Genes and proteins expressed at different life cycle stages in the model protist Euplotes vannus revealed by both transcriptomic and proteomic approaches.

Sexual reproduction first appeared in unicellular protists and has continued to be an essential biological process in almost all eukaryotes. Ciliated protists, which contain both germline and somatic genomes within a single cell, have evolved a special form of sexual reproduction called conjugation that involves mitosis, meiosis, fertilization, nuclear differentiation, genome rearrangement, and the development of unique cellular structures. The molecular basis and mechanisms of conjugation vary dramatically among ciliates, and many details of the process and its regulation are still largely unknown. In order to better comprehend these processes and mechanisms from an evolutionary perspective, this study provides the first comprehensive overview of the transcriptome and proteome profiles during the entire life cycle of the newly-established marine model ciliate Euplotes vannus. Transcriptome analyses from 14 life cycle stages (three vegetative stages and 11 sexual stages) revealed over 26,000 genes that are specifically expressed at different stages, many of which are related to DNA replication, transcription, translation, mitosis, meiosis, nuclear differentiation, and/or genome rearrangement. Quantitative proteomic analyses identified 338 proteins with homologs associated with conjugation and/or somatic nuclear development in other ciliates, including dicer-like proteins, Hsp90 proteins, RNA polymerase II and transcription elongation factors, ribosomal-associated proteins, and ubiquitin-related proteins. Four of these homologs belong to the PIWI family, each with different expression patterns identified and confirmed by RT-qPCR, which may function in small RNA-mediated genome rearrangement. Proteins involved in the nonhomologous end-joining pathway are induced early during meiosis and accumulate in the developing new somatic nucleus, where more than 80% of the germline sequences are eliminated from the somatic genome. A number of new candidate genes and proteins likely to play roles in conjugation and its related genome rearrangements have also been revealed. The gene expression profiles reported here will be valuable resources for further studies of the origin and evolution of sexual reproduction in this new model species.

Process parameters optimization of EDM for hybrid aluminum MMC using hybrid optimization technique.

This study explores how machining parameters affect Surface Roughness (SR), Tool Wear Rate (TWR), and Material Removal Rate (MRR) during Electrical Discharge Machining (EDM) of a hybrid aluminum metal matrix composite (AMMC). The composite includes 6 % Silicon carbide (SiC) and 6 % Boron carbide (B4C) in an Aluminum 7075 (Al7075) matrix. A combined optimization approach was used to balance these factors, evaluating Pulse ON time, Current, Voltage, and Pulse OFF time. Response Surface Methodology (RSM) optimized single responses, while multi-response optimization employed a hybrid method combining the Entropy Weight Method (EWM), Taguchi approach, TOPSIS, and GRA. Analysis of Variance (ANOVA) assessed parameter significance, revealing substantial impacts on SR, MRR, and EWR. Based on TOPSIS and GRA, optimized parameters achieved a desirable balance: high MRR (0.4172, 0.5240 mm³/min), minimal EWR (0.0068, 0.0103 mm³/min), and acceptable SR (10.3877, 9.1924 µm) based on EWM-weighted priorities. Confirmation experiments validated a 15 % improvement in the closeness coefficient, and a 16 % improvement in the Grey relational grade, which considers combined SR, MRR, and EWR performance. Scanning Electron Microscope (SEM) analysis of surfaces machined with optimal parameters showed minimal debris, cracks, and no recast layer, indicating high surface integrity. This research enhances EDM optimization for AMMC, achieving efficiency in machining, minimizing tool wear, and meeting surface quality requirements.

Findings on three endocommensal scuticociliates (Protista, Ciliophora) from freshwater mollusks, including their morphology and molecular phylogeny with descriptions of two new species.

Species of the ciliate genera Myxophyllum and Conchophthirus are found as endocommensals of terrestrial and freshwater mollusks, respectively. So far, there have been few studies of these genera and morphological data for most members are often incomplete. In the present work, two new species, Myxophyllum weishanense sp. nov. and Conchophthirus paracurtus sp. nov., and a known species, Conchophthirus lamellidens, were isolated from hosts in Lake Weishan Wetland, China. Taxonomic studies indicate that M. weishanense sp. nov. can be recognized mainly by the combination of about 60 somatic kineties on both ventral and dorsal sides and the presence of caudal cilia. Conchophthirus paracurtus sp. nov. differs from congeners in its body shape and size, having a glabrous area on the posterior right side, and having fewer somatic kineties. In addition, differences in their ITS2 (Internally Transcribed Spacer 2) secondary structures support the discrimination of the two new species from their highly similar congeners. An improved diagnosis for the poorly known species, C. lamellidens is also provided. Phylogenetic analyses reveal that members of the genus Myxophyllum belong to a fully supported clade that is sister to a large, poorly supported clade consisting of Hemispeiridae, Ancistridae, and several lineages of the nonmonophyletic Cyclidiidae. The Myxophyllum clade also includes Protophyra ovicola JQ956552, a possible misidentification. Sequences of the two new Conchophthirus species cluster with other congeners in a fully supported clade that is unrelated to either the 'typical' thigmotrichs or to pleuronematids, thus conflicting with the traditional classification, and may represent an orphan scuticociliate lineage. Supplementary Information: The online version contains supplementary material available at 10.1007/s42995-024-00230-4.

