The interplay of nested biotic interactions and the abiotic environment regulates populations of a hypersymbiont.

The role of biotic interactions in shaping the distribution and abundance of species should be particularly pronounced in symbionts. Indeed, symbionts have a dual niche composed of traits of their individual hosts and the abiotic environment external to the host, and often combine active dispersal at finer scales with host-mediated dispersal at broader scales. The biotic complexity in the determinants of species distribution and abundance should be even more pronounced for hyper symbionts (symbionts of other symbionts). We use a chain of symbiosis to explore the relative influence of nested biotic interactions and the abiotic environment on occupancy and abundance of a hypersymbiont. Our empirical system is the epibiont ciliate Lagenophrys discoidea, which attaches to an ostracod that is itself ectosymbiotic on crayfish (the basal host). We applied multimodel selection and variance partitioning for GLMM to assess the relative importance of (a) traits of symbiotic hosts (ostracod sex and abundance), (b) traits of basal hosts (crayfish body weight, abundance and intermoult stage), (c) the abiotic environment (water chemistry and climate) and (d) geospatial autocorrelation patterns (capturing potential effects of crayfish dispersal among localities). Our models explained about half of the variation in prevalence and abundance of the hypersymbiont. Variation in prevalence was partly explained, in decreasing order of importance (18%-4%) by shared effects of symbiotic host traits and the abiotic environment, pure fixed effects of symbiotic hosts, abiotic environment and geospatial patterns (traits of basal hosts were not relevant). Hypersymbiont abundance was most strongly explained by random effects of host traits (mainly the symbiotic host), in addition to weaker fixed effects (mostly abiotic environment). Our results highlight the major role of the interplay between abundance of symbiotic hosts and water physico-chemistry in regulating populations of a hypersymbiotic ciliate, which is likely critical for dispersal dynamics, availability of attachment resources and suitability of on-host living conditions for the ciliate. We also found moderate signal of regulation by the basal host, for which we propose three mechanisms: (a) modulation of microhabitat suitability (crayfish-created water currents); (b) concentration of symbiotic hosts within crayfish; and (c) dispersal mediated by crayfish.

Phylogeny of the families Zoothamniidae and Epistylididae (Protozoa: Ciliophora: Peritrichia) based on analyses of three rRNA-coding regions.

Peritrichs are a major group of ciliates with worldwide distribution, and they play important roles in many habitats. Intrafamilial phylogeny of some peritrichs was investigated using information from three genes, which provided more robust interpretations than single-gene analyses. Sixty-seven new sequences including SSU rDNA, ITS1-5.8S-ITS2 and LSU rDNA were aligned with available sequences in GenBank to infer phylogenetic relationships within the families Zoothamniidae and Epistylididae. Results reveal the following relationships: (1) Epistylididae is polyphyletic, consisting of two clades that nest within the Zoothamniidae as part of the crown clade of peritrichs (order Vorticellida) and a third one that is part of the basal clade of peritrichs (order Opercularida); (2) Epistylis elongata falls within one of the clades of Zoothamnium rather than with congeners; (3) Zoothamnium is probably paraphyletic, consisting of three divergent clades, with the genera Myoschiston and Zoothamnopsis intermingled with species of Zoothamnium. The following evolutionary hypotheses can be inferred from these results: (1) the contractile stalk of Zoothamnium is plesiomorphic. (2) Myoschiston, Zoothamnopsis and clade II of Epistylididae are derived from the Zoothamnium morphotype by partial or incomplete development of the spasmoneme that forms the contractile center of the stalk around which the rigid cortex is secreted. (3) Clade I of the Epistylididae, which are primarily colonial forms that appear never to have evolved a spasmoneme of any sort, may represent the ancestral morphotype of peritrichs.

Beyond the "Code": A Guide to the Description and Documentation of Biodiversity in Ciliated Protists (Alveolata, Ciliophora).

