Checklist of species in the genus Epistylis (Ciliophora, Peritrichia): the first step for a taxonomic revision.

The subclass Peritrichia includes sessile ciliates that attach to aquatic non-living or living substrates. In the case of the latter, they live in a relationship called epibiosis. The genus Epistylis Ehrenberg,1830 is the second-most speciose genus within Sessilida, being characterized by its colonial lifestyle and having a non-contractile stalk. Species of Epistylis have been described from a wide range of aquatic habitats worldwide. The main goal of the present study is to provide a global checklist of the valid species of Epistylis, which may serve as a first step towards a taxonomic review of the genus.

The tale of an endemic shrimp's exceptional osmoregulation and the ancient Athalassic mangrove oasis.

The hyperarid mangrove in the Middle East is characterised by the absence of rivers or freshwater inputs and is one of the most extreme settings of this ecosystem on Earth. Endemic to Qatar's hyperarid mangroves, a Palaemon shrimp is uniquely confined to a sole mangrove site in the Arabian Gulf. Within these mangrove channels, we unveiled brine groundwater sources exceeding 70 ppt salinity, contrasting the local marine standard of 42 ppt. Concurrently, a mysid species typically linked to salt pans and groundwater coexists. Stable isotopic analysis implied the existence of a predator-prey dynamic between this mysid species and the studied shrimp. Then, investigating the endemic shrimp's adaptation to extreme salinity, we conducted osmolarity experiments and phylogenetic studies. Our findings demonstrate that this shrimp transitions from hypo- to hyper-osmoregulation, tolerating salinities from 18 to 68 ppt-an unprecedented osmoregulatory capacity among caridean shrimps. This speciation pattern likely arises from the species osmolarity adaptation, as suggested for other Palaemon congeners. Phylogenetic analysis of the studied Palaemon, along with the mangrove's geological history, suggests a profound evolutionary interplay between the ecosystem and the shrimp since the Eocene. This study proposes the hyperarid mangrove enclave as an Athalassic mangrove oasis-a distinctive, isolated ecosystem within the desert landscape.

Structure and Dynamics of Periphyton in a Neotropical Freshwater Lake, with Emphasis on Ciliates and Their Relationships with Bacterial Taxa.

Periphyton communities in freshwater systems play an essential role in biogeochemical processes, but knowledge of their structure and dynamics lags far behind other environments. We used eDNA metabarcoding of 16S and 18S rRNA markers to investigate the formation and establishment of a periphytic community, in addition to a morphology-based approach for peritrich ciliate determinations, its most abundant group. We sampled two nearby sites within a large Neotropical lake at four time points, aiming to assess whether periphyton establishment can be replicated on this local scale. Producers and denitrifiers were abundant in the community, illustrating the relevant role of biofilms in freshwater nutrient recycling. Among microeukaryotes, peritrich ciliates dominated the community, with genera Epistylis and Vorticella being the most abundant and showing a clear succession at both sites. Other ciliates were morphologically identified and, in some cases, their occurrence was strongly related to bacterial abundance. The structure of both prokaryotic and eukaryotic components of periphyton was not different, while the turnover dynamics differed between the two sites, in spite of their adjacent locations and similar abiotic properties. This indicates that the establishment of these communities can vary even on a local scale within a lake ecosystem.

"Hitchhicking with invertebrates": two reports of epibiosis by peritrich ciliates on ostracods and hydrachnid mites in tanks of epiphytic bromeliads from south Brazil.

