The mitochondrial citrate synthase (mCS) purified from the ciliate Tetrahymena thermophila has been reported to form intermediate-filament-like structures during conjugation and to self-assemble into fibers when recombinantly expressed. This would represent a rare example of a tractable and recent origin of a novel cytoskeletal element. In an attempt to investigate the evolutionary emergence of this behavior, we re-investigated the ability of Tetrahymena's mCS to form filaments in vivo. Using strep-tagged mCS in Tetrahymena and monoclonal antibodies, we found no evidence of filamentous structures during conjugation or starvation. Extensive biochemical characterization of mCS revealed that the self-assembly of recombinant protein is triggered by a specific chemical moiety shared by MES and HEPES buffers used in previous studies. The absence of indicative phenotypes in fiber-deficient GFP-tagged mutants indicates that Tetrahymena mCS did not evolve a structural role in sexual reproduction or metabolic regulation.

The paper describes a new myxomycete species, Trichia tuberculata, from the Ecuadorian cloud forest. The phylogeny constructed with nuclear 18S rDNA and mitochondrial 17S rDNA sequences indicates that the taxon is closely related to recently described species T. acetocorticola, T. nubila, and T. pinicola, all within the T. botrytis complex. The morphology of the inner peridium surface and capillitium also aligns T. tuberculata with these species. However, the distinctive tuberculate peridium, short elater tips, and perhaps the distribution range (so far found in the equatorial zone only) distinguish T. tuberculata from related taxa.

Ciliates of the class Prostomatea show a broad spectrum of feeding strategies and often occur abundantly in various aquatic habitats, playing a vital role in the biogeochemical cycle. Due to their small cell size and simple structure, prostomateans were considered to be a group with a lower degree of morphological differentiation for a long time. However, recent research suggests that the diversity of this group of ciliates is higher than previously thought. In the present study, three prostomateans, collected from different localities in China and classified into three families, were examined using morphological and phylogenetic techniques. Our analyses revealed two new species, Metacystis multitricha sp. nov. and Apolagynus spiralis sp. nov., and suggested a transfer of Prorodon abietum Penard, 1922 to the genus Pelagothrix. Phylogenetic analyses corroborated the morphological classifications of the three species into the families Metacystidae, Lagynusidae, and Holophryidae, respectively, demonstrating that their diagnostic characteristics bear an evolutionary signal at the family level.

Rumen ciliates constitute a highly diverse group within the subclass Trichostomatia. They are known for their symbiotic relationship with ruminant hosts and their pivotal role in digestive metabolism. This review presents a historical analysis of research on rumen ciliate metabolism, since the earliest studies, based on in vitro and in situ experiments, as well as molecular studies, initially relying on Sanger sequencing, and more recently, next-generation sequencing techniques. Finally, the paper discusses future approaches that may be useful for elucidating the metabolic profile of various taxa of rumen ciliates.

The Kingdom of Bhutan in the Eastern Himalayas harbors a rich biodiversity. However, its detailed documentation still needs to be created, including not only macroorganisms but also protists. In the present study, as many as 105 taxa of testate amoebae were identified and directly counted in soil suspension samples collected from the wetlands of the Gangtey-Phobji Valley (generally Phobjikha), Wangdue Phodrang district. The most dominant were cosmopolitan species, e.g., Euglypha rotunda, Trinema lineare, T. complanatum, T. enchelys, Phryganella acropodia, and Plagiopyxis declivis. However, some taxa such as Centropyxis deflandriana, C. stenodeflandriana, Hoogenraadia cf. humicola, Pareuglypha reticulata, and Sphenoderia chardezi have a geographically restricted distribution and/or are considered rare. Some specimens could not be assigned to already described species (Centropyxis dentata sp. nov. and Difflugia fusiforma sp. nov.) or were highly similar to known species but differed in size or other taxonomically important characteristics (e.g., Hoogenraadia cf. humicola and Planhoogenraadia sp.). To our knowledge, this study is the first to present a comprehensive list of testate amoebae from Bhutan, including some new and rare species. The discovery of a remarkable diversity of testate amoebae in only a few soil samples suggests that this remote and largely unexplored region likely harbors many more intriguing species.

