Overview of the Diversity, Phylogeny and Biogeography of Strombidiid Oligotrich Ciliates (Protista, Ciliophora), With a Brief Revision and a Key to the Known Genera

Strombidiids are common free-living ciliates that have colonized coastal and open oceanic waters across the world. In recent years, numerous new taxa and gene sequences of strombidiids have been reported, revealing a large diversity of both their morphologic and genetic features. Here, we compare the taxonomic characters of all genera in the family Strombidiidae, provide a key to their identification, and investigate their molecular phylogeny. In addition, we analyze their regional distribution based on faunal data accumulated in China and attempt to infer their global distribution based on SSU rRNA gene sequence data. The current work revises the systematics of strombidiids based on morphologic, phylogenetic, and biogeographic evidence and provides a genus-level review of marine strombidiids.

Prevalence and Characterization of Cryptosporidium Species in Tibetan Antelope (Pantholops hodgsonii)

Cryptosporidium is an enteric apicomplexan parasite, which can infect multiple mammals including livestock and wildlife. Tibetan Antelope (Pantholops hodgsonii) is one of the most famous wildlife species, that belongs to the first class protected wild animals in China. However, it has not been known whether Tibetan Antelope is infected with Cryptosporidium so far. The objective of the present study was to determine the prevalence and characterization of Cryptosporidium species infection in Tibetan Antelope and the corresponding species by using molecular biological method. In the current study, a total of 627 fecal samples were randomly collected from Tibetan Antelope in the Tibet Autonomous Region (2019–2020), and were examined by PCR amplification of the small subunit ribosomal RNA (SSU rRNA) gene. Among 627 samples, 19 (3.03%, 19/627) were examined as Cryptosporidium-positive, with 7 (2.33%, 7/300) in females and 12 (3.67%, 12/327) in males. The analysis of SSU rRNA gene sequence suggested that only two Cryptosporidium species, namely, C. xiaoi and C. ubiquitum, were identified in this study. This is the first evidence for an existence of Cryptosporidium in Tibetan Antelope. These findings extend the host range for Cryptosporidium spp. and also provide important data support for prevention and control of Cryptosporidium infection in Tibetan Antelope.

Genetic Diversity of Cryptosporidium in Bactrian Camels (Camelus bactrianus) in Xinjiang, Northwestern China

Cryptosporidium species are ubiquitous enteric protozoan pathogens of vertebrates distributed worldwide. The purpose of this study was to gain insight into the zoonotic potential and genetic diversity of Cryptosporidium spp. in Bactrian camels in Xinjiang, northwestern China. A total of 476 fecal samples were collected from 16 collection sites in Xinjiang and screened for Cryptosporidium by PCR. The prevalence of Cryptosporidium was 7.6% (36/476). Six Cryptosporidium species, C. andersoni (n = 24), C. parvum (n = 6), C. occultus (n = 2), C. ubiquitum (n = 2), C. hominis (n = 1), and C. bovis (n = 1), were identified based on sequence analysis of the small subunit (SSU) rRNA gene. Sequence analysis of the gp60 gene identified six C. parvum isolates as subtypes, such as If-like-A15G2 (n = 5) and IIdA15G1 (n = 1), two C. ubiquitum isolates, such as subtype XIIa (n = 2), and one C. hominis isolate, such as Ixias IkA19G1 (n = 1). This is the first report of C. parvum, C. hominis, C. ubiquitum, and C. occultus in Bactrian camels in China. These results indicated that the Bactrian camel may be an important reservoir for zoonotic Cryptosporidium spp. and these infections may be a public health threat in this region.

