Genetic Diversity and Phylogeny of the Genus Euplotes (Protozoa, Ciliophora) Revealed by the Mitochondrial CO1 and Nuclear Ribosomal Genes

Nuclear ribosomal and mitochondrial genes have been utilized individually or in combination to identify known species and discriminate closely related species. However, compared with metazoans, genetic diversity within the ciliate order Euplotida is poorly known. The aim of this study is to investigate how much nucleotide sequence divergence occurs within Euplotes. A total of 14 new gene sequences, comprising four SSU rDNA and 10 CO1 (including three species for the first time) were obtained. Phylogenetic analyses were carried out based on sequences of two DNA fragments from the same 27 isolates. We found that CO1 revealed a larger interspecific divergence than the SSU rRNA gene, thus demonstrating a higher resolution for separating congeners. Genetic distances differ significantly at the species level. Euplotes balteatus was revealed to have a large intraspecific variation at two loci, while E. vannus showed different levels of haplotype variability, which appeared as a polyphyletic cluster on the CO1 tree. These high genetic divergences suggest the presence of more cryptic species. By contrast, the CO1 gene showed low variability within E. raikovi, appearing as monophyletic clusters, which indicates that this species could be identified based on this gene. Conclusively, CO1 is a suitable marker for the study of genetic diversity within Euplotes, and increased taxon sampling gives an opportunity to screen relationships among members of this genus. Additionally, current data present no clear biogeographical pattern for Euplotes.

A New Record of Oxytricha granulifera granulifera Foissner and Adam, 1983 (Protozoa, Ciliophora, Oxytrichidae) From a Hot Spring in Iceland, With Notes on Its Abnormal Form During Cultivation

We isolated a population of Oxytricha granulifera granuliferaFoissner and Adam (1983) from a hot spring in Iceland. The pure culture of this isolate was established at room temperature in the laboratory. This allowed for a detailed investigation, informed by integrated approaches, of the isolate’s morphology and morphogenesis, as well as molecular phylogeny. Results showed that the morphological and morphogenetic characteristics of the Iceland population are consistent with those of other populations. During the 3-year long period of laboratory cultivation, some abnormal individuals appeared repeatedly in the culture system. Interestingly, the morphological characteristics of these abnormal cells were rather stable, and were as follows: 1) body slender and elliptical-shaped; 2) remarkably shortened adoral zone and significantly reduced number of adoral membranelles; and 3) loss of undulating membranes. Resting cysts, binary fission and conjugate reproduction were not found in abnormal specimens. Although the morphology of abnormal individuals changed significantly, the sequences of the SSU rDNA of the normal and abnormal morphotypes were the same. Phylogenetic analyses showed that the two morphotypes clustered in a clade with other populations of O. granulifera granulifera.

Taxonomy of Three Oxytrichids (Protozoa, Ciliophora, Hypotrichia), With Establishment of the New Species Rubrioxytricha guangzhouensis spec. nov.

Two oxytrichid ciliates collected from China, Oxytricha lithofera Foissner, 2016 and Rubrioxytricha haematoplasma (Blatterer and Foissner, 1990) Berger, 1999 were investigated based on living observations, protargol preparations, and molecular analyses. The Chinese population of O. lithofera shares highly distinctive features with the type population, e.g., lithosomes, abutting macronuclear nodules, straight undulating membranes, and long dorsal bristles. The morphology and morphogenesis of our new isolate of R. haematoplasma corresponds well with the type population. The 18S rRNA gene sequences of both species were obtained, and the molecular phylogeny of the genera Oxytricha Bory de Saint-Vincent in Lamouroux et al., 1824 and Rubrioxytricha Berger, 1999 was analyzed. In addition, we found that the Guangzhou population of R. haematoplasma described by Chen et al. (2015) can be distinguished from the type population and our isolate by the reddish-brown cortical granules (vs. lemon yellowish to greenish) and mitochondria-like granules present (vs. absent), as well as the slightly red cytoplasm (vs. slightly orange) and the marine or brackish water habitat (vs. freshwater). In addition, a 20-bp divergence in their 18S rRNA gene sequences indicates that they are not conspecific. Thus, we establish a new species, Rubrioxytricha guangzhouensis, for R. haematoplasma sensu Chen et al. (2015).

