SSU rRNA Gene Sequences Research Articles

Pseudodesulfovibrio thermohalotolerans sp. nov., a novel obligately anaerobic, halotolerant, thermotolerant, and sulfate-reducing bacterium isolated from a western offshore hydrocarbon reservoir in India

ObjectiveCharacterization and documentation of strain MCM B-1480T, a novel sulfate-reducing bacterium isolated from produced water of India's western offshore hydrocarbon reservoir. MethodStrain MCM B-1480T was unequivocally identified using a polyphasic approach routinely followed in bacterial systematics. The morphological and biochemical characterization of strain MCM B-1480T was carried out using standard microbiological techniques. ResultsMCM B-1480T was a Gram-stain-negative, motile, non-spore-forming, curved-rod-shaped bacterium. MCM B-1480T could grow at temperatures between 20 and 60 °C (optimum 37 °C), pH 6–8 (optimum 7), and required 1–6% NaCl (optimum 3%) for growth. Strain MCM B-1480T was reducing sulfate to produce hydrogen sulfide during growth. This strain used lactate and pyruvate as prominent electron donors, whereas sulfate, sulfite, thiosulfate, and nitrate served as electron acceptors. MCM B-1480T shared maximum 16S rRNA gene sequence homology of 98.65% with the members of the genus Pseudodesulfovibrio. The G + C content of the 3.87 Mb MCM B-1480T genome was 60.39%. Digital DDH (27.7%) and average nucleotide identity (ANI 84%) with the closest phylogenetic affiliate (less than 70% and 95%, respectively) reaffirmed its distinctiveness. The major cellular fatty acids components, namely iso-C15:0, anteiso-C15:0, C16:0, and anteiso-C17:0, differentiated strain MCM B-1480T from other species of Pseudodesulfovibrio. Genome annotation revealed the presence of genes encoding dissimilatory sulfate reduction and nitrate reduction in strain MCM B-1480T. ConclusionThe polyphasic studies, including SSU rRNA gene sequencing, average nucleotide identity, Digital DNA-DNA hybridization, cell wall fatty acids analysis, etc., identified strain MCM B-1480T as a novel taxon and Pseudodesulfovibrio thermohalotolerans sp. nov. was proposed (= JCM 39269T = MCC 4711T).

Three new species of Gromia (Protista, Rhizaria) identified from the Romanian Black Sea shelf

The protist genus Gromia was first described in 1835 by Dujardin and while gromiids are prominent in the marine environment, Gromia oviformis was, for a long time, the only valid species regularly recorded. To date, 16 species that are morphologically and/or genetically distinct have been described. While recent studies are documenting their diversity and their ecological importance, G. oviformis has been the sole gromiid species identified in the Black Sea, although unnamed Gromia species have also been recorded. We collected sediment samples from the Romanian continental shelf at varying depths (48 – 58 m) to study the morphological and genetic diversity of gromiids in this part of the Black Sea. Three new species, Gromia bugnae sp. nov., Gromia dianae sp. nov. and Gromia fabi sp. nov., were identified based on an integrative taxonomic approach, thus bringing the total described gromiid species to 19. Analysis of partial SSU rRNA gene sequences confirms that these are distinct species. Additionally, an undescribed species is represented by a sequence from the northern part of the Black Sea (Sevastopol, Kazachya Bay). The study provides further evidence of the diversity of gromiids in the Black Sea and underlines the importance of this little-known group in marginal seas.

Differential diagnosis of human Entamoeba infections: Morphological and molecular characterization of new isolates in Argentina

Entamoeba infections occur worldwide, with higher frequency in countries of low socioeconomic status and poor public health. Since Entamoeba histolytica has long been recognized as the only pathogenic species, making a differential diagnosis of other morphologically identical Entamoeba is important. This study aimed to determine the prevalence of Entamoeba species in two populations from Argentina, make a differential diagnosis by PCR and characterize Entamoeba isolates at the SSU rRNA gene. A total of 493 serial fecal samples were obtained from individuals in the provinces of Buenos Aires (n=210) and Misiones (n=283). Samples were examined by conventional methods (formalin–ethyl acetate and Willis flotation) and specific PCRs to differentiate Entamoeba species. Entamoeba isolates were characterized by sequencing a fragment of the SSU rRNA gene. The overall prevalence of Entamoeba infection was 12.4%, being more prevalent in Buenos Aires than in Misiones (14.8% vs. 10.6%). A case of E. histolytica confirmed by PCR and sequence analysis was reported for the first time in Buenos Aires. Moreover, new genetic data on Entamoeba coli and Entamoeba dispar were recorded. The phylogenetic analysis revealed a congruence between morphological characteristics and SSU rRNA gene sequences. This study increases the amount of information on the distribution of these species in Argentina and the region of the Americas.