Recent advances in molecular technology have revolutionized research on all aspects of the biology of organisms, including ciliates, and created unprecedented opportunities for pursuing a more integrative approach to investigations of biodiversity. However, this goal is complicated by large gaps and inconsistencies that still exist in the foundation of basic information about biodiversity of ciliates. The present paper reviews issues relating to the taxonomy of ciliates and presents specific recommendations for best practice in the observation and documentation of their biodiversity. This effort stems from a workshop that explored ways to implement six Grand Challenges proposed by the International Research Coordination Network for Biodiversity of Ciliates (IRCN-BC). As part of its commitment to strengthening the knowledge base that supports research on biodiversity of ciliates, the IRCN-BC proposes to populate The Ciliate Guide, an online database, with biodiversity-related data and metadata to create a resource that will facilitate accurate taxonomic identifications and promote sharing of data.

An ITS-based phylogenetic framework for the genus Vorticella: finding the molecular and morphological gaps in a taxonomically difficult group.

Vorticella includes more than 100 currently recognized species and represents one of the most taxonomically challenging genera of ciliates. Molecular phylogenetic analysis of Vorticella has been performed so far with only sequences coding for small subunit ribosomal RNA (SSU rRNA); only a few of its species have been investigated using other genetic markers owing to a lack of similar sequences for comparison. Consequently, phylogenetic relationships within the genus remain unclear, and molecular discrimination between morphospecies is often difficult because most regions of the SSU rRNA gene are too highly conserved to be helpful. In this paper, we move molecular systematics for this group of ciliates to the infrageneric level by sequencing additional molecular markers-fast-evolving internal transcribed spacer (ITS) regions-in a broad sample of 66 individual samples of 28 morphospecies of Vorticella collected from Asia, North America and Europe. Our phylogenies all featured two strongly supported, highly divergent, paraphyletic clades (I, II) comprising the morphologically defined genus Vorticella. Three major lineages made up clade I, with a relatively well-resolved branching order in each one. The marked divergence of clade II from clade I confirms that the former should be recognized as a separate taxonomic unit as indicated by SSU rRNA phylogenies. We made the first attempt to elucidate relationships between species in clade II using both morphological and multi-gene approaches, and our data supported a close relationship between some morphospecies of Vorticella and Opisthonecta, indicating that relationships between species in the clade are far more complex than would be expected from their morphology. Different patterns of helix III of ITS2 secondary structure were clearly specific to clades and subclades of Vorticella and, therefore, may prove useful for resolving phylogenetic relationships in other groups of ciliates.

Molecular and morphological characterization of a poorly known marine ciliate, Myoschiston duplicatum precht 1935: implications for phylogenetic relationships between three morphologically similar genera -- Zoothamnium, Myoschiston, and Zoothamnopsis (Ciliophora, Peritrichia, Zoothamniidae).

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

Evolution of variations in the common pattern of stomatogenesis in peritrich ciliates: evidence from a comparative Study including a new description of stomatogenesis in Pseudepistylis songi Peng et al., 2007.

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

Molecular phylogeny of the family Vorticellidae (Ciliophora, Peritrichia) using combined datasets with a special emphasis on the three morphologically similar genera Carchesium, Epicarchesium and Apocarchesium.

Little is known about the phylogeny of the family Vorticellidae at the generic level because few comprehensive analyses of molecular phylogenetic relationships between members of this group have, so far, been done. As a result, the phylogenetic positions of some genera that were based originally on morphological analyses remain controversial. In the present study, we performed phylogenetic analyses of vorticellids based on the sequence of the small-subunit (SSU) rRNA gene, including one species of the genus Apocarchesium, for which no sequence has previously been reported. Phylogenetic trees were reconstructed with SSU rRNA gene sequences by using four different methods (Bayesian analysis, maximum-likelihood, neighbour-joining and maximum-parsimony) and had a consistent branching pattern. Members of the genera Vorticella (except V. microstoma) and Carchesium formed a clearly defined, well supported clade that was divergent from the clade comprising members of the genera Pseudovorticella and Epicarchesium, suggesting that the differences in the silverline system (transverse vs reticulate) among vorticellids may be the result of genuine evolutionary divergence. Members of the newly established genus Apocarchesium clustered within the family Vorticellidae basal to the clade containing members of the genera Pseudovorticella and Epicarchesium and were distinct from members of the genus Carchesium, supporting the validity of Apocarchesium as a novel genus. Additional phylogenetic analyses of 21 strains representing seven genera from the families Vorticellidae and Zoothamniidae were performed with single datasets (ITS1-5.8S-ITS2, ITS2 alone) and combined datasets (SSU rRNA+ITS1-5.8S-ITS2, SSU rRNA+ITS2) to explore further the phylogenetic relationship between the three morphologically similar genera Carchesium, Epicarchesium and Apocarchesium, using characteristics not included in previous analyses. The phylogenetic trees reconstructed with combined datasets were more robust and therefore more reliable than those based on single datasets and supported the results of trees based on SSU rRNA sequences.