A new urostylid ciliate, Holostichides (Holostichides) parachardezi, collected from a Chinese tropical city, was studied using morphological and molecular methods. Diagnostic features of the new species include: size 140-180 × 30-60 μm in vivo; 25-43 macronuclear nodules; colourless cortical granules; three to six frontoterminal cirri; a midventral complex composed of seven to eleven cirral pairs and one midventral row of five to eleven cirri; four bipolar dorsal kineties and six to nine caudal cirri in four groups; and a terrestrial habitat. Its ontogenesis follows the typical Holostichides mode in that (1) the proximal portion of the parental adoral zone of membranelles is partly renewed and the opisthe's oral primordium originates very close to the midventral complex; (2) the last anlage n generates the frontoterminal cirral row, and the midventral complex is formed from anlage IV to n - 1; and (3) both marginal rows and dorsal kinety anlagen develop within the parental rows. The present SSU rDNA phylogenies corroborated the monophyly of the genus Holostichides.

The fine-structural organization of the protoperidiniacean Kolkwitziella acuta was examined by SEM and TEM. Serial sections of five cells of K. acuta were used to study the architecture of basal bodies and associated roots, the pusular system, and the feeding apparatus. The basal bodies were inserted 1 µm apart at an angle of ca. 80°, and displayed the typical peridinioid features of associating with two roots each and having a layered connective linking the longitudinal microtubular root to the transverse striated root. The transverse flagellar canal was associated with a 'sac pusule', while the longitudinal flagellar canal was linked, via a pusule canal surrounded by a conspicuous layer of striated material, to a collecting chamber from which ca. 40 pusular tubes radiated. An extruded pallium was present, associated with a microtubular strand (the MSP) that extended anteriorly and progressively separated into six or seven groups of microtubules, with electron-opaque vesicles present along the MSP. A prominent striated collar surrounded the exit area of the pallium and was connected to the striated collars of both the transverse and longitudinal flagellar canals. The partial nuclear-encoded LSU rDNA sequence confirmed the identification as K. acuta. The currently known fine-structural features of pallium-feeding Protoperidiniaceae are summarized.

Short-branch Microsporidia were previously shown to form a basal grade within the expanded Microsporidia clade and to branch near the classical, long-branch Microsporidia. Although they share simpler versions of some morphological characteristics, they do not show accelerated evolutionary rates, making them ideal candidates to study the evolutionary trajectories that have led to long-branch microsporidian unique characteristics. However, most sequences assigned to the short-branch Microsporidia are undescribed, novel environmental lineages for which the identification requires knowledge of where they can be found. To direct future isolation, we used the EukBank database of the global UniEuk initiative that contains the majority of the publicly available environmental V4 SSU rRNA gene sequences of protists. The curated OTU table and corresponding metadata were used to evaluate the occurrence of short-branch Microsporidia across freshwater, hypersaline, marine benthic, marine pelagic, and terrestrial environments. Presence-absence analyses infer that short-branch Microsporidia are most abundant in freshwater and terrestrial environments, and alpha- and beta-diversity measures indicate that focusing our sampling effort on these two environments would cover a large part of their overall diversity. These results can be used to coordinate future isolation and sampling campaigns to better understand the enigmatic evolution of microsporidians' unique characteristics.

Small chrysomonads are important bacterivores in aquatic ecosystems with a high molecular diversity compared to low morphological differences observed by light microscopy. The high diversity of these morphologically almost indistinguishable species leads to the question to which extent their functional role in ecosystems differs and how their ecological traits can be defined. The present study investigates the prey size and population growth rate of different chrysomonad species. Eleven phylogenetically well-defined strains representing seven strains of heterotrophic and four strains of mixotrophic chrysomonads were compared. All investigated strains belonged to the same functional group of bacterivorous flagellates, feeding on the same bacteria size range, while population growth rates of chrysomonads depended on nutritional strategy and species-specific differences. We observed a high individual variability of growth rates within a population. Our results point to the necessity to consider not only differences in ecological traits among species but also among specimens within a population.