Residual phosphorus availability after soil application of different organic waste in varying soil P status soils.

Transformations of applied phosphorus (P) fertilizer to inaccessible residual soil P is the main cause of inadequate P availability to plants in the majority of the cultivated soils. This study investigated the effect of organic wastes (rice-residue biochar, farmyard manure (FYM), poultry manure (PM), green manure (GM), and wheat straw (WS) on residual-P mobilization and its bioavailability in maize crops under different P status soils. Surface soil samples of 'medium-P' (12.5-22.5 kg P ha-1) and 'high-P' (22.5-50.0 kg P ha-1) status soils were collected from a long-term differential P fertilization experiment on maize-wheat rotation and were subjected to examine P adsorption/desorption, phosphatase activity and microbial biomass P (MBP) after incubation with organic amendments of varying elemental composition. The incorporation of organic manures decreases P sorption with maximum decrease in FYM-treated soils, indicating increased P concentration in soil solution. In contrast, WS due to its wider C/P ratio increased P sorption and did not produce any significant impact on the bioavailability of P. High-P status soils witnessed lower P sorption than medium-P soils. The MBP increased in the order of PM > FYM > GM > WS > biochar irrespective of soil P status. The availability and mobility of residual-P with FYM and PM was significantly higher than that of residual-P from biochar, GM and WS. Organics with wider C/P ratio immobilize bioavailable P in the short term regardless of soil P status.

Acid etching post-treatment enhanced fungal sterilization performance of copper-manganese-cerium oxide in liquid and aerosol: Materials and molecular biological mechanisms.

Bioaerosol is one of the main ways to spread respiratory infectious diseases. In order to further improve the sterilization efficiency of copper-manganese-cerium oxide (CuMnCeOx), the post-treatment method based on acid etching was adopted. The results showed that sterilization efficiency of the treated CuMnCeOx could reach 99% in aerosol with space velocity of 1400 h-1. L(+)-ascorbic acid successfully promoted the formation of Cu+, oxygen vacancies and the generation of reactive oxygen species (ROS) on the surface of the treated CuMnCeOx. During sterilization in liquid system, the transcriptome identified 316 differentially expressed genes, including 270 up-regulated genes and 46 down-regulated genes. Differentially expressed genes were significantly enriched in cell wall (GO:0005618) and external encapsulating structure (GO:0030312). Up-regulated genes were shown in regulation of reactive oxygen species biosynthetic processes (GO:1903409, GO:1903426, GO:1903428) and positive regulation all of reactive oxygen species metabolic process (GO:2000379), indicating that ROS induced cell death by destroying cell wall.

An overview on the key advantages and limitations of batch and dynamic modes of biosorption of metal ions.

Water purification using adsorption is a crucial process for maintaining human life and preserving the environment. Batch and dynamic adsorption modes are two types of water purification processes that are commonly used in various countries due to their simplicity and feasibility on an industrial scale. However, it is important to understand the advantages and limitations of these two adsorption modes in industrial applications. Also, the possibility of using batch mode in industrial scale was scrutinized, along with the necessity of using dynamic mode in such applications. In addition, the reasons for the necessity of performing batch adsorption studies before starting the treatment on an industrial scale were mentioned and discussed. In fact, this review article attempts to throw light on these subjects by comparing the biosorption efficiency of some metals on utilized biosorbents, using both batch and fixed-bed (column) adsorption modes. The comparison is based on the effectiveness of the two processes and the mechanisms involved in the treatment. Parameters such as biosorption capacity, percentage removal, and isotherm models for both batch and column (fixed bed) studies are compared. The article also explains thermodynamic and kinetic models for batch adsorption and discusses breakthrough evaluations in adsorptive column systems. The review highlights the benefits of using convenient batch-wise biosorption in lab-scale studies and the key advantages of column biosorption in industrial applications.

Multifaceted Anticancer Potential of Gnidia glauca (Fresen.) Gilg Leaf Alkaloids: Impact on Multiple Cellular Targets.