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

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

Description of Paratetrahymena parawassi n. sp. using morphological and molecular evidence and a phylogenetic analysis of Paratetrahymena and other taxonomically ambiguous genera in the order Loxocephalida (Ciliophora, Oligohymenophorea).

The marine scuticociliate Paratetrahymena parawassi n. sp. is described on the basis of morphology, especially infraciliature, and the sequence of its small subunit (SSU) rRNA gene to become the second known member of its genus. Paratetrahymena and other ciliates in the order Loxocephalida possess a mixture of morphological and morphogenetic features characteristic of the subclasses Hymenostomatia and Scuticociliatia. Accordingly, we used SSU rRNA sequences to analyze the phylogeny of Paratetrahymena and three other loxocephalid genera. Paratetrahymena and Cardiostomatella vermiformis formed a moderately well-supported clade that diverged at a deep level from all other scuticociliates, supporting separation of loxocephalids from other scuticociliates as a suprafamilial taxon. Sathrophilus holtae was a sister taxon to Paratetrahymena and Cardiostomatella in a poorly supported, unresolved relationship; nevertheless, association of all three genera into a single clade was supported by an approximately unbiased (AU) test. Any association of these genera singly or as a group with the Hymenostomatia was rejected decisively by AU tests and by a complete absence in the loxocephalids of the unique nucleotide identities that distinguish hymenostomes. Therefore, the morphological and morphogenetic similarities of loxocephalids to hymenostomes may be plesiomorphies, and the conflicting mix of scuticociliate and hymenostome characteristics seen in loxocephalids may result from differing rates of character evolution. Dexiotrichides pangi and Urocentrum, which is currently classified as a peniculid, formed a small clade that associated with hymenostomes and peritrichs. Monophyly of the Loxocephalida with Dexiotrichides and/or Urocentrum included was not rejected by AU; however, inclusion of Urocentrum in the Peniculia was rejected by AU tests. A hypothesis is offered to explain the lack of resolution of loxocephalid ciliates and Urocentrum in phylogenetic trees, namely that their phylogenetic positions are influenced by a combination of heterogeneous data and long-branch attraction caused by poor representation of taxa in analyses. The well-known genus Cyclidium, a member of the order Pleuronematida, was revealed to be polyphyletic as a byproduct of our analyses of loxocephalids. In particular, Cyclidium porcatum appears to fall outside the clade containing typical members of the subclass Scuticociliatia and thus invites investigation as a possible member of the order Loxocephalida.

Reconsideration of systematic relationships within the order Euplotida (Protista, Ciliophora) using new sequences of the gene coding for small-subunit rRNA and testing the use of combined data sets to construct phylogenies of the Diophrys-complex.

Comprehensive molecular analyses of phylogenetic relationships within euplotid ciliates are relatively rare, and the relationships among some families remain questionable. We performed phylogenetic analyses of the order Euplotida based on new sequences of the gene coding for small-subunit RNA (SSrRNA) from a variety of taxa across the entire order as well as sequences from some of these taxa of other genes (ITS1-5.8S-ITS2 region and histone H4) that have not been included in previous analyses. Phylogenetic trees based on SSrRNA gene sequences constructed with four different methods had a consistent branching pattern that included the following features: (1) the "typical" euplotids comprised a paraphyletic assemblage composed of two divergent clades (family Uronychiidae and families Euplotidae-Certesiidae-Aspidiscidae-Gastrocirrhidae), (2) in the family Uronychiidae, the genera Uronychia and Paradiophrys formed a clearly outlined, well-supported clade that seemed to be rather divergent from Diophrys and Diophryopsis, suggesting that the Diophrys-complex may have had a longer and more separate evolutionary history than previously supposed, (3) inclusion of 12 new SSrRNA sequences in analyses of Euplotidae revealed two new clades of species within the family and cast additional doubt on the present classification of genera within the family, and (4) the intraspecific divergence among five species of Aspidisca was far greater than those of closely related genera. The ITS1-5.8S-ITS2 coding regions and partial histone H4 genes of six morphospecies in the Diophrys-complex were sequenced along with their SSrRNA genes and used to compare phylogenies constructed from single data sets to those constructed from combined sets. Results indicated that combined analyses could be used to construct more reliable, less ambiguous phylogenies of complex groups like the order Euplotida, because they provide a greater amount and diversity of information.