Catalases (CATs) are ubiquitous antioxidant enzymes that prevent cellular oxidative damage through the decomposition of HO. However, there is relatively little information on CAT in the worldwide-distributed freshwater green alga Closterium ehrenbergii. Here, we cloned the full-length catalase cDNA from C. ehrenbergii (CeCAT) and characterized its structural features and expressional responses against aquatic contaminants. The open reading frame of CeCAT was determined to be 1476 bp, encoding 491 amino acids with a theoretical molecular mass of 56.1 kDa. The CeCAT protein belongs to the NADPH-binding CAT family and might be located in the cytosol. BLAST and phylogenetic results showed that CeCAT had a high identity with CAT proteins from other microalgae and the water lily Nymphaea colorata (Streptophyta). The transcriptional levels of CeCAT were significantly upregulated by the metal copper and herbicide atrazine, but little affected by other tested metals (Ni and Cr) and endocrine-disrupting chemicals (polychlorinated biphenyl, PCB). The maximum expression was registered under 0.1 mg/L CuCl and 0.2 mg/L CuSO exposures. In addition, excess copper considerably increased production of reactive oxygen species in the cells. These results suggest that CeCAT may function to defend against oxidative stress in green algae and can respond specifically to different kinds of metals and herbicides.

Euplotes baugilensis n. sp. was discovered in a temporary puddle that formed after rainfall on a mountain footpath near Gangneung-Wonju National University in Gangneung, South Korea. After isolation, a pure culture was established, and the new species was examined using live observation, silver-impregnation (protargol and 'wet' silver nitrate), scanning electron microscopy, and the analysis of the 18S rRNA gene sequence. Morphologically, E. baugilensis n. sp. is characterized by small body size (on average 49 × 31 µm in vivo), 9 ordinary fronto-ventral cirri (cirrotype-9) with one reduced cirrus V/2 (composed of four non-ciliated basal bodies), 5 transverse cirri, 7 or 8 dorsolateral kineties, 6 dorsal prominent ridges, and a dargyrome (silverline system) of double type. In this study, we have used a combination of morphological and molecular techniques to characterize E. baugilensis n. sp. and determine its phylogenetic position within the genus Euplotes. Molecular analysis using 18S rRNA gene sequences indicated that E. baugilensis n. sp. is most closely related to E. curdsi (with a sequence identity of 96.8 %).

This study investigated the dynamics of reciprocal phenotypic plasticity entailing inducible defense and offense in freshwater ciliate communities in response to altered resource supply and the extent of intraspecific trait variation. Communities consisted of Euplotes octocarinatus (intraguild prey) capable of inducible defense to escape predation, Stylonychia mytilus (intraguild predator) capable of inducible offense to expand its prey spectrum, and Cryptomonas sp. (algal resource). The extent of inducible defense was tested in ten different Euplotes strains in response to freeze-killed Stylonychia concentrate, revealing significant differences in their width and length development. In a subsequent 30-day experiment, four strains were incubated in monoculture and mixture with Stylonychia under continuous and pulsed microalgae supply. The polyclonal Euplotes population outperformed the monoclonal populations, except one, which developed the most pronounced inducible defense and retained the highest biovolume. Stylonychia fluctuated in size, but dominated all communities irrespective of clonal composition. Pulsed resource supply promoted biovolume production of both species. However, periods of resource depletion resulted in more Stylonychia resting cysts, allowing Euplotes to resume growth. Our study provides new insights into interactions of induced defense and intraguild predation under variable environmental conditions, emphasizing the relevance of intraspecific trait variation for predator-prey interactions and community dynamics.

After Gulielmina was erected and Ophiotheca was resurrected based on some species originally included in Perichaena (Trichiaceae, Trichiales, Myxomyxetes), some specimens from the Herbarium of Fungi of Nanjing Normal University previously identified as Perichaena species were reexamined from morphological and two-gene (nuclear 18S rDNA and elongation factor-1 alpha) phylogenetic perspectives. In this study, two new myxomycete species, Gulielmina subreticulospora and Ophiotheca dictyospora, are described. Gulielmina subreticulospora shows the following character combination: branched plasmodiocarps, single peridium with circular protrusions in the inner surface, capillitium (2.4-)2.8-3.0(-3.4) μm in diameter, spores (7.4-)8.0-8.5(-9.0) μm in diameter and sub-reticulated. Ophiotheca dictyospora shows the following character combination: sessile sporocarps to short plasmodiocarps, single peridium with a densely irregular network and protrusions in the inner surface, capillitium (2.7-)3.5-5.0(-7.1) μm in diameter, uneven, decorated with spines of uneven size, spores (7.7-)8.2-8.6(-9.4) μm in diameter including obviously complete cristate reticulation with serrated edges, with deep and clear grids. Both new taxa were compared with related species and their genetic isolation was statistically tested. Moreover, a comprehensive morphological description and a detailed figure plate are provided for Perichaena verrucifera, and its phylogenetic position is determined.