First redescription and molecular phylogeny of Trimyema claviforme Kahl, 1933 with the description of a Chinese population of Plagiopyla nasuta Stein, 1860 (Ciliophora, Plagiopylea)

Ciliates belonging to the class Plagiopylea are obligate anaerobes that are often neglected due to their cryptic lifestyles, difficulty of observation, and overall under-sampling. Here, we investigate two species, namely Trimyema claviforme Kahl, 1933 and Plagiopyla nasuta Stein, 1860, collected in China from marine and freshwater anaerobic sediments, respectively. A complete morphological dataset, together with SSU rRNA gene sequence data were obtained and used to diagnose the species. No molecular sequencing had ever been performed on Trimyema claviforme, with its ciliature also previously unknown. Based on these novel data presented here, the ciliate is characterized by a claviform cell shape, with a size of 35–45 × 10–20 μm in vivo, 28–39 longitudinal somatic ciliary rows forming five ciliary girdles (four complete girdles and a shorter one), two dikinetids left to anterior end of oral kinety 1, and an epaulet. A Chinese population of the well-known ciliate P. nasuta was investigated, and morphological comparisons revealed phenotypic stability of the species. The phylogenetic analyses supported previous findings about the monophyly of the families Trimyemidae and Plagiopylidae, with Trimyema claviforme branching off early in the genus Trimyema. The Chinese population of P. nasuta clusters together with two other populations with full support corroborating their conspecificity.

Taxonomic contributions to Hapalidiales (Corallinophycidae, Rhodophyta): Boreolithothamnion gen. nov., Lithothamnion redefined and with three new species and Roseolithon with new combinations.

Phylogenetic analyses of rbcL gene sequences and of concatenated rbcL, psbA, and nuclear SSU rRNA gene sequences resolved the generitype of Lithothamnion, L. muelleri, in a clade with three other southern Australian species, L. kraftii sp. nov., L. saundersii sp. nov., and L. woelkerlingii sp. nov. Cold water boreal species currently classified in Lithothamnion and whose type specimens have been sequenced are transferred to Boreolithothamnion gen. nov., with B. glaciale comb. nov. as the generitype. The other species are B. giganteum comb. nov., B. phymatodeum comb. nov., and B. sonderi comb. nov., whose type specimens are newly sequenced, and B. lemoineae comb. nov., B. soriferum comb. nov., and B. tophiforme comb. nov., whose type specimens were already sequenced. Based on rbcL sequences from the type specimens of Lithothamnion crispatum, L. indicum, and L. superpositum, each is recognized as a distinct species and transferred to the recently described Roseolithon as R. crispatum comb. nov., R. indicum comb. nov., and R. superpositum com. nov., respectively. To correctly assign species to these three genera based only on morpho-anatomy, specimens must have multiporate conceptacles and some epithallial cells with flared walls. The discussion provides examples demonstrating that only with phylogenetic analyses of DNA sequences can the evolution of morpho-anatomical characters of non-geniculate corallines be understood and applied at the correct taxonomic rank. Finally, phylogenetic analyses of DNA sequences support recognition of the Hapalidiales as a distinct order characterized by having multiporate tetra/bisporangial conceptacles, and not as a suborder of Corallinales whose tetra/bisporangial conceptacles are uniporate.

New Parabasalia symbionts Snyderella spp. and Daimonympha gen. nov. from South American Rugitermes termites and the parallel evolution of a cell with a rotating "head".