The first case of microsporidiosis in Paramecium

A new microsporidian species, Globosporidium paramecii gen. nov., sp. nov., from Paramecium primaurelia is described on the basis of morphology, fine structure, and SSU rRNA gene sequence. This is the first case of microsporidiosis in Paramecium reported so far. All observed stages of the life cycle are monokaryotic. The parasites develop in the cytoplasm, at least some part of the population in endoplasmic reticulum and its derivates. Meronts divide by binary fission. Sporogonial plasmodium divides by rosette-like budding. Early sporoblasts demonstrate a well-developed exospore forming blister-like structures. Spores with distinctive spherical shape are dimorphic in size (3.7 ± 0.2 and 1.9 ± 0.2 μm). Both types of spores are characterized by a thin endospore, a short isofilar polar tube making one incomplete coil, a bipartite polaroplast, and a large posterior vacuole. Experimental infection was successful for 5 of 10 tested strains of the Paramecium aurelia species complex. All susceptible strains belong to closely related P. primaurelia and P. pentaurelia species. Phylogenetic analysis placed the new species in the Clade 4 of Microsporidia and revealed its close relationship to Euplotespora binucleata (a microsporidium from the ciliate Euplotes woodruffi), to Helmichia lacustris and Mrazekia macrocyclopis, microsporidia from aquatic invertebrates.

Morphological redescriptions and neotypification of two poorly known tintinnine ciliates (Alveolata, Ciliophora, Tintinnina), with a phylogenetic investigation based on SSU rRNA gene sequences

Two poorly known tintinnine ciliates collected from the coastal waters of PR China, viz., Codonellopsis mobilis Wang, 1936 and Tintinnopsis chinglanensis Nie & Ch’eng, 1947, were redescribed and neotypified using live observation, protargol staining and SSU rRNA gene sequencing. Ciliature information and SSU rRNA gene sequence data of both species were revealed for the first time and improved diagnoses were given based on the original descriptions and data from the present study. Further phylogenetic analyses inferred from SSU rRNA gene sequences and morphological data suggested that the genus Tintinnopsis is polyphyletic and that the genus Codonellopsis is non-monophyletic. The approximately unbiased test, however, does not reject the possibility that Codonellopsis is monophyletic.

Morphological Redescriptions and Molecular Phylogeny of Three Stentor Species (Ciliophora: Heterotrichea: Stentoridae) from Korea

The morphologies of the three freshwater stentorid ciliates in Korea, Stentor coeruleus (Pallas, 1766); Stentor muelleri Ehrenberg, 1831, and Stentor tartari Murthy & Bai, 1974, were investigated based on live observations and protargol impregnation. The Korean population of S. tartari exhibits the following characteristics: body size 200–355 × 85–135 µm in vivo, 62–106 somatic kineties, 8–13 peristomial kineties, 110–180 adoral membranelles, mostly two macronuclear nodules and 5–18 micronuclei, reddish and colorless cortical granules and the presence of symbiotic algae. We identified S. tartari based on unique characteristics compared to close congeners. Korean populations of S. coeruleus and S. muelleri are congruent with previously described populations in most aspects of their morphologies. Here, for the first time, we report molecular gene sequence information for S. tartari. Small subunit (SSU) rRNA gene sequence-based phylogeny indicates that S. tartari, which has multiple macronuclei, forms a monophyletic group with other Stentor species having a single macronucleus. Our findings based on morphology and SSU rRNA gene sequence information corroborate the hypothesis that the elongated macronucleus evolved from the compact single or multi macronucleus state.

Entamoeba chiangraiensisn. sp. (Amoebozoa: Entamoebidae) isolated from the gut of Asian swamp eel (Monopterus albus) in northern Thailand

The genusEntamoebacomprises mostly gut parasites and commensals of invertebrate and vertebrate animals including humans. Herein, we report a new species ofEntamoebaisolated from the gut of Asian swamp eels (Monopterus albus) in northern Thailand. Morphologically, the trophozoite is elongated and has a single prominent pseudopodium with no clear uroid. The trophozoite is actively motile, 30–50µm in length and 9–13µm in width. Observed cysts were uninucleate, ranging in size from 10 to 17.5µm in diameter. Chromatin forms a fine, even lining along the inner nuclear membrane. Fine radial spokes join the karyosome to peripheral chromatin. Size, host and nucleus morphology set our organism apart from other members of the genus reported from fish. The SSU rRNA gene sequences of the new isolates are the first molecular data of anEntamoebaspecies from fish. Phylogenetic analysis places the new organism as sister toEntamoeba invadens. Based on the distinct morphology and SSU rRNA gene sequence we describe it as a new species,Entamoeba chiangraiensis.

