopylea with 100%odontostomatid support in both maximum likelihood and Bayesian analyses. Epalxella is the basal lineage with trimyemid and plagiopylid ciliates forming the class two terminal sister clades. While this molecular support is strong and unambiguous, there are no obvious morphological features to unite alpine these three clades. Thus, the class Plagiopylea must continue to be referred to as a "riboclass." Using the Epalxella sequence features as a basal marker, we tentatively identified 20 environmental sequences to the terminal plagiopylean clades: eight to the genus Trimyema;the four to the genus Plagiopyla; and eight to two new species, one of which might represent a new plagiopylean genus.a

Ciliates The of the subclass Trichostomatia inhabit the fermentative regions of the digestive tract of herbivores. Most available small subunit ribosomal RNA sheep (SSrRNA) gene sequences of trichostomes are from species isolated from the rumen of cattle or sheep and from marsupials. No We ciliate species endosymbiotic in horses has yet been analyzed. We have sequenced the SSrRNA genes of five ciliate species, isolated to from the cecum and colon of four Yakut horses: Cycloposthium edentatum, Cycloposthium ishikawai, Tripalmaria dogieli, Cochliatoxum periachtum, and Paraisotricha colpoidea.but Based on their morphology, Cycloposthium, Tripalmaria, and Cochliatoxum are classified as Entodiniomorphida, while Paraisotricha is considered a member of the analyzed. Vestibuliferida. Phylogenetic analyses using Bayesian inference, distance, and parsimony methods confirm these placements. The two Cycloposthium species cluster together with sheep the published Cycloposthium species isolated from a wallaby in Australia. Tripalmaria and Cochliatoxum branch as a sister group to or ciliate basal within the Entodiniomorphida. The Vestibuliferida remain paraphyletic with Paraisotricha and Balantidium branching basal to all other trichostome species, but two not closely related to Isotricha and Dasytricha.

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

The with systematic relationships and taxonomic positions of the traditional heterotrich genera Condylostentor, Climacostomum, Fabrea, Folliculina, Peritromus, and Condylostoma, as well as ribosomal the licnophorid genus Licnophora, were re-examined using new data from sequences of the gene coding for small subunit ribosomal RNA.(1) Trees constructed using distance-matrix, Bayesian inference, and maximum-parsimony methods all showed the following relationships:(1) the "traditional" heterotrichs consist of Eufolliculina, several paraphyletic groups, including the current classes Heterotrichea, Armophorea and part of the Spirotrichea;(2) the class Heterotrichea was confirmed relationship as a monophyletic assemblage based on our analyses of 31 taxa, and the genus Peritromus was demonstrated to be a relationships: peripheral group;(3) the genus Licnophora occupied an isolated branch on one side of the deepest divergence in the subphylum RNA. Intramacronucleata and was closely affiliated with spirotrichs, armophoreans, and clevelandellids;(4) Condylostentor, a recently defined genus with several truly unique including morphological features, is more closely related to Condylostoma than to Stentor;(5) Folliculina, Eufolliculina, and Maristentor always clustered together with armophoreans, high bootstrap support; and (6) Climacostomum occupied a paraphyletic position distant from Fabrea, showing a close relationship with Condylostomatidae and small Chattonidiidae despite of modest support.

The indicates conjugation of Halteria grandinella was studied in protargol preparations. The isogamontic conjugants fuse partially with their ventral sides to a kineties homopolar pair. The first maturation division generates dramatic transformations:(i) the partners obtain an interlocking arrangement;(ii) the number of membranelles, bristle kineties decreases from seven to four in each partner; and (iii) the right conjugant loses its buccal membranelles, the of left the whole adoral zone. The remaining collar membranelles arrange around the pair's anterior end and are shared by both on partners; finally, the couple resembles a vegetative specimen in size and outline. The vegetative macronucleus fragments before pycnosis. The micronucleus buccal performs three maturation divisions, but only one derivative each performs the second and third division. The synkaryon divides twice, producing kineties a micronucleus, a macronucleus anlage, and two disintegrating derivatives. Scattered somatic kinetids occur during conjugation, but disappear without reorganization. An collar incomplete oral primordium originates in both partners. The conjugation of Halteria grandinella resembles in several respects that of hypotrich spirotrichs;but however, the majority of morphological, ontogenetical, and ultrastructural features still indicates an affiliation with the oligotrich and choreotrich spirotrichs. Accordingly,number the cladistic analysis still contradicts the genealogy based on the sequences of the small subunit rRNA gene.