Temporary waters are common environments found in physical and biological substrates. Among them, some bromeliads species are known to hold water in their tanks, in a habitat called phytotelmata. Phytotelmata serve as habitats for several organisms, from bacteria and protists to arthropods and anurans. Peritrich ciliates are often found as epibionts on aquatic invertebrates in these environments. Here, we report two cases of epibiosis involving Lagenophrys sp. attached to ostracods (Elpidium spp.) and Rhabdostyla sp. colonizing hydrachnid mites in the tanks of two bromeliad species. In our analysis, we measured the frequency of epibiosis considering the presence of both basibiont and epibiont in the samples. The results shown a significant difference between Elpidium sp. and Lagenophrys sp. compared to hydrachnid mites and Rhabdostyla sp. (87.5% and 19%, respectively), supported by the Kruskal-Walis test (p = 0.0003, Chi-square = 9.687). These reports are important since the knowledge of phytotelmata communities from tropical and subtropical areas is incipient, although it has been increasing over the last years. It also shows that epibiosis doesn't always represent a beneficial relationship. These two epibiosis systems found in bromeliad tanks raise questions about organism's dispersal throughout other phytotelmata and other temporary water habitats.

Symposium on Ciliates in Memory of Denis Lynn.

Denis Lynn (1947-2018) was an outstanding protistologist, applying multiple techniques and data sources and thus pioneering an integrative approach in order to investigate ciliate biology. For example, he recognized the importance of the ultrastructure for inferring ciliate phylogeny, based on which he developed his widely accepted classification scheme for the phylum Ciliophora. In this paper, recent findings regarding the evolution and systematics of both peritrichs and the mainly marine planktonic oligotrichean spirotrichs are discussed and compared with the concepts and hypotheses formulated by Denis Lynn. Additionally, the state of knowledge concerning the diversity of ciliates in bromeliad phytotelmata and amitosis in ciliates is reviewed.

Remarkably Complex Microbial Community Composition in Bromeliad Tank Waters Revealed by eDNA Metabarcoding.

To investigate patterns of biotic community composition at different spatial scales and biological contexts, we used environmental DNA metabarcoding to characterize eukaryotic and prokaryotic assemblages present in the phytotelmata of three bromeliad species (Aechmea gamosepala, Vriesea friburgensis, and Vriesea platynema) at a single Atlantic Forest site in southern Brazil. We sampled multiple individuals per species and multiple tanks from each individual, totalizing 30 samples. We observed very high levels of diversity in these communities, and remarkable variation across individuals and even among tanks from the same individual. The alpha diversity was higher for prokaryotes than eukaryotes, especially for A. gamosepala and V. platynema samples. Some biotic components appeared to be species-specific, while most of the biota was shared among species, but varied substantially in frequency among samples. Interestingly, V. friburgensis communities (which were sampled at nearby locations) tended to be more heterogeneous across samples, for both eukaryotes and prokaryotes. The opposite was true for V. platynema, whose samples were more broadly spaced but whose communities were more similar to each other. Our results indicate that additional attention should be devoted to within-individual heterogeneity when assessing bromeliad phytotelmata biodiversity, and highlight the complexity of the biotic assemblages gathered in these unique habitats.

Quadrigyrus torquatus cystacants Van Cleave, 1920 (Acanthocephala: Quadrigyridae) parasitizing species of Astyanax (Characiformes: Characidae) from southern Brazil / Cistacantos de Quadrigyrus torquatus Van Cleave, 1920 (Acanthocephala: Quadrigyridae) parasitando espécies de Astyanax (Characiformes: Characidae) no sul do Brasil

Abstract: The acanthocephalan species Quadrigyrus torquatus Van Cleave, 1920 has been found in different hosts from South America, but recent papers have not focused on its morphology traits. Different species of Astyanax Baird & Girard, 1854 were collected with seine nets between January 2017 and October 2017 in Pintada Island, Lake Guaíba, Porto Alegre city (30º17'11''S and 51º18'01"W), Rio Grande do Sul State, Brazil. Acanthocephalans found in cysts were processed according to the techniques for the group. Specimens of Q. torquatus were identified based mainly in the morphology of proboscis hooks and trunk spines, and by the measurements made using light microscopy. Supplemental observations on the morphology of Q. torquatus were made for the first time considering the morphology and amount of hooks and spines. The amplitude of intensity of infections was 1 (A. aff. fasciatus) and 1‒3 (A. lacustris) helminths. This difference could be related to the number of hosts examined, the feeding habit of the hosts or related to aspects in the parasite life cycle. This paper is the first report of Q. torquatus in A. aff. fasciatus and A. lacustris from Lake Guaíba, extending the distribution to the southernmost state of Brazil, thus contributing to the knowledge of acanthocephalans in freshwater fishes from South America.