Most Parabasalia are symbionts in the hindgut of "lower" (non-Termitidae) termites, where they widely vary in morphology and degree of morphological complexity. Large and complex cells in the class Cristamonadea evolved by replicating a fundamental unit, the karyomastigont, in various ways. We describe here four new species of Calonymphidae (Cristamonadea) from Rugitermes hosts, assigned to the genus Snyderella based on diagnostic features (including the karyomastigont pattern) and molecular phylogeny. We also report a new genus of Calonymphidae, Daimonympha, from Rugitermes laticollis. Daimonympha's morphology does not match that of any known Parabasalia, and its SSU rRNA gene sequence corroborates this distinction. Daimonympha does however share a puzzling feature with a few previously described, but distantly related, Cristamonadea: a rapid, smooth, and continuous rotation of the anterior end of the cell, including the many karyomastigont nuclei. The function of this rotatory movement, the cellular mechanisms enabling it, and the way the cell deals with the consequent cell membrane shear, are all unknown. "Rotating wheel" structures are famously rare in biology, with prokaryotic flagella being the main exception; these mysterious spinning cells found only among Parabasalia are another, far less understood, example.

Prevalence and Molecular Characterization of Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi in Diarrheic and Non-Diarrheic Calves from Ningxia, Northwestern China.

Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi are significant parasitic gastrointestinal pathogens with global distribution in humans and domestic animals, including calves. The main symptoms of calf infection are severe diarrhea, dehydration, growth retardation, and sometimes even death. To date, there has been limited information on the prevalence of Cryptosporidium spp., G. duodenalis, and E. bieneusi infections in calves in Ningxia, China, especially between diarrheic and non-diarrheic calves. A total of 438 fecal samples were collected from diarrheic (201) and non-diarrheic (237) calves in Ningxia. PCR and DNA sequencing were used to find the prevalence of Cryptosporidium spp. at 46.8% (205/438), G. duodenalis at 16.9% (74/438), and E. bieneusi at 10.0% (44/438). The prevalence of Cryptosporidium spp. infection in diarrheic and non-diarrheic calves was 54.0% (128/237) and 38.3% (77/201), respectively, and statistical analysis showed a positive correlation between the prevalence of Cryptosporidium spp. infection and calf diarrhea (p < 0.01). However, in this study, there was no statistical correlation between the prevalence of G. duodenalis infection as well as E. bieneusi infection and calf diarrhea (p > 0.05). Furthermore, four known Cryptosporidium species were successfully identified by comparing them with SSU rRNA gene sequences, including C. parvum, C. bovis, C. ryanae, and C. andersoni. In addition, all 74 G. duodenalis-positive samples were identified as assemblage E by comparative analysis of bg gene sequences. Among the 44 E. bieneusi-positive samples sequenced in the present study, 4 distinct E. bieneusi genotypes were successfully identified by comparative analysis of ITS sequences, including 3 known genotypes (J, BEB4, and N) and 1 novel genotype, the latter of which was identified and designated as NX1. These findings indicated that the high genetic diversity and complex population structures of Cryptosporidium spp., G. duodenalis, and E. bieneusi in Ningxia diarrhea calves and non-diarrhea calves, which provide new data for understanding the epidemiological status of Cryptosporidium spp., G. duodenalis, and E. bieneusi in Ningxia calves.

Identification and validation of novel Blastocystis subtype ST41 in a Colombian patient undergoing colorectal cancer screening.

Blastocystis sp. is among the most frequent intestinal protists identified in humans globally. However, characterization of Blastocystis subtype diversity in humans is ongoing. We report here the identification of novel Blastocystis subtype ST41 in a Colombian patient undergoing colorectal cancer screening involving colonoscopy and fecal testing (microscopy, culture, PCR). The full-length ssu rRNA gene sequence of the protist was generated using MinION long-read sequencing technology. The validity of the novel subtype was confirmed via phylogenetic and pairwise distance analyses of the full-length ST41 sequence and all other valid subtypes. The study provides reference material essential for conducting subsequent experimental studies.

Distribution and Phylogenetic Analysis of Subtypes and Alleles of Blastocystis sp. in the Stool Samples Collected from Patients with Gastrointestinal Complaints in İzmir, Turkey.