Further consideration of the phylogeny of some "traditional" heterotrichs (Protista, Ciliophora) of uncertain affinities, based on new sequences of the small subunit rRNA gene.

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

Phylogeny of six genera of the subclass Haptoria (Ciliophora, Litostomatea) inferred from sequences of the gene coding for small subunit ribosomal RNA.

The small subunit ribosomal RNA genes of nine species belonging to six genera of litostome ciliates, namely Amphileptus aeschtae, Chaenea teres, Chaenea vorax, Lacrymaria marina, Litonotus paracygnus, Loxophyllum sp.-GD-070419, Loxophyllum jini, Loxophyllum rostratum, and Phialina salinarum, were sequenced for the first time. Phylogenetic trees were constructed using different methods to assess the inter- and intra-generic relationships of haptorians, of which Chaenea, Lacrymaria, Litonotus, and Phialina were analyzed for the first time based on molecular data. Monophyly of the order Pleurostomatida was strongly confirmed, and the two existing families of pleurostomatids, created on the basis of morphology, were confirmed by molecular evidence. Within the Pleurostomatida, Siroloxophyllum utriculariae occupied a well-supported position basal to the Loxophyllum clade, supporting the separation of these genera from one another. Both the subclass Haptoria and the order Haptorida were partially unresolved, possibly paraphyletic assemblages of taxa in all analyses, creating doubts about the traditional placement of some haptorid taxa. The existing sequence of L. rostratum in GenBank (DQ411864) was conspicuously different from that of the isolate from Qingdao, China sequenced in the present work, indicating that they are different species. The isolate from Qingdao was verified as L. rostratum by morphological analysis, and the published morphology of existing GenBank record of L. rostratum is different from it. Based on both morphological and molecular evidence, the latter may be congeneric with an undescribed species of Loxophyllum from Guangdong Province, China.

Morphology, morphogenesis, and phylogeny of an Anteholosticha intermedia (Ciliophora, Urostylida) population from the United States.

A distinct population of Anteholosticha intermedia was isolated from soil in the Great Smoky Mountains of North Carolina, USA, and its morphology, morphogenesis and molecular phylogeny investigated by microscopic observations of live and protargol-prepared specimens and analyses of the sequence of small subunit (SSU) rDNA. Our population closely resembles the populations from Austria and Korea. Members of the genus Anteholosticha have been regarded as ontogenetically diverse, which is confirmed by the present work. The most noteworthy ontogenetic feature of the American population of A. intermedia is that the oral primordium in the proter appears apokinetally at the posterior end of the undulating membranes anlage at the beginning of division and then dedifferentiates midway through morphogenesis. Molecular phylogenetic analyses demonstrate, with high support, that the American population of A. intermedia is clearly distinct from congeners and branches as part of a sister lineage to the Bakuella-Urostyla clade that belongs to the major clade comprising the order Urostylida.

Investigating the biodiversity of ciliates in the 'Age of Integration'.