Resumo: A espécie de acantocéfalo Quadrigyrus torquatus Van Cleave, 1920, tem sido encontrada em diferentes hospedeiros na América do Sul, contudo os estudos recentes não focam no estudo dos caracteres morfológicos. Diferentes espécies de Astyanax Baird & Girard, 1854 foram coletadas com redes do tipo picaré entre janeiro e outubro de 2017, na Ilha da Pintada, Lago Guaíba, Cidade de Porto Alegre (30º17'11''S e 51º18'01"W), Estado do Rio Grande do Sul, Brasil. Os acantocéfalos encontrados encistados foram processados de acordo com as técnicas para o grupo. Os espécimes de Q. torquatus foram identificados principalmente pela morfologia dos ganchos da probóscide, espinhos do tronco e, pelas medidas realizadas através de microscopia de luz. Observações suplementares na morfologia de Q. torquatus foram realizadas pela primeira vez, considerando a morfologia e o número de ganchos e espinhos. A amplitude das infecções foi 1 (A. aff. fasciatus) e 1‒3 (A. lacustris) helmintos. Esta diferença pode estar associada com o número de hospedeiros examinados, com os hábitos alimentares dos hospedeiros, ou relacionada com aspectos no ciclo de vida do parasito. Este estudo é o primeiro registro de Q. torquatus em A. aff. fasciatus e A. lacustris do Lago Guaíba, estendendo a distribuição do parasito para o sul do Brasil, contribuindo com o conhecimento dos acantocéfalos parasitos de peixes de água doce na América do Sul.

Description of Two New Species of the Genus Vorticella (Ciliophora: Peritrichia) Epibionts on Pomacea canaliculata (Mollusca: Ampullariidae: Gastropoda) in Southern Brazil.

The peritrich ciliates Vorticella veloxiiforme n.sp. and Vorticella ampullaria n.sp. were found as epibionts on the ampullarid mollusk Pomacea canaliculata collected from Patos Lagoon, Rio Grande do Sul state, Brazil. The detailed morphology of both species was investigated using live and stained specimens, as well as scanning electron microscopy. Zooids of Vorticella veloxiiforme are cup shaped measuring 57 X 41 µm on average. A J-shaped macronucleus lies in the upper half of the cell. One contractile vacuole was observed in the upper part of the cell. V. ampullaria presents an elongate zooid measuring 70 X 25 µm on average. A C-shaped macronucleus lies in the middle of the cell. Two contractile vacuoles are present: one close to the peristome and the other in the lower half of the cell. The pattern of oral polykinetids was typical of the genus, with all infundibular polykinetids composed by three rows of kinetosomes each. SEM revealed a regular pattern of pores present on the zooid membrane of both species. Analyses of the 18s rDNA placed the two new species in the order Vorticellida close to described species of Vorticella.

Characterization of ciliate diversity in bromeliad tank waters from the Brazilian Atlantic Forest.

Bromeliads are a diverse group of plants that includes many species whose individuals are capable of retaining water, forming habitats called phytotelmata. These habitats harbor a diversity of organisms including prokaryotes, unicellular eukaryotes, metazoans, and fungi. Among single-celled eukaryotic organisms, ciliates are generally the most abundant. In the present study, we used Illumina DNA sequencing to survey the eukaryotic communities, especially ciliates, inhabiting the tanks of the bromeliads Aechmea gamosepala and Vriesea platynema in the Atlantic Forest of southern Brazil. Filtered sequences were clustered into distinct OTUs using a 99% identity threshold, and then assigned to phylum and genus using a BLAST-based approach (implemented in QIIME) and the SILVA reference database. Both bromeliad species harbored very diverse eukaryotic communities, with Arthropoda and Ciliophora showing the highest abundance (as estimated by the number of sequence reads). The ciliate genus Tetrahymena was the most abundant among single-celled organisms, followed by apicomplexan gregarines and the ciliate genus Glaucoma. Another interesting finding was the presence and high abundance of Trypanosoma in these bromeliad tanks, demonstrating their occurrence in this type of environment. The results presented here demonstrate a hidden diversity of eukaryotes in bromeliad tank waters, opening up new avenues for their in-depth characterization.