Blastocystis sp. is one of the most prevalent intestinal protozoa found in humans and many other animals. The present study aimed to examine the distribution and genetic diversity of Blastocystis sp. in stool samples from patients with gastrointestinal complaints in İzmir, Turkey. All stool samples of 439 patients with gastrointestinal complaints were examined by native-Lugol and trichrome staining. To investigate the presence of Blastocystis sp. in stool samples, DNA was isolated, and PCR was performed with the barcode region in the SSU rRNA gene. PCR positive samples were sequenced to identify subtypes and alleles of Blastocystis sp. The prevalence of Blastocystis sp. was found to be 16.6% (73/439) in patients with gastrointestinal complaints in İzmir, Turkey. Three different Blastocystis sp. subtypes were identified. ST3 (28/55; 51.0%) was the most common subtype followed by ST2 (19/55; 34.5%) and ST1 (8/55; 14.5%). Itching and diarrhea were the most prominent clinical symptoms in Blastocystis sp. positive patients. When clinical symptoms and subtypes were compared, diarrhea was found in 62.5%, 47.4%, and 46.4% of patients with ST1, ST2, and ST3 subtypes, respectively. In addition, itching was found in 37.5%, 32.1%, and 21.1% of patients with ST1, ST3, and ST2, respectively. Six distinct alleles were identified by allele analysis of Blastocystis 18S rRNA gene: allele 4 for ST1, alleles 9, 11, and 12 for ST2, and alleles 34 and 36 for ST3. In this study, Blastocystis sp. was detected in 16 of 21 districts, including the central and rural districts of İzmir. Although ST1 was detected in central districts, it was not found in rural districts. This study provides comprehensive data on the prevalence and molecular epidemiology of the genetic diversity at the level of subtypes and alleles of Blastocystis sp. in different districts of İzmir province in Turkey. To the best of our knowledge, this is the first study which evaluates the distribution of subtypes and alleles of Blastocystis sp. according to PCR and SSU rRNA gene sequencing in patients with gastrointestinal complaints in different districts of İzmir province in Turkey.

Identification and Molecular Characterization of Four New Blastocystis Subtypes Designated ST35-ST38.

Three recent studies of Blastocystis epidemiology in mammalian hosts identified four novel sequences that appeared to share B. lapemi as the most similar sequence. However, full-length ssu rRNA gene sequences were not available to confirm the validity of these new subtypes. In the present study, Nanopore MinION sequencing was used to obtain full-length reference sequences for each of the new subtypes. Additionally, phylogenetic analyses and pairwise distance comparisons were performed to confirm the validity of each of these new subtypes. We propose that the novel sequences described in this study should be assigned the subtype designations ST35-ST38. The full-length reference sequences of ST35-ST38 will assist in accurate sequence descriptions in future studies of Blastocystis epidemiology and subtype diversity.

Gonyaulax geomunensis sp. nov. and two allied species (Gonyaulacales, Dinophyceae) from Korean coastal waters and East China Sea: morphology, phylogeny and growth response to changes in temperature and salinity

ABSTRACT Six strains of three different Gonyaulax species were established by isolating cells from the Korean coastal area and the East China Sea, and their morphologies and molecular phylogenies based on SSU and LSU rRNA gene sequences were examined. In addition, the growth responses of the Gonyaulax species to changes in temperature and salinity were investigated. Based on morphological features and phylogenetic positions, Gonyaulax whaseongensis and G. polygramma were identified, and G. geomunensis sp. nov. is proposed in this study. These species displayed the plate formula typical for Gonyaulax, but G. polygramma and G. geomunensis sometimes showed a small intercalary plate (1a) surrounded by plates 2’, *3’ and 3’’. G. geomunensis was morphologically characterized by an S-type ventral organization, descending with a displacement of one cingulum width and bearing one, two or three prominent antapical spines of similar size. The cell surface, which was distinct from other Gonyaulax species, was thick and heavily reticulated into numerous polygonal areas. The reticulation was deeply excavated. The phylogenetics revealed that G. geomunensis and G. whaseongensis belong to different clades, and that there are two ribotypes of G. polygramma, which were morphologically indistinguishable. These species had a close phylogenetic relationship to G. hyalina, and all of them were characterized by dextral torsion. G. whaseongensis, G. polygramma and G. geomunensis had different growth responses to changes of temperature and salinity, respectively, indicating that morphological and phylogenetic identification of Gonyaulax species can be supported by ecological niches.

Phylogeny of the anaerobic ciliate genus Sonderia (Protista: Ciliophora: Plagiopylea), including the description of three novel species and a brief revision of the genus.