The phylogeny of brown algae (Phaeophyceae) has undergone extensive changes in the recent past due to regular new scientific insights. We used nuclear 18S rDNA with an extensive dataset, aiming to increase the accuracy and robustness of the reconstructed phylogenetic trees using a simultaneous sequence-structure approach. Individual secondary structures were generated for all 18S rDNA sequences. The sequence-structure information was encoded and used for an automated simultaneous sequence-structure alignment. Neighbor-joining and profile neighbor-joining trees were calculated based on 186 phaeophycean sequence-structure pairs. Additionally, sequence-structure neighbor-joining, maximum parsimony and maximum likelihood trees were reconstructed on a representative subset. Using a similar approach, ITS2 rDNA sequence-structure information was used to reconstruct a neighbor-joining tree including 604 sequence-structure pairs of the Laminariales. Our study results are in significant agreement with previous single marker 18S and ITS2 rDNA analyses. Moreover, the 18S results are in wide agreement with recent multi-marker analyses. The bootstrap support was significantly higher for our sequence-structure analysis in comparison to sequence-only analyses in this study and the available literature. This study supports the simultaneous inclusion of sequence-structure data at least for 18S to obtain more accurate and robust phylogenetic trees compared to sequence-only analyses.

Protists can endure challenging environments sustaining key ecosystem processes of the microbial food webs even under aridic or hypersaline conditions. We studied the diversity of protists at different latitudes of the Atacama Desert by massive sequencing of the hypervariable region V9 of the 18S rRNA gene from soils and microbial mats collected in the Andes. The main protist groups in soils detected in active stage through cDNA were cercozoans, ciliates, and kinetoplastids, while the diversity of protists was higher including diatoms and amoebae in the microbial mat detected solely through DNA. Co-occurrence networks from soils indicated similar assemblages dominated by amplicon sequence variants (ASVs) identified as Rhogostoma, Euplotes, and Neobodo. Microbial mat networks, on the other hand, were structured by ASVs classified as raphid-pennate diatoms and amoebae from the genera Hartmannella and Vannella, mostly negatively correlated to flagellates and microalgae. Additionally, our phylogenetic inferences of ASVs classified as Euplotes, Neobodo, and Rhogostoma were supported by sequence data of strains isolated during this study. Our results represent the first snapshot of the diversity patterns of culturable and unculturable protists and putative keystone taxa detected at remote habitats from the Atacama Desert.

Cyphoderia compressa has only been described from supralittoral environments, as a psammobiont, with salinities from 1.33 to 36.00 ‰. Other Cyphoderia species such as those in the C. ampulla species complex are more ecologically and environmentally widespread, occurring as free-living individuals within water bodies or in association with vegetation, and over a wider salinity range, including freshwater. We postulate that C. compressa may not be as restricted in terms of its environmental or ecological distribution. To this end, we examined a variety of water and sediment samples from Scottish localities, ranging from supralittoral to inland freshwater environments. The Scottish material occurs as a psammobiont within supralittoral beach sands and is newly recorded within sands from freshwater to brackish stream sections and along the margin of the freshwater Loch Lomond. It is also recorded from freshwater to brackish settings as part of the stream and pond water biota, associated biofilm and vegetative material. Test morphology is more variable than previously appreciated, including those with a papillate fundus, and many that are not as laterally compressed as typified by the species. Differences in plate morphology and size were also noted, as well as a novel arcuate cross-cutting ridged cement structure, that is restricted to C. compressa.

The morphology and molecular phylogeny of a new ciliate, Conchophthirus sinanodontae n. sp., which was discovered in the freshwater mussel Sinanodonta woodiana (Lea, 1834) from the Chilsancheon River, Buyeo-gun, South Korea, were investigated. The new species was characterized and could be distinguished from congeners by a combination of characters including the ovate body outline, four to six oral polykinetids deeply embedded in the upper wall of the buccal cavity, six to ten vestibular kineties, 34-49 ventral and 36-53 dorsal somatic kineties. The genetic differences among C. sinanodontae n. sp. and other congeners with available 18S rDNA sequences further support its distinctness. Moreover, the phylogenetic analyses based on the 18S rDNA sequences show that the new species clusters with other congeners, corroborating the monophyly of the genus Conchophthirus. The Conchophthirus clade nests within the cluster of Dexiotricha spp., Loxocephalus luridus, and Haptophrya spp.