Epistylis portoalegrensis n. sp. (Ciliophora, Peritrichia): A New Freshwater Ciliate Species from Southern Brazil.

The peritrich ciliate Epistylis portoalegrensis n. sp. was found in two bodies of freshwater located in Porto Alegre, Southern Brazil. Morphological features were investigated using live and protargol-stained specimens. The zooids presented a vase to cylindrical shape narrowed at the scopula, and a mean size of 131 × 37 µm in vivo. A C-shaped macronucleus lay in the middle of the cell close to a single contractile vacuole. The oral infraciliature was typical for the genus, with all infundibular polykineties composed by three distinct rows of kinetosomes. Colonies are often nonbranched with no lateral stalk, carrying several zooids stemming from a single point. Specimens from the two sampling sites showed identical arrangement of the infraciliature, similar morphometry, identical 18S rDNA sequences, and a single nucleotide difference across the more variable ITS regions. Molecular phylogenetic analyses placed E. portoalegrensis in a well-supported clade containing other Epistylis species, within the order Vorticellida.

Epistylis smalli (Ciliophora: Peritrichia) a new peritrich from Guaiba Lake, Southern Brazil.

Epistylis smalli n. sp., a freshwater colonial peritrich, was collected in Guaíba Lake, Southern Brazil. Its morphology was investigated using in vivo observations and protargol stained specimens. E. smalli possess an elongate zooid that measures in vivo, on average, 173 µm in length and 50 µm in width. A C-shaped macronucleus that surrounds the infundibulum and a single contractile vacuole could be easily observed in the living cell. The oral infraciliature observed in silver-stained specimens was typical of peritrich ciliates, with three infundibular polykinetids bearing three rows of kinetosomes. A detailed description of the live and stained zooids is given.

Effects of Seasonality and Dispersal on the Ciliate Community Inhabiting Bromeliad Phytotelmata in Riparian Vegetation of a Large Tropical River.

This study evaluated the influence of rainfall amount on the abundance, species richness, and species occurrence and abundance distribution of the ciliate community associated with the bromeliad Aechmea distichantha. The plants were collected from a rock wall of about 10-km long at the left bank of Paraná River. We assessed the effects of both spatial and temporal variables on the community attributes, as well as whether plants geographically closer have a similar abundance distribution and species composition. The ciliate community was substantially distinct between both hydrological periods, with greater values of species richness and abundance in the rainy period. No spatial structuring (differences in the species occurrence and abundance distribution among strata) or geographical similarity (similarity in ciliate species composition among the plants) was found. Multiple regression analysis showed a positive relationship only between the ciliate abundances and water volumes for both periods. Although few of the formulated predictions were confirmed, our study provides valuable information on the ecological aspects of the ciliate community inhabiting bromeliad phytotelmata.

Description of Epistylis riograndensis n.sp. (Ciliophora: Peritrichia) found in an artificial lake in Southern Brazil.

Epistylis riograndensis n. sp., a freshwater peritrich hosting symbiotic algae in its cytoplasm, was collected from an artificial lake, in a Botanical garden in Southern Brazil. Its detailed morphology was investigated using live and silver-stained specimens. The colonial sessile E. riograndensis has elongate zooids measuring, on average, 162 µm in length and 45 µm in width. A single contractile vacuole located near the infundibulum and a C-shaped macronucleus located transversely in the adoral half of the cell were also observed. The oral infraciliature revealed in silver-stained specimens was typical of peritrich ciliates. Three infundibular polykineties consisting of 3 rows of kinetosomes were observed. Molecular analyses of 18s rDNA placed E. riograndensis among other Epistylis species in the Order Vorticellida.