Gnidia glauca (Fresen.) Gilg has demonstrated significant anticancer potential through multiple mechanisms, including apoptosis induction, as shown by the TUNEL assay against MCF-7 cells, modulation of tubulin polymerization, preservation of mitochondrial function indicated by the JC-1 assay, and inhibition of DNA polymerase α and ß activities. Rationale for the present study is to investigate the potential anticancer properties of G. glauca leaf alkaloid extract. Fresh and healthy G. glauca leaves were cleaned, shade-dried, and the powder was defatted, extracted with 10% acetic acid in ethanol, and subjected for alkaloid extraction. The partially purified G. glauca leaf alkaloid extract was evaluated for its effects on tubulin polymerization, DNA polymerase activity, mitochondrial membrane potential, and apoptosis studies using human breast cancer (MCF-7) cells by flow cytometry. The extract was found to affect microtubule assembly in a concentration-dependent manner (15.125-250 µg/mL), indicating presence of alkaloids that function as spindle poison agents. Leaf alkaloid extract of G. glauca was also found to affect the mitochondrial membrane potential with IC50 value 144.51 µg/mL, and inhibited DNA polymerase α and ß activities dose dependently, thus potentially interfering with DNA replication and repair processes. Leaf alkaloid extract also showed the potential to induce DNA damage of 53.6%, albeit somewhat less than the standard drug camptothecin (64.94%) as confirmed by the TUNEL assay. Additionally, the GgLAE (IC50 144.51 µg/mL) showed significant inhibition of MCF-7 cells proliferation after 24 h, revealing phase arrests in sub G0/G1, S, and G2/M. These findings suggest that G. glauca leaf alkaloid extract contains alkaloids that possess anticancer properties with multiple targets, making the plant a natural source for a promising phytochemical drug candidates for further evaluation in pre-clinical and clinical studies. Further investigations are warranted to determine the efficacy, safety, identification and characterization of the alkaloids, and evaluate and determine their potential applications in cancer therapy.

Single-cell transcriptomic analysis reveals genome evolution in predatory litostomatean ciliates.

Many ciliated protists prey on other large microbial organisms, including other protists and microscopic metazoans. The ciliate class Litostomatea unites both predatory and endosymbiotic species. The evolution of predation ability in ciliates remains poorly understood, in part, due to a lack of genomic data. To fill this gap, we acquired the transcriptome profiles of six predatory litostomateans using single-cell sequencing technology and investigated their transcriptomic features. Our results show that: (1) in contrast to non-predatory ciliates, the predatory litostomateans have expanded gene families associated with transmembrane activity and reactive oxidative stress response pathways, potentially as a result of cellular behaviors such as fast contraction and extension; (2) the expansion of the calcium-activated BK potassium channel gene family, which hypothetically regulates cell contractility, is an ancient evolutionary event for the class Litostomatea, suggesting a rewired metabolism associated with the hunting behavior of predatory ciliates; and (3) three whole genome duplication (WGD) events have been detected in litostomateans, with genes associated with biosynthetic processes, transmembrane activity, and calcium-activated potassium channel activity being retained during the WGD events. In addition, we explored the evolutionary relationships among 17 ciliate species, including eight litostomateans, and provided a rich foundational dataset for future in-depth phylogenomic studies of Litostomatea. Our comprehensive analyses suggest that the rewired cellular metabolism via expanded gene families and WGD events might be the potential genetic basis for the predation ability of raptorial ciliates.

Highly divergent morphology but a close molecular phylogenetic relationship between two little-known ciliate genera Actinobolina and Papillorhabdos (Protozoa: Ciliophora: Litostomatea) with description of two new species.

Free-living litostomatean ciliates, prominent microeukaryote predators commonly encountered in freshwater and marine habitats, play vital roles in maintaining energy flow and nutrient cycles. Nevertheless, understanding their biodiversity and phylogenetic relationships remains challenging due to insufficient morphological information and molecular data. As a new contribution to this group, three haptorian ciliates, including two new species (Actinobolina bivacuolata sp. nov. and Papillorhabdos foissneri sp. nov.) and the insufficiently described type species, Actinobolina radians, were isolated from wetlands around Lake Weishan, China and investigated by a combination of living morphology, stained preparations, and 18S rRNA gene sequence data. An illustrated key of the valid species within the two genera is provided. In addition, we reveal the phylogenetic positions of these two genera for the first time. Although they differ in all key morphologic characters such as general appearance (ellipsoidal with numerous tentacles vs. cylindrical), extrusomes (stored in tentacles vs. anchored to pellicle), circumoral kinety (present vs. absent), composition of somatic kineties (kinetosome clusters vs. monokinetids), and number of dorsal brush rows (1 vs. 4), they both cluster in a fully supported clade in the phylogenetic tree, which indicates that the biodiversity and additional molecular markers of this group need further exploration.

Morphogenesis of an anaerobic ciliate Heterometopus palaeformis (Kahl, 1927) Foissner, 2016 (Ciliophora, Armophorea) with notes on its morphological and molecular characterization.