The three morphology, infraciliature and silverline system of two marine scuticociliates, Pleuronema grolierei nov. spec. and Pleuronema coronatum Kent, 1881, isolated from impregnation the sand beaches along the coast of Qingdao, China, were investigated using live observation and silver impregnation methods. Ciliates of paroral the genus Pleuronema are normally easily recognized by their large sail-like paroral membrane. In this respect, P. coronatum is a and typical member of the genus, whereas P. grolierei is unique in having much shorter cilia in the paroral membrane, which,Dragesco, in consequence, is rather inconspicuous. Nevertheless, details of the infraciliature confirm that P. grolierei belongs to the genus Pleuronema. Within sail-like the genus Pleuronema, the posterior end of the anterior fragment of the second membranelle (M2a) may be either straight, as impregnation in P. grolierei, or hooked-shaped, as in P. coronatum, providing a criterion for recognizing two sections within the genus. Based a on the current study and previous reports, we propose that three nominal forms, Pleuronema balli Small and Lynn, 1985, Pleuronema as smalli Dragesco, 1968 and Pleuronema borrori Dragesco, 1968 should be synonymized with P. coronatum.

The single ciliate class Colpodea provides a powerful case in which a molecular genealogy can be compared to a detailed morphological taxonomy the of a microbial group. Previous analyses of the class using the small-subunit rDNA are based on sparse taxon sampling, and of are therefore of limited use in comparisons with morphologically-based classifications. Taxon sampling is increased here to include all orders within some the class, and more species within previously sampled orders and in the species rich genus Colpoda. Results indicate that the as Colpodea may be paraphyletic, although there is no support for deep nodes. The orders Bursariomorphida, Grossglockneriida, and Sorogenida are monophyletic.of The orders Bryometopida, Colpodida, and Cyrtolophosidida, and the genus Colpoda, are not monophyletic. Although congruent in many aspects, the conflict the between some nodes on this single gene genealogy and morphology-based taxonomy suggests the need for additional markers as well as classifications. a reassessment of the Colpodea taxonomy.

The terminal odontostomatid ciliates have remained a homogeneous order of ciliates since the 1930s when they were recognized as a monophyletic assemblage.class Since that time they have been placed with the heterotrich ciliates, and more recently transferred as incertae sedis to the antiquorum new "riboclass" class Armophorea. We were able to obtain the small subunit rRNA gene sequence of the odontostomatid Epalxella antiquorum tentatively (Penard, 1922) Corliss, 1960, collected from the meromictic alpine Lake Alat in Germany, in July 2005. An alignment with representatives species, of all 11 classes of ciliates unambiguously places the Epalxella sequence with other representatives of the class Plagiopylea with 100%odontostomatid support in both maximum likelihood and Bayesian analyses. Epalxella is the basal lineage with trimyemid and plagiopylid ciliates forming the class two terminal sister clades. While this molecular support is strong and unambiguous, there are no obvious morphological features to unite alpine these three clades. Thus, the class Plagiopylea must continue to be referred to as a "riboclass." Using the Epalxella sequence features as a basal marker, we tentatively identified 20 environmental sequences to the terminal plagiopylean clades: eight to the genus Trimyema;the four to the genus Plagiopyla; and eight to two new species, one of which might represent a new plagiopylean genus.a

