New evidence of consistency between phylogeny and morphology for two taxa in ciliated protists, the subclasses Oligotrichia and Choreotrichia (Protista, Ciliophora).

Marine planktonic ciliates are largely oligotrichs and choreotrichs, which are two subclasses of the class Spirotrichea. The current phylogenetic assignments of oligotrichs and choreotrichs are inconsistent with previous results based on morphological features, probably hindered by the limited information from a single gene locus. Here we provide 53 new sequences from small subunit ribosomal RNA (SSU rDNA), ITS1-5.8S rDNA-ITS2, and large subunit ribosomal RNA (LSU rDNA) gene loci in 25 oligotrich and choreotrich species. We also predict RNA secondary structures for the ITS2 regions in 55 species, 48 species of which are reported for the first time. Based on these novel data, we make a more comprehensive phylogenetic reconstruction, revealing consistency between morphological taxonomy and an updated phylogenetic system for oligotrichs and choreotrichs. With the addition of data from ciliature patterns and genes, the phylogenetic analysis of the subclass Oligotrichia suggests three evolutionary trajectories, among which: 1) Novistrombidium asserts an ancestral ciliary pattern in Oligotrichia; 2) the subgenera division of Novistrombidium and Parallelostrombidium are fully supported; 3) the three families (Tontoniidae, Pelagostrombidiidae and Cyrtostrombidiidae) all evolved from the most diverse family Strombidiidae, which explains why strombidiids consistently form polyphyletic clades. In the subclass Choreotrichia, Strombidinopsis likely possesses an ancestral position to other choreotrichs, and both phylogenetic analysis and RNA secondary structure prediction support the hypothesis that tintinnids may have evolved from Strombidinopsis. The results presented here offer an updated hypothesis for the evolutionary history of oligotrichs and choreotrichs based on new evidence obtained by expanding sampling of molecular information across multiple gene loci.

The First Record for the Americas of Loxodes rex, a Flagship Ciliate with an Alleged Restricted Biogeography.

As the foundations of food webs, protozoa are essential to the success of an ecological system. These organisms are often overlooked, and research in the Americas is sparse. Recent samplings conducted in freshwater canals and ponds in Florida, USA, have revealed Loxodes rex, an alleged endemic ciliate species. Originally described as endemic to tropical Africa, L. rex has been considered a prime candidate for proof of microbial endemism. Our studies have shown this giant, non-encysting ciliate to be thriving in subtropical Florida. Our observations are novel and include both the first record of occurrence for the Americas and the first high-quality in vivo images for this charismatic species.

Exploring the biogeography, morphology, and phylogeny of the condylostomatid ciliates (Alveolata, Ciliophora, Heterotrichea), with establishment of four new Condylostoma species and a revision including redescriptions of five species found in China.

Species of the ciliate class Heterotrichea Stein, 1859 are a cosmopolitan group of unicellular eukaryotic microorganisms, many of which have been widely used as models in various fields of research such as regenerative biology, functional ecology, environmental toxicology, and symbiotic behavior. However, species identification in the heterotrich family Condylostomatidae, especially the most species-rich and type genus Condylostoma Bory de Saint-Vincent, 1824, remains challenging due to incomplete original descriptions, few reliable distinguishing characters, and overlapping features between different species. This study presents an updated revision of Condylostoma and its related genus Condylostomides da Silva Neto, 1994 based on descriptions of five species, including nine populations collected from China, using both morphological and molecular methods. The main findings are as follows: (1) 43 nominal species and about 130 populations are reviewed, resulting in the recognition of 30 valid species of Condylostoma and eight valid species of Condylostomides; (2) keys, synonyms, biogeographic distributions and amended/improved diagnoses of all valid species are provided; (3) based on the available data, four new Condylostoma species (C. marinum sp. nov., C. petzi sp. nov., C. villeneuvei sp. nov., and C. microstomum sp. nov.), one new combination (Condylostomides minimus (Dragesco, 1954) comb. nov. & nom. corr.), and two corrected names (Condylostoma ancestrale Villeneuve-Brachon, 1940 nom. corr. and Condylostomides nigrus (Dragesco, 1960) nom. corr.) are suggested; (4) cryptic species are detected and proposed for the first time to form the Condylostoma curvum species complex; (5) three highly confusing Condylostoma species, C. kris, C. spatiosum, and C. minutum, are redefined for the first time based on modern taxonomic methods; (6) a 'flagship' species, Condylostomides coeruleus, is recorded for the first time from the continent of Asia, substantially expanding its biogeography; (7) ciliature adjacent to the distal end of the paroral membrane within the family Condylostomatidae is uniformly defined as frontal membranelles and is classified into three patterns according to the arrangement of kinetosomes, which serve as important key features. Supplementary Information: The online version contains supplementary material available at 10.1007/s42995-024-00223-3.