The morphology, morphogenesis, and molecular phylogeny of Heterometopus palaeformis (Kahl, 1927) Foissner, 2016 were studied using microscopical observations on live and protargol-stained specimens as well SSU rRNA gene sequencing. The morphogenetic data for the genus are presented for the first time. Compared to other metopids, the morphogenesis of H. palaeformis is distinct since its (1) perizonal stripe rows 4 and 5 are involved in the formation of the opisthe's adoral polykinetids; (2) perizonal stripe rows 3-5 and two adjacent preoral dome kineties contribute to most of the opisthe's paroral membrane while perizonal stripe rows 1 and 2 contribute very little; (3) four kinety rows are formed to the left of the opisthe's adoral zone of polykinetids. The Chinese population resembles the original and neotype populations well in terms of general morphology - characterized by a life size of 55-120 × 10-20 µm, an elongate ellipsoidal body with a hardly spiralized flat preoral dome, about 18 somatic kineties and 20 adoral polykinetids. The SSU rDNA sequence of the present population exhibits a disparity of 1.33%-2.22% divergence from sequences of other populations. Nevertheless, phylogenetic analysis reveals that populations of H. palaeformis form a separate, stable cluster within the paraphyletic Metopidae clade.

Invasion by exogenous RNA: cellular defense strategies and implications for RNA inference.

Exogenous RNA poses a continuous threat to genome stability and integrity across various organisms. Accumulating evidence reveals complex mechanisms underlying the cellular response to exogenous RNA, including endo-lysosomal degradation, RNA-dependent repression and innate immune clearance. Across a variety of mechanisms, the natural anti-sense RNA-dependent defensive strategy has been utilized both as a powerful gene manipulation tool and gene therapy strategy named RNA-interference (RNAi). To optimize the efficiency of RNAi silencing, a comprehensive understanding of the whole life cycle of exogenous RNA, from cellular entry to its decay, is vital. In this paper, we review recent progress in comprehending the recognition and elimination of foreign RNA by cells, focusing on cellular entrance, intracellular transportation, and immune-inflammatory responses. By leveraging these insights, we highlight the potential implications of these insights for advancing RNA interference efficiency, underscore the need for future studies to elucidate the pathways and fates of various exogenous RNA forms, and provide foundational information for more efficient RNA delivery methods in both genetic manipulation and therapy in different organisms.

Design, Synthesis, Molecular Docking, and In Vitro Antibacterial Evaluation of Benzotriazole-Based ß-Amino Alcohols and Their Corresponding 1,3-Oxazolidines.

In the present study, a series of benzotriazole-based ß-amino alcohols were efficiently synthesized in excellent yields via aminolysis of benzotriazolated epoxides under catalyst- and solvent-free conditions. Further these ß-amino alcohols were successfully utilized to synthesize the corresponding benzotriazole-based oxazolidine heterocyclic derivatives. All the synthesized compounds were characterized by various spectroscopic techniques such as 1H NMR, 13C NMR, and mass spectroscopy for structure elucidation. The compounds were subjected to a microtiter plate-based antimicrobial assay. The antimicrobial activity results reveal that the compounds 4a, 4e, and 5f were found to be active against Staphylococcus aureus (ATCC-25923) with minimum inhibitory concentrations (MICs) of 32, 8, and 64 µM, respectively. Also, the compounds 4a, 4e, 4k, 4i, 4m, 4n, 4o, 5d, 5e, 5f, 5g, and 5h showed effective activity against Bacillus subtilis (ATCC 6633) with MICs of 64, 16, 16, 16, 64, 16, 64, 64, 32, 64, 8, and 16 µM, respectively. A biological investigation was conducted, including molecular docking of two compounds with several receptors to identify and confirm the best ligand-protein interactions. Hence, this study found a significant strategy to diversify the chemical molecules. The synthesized compounds play a potential role as an antibacterial intensifier against some pathogenic bacteria for the development of antibacterial substances.

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.

Effectiveness of Opuntia ficus indica (cactus) fruit juice for sludge conditioning.

The possibility of using Opuntia ficus indica fruit juice (OFIFJ) as a bioflocculant for conditioning the synthetic kaolin sludge and sewage sludge (region Oran, Algeria, and Pau, France) was studied. Turbidity of the supernatant, dryness of the sludge cake, and total time of filtration (TTF) were examined parameters. Using vacuum filtration, lime was also tested as a chemical conditioner and gives good results on Lescar (France) sewage sludge in terms of cake's dryness, filtrate quality, turbidity (13.54%), and total time of filtration (TTF = 85.29%), comparing to the industrial polymer (Sedifloc 408C; turbidity; 8.33% and TTF: 2.