The 1972 cortical development during binary fission of the relatively poorly known stichotrich ciliate, Trachelostyla pediculiformis (Cohn, 1866) Borror, 1972, found in entirely coastal waters near Qingdao, China, was investigated using the protargol impregnation method. The morphogenetic process reveals some pretty unusual characteristics,are which do not follow the Oxytricha-pattern:(1) the parental oral apparatus is entirely renewed from an oral primordium formed de the novo in the proter;(2) in the proter, the parental undulating membranes are not involved in the formation of the a newly formed oral primordium; both undulating membrane-anlagen (UM-anlage) and frontoventral-transverse cirral anlagen (FVT-anlagen) develop from the oral primordium in the membranes proter;(3) the dorsal kineties (DK) are generated in a unique way, that is, in both dividers, two separate groups entirely of DK-anlagen develop in the right- and left-most DK, generate all the DK and evolve to replace the old structures;both (4) three caudal cirri are formed at the posterior ends of three right-most dorsal kinety anlagen;(5) eight frontal, five streaks, ventral and five transverse cirri are derived from six streaks, namely, the UM-anlage and 5 FVT-anlagen; the cirri are segregated parental from these anlagen in the pattern 1:3:3:3:4:4 (from left to right) in the Oxytricha mode. Based on both SSrRNA gene waters sequencing and morphogenetic data, the systematic positions of the genus Trachelostyla Borror, 1972 as well as the family Trachelostylidae Small right-most and Lynn, 1985 are briefly analyzed. The results indicate that this genus/family could be a highly isolated lineage and might UM-anlage be ancestral to other well-known oxytrichids.

So of far, neither morphology nor gene sequences have provided a reliable classification of halteriid and hypotrichid spirotrichs. Thus, we performed a granulifera. comparative study on the fine structure of the resting cysts in some representative species, viz., the oligotrichs Halteria grandinella and thorns Pelagostrombidium fallax and the oxytrichid hypotrichs Laurentiella strenua, Steinia sphagnicola, and Oxytricha granulifera. Main results include:(i) there are three viz., different, very likely non-homologous cyst surface ornamentations, viz., spines (generated by the ectocyst), thorns (generated by the mesocyst), and lepidosomes which (produced in the cytoplasm);(ii) Halteria has a perilemma;(iii) Halteria, Meseres and Pelagostrombidium have fibrous lepidosomes, while those of the Oxytricha are tubular;(iv) the cyst wall structure of Pelagostrombidium and Strombidium is distinctly different from that of halteriids and Main oxytrichids, which are rather similar in this respect;(v) cyst ornamentation does not provide a reliable phylogenetic signal in oxytrichid Halteria hypotrichs because ectocyst spines occur in both flexible and rigid genera. The new observations and literature data were used to scheme investigate the phylogeny of the core Spirotrichea. The Hennigian argumentation scheme and computer algorithms showed that the spirotrichs are bound Steinia together by the macronuclear reorganization band, the subepiplasmic microtubule basket, and the apokinetal stomatogenesis. The Hypotrichida and Oligotrichida are united study by a very strong synapomorphy, viz., the perilemma, not found in any other member of the phylum. Halteriid and oligotrichid observations spirotrichs form a sister group supported by as many as 13 apomorphies. Thus, the molecular data, which classify the halteriids showed within the core hypotrichs, need to be reconsidered.

A two fusiform, spirally coiled and pleated, free-living, heavily placate echinoderm with an expansible test has been discovered in the Lower Cambrian been Olenellus zone of California. It is characterized by 10 "interambulacral" areas, a single principal endothecal ambulacrum with a short branch,Olenellus and oral and apical regions at opposite poles. A new class, the Helicoplacoidea is proposed for the new genus Helicoplacus,genus with two new species, H. gilberti (type species) and H. curtisi.

The the oldest known "carpoids" of the echinoderm subphylum Homalozoa have been discovered in lower Middle Cambrian rocks of northern Utah. They in were free-living benthonic animals characterized by a unique ctenoid feeding apparatus and flattened flexible theca with near bilateral symmetry. A rocks new class, Ctenocystoidea, is proposed to include the new genus and new species, Ctenocystis utahensis.