Anaerobic protists in general, and ciliates in particular, are important components of anoxic or hypoxic environments, however, their diversity remains underestimated. Sonderia is a poorly studied genus that is distributed worldwide and is commonly found in anaerobic environments. In the present study, the taxonomy and phylogeny of three new species, namely Sonderia aposinuata sp. nov., Sonderia paramacrochilus sp. nov. and Sonderia steini sp. nov., collected from China, were investigated based on microscopic observations and SSU rRNA gene sequencing methods. Sonderia aposinuata sp. nov. is diagnosed mainly by having a relatively large body size, a crescent-shaped oral opening, numerous slender extrusomes, one suture on the ventral side and two on the dorsal side, and a buccal cavity that occupies the anterior third of the cell. Sonderia paramacrochilus sp. nov. closely resembles S. macrochilus but differs mainly by its oral opening being located closer to the anterior cell margin and its spindle-shaped extrusomes. Sonderia steini sp. nov. is a freshwater species that can be recognized by its shallow buccal cavity, sparsely distributed rod-shaped extrusomes, and having 68-79 monokinetidal somatic kineties that form sutures on both sides of the body. Phylogenetic analyses based on small subunit ribosomal RNA (SSU rRNA) gene sequence data support the monophyly of the family Sonderiidae, however, Sonderia is paraphyletic. The genus Sonderia is briefly revised and a key to the identification of species belonging to this genus is supplied.

Analysis of autapomorphic point mutations provides a key for the tangled taxonomic distinction of the closely related species, Euplotes crassus, E. minuta and E. vannus (Ciliophora, Euplotida)

A well-defined clade of the Euplotes phylogenetic tree is represented by marine species characterized by a single-type dargyrome and ten fronto-ventral cirri. Three of them, namely Euplotes crassus, E. minuta and E. vannus, form a complex of closely related species of large use in experimental ciliatology. Despite morphometric and genetic analyses having substantiated their taxonomic separation, ambiguities still persist in strain assignments to one or another species. In addition to objective reasons intrinsic to significant overlapping of most morphological parameters, ambiguities also result from divergences (inherited from past literature) in deciding which of the two morphotypes, E. crassus or E. vannus, is characterized by a larger or a medium cell body size (E. minuta being clearly distinct by a smaller morphotype). By analysing nuclear SSU-rRNA gene and ITS region sequences from 37 strains, previously assigned to E. crassus, E. minuta and E. vannus based on conventional taxonomic parameters, we identified and used ITS autapomorphic point mutations to design three species-specific primers. In combination with an Euplotes-generic primer, they proved to be very effective in running polymerase chain reactions that produce amplicons of species-specific size that reliably resolve ambiguities in assigning strains to E. crassus, E. minuta or E. vannus.

Diversity of Blastocystis Subtypes in Horses in Colombia and Identification of Two New Subtypes

Blastocystis is a common intestinal protist in humans and animals worldwide. Wild and domestic animals are thought to be reservoirs of Blastocystis subtypes that also infect humans. There are limited studies on the prevalence and subtype distribution of Blastocystis in horses. In this study, 185 fecal samples were collected from horses (1 month to 17 years of age) in four regions of Colombia (Sabana de Bogotá, Costa Atlántica, Llanos Orientales, and Bogotá D.C.). Blastocystis presence and subtypes were determined by PCR and next generation amplicon sequencing. Eighty-one (43.8%) horses were positive for Blastocystis, with positive horses in all four regions. Molecular characterization identified 12 Blastocystis subtypes, 10 known subtypes (ST1, ST3–ST6, ST10, ST14, ST25, ST26), and 2 novel subtypes (ST33 and ST34). The validity of the novel subtypes was confirmed via phylogenetic and pairwise distance analyses of the full-length SSU rRNA gene sequences. Mixed subtype infections were common (55.6% of Blastocystis-positive horses). ST10 was the most prevalent subtype, present in 82.8% of Blastocystis-positive horses. Potentially zoonotic subtypes were identified in 88.9% of the Blastocystis-positive horses. This constitutes the most comprehensive study of Blastocystis in horses. Our findings indicate that horses harbor potentially zoonotic subtypes and could contribute to the transmission of Blastocystis to humans.