Multi-gene-based investigation on the molecular phylogeny of the hypotrichous family Strongylidiidae (Protista, Ciliophora), with notes on the ontogeny of a new genus and new species.

Ciliates in the subclass Hypotrichia have long been difficult to classify as they are one of the most polymorphic and highly differentiated groups, leading to their systematics remaining unresolved. Phylogenetic relationships within the hypotrich family Strongylidiidae have been ambiguous due to discordance between the morphological and genetic data. In this study, a new strongylidiid genus Heterouroleptus is established, mainly based on the novel mode of origin of the ventral cirral rows: left ventral cirral row (LVR) originates from frontal-ventral-transverse cirral anlagen (FVTA) III (anterior portion), IV (middle portion), and V (rear portion); right ventral cirral row comes from the entire FVTA VI. A new species, Heterouroleptus weishanensis gen. nov., sp. nov., is investigated along with the morphometric and molecular data from a population of Strongylidium wuhanense. Eight new sequences and nuclear gene markers (single-gene and multi-gene) are provided to analyze the phylogenetic relationships of strongylidiids, with the COI gene utilized to uncover further genetic information at species level and below. The results reveal that: (1) Strongylidiidae is monophyletic and has a close relationship with Dorsomarginalia; (2) Heterouroleptus gen. nov. forms a clade that is sister to all the other strongylidiids; (3) Hemiamphisiella Foissner, 1988 and Pseudouroleptus Hemberger, 1985 should not be synonyms, and both genera should be subdivided due to their variable morphological characteristics; (4) LVR originating from three anlagen is a plesiomorphy of Strongylidiidae. The discovery of the origin of the LVR not only contributes to the establishment of the genus Heterouroleptus, but also helps to improve the diagnosis of the family Strongylidiidae. Supplementary Information: The online version contains supplementary material available at 10.1007/s42995-024-00243-z.

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.

Morphological and molecular examination of the ciliate family Lagynusidae (Protista, Ciliophora, Prostomatea) with descriptions of two new genera and two new species from China.

Ciliates in the class Prostomatea play an important role in the global microbial loop due to their significant abundances and broad feeding strategies at the foundation of food webs. Despite their importance in ecosystems, the taxonomy and systematics of this group of ciliates has long been poorly understood, with this being especially true for members of the family Lagynusidae. Here we examine four lagynusids collected from sandy beaches in China, using silver-staining and 18S rRNA gene sequencing techniques. These investigations revealed two new genera and two new species and provided details for two little known forms: Penardella marina gen. nov., sp. nov., Apolagynus cucumis (as reported by Penard. Études sur les infusoires d'eau douce. Georg and Cie, Genève, 1922) gen. nov., comb. nov., Lagynus minutus sp. nov., and Lagynus elegans (Engelmann in Z Wiss Zool 11:347-393, 1862) Quennerstedt (Acta Univ Lund 4:1-48, 1867). Penardella gen. nov. can be morphologically distinguished by having more than three dikinetidal perioral kineties. Apolagynus gen. nov. differs from the closely related genus Lagynus in the absence of a conspicuous neck-like region. The ciliature of Apolagynus cucumis is revealed here for the first time, which demonstrates the classification of this species within Lagynusidae. Furthermore, Apolagynus binucleatus (Jiang et al., 2021) comb. nov. is established according to the new finding. The results of our phylogenetic analyses based on the 18S rRNA gene support the establishment of two new genera and indicate that Lagynusidae is monophyletic, which further strengthens its valid taxonomic status.

Taxonomy and molecular phylogeny of pleurostomatid ciliates from China with a description of two new species.