Multiple lines of evidence shed light on the occurrence of paramecium (ciliophora, oligohymenophorea) in bromeliad tank water.

Phytotelmata are vegetal structures that hold water from the rain, and organic matter from the forest and the soil, resulting in small, compartmentalized bodies of water, which provide an essential environment for the establishment and development of many organisms. These microenvironments generally harbor endemic species, but many organisms that are found in lakes and rivers, are also present. Here, we report, for the first time, the occurrence of the ciliate genus Paramecium in the tank of the bromeliad species Aechmaea distichantha. The identification of the Paramecium species was performed based on live observations, protargol impregnation, scanning electronic microscopy, and sequencing of the 18s rRNA. The absence of Paramecium from bromeliad tank water was highlighted in several earlier investigations, and may be due to the fact that this species is unable to make cysts. The occurrence of Paramecium multimicronucleatum in our samples may be explained by the proximity between the bromeliads and the river, a potential source of the species. Further, we also believe that the counting methodology used in our study provides a more accurate analysis of the species diversity, since we investigated all samples within a maximum period of 6 h after sampling, allowing minimum loss of specimens.

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.

Morphology and morphogenesis of a freshwater ciliate, Epistylis chlorelligerum Shen, 1980 (Ciliophora, Peritrichia).

An oligohalobic peritrichous ciliate, Epistylis chlorelligerum Shen, 1980, was collected from a ditch in Hangzhou, China. The morphology, oral infraciliature, and morphogenesis of the species were studied using living and protargol-impregnated specimens. Zooids of E. chlorelligerum are 160-230 × 50-60 µm in vivo, and characterized by green-colored endoplasm containing symbiotic algae. The oral infraciliature presents a well-developed filamentous reticulum linked to the circular fiber of the cytostome; the outer two rows of P3 extend adstomally over P1 and usually enfold it. During binary fission, one daughter cell inherits most part of the old buccal apparatus and the reorganized haplokinety and germinal kinety (Hk' and G'), and new buccal apparatus of the other daughter cell is mostly developed from the original germinal kinety (G) and haplokinety (Hk): new peniculi 2, 3 (2P2, 2P3), new haplokinety (2Hk), and new germinal kinety (2G) are formed from G, while the new peniculus 1 (2P1) and its peristomial extention (2Pk) originate from Hk. The epistomial membrane can be observed until the two sets of buccal apparatus begin to separate from each other.

Expanded phylogenetic representation of genera Opercularia and Epistylis sheds light on the evolution and higher-level taxonomy of peritrich ciliates (Ciliophora: Peritrichia).

We have generated 18S rRNA sequences for peritrichs collected in Brazil, including four Opercularia species, two different populations of Epistylis plicatilis (one epibiont and another free-living), and one additional Epistylis species. Our Opercularia species clustered with the previously available Opercularia microdiscum, corroborating the monophyly of this genus. The Epistylis sampled here clustered with previously sequenced species of this genus. The two populations of E. plicatilis collected in Brazil clustered closely together despite their different ecological contexts, whereas both were very divergent from the sample assigned to the same species previously sampled in China. If affirmed by additional morphological corroboration of species assignment, this observation would indicate that samples from different continents morphologically allocated in the same species may in fact belong to distant evolutionary lineages. More broadly, our results support the recognition of two major clades within Peritrichia. Given the robustness of their support, we suggest that these two clades should be formally recognized as orders, and propose the names Vorticellida and Operculariida to designate them. Furthermore, Epistylis species occurred in both orders, tending to occupy basal positions. This suggests that characters used to define this genus may be plesiomorphic for Peritrichia, so that Epistylis may in fact represent an assemblage of basal species retaining ancestral features.