SSUnique: Detecting Sequence Novelty in Microbiome Surveys

ABSTRACT Extensive SSU rRNA gene sequence libraries, constructed from DNA extracts of environmental or host-associated samples, often contain many unclassified sequences, many representing organisms with novel taxonomy (taxonomic “blind spots”) and potentially unique ecology. This novelty is poorly explored in standard workflows, which narrows the breadth and discovery potential of such studies. Here we present the SSUnique analysis pipeline, which will promote the exploration of unclassified diversity in microbiome research and, importantly, enable the discovery of substantial novel taxonomic lineages through the analysis of a large variety of existing data sets. High-throughput sequencing of small-subunit (SSU) rRNA genes has revolutionized understanding of microbial communities and facilitated investigations into ecological dynamics at unprecedented scales. Such extensive SSU rRNA gene sequence libraries, constructed from DNA extracts of environmental or host-associated samples, often contain a substantial proportion of unclassified sequences, many representing organisms with novel taxonomy (taxonomic “blind spots”) and potentially unique ecology. Indeed, these novel taxonomic lineages are associated with so-called microbial “dark matter,” which is the genomic potential of these lineages. Unfortunately, characterization beyond “unclassified” is challenging due to relatively short read lengths and large data set sizes. Here we demonstrate how mining of phylogenetically novel sequences from microbial ecosystems can be automated using SSUnique, a software pipeline that filters unclassified and/or rare operational taxonomic units (OTUs) from 16S rRNA gene sequence libraries by screening against consensus structural models for SSU rRNA. Phylogenetic position is inferred against a reference data set, and additional characterization of novel clades is also included, such as targeted probe/primer design and mining of assembled metagenomes for genomic context. We show how SSUnique reproduced a previous analysis of phylogenetic novelty from an Arctic tundra soil and demonstrate the recovery of highly novel clades from data sets associated with both the Earth Microbiome Project (EMP) and Human Microbiome Project (HMP). We anticipate that SSUnique will add to the expanding computational toolbox supporting high-throughput sequencing approaches for the study of microbial ecology and phylogeny. IMPORTANCE Extensive SSU rRNA gene sequence libraries, constructed from DNA extracts of environmental or host-associated samples, often contain many unclassified sequences, many representing organisms with novel taxonomy (taxonomic “blind spots”) and potentially unique ecology. This novelty is poorly explored in standard workflows, which narrows the breadth and discovery potential of such studies. Here we present the SSUnique analysis pipeline, which will promote the exploration of unclassified diversity in microbiome research and, importantly, enable the discovery of substantial novel taxonomic lineages through the analysis of a large variety of existing data sets.

Morphological and phylogenetic studies on three members of the genus Pseudochilodonopsis (Ciliophora, Cyrtophoria) isolated from brackish waters in China, including a novel species, Pseudochilodonopsis quadrivacuolata sp. nov.

Three cyrtophorian ciliates isolated from brackish biotopes in China, Pseudochilodonopsis quadrivacuolata sp. nov., Pseudochilodonopsis fluviatilis Foissner, 1988 and Pseudochilodonopsis mutabilis Foissner, 1981, were investigated using living observation and protargol-staining methods. P. quadrivacuolata sp. nov. can be characterized as follows: cell size 50–70 × 30–40 μm in vivo; body oval with posterior end rounded; four tetragonally positioned contractile vacuoles; 12–15 nematodesmal rods; five right and six left somatic kineties; terminal fragment positioned apically on dorsal side, consisting of 11–14 basal bodies; four or five fragments in preoral kinety. P. fluviatilis and P. mutabilis were generally consistent with previous descriptions. In addition, a brief revision and a key to Pseudochilodonopsis are presented. The small-subunit (SSU) rRNA gene was also sequenced to support the identification of these species. Phylogenetic analyses based on molecular data indicate that the genera Pseudochilodonopsis and Chilodonella are closely related and both are well outlined; that is, all known congeners for which SSU rRNA gene sequence data are available group together, forming the core part of the family Chilodonellidae.