Ciliates in the order Pleurostomatida are found free-living in many habitats including within biofilms, but some (e.g. Pseudoamphileptus spp.) are ectocommensal on various hosts. Due to issues involving overall undersampling, the exact diversity and molecular phylogeny of this group remain largely underexplored. To combat this deficiency, detailed investigations were undertaken in northern China. As a result of these studies, we provide the morphological descriptions of two new species. Pseudoamphileptus apomacrostoma sp. nov., a new ectocommensal species, is characterized by the broadly oval cell shape, numerous scattered contractile vacuoles, and unique densely bounded extrusomes; Amphileptus qingdaoensis sp. nov., a marine form, is characterized by possessing oblong extrusomes with a conical anterior end, a single contractile vacuole and 5-7 left and 18-23 right kineties. In addition, a new population of Amphileptus orientalis Zhang et al., 2022, a freshwater representative, was documented and an improved diagnosis is provided. The phylogenetic analyses based on the SSU rDNA sequences imply that the genus Pseudoamphileptus is monophyletic whereas the genus Amphileptus is paraphyletic. The new molecular sequences presented here further support the establishment of two new species.

First Records of 'Flagship' Soil Ciliates in North America.

'Flagship' ciliates were investigated from soil samples collected in Florida, USA. This was undertaken to determine if species thought to be restricted to a given world region could be uncovered from similar habitats in a novel location, e.g. another continent. Two species of Condylostomides were discovered, and recorded from the North American continent for the first time. Condylostomides etoschensis was known only from Africa, but was found to be thriving in a Florida study site. An 18S rDNA sequence for this species was determined for the first time. Also discovered from the same study site was the ciliate Condylostomides coeruleus, previously known only from Central and South America. These two 'flagship' ciliates were found in the same habitat, from a continent well outside of their previously recorded biogeographies. Molecular sequencing and microscopy investigations were conducted to form the baseline for future work within this genus. Soil ciliates can obtain large population numbers and form cysts and are therefore likely able to disperse globally. These new records provide additional evidence that large distances, even between continents, do not hinder microbes from thriving globally. The absence of these conspicuously-colored gold and blue ciliates from previous studies is likely due to undersampling, rather than to any physical barriers.

Protistological science dissemination.

It is rare to meet protistologists who are not passionate about their study subject. The vast majority of people, however, never get the chance to hear about the work of these researchers. Although every researcher working on protists is likely to be aware of this situation, efforts made and tools employed for dissemination of knowledge are rarely documented. Following a proposal by the Italian Society of Protistology, a workshop at the 2019 VIII European Congress of Protistology in Rome, Italy, was dedicated to protistological knowledge dissemination. Through the many interventions, we discovered the diversity of efforts to reveal the protistan world to the general public, including museum exhibitions and activities, public understanding of science events, citizen science projects, specific book publications, the use of protists in teaching at all levels from primary school children to university undergraduate students, and to a global audience via social media. The participation of the workshop delegates in the discussions indicated that presentations on the wonderful world of protists to the public not only increase the visibility and accessibility of protistology research but are also very important for the scientific community. Here we report on some of the key aspects of the presentations given in the dissemination workshop.

Cell-fies: sharing microbiology with global audiences through Instagram.

For the dissemination of microbiology and science in general, social media is an underutilized but increasingly powerful tool, and one that transcends traditional avenues for scientific outputs. In order to reach and potentially inspire a deeply interested and general public, scientists should engage with platforms such as Instagram to share their content to an increasingly global audience.

Molecular Investigation of the Ciliate Spirostomum semivirescens, with First Transcriptome and New Geographical Records.

The ciliate Spirostomum semivirescens is a large freshwater protist densely packed with endosymbiotic algae and capable of building a protective coating from surrounding particles. The species has been rarely recorded and it lacks any molecular investigations. We obtained such data from S. semivirescens isolated in the UK and Sweden. Using single-cell RNA sequencing of isolates from both countries, the transcriptome of S. semivirescens was generated. A phylogenetic analysis identified S. semivirescens as a close relative to S. minus. Additionally, rRNA sequence analysis of the green algal endosymbiont revealed that it is closely related to Chlorella vulgaris. Along with the molecular species identification, an analysis of the ciliates' stop codons was carried out, which revealed a relationship where TGA stop codon frequency decreased with increasing gene expression levels. The observed codon bias suggests that S. semivirescens could be in an early stage of reassigning the TGA stop codon. Analysis of the transcriptome indicates that S. semivirescens potentially uses rhodoquinol-dependent fumarate reduction to respire in the oxygen-depleted habitats where it lives. The data also shows that despite large geographical distances (over 1,600km) between the sampling sites investigated, a morphologically-identical species can share an exact molecular signature, suggesting that some ciliate species, even those over 1mm in size, could have a global biogeographical distribution.