Morphology and molecular phylogeny of two new Aspidisca species (Ciliophora, Spirotrichea, Euplotida) collected from subtropical coastal waters in China

Ciliates are morphologically diverse and highly specialized unicellular eukaryotes that constitute an important component of the microbial food web. Aspidisca Ehrenberg, 1830, is a highly speciose genus that plays key ecological roles in a wide range of environments, and its species diversity has always been a hot spot in protozoan research. In this study, we investigate the living morphology, infraciliature, silverline system, and ribosomal small subunit (SSU rRNA) gene sequences of two new Aspidisca species collected from subtropical coastal waters of China using standard methods. Aspidisca spina sp. n. is characterized by having an obvious peristomial spur, two transparent posterior protrusions, seven frontoventral cirri in “polystyla-arrangement”, and six dorsal ridges. It can be distinguished from the most similar congener, A. magna Kahl, 1932, by the presence of posterior protrusions, more membranelles in the posterior part of adoral zone (AZM2), and different SSU rRNA gene sequences. A. shini sp. n. is smaller in body size, only 35–40 × 25–30 µm in vivo, with four prominent ridges on the arched dorsal side, very similar with the “well-known” species A. steini Buddenbrock, 1920, in morphological characteristics, but it can be distinguished by the arrangement of frontoventral cirri and 121 nucleotide difference in the SSU rRNA gene sequences. Phylogenetic analyses based on the SSU rRNA gene sequences revealed the systematic positions of two new taxa and supported the validity of them as distinct species.

From Argentine Abyssal Plain to farmed turbot in Spain: A ubiquitous amoeba species Vannella robusta sp. nov. (Amoebozoa, Vannellida)

A strain with the characters of the genus Vannella was isolated from the water layer immediately above the deep-sea sediment collected in the south-western Atlantic Ocean, ca. 4.6 km deep. Small-subunit ribosomal RNA (SSU rRNA) and cytochrome c oxidase (Cox1) gene phylogenetic analyses showed that the new strain branches within the clade of previously isolated unnamed Vannella strains from different marine fish and invertebrate hosts. Although the SSU rRNA gene sequences of these strains show variability within 2% of all nucleotide positions without any regular pattern, the available Cox1 gene sequences from within this clade are identical. Given the morphological homogeneity of the revealed clade, all of its strains can be assigned under the same species name, and the variation of their SSU rRNA is comparable to its intragenomic variation, as shown by molecular cloning of the PCR amplicons. High variability of the SSU rRNA gene sequences within and between independently isolated morphologically identical strains in combination with highly conserved Cox1 gene sequences may be a feature in some clades of Vannella, but is not a general rule for this genus, as SSU rRNA genes conserved between different morphospecies occur in several other clades within Vannella.

Morphology and SSU rRNA gene-based phylogeny of three peniculid ciliates (Ciliophora, Oligohymenophorea) from China, including a new Frontonia species

In the present study, a new freshwater peniculid species, Frontonia apoelegans sp. nov., and two other peniculid species, Frontonia atra (Ehrenberg, 1833) Bütschli, 1889 and Stokesia vernalis Wenrich, 1929, were isolated from Lake Weishan wetland, northern China. Their morphology and infraciliature are described based on live observations and silver staining methods. The SSU rRNA gene sequences are also provided. Frontonia apoelegans sp. nov. is recognized by the following combination of characteristics: two contractile vacuoles located right-dorsally, without collecting canals; peniculi 1 and 2 four-rowed, peniculus 3 three-rowed with leftmost row containing only one kinetosome; 62–76 somatic kineties; three ophryokineties; and four or five postoral kineties. We also provide improved diagnoses for Frontonia atra and Stokesia vernalis based on current and previous reports. Comparisons with sequences of morphologically similar species clearly support the validity of the new species. Phylogenetic analyses based on SSU rRNA gene sequence data reveal that Frontonia species with two contractile vacuoles cluster in a single clade, indicating these species may have a common origin. The family Frontoniidae is non-monophyletic whereas the family Stokesiidae remains monophyletic according to our analyses.