Biology is now turning toward a more integrative approach to research, distinguished by projects that depend on collaboration across hierarchical levels of organization or across disciplines. This trend is prompted by the need to solve complex, large-scale problems and includes disciplines that could be defined as integrative biodiversity. Integrative biodiversity of protists, including that of ciliates, is still partially in its infancy. This is the result of a shortage of historical data resources such as curated museum collections. Major areas of integrative biodiversity of ciliates that have begun to emerge can be categorized as integrative systematics, phenotypic plasticity, and integrative ecology. Integrative systematics of ciliates is characterized by inclusion of diverse sources of data in treatment of taxonomy of species and phylogenetic investigations. Integrative research in phenotypic plasticity combines investigation of functional roles of individual species of ciliates with genetic and genomic data. Finally, integrative ecology focuses on genetic identity of species in communities of ciliates and their collective functional roles in ecosystems. A review of current efforts toward integrative research into biodiversity of ciliates reveals a single, overarching concern-rapid progress will be achieved only by implementing a comprehensive strategy supported by one or more groups of active researchers.

An annotated checklist of species in the family Lagenophryidae (Ciliophora, Oligohymenophorea, Peritrichia), With a brief review of their taxonomy, morphology, and biogeography.

The genera Lagenophrys Stein, 1852, Paralagenophrys Clamp, 1987, Clistolagenophrys Clamp, 1991, and Operculigera Kane, 1969 make up the family Lagenophryidae and, together, contain more than 80 species worldwide that live mainly as ectosymbionts of crustaceans. Lagenophryids are characterized by possession of a lorica, but the lorica aperture and the associated parts of the body differ between genera. Despite their widespread distribution, relatively few papers have been published on lagenophryids in recent years, and the present paper is intended to promote research on lagenophryids by providing an annotated checklist of all known species and a brief review of the family, including data of their geographic distribution and their hosts.

A new Peritrich Ciliate from a Hypersaline Habitat in Northern China.

A new peritrichous ciliate, Cothurnia salina n. sp., collected from a brine pond of a salt factory in Yantai, China, was investigated based on live observations, silver staining method and molecular phylogenetic analysis. The diagnosis for this new taxon: body elongated columnar, in vivo 80-98 × 12-19 µm; lorica barrel-shaped, with aboral part heavily thickened; stalk extremely short, with approximately ½ of its length within the lorica; macronucleus wormlike, longitudinally oriented; single contractile vacuole ventrally located; pellicle with conspicuous parallel transverse striations, 62-73 from aboral trochal band to peristome and 32-38 from aboral trochal band to scopula; infundibular polykinety 3 (P3) consisting of two ciliary rows, which are equal length, parallel to each other and terminate adstomally between P1 and P2. Small subunit (SSU) rRNA gene trees revealed that the new species clustered with other members of the family Vaginicolidae as expected.

Occurrence of Epistylis anastatica (Linnaeus, 1767) (Ciliophora: Peritrichia) on Mesocyclops isabellae Dussart & Fernando (Crustacea: Copepoda: Cyclopoida) in India, with an annotated checklist of species of Epistylis reported as Epibionts of Cyclopoid Copepods and resources for their identification.

The colonial peritrich ciliate Epistylis anastatica (Linnaeus, 1767) was discovered living as an epibiont on Mesocyclops isabellae Dussart & Fernando, 1988 collected from a pond in Andhra Pradesh state, India. This is the first report of Epistylis on this host, and a description of its morphological characteristics is provided. The biomass of colonies of E. anastatica relative to that of their hosts was relatively high, being greater than that of the host in some cases (8.11 mm3 vs. 1.14 mm3 or less), and this epibiotic burden has the potential to have an impact on the host's activities. Relationships between epibiotic peritrichs and microcrustaceans like cyclopoid copepods may be much more complex than previously supposed and merit more thorough investigation. Species of Epistylis are commonly found on cyclopoid copepods, but resources for taxonomic identification are limited and scattered; therefore, an annotated checklist of all species reported from cyclopoid copepods and a summary of the diagnostic characteristics of each one is provided as an aid to investigators. Some records of Epistylis spp. from cyclopoid copepods that are accompanied by documentary evidence are misidentifications, raising the suspicion that many other undocumented records also may be erroneous; therefore, it is suggested that authors provide supplementary descriptions and figures.

Phylogenetic and Taxonomic Revision of an Enigmatic Group of Haptorian Ciliates, with Establishment of the Kentrophyllidae fam. n. (Protozoa, Ciliophora, Litostomatea, Pleurostomatida).