Morphological characterization and intraspecific variation of Zoothamnium intermedium Precht, 1935 (Ciliophora, Peritrichia) attached to calanoid copepods in the Chesapeake Bay, USA.

A redescription of Zoothamnium intermedium Precht, 1935, a peritrich epibiont on copepods, is provided using specimens colonizing Acartia tonsa and Eurytemora affinis in Chesapeake Bay, USA. Bell-shaped zooids of Z. intermedium ranged in size from 31.2-54.7 microm x 16.7-31.3 microm in vivo. A single contractile vacuole and a "C shaped" macronucleus lie in the upper half of the body. Colonies had up to 30 zooids, but most presented two to four zooids. The single myoneme was continuous from the zooids through the lateral branches and basal stalk, terminating 4-73.2 mm before the attachment point of the colony. The ciliature of Z. intermedium was typical of sessile peritrichs, consisting of an outer haplokinety and an inner polykinety 1 (PK1) that made approximately turns around the peristomial disk before entering the infundibulum. Within the infundibulum, PK 1 was accompanied by polykinetid 2 (PK2) and polykinetid 3 (PK3), each consisting of three kinetosomal rows: PK2 of Z. intermedium terminated adoral to the aboral end of PK1 and had a central row of kinetosomes shorter than the lateral rows. Scanning electron microscopy revealed an annular pattern of pellicular ridges with pellicular pores. Morphological features used in peritrich species identification are quantified for specimens attached to both hosts and compared with other species of Zoothamnium. Statistically significant differences found between specimens attached to A. tonsa and E. affinis may reflect phenotypic plasticity rather than infestation by multiple species of Zoothamnium.

Molecular phylogenetics of subclass Peritrichia (Ciliophora: Oligohymenophorea) based on expanded analyses of 18S rRNA sequences.

Phylogenetic relationships among peritrich ciliates remain unclear in spite of recent progress. To expand the analyses performed in previous studies, and to statistically test hypotheses of monophyly, we analyzed a broad sample of 18s rRNA sequences (including 15 peritrich genera), applying a conservative alignment strategy and several phylogenetic approaches. The main results are that: (i) the monophyly of Peritrichia cannot be rejected; (ii) the two main clades of Sessilida do not correspond to formally recognized taxa; (iii) the monophyly of genera Vorticella and Epistylis is significantly rejected; and (iv) morphological structures commonly used in peritrich taxonomy may be evolutionarily labile.

Spatial and temporal patterns in the occurrence of peritrich ciliates as epibionts on calanoid copepods in the Chesapeake Bay, USA.

We investigated temporal and spatial patterns of distribution in two peritrich ciliates (i.e. Zoothamnium intermedium and Epistylis sp.) living as epibionts on calanoid copepods (i.e. Acartia tonsa and Eurytemora affinis) in Chesapeake Bay. Net tow samples collected along the main axis of the Bay were analyzed to estimate the occurrence of epibionts on copepods and to explore relationships among infestation prevalence, host abundance, and environmental variables. Zoothamnium intermedium and Epistylis sp. colonized populations of A. tonsa during spring and summer months, while only Z. intermedium colonized E. affinis during spring. Occurrence of epibionts on copepods showed high interannual variation, marked seasonality, and geographic heterogeneity. Extensive statistical analyses rejected simple scenarios of interactions between epibiosis, environmental variables, and host density, suggesting a more complex dynamics for the system. Analyses of epibiont colonies and zooids per host area (i.e. the sum of width and length of the body including antennae and swimming legs calculated assuming a cylindrical shape) were also performed. Overall, epibiont infestation prevalence (i.e. colonies/host area) and load (i.e. zooids/host area) were higher on copepodites than on adults for both host species, suggesting a preferential attachment to juveniles, or a higher predation pressure on adult stages. Infestation density and loads of both epibiont species were higher on the cephalothorax and abdomen of A. tonsa and E. affinis in comparison to the antennae and swimming legs, suggesting that ciliates can more easily colonize less active parts of the host.