Rustic aquaculture systems represent a sustainable production alternative in the world, providing appropriate conditions for the establishment of epibiotic ciliates on crayfish. However, the impact of epibiotic ciliates on the aquaculture production has been insufficiently studied hitherto. The objective of this work was to identify the epibiotic ciliates on the surface of the crayfish Procambarus (Austrocambarus) sp. and analyze whether there is any correlation between seasonality and environmental conditions in an artificial pond in Chiapas, Mexico. The crayfish were collected during dry and rainy seasons to investigate their ciliate epibionts. Some physical and chemical parameters of the sampled water were measured and correlation matrix analyses were applied to detect possible relationships between the presence of ciliates and environmental parameters. Thirteen species belonging to the subclasses Peritrichia and Suctoria were identified. Significant differences were found for some environmental parameters and species recorded in both seasons. The present data could help to better understand the relationship bewteen the epibiotic ciliate community on crayfish and some environmental parameters in aquaculture systems.

The ciliate Paramecium bursaria possesses cell organelles called trichocysts that have defensive functions. Paramecium bursaria is capable of symbiosis with Chlorella variabilis, and the symbiotic algae are situated in close proximity to the trichocysts. To clarify the relationship between trichocysts in P. bursaria and the presence or absence of the intracellular symbiotic C. variabilis, this study compared the regeneration capacity of trichocysts in alga-free and algae-bearing P. bursaria. In addition, trichocyst protein abundance was measured when alga-free P. bursaria specimens were artificially infected with Chlorella. After completely removing trichocysts from P. bursaria cells by treatment with lysozyme and observing them after 24 h, the percentage of regenerating trichocysts in the entire cell was significantly higher in alga-free cells than that in algae-bearing cells. We also developed a simple method for the isolation of high-purity trichocysts to quantify trichocyst protein amounts. There was a significant difference in the trichocyst protein abundance of P. bursaria before and one week after mixing with Chlorella (i.e., after the establishment of symbiosis with algae). This study shows the importance of trichocysts in alga-free P. bursaria as well as their competition with symbiotic C. variabilis for attachment sites during the algal infection process.

The subclass Trichostomatia (Ciliophora, Litostomatea) constitutes a well-supported monophyletic group, which includes ciliates exclusively found as symbionts of vertebrates, primarily herbivorous mammals. Recent molecular analyses reinforce the subclass monophyly, though almost all orders, suborders, families, and genera are found to be non-monophyletic. Here, we reconstructed the evolutionary history of the subclass Trichostomatia using a phylogenomic approach and discussed some systematic inconsistencies. We propose a new Ophryoscolecidae genus, Dagostonium, to include Diplodinium polygonale. Monoposthium cynodontum is transferred to the genus Cycloposthium.

The function of recirculating aquaculture systems (RAS) relies on microbial communities, which convert toxic, fish-excreted ammonia into substances that can provide nutrients to plants as in the case of aquaponics systems. In the present study, heterotrophic protist communities of experimental sea water RAS and freshwater aquaponics systems were investigated using microscopy to characterize their diversity, natural abundance, and potential growth rates. Heterotrophic protist abundance was low (732 ± 21 to 5451 ± 118 ciliates L and 58 ± 8 to 147 ± 18 nanoflagellates mL in the aquaponics system and 78 ± 28 to 203 ± 48 ciliates L in the RAS), which is in line with values typically reported for rivers. In the aquaponics system, ciliates grew faster in the fish rearing tanks (1.9 ± 0.01 to 1.21 ± 0.03 d compared to 0.54 ± 0.03 to 0.79 ± 0.05 d in the other compartments), while heterotrophic nanoflagellates grew slower in drain tanks downstream of the hydroponics compartment (0.5 ± 0.3 to 1.37 ± 0.05 d and 4.09 ± 0.11 d to 6.03 ± 0.34 din the other compartments). Results indicated distinct niches and reduced microeukaryotic diversity at the end of the system's operation cycle.