Haptorian ciliates in the closely similar genera Kentrophyllum and Epiphyllum possess a unique pattern of ciliature and are distinguished from one another only by the presence or absence of cytoplasmic spines projecting from the margin of the cell. Phylogenetic analyses based on SSU rDNA sequences of three new samples from coastal habitats in China revealed that species in the two genera clustered together indiscriminately (i.e. forms of neither genus clustered into an independent clade) as a maximally supported, monophyletic clade that branches basally to all other clades in the order Pleurostomatida and is strongly divergent from other members of the family in which the genera have been placed. As a result, we propose that Epiphyllum be synonymized with Kentrophyllum and that a new family Kentrophyllidae fam. n. be established for the genus. We hypothesize that the two-sutures of Kentrophyllum is a plesiomorphy within the Pleurostomatida and the unique peripheral kinety might represent an autapomorphy of Kentrophyllum. In addition, we provide a taxonomic revision of Kentrophyllum including description of three new species (K. bispinum sp. n., K. strumosum sp. n., and K. qingdaoense sp. n.), redescription of K. verrucosum (Stokes, 1893) Petz et al., 1995, and three new combinations (K. soliforme (Fauré-Fremiet, 1908) comb. n., K. hohuensis (Wang and Nie, 1933) comb. n. and K. shenzhenense (Pan et al., 2010) comb. n.). The surface ultrastructure of the genus Kentrophyllum is recorded for the first time. And a key to all known species of Kentrophyllum was also suggested.

Two New Genera of Planktonic Ciliates and Insights into the Evolution of the Family Strombidiidae (Protista, Ciliophora, Oligotrichia).

Oligotrich ciliates are common marine microplankters, but their biodiversity and evolutionary relationships have not been well-documented. Morphological descriptions and small subunit rRNA gene sequences of two new species representing two new strombidiid genera, Sinistrostrombidium cupiformum gen. nov., sp. nov. and Antestrombidium agathae gen. nov., sp. nov. are presented, and their taxonomy and molecular phylogeny are analyzed. Sinistrostrombidium gen. nov. is characterized by a sinistrally spiraled girdle kinety and a longitudinal ventral kinety. Antestrombidium gen. nov. is distinguished by tripartite somatic kineties (circular and ventral kineties plus dextrally spiraled girdle kinety). Sinistrostrombidium and Antestrombidium branched separately from one another in phylogenetic trees, clustering with different clades of strombidiids. The new genera added to the diversities of ciliary patterns and small subunit rRNA gene sequences in strombidiids leads to presentation of a new hypothesis about evolution of the 12 known strombidiid genera, based on ciliary pattern and partly supported by molecular evidence. In addition, our new morphological and molecular analyses support establishment of a new order Lynnellida ord. nov., characterized by an open adoral zone of membranelles without differentiation of anterior and ventral membranelles, for Lynnella, but we remain unable to assign the genus to a subclass with confidence.

Morphology and molecular phylogeny of two new brackish-water species of Amphisiella (Ciliophora, Hypotrichia), with notes on morphogenesis.

Two new species of the taxonomically confused genus Amphisiella were isolated from brackish-water habitats in southern China, and their morphology and morphogenesis were investigated. The genes coding for small subunit (SSU) rRNA were sequenced in each species and included in a phylogenetic analysis with all data available from nominal species of Amphisiella and related hypotrichs. Both new species are diagnosed by an elongate body shape, yellow-brown/grey-brown cell color, one kind of cortical granule, and two macronuclear nodules. The cortical granules in A. pulchra sp. n. are grouped in small clusters which are irregularly distributed both ventrally and dorsally; in A. candida sp. n., they are irregularly and sparsely distributed ventrally and form longitudinal stripes dorsally. The ontogenetic processes of both species show similarities to those of other congeners. Phylogenetic trees based on SSU rRNA gene sequences suggest that the family Amphisiellidae is a paraphyletic assemblage. The results further demonstrate that two isolates identified as Amphisiella annulata (DQ832260 and GU170843) are very likely cryptic species, and a sequence identified as A. milnei (DQ845293) may belong to a genus other than Amphisiella.