Transcriptome Profiling Revealed Multiple rquA Genes in the Species of Spirostomum (Protozoa: Ciliophora: Heterotrichea)

Adaptation to life at different oxygen tensions plays a role in protozoan ecology and controls the distribution of different species in anoxic habitats. The ciliate genus Spirostomum inhabiting fresh or low salinity water globally where these species are considered as bioindicators. Under anaerobic or low oxygen conditions, the rhodoquinol-dependent pathway has been reported in the species from the class Heterotrichea. With the help of RNA sequencing (RNAseq) data, Spirostomum spp., are suitable for deep molecular investigations on rquA for rhodoquinone (RQ) biosynthesis. In this study, Spirostomum ambiguum, Spirostomum subtilis, and Spirostomum teres collected from densely vegetated freshwater habitat in Fuzhou, China, explored the evidence of rquA. Based on transcriptome analysis, two to three RquA proteins were identified in S. ambiguum, S. teres, and S. subtilis, respectively. The presence of a key Motif-I of RquA and mitochondrial targeting signals (MTS), also confirmed the identity of these as RquA. Furthermore, Spirostomum RquA proteins could be sorted into two groups based on their conserved amino acid (CAA) residues. Phylogenetic analysis also exhibited RquA division into two subclades contained RquA1 and RquA2/RquA3 and supports two to three paralogs of rquA genes in the genomes Spirostomum spp. Additional transcriptomes and genomes analysis of Blepharisma spp., and Stentor spp., respectively, also revealed at least two paralogs of rquA in members of the class Heterotrichea. The present study provides evidence for the presence of RquA and rhodoquinol dependent fumarate reduction pathway in Spirostomum species potentially use to respire in the oxygen-depleted habitats and two to three diverse rquA genes.

Taxonomy and molecular phylogeny of four karyorelictid species belonging to the genera Apotrachelocerca and Tracheloraphis (Protozoa: Ciliophora), with descriptions of two new species

The taxonomy and phylogeny of four trachelocercid ciliates, Apotrachelocerca orientalis sp. nov., Tracheloraphis apoligostriata sp. nov., T. oligostriata and T. prenanti, isolated from marine habitats in Qingdao, China, were studied. The new species, A. orientalis sp. nov., can be distinguished from its most closely related congener, A. arenicola, by the absence of cortical granules and its conspicuously different SSU rDNA sequence. Tracheloraphis apoligostriata sp. nov. differs from the similar T. oligostriata by its body shape, distribution pattern of the cortical granules and SSU rDNA sequence. Based on available data, we suggest that T. prenanti can be defined by its colourless, globular cortical granules, which indicates that this species should be removed from the T. phoenicopterus-complex. Phylogenetic analyses based on SSU rDNA sequence data indicates that: A. orientalis sp. nov. groups with A. arenicola with full support, together sister to all other trachelocercid species; and T. cf. phoenicopterus, T. prenanti, T. oligostriata and T. apoligostriata sp. nov. form a clearly outlined clade.

Taxonomy and Molecular Phylogeny of Three Species of Scuticociliates From China: Citrithrix smalli gen. nov., sp. nov., Homalogastra binucleata sp. nov. and Uronema orientalis Pan et al., 2015 (Protozoa, Ciliophora, Oligohymenophorea), With the Proposal of a New Family, Citrithrixidae fam. nov.

The morphology and taxonomy of three scuticociliates found in China, viz. Citrithrix smalli sp. nov., Homalogastra binucleata sp. nov., and Uronema orientalis Pan et al., 2015, were investigated. The small subunit ribosomal RNA (SSU rRNA) gene of these species, and the cytochrome c oxidase subunit I (COI) gene of Uronema orientalis, were sequenced and compared with those of related taxa to determine their systematic positions. The new monotypic genus Citrithrix gen. nov. is characterized by its lemon-shaped body, posteriorly located cytostome, dominant oral groove, and the compact structure of its multi-rowed membranelles 1 and 2 (M1, M2). Based on both morphological and molecular data, this new genus cannot be assigned to any known family and thus, a new family, Citrithrixidae fam. nov., is proposed within the order Philasterida. Homalogastra binucleata sp. nov., a brackish water form (salinity 2‰), differs from all congeners in having two macronuclear nodules. Uronema orientalis closely resembles the type population in all respects other than having fewer somatic kineties.This article is registered in ZooBank under: urn:lsid:zoobank.org:pub:5727F18E-5421-446D-B22C-774783539FE4.