Taxonomy and SSU rRNA gene-based phylogeny of two new Euplotes species from China: E. chongmingensis n. sp. and E. paramieti n. sp. (Protista, Ciliophora)

BackgroundThe genus Euplotes Ehrenberg, 1830, one of the most complicated and confused taxa, contains about 160 nominal species. It was once proposed to be divided into four genera, two of which were proved to be non-monophyletic. At least 19 new species have been discovered in the past decade, implying that there is a large undiscovered diversity of this genus.ResultsThe morphology of two new freshwater euplotid ciliates, Euplotes chongmingensis n. sp. and E. paramieti n. sp., isolated from Shanghai, China, were investigated using live observations, protargol staining, and Chatton-Lwoff silver staining method. Euplotes chongmingensis is characterized by its small size (40–50 × 25–35 μm), about 24 adoral membranelles, 10 frontoventral cirri, two marginal and two caudal cirri, eight dorsolateral kineties with 11–16 dikinetids in the mid-dorsolateral kinety and a double type of silverline system. Euplotes paramieti n. sp. is 180–220 × 110–155 μm in vivo and strongly resembles E. amieti but having a difference of 57 bp in their SSU rRNA gene sequences. Phylogenetic analyses based on SSU rRNA gene sequence data were used to determine the systematic positions of these new taxa.ConclusionsThe description of two new freshwater taxa and their SSU rRNA gene sequences improve knowledge of biodiversity and enrich the database of euplotids. Furthermore, it offers a reliable reference for environmental monitoring and resource investigations.

Naidispora caidianensis n. gen. n. sp. infecting coelomocytes of oligochaete Branchiura sowerbyi (Oligochaeta: Naididae) in China

A new microsporidian parasite, Naidispora caidianensis n. gen. n. sp. was found in coelomocytes of oligochaete Branchiura sowerbyi Beddard, 1892 from Wuhan city, Hubei Province, China. Opaque, hypertrophied coelomocytes (0.17–0.24 mm in diameter) depicted clinical signs of infection. Electron microscopy revealed a microsporidian with monokaryotic life stages. Rounded uninucleate meronts subsequently transformed into multinucleate merogonial plasmodia with masses of electron-dense projections on their plasma membrane. Electron–dense sporogonial plasmodia separated into uninucleate sporonts through rosette-like budding, and further developed into sporoblasts, enclosed by a sporophorous vesicle. Uninucleate mature spores were pyriform, measured 4.1 ± 0.1 (3.9–4.3) μm × 2.1 ± 0.07 (1.9–2.2) μm (average ± SE, range, n = 50), and contained a mushroom-like anchoring disk, bipartite polaroplast, electron-lucent posterior vacuole, trilaminar spore wall and 12–14 turns of an isofilar polar filament arranged in 2–3 ranks. The 1542 bp sequence from B. soweryi showed the highest similarity (below 80 %) with Nematocenator marisprofundi (accession number: JX463178). SSU rRNA gene-based phylogenetic analysis demonstrated that the novel taxon formed an independent clade from known microsporidian parasites. Based on the ultrastructural features and SSU rRNA gene sequence, we propose the establishment of a novel genus (Naidispora n. gen.) and species (Naidispora caidianensis n. sp.) to contain this parasite.

A new “flagship” ciliate, Pseudostylonychia obliquocaudata n. gen., n. sp. (Ciliophora, Hypotricha), from South Korea

The morphology, ontogenesis, and molecular phylogeny of a new ciliate, Pseudostylonychia obliquocaudata n. gen., n. sp., discovered in a semiterrestrial habitat in South Korea, were investigated. The new species is characterized by an obovate or triangular body with pointed posterior end. The caudal area is distinctly obliquely truncated rightwards. Pseudostylonychia n. gen. is established and characterized by the obovate or triangular body, Stylonychia-like infraciliature and macronuclear pattern, and undulating membranes in Laurentiella pattern. It is also characterized by the involvement of cirri III/2 and IV/3 in primordia formation. SEM investigations show that the resting cyst has high, spirally arranged ridges. Phylogenetic analyses based on SSU rRNA gene sequences show that the new species is placed as a sister to the clade containing sequences of Stylonychia mytilus complex, Stylonychia (Metastylonychia), and Coniculostomum. Furthermore, Stylonychia koreana Kumar et al., 2016 nov. stat. is raised to species level based on the differences in the cyst morphology between the two subspecies of S. ammermanni. Also, Metastylonychia Kumar and Foissner, 2017 nov. stat. is raised to genus rank based on the new molecular data and the morphological and ontogenetic differences to Coniculostomum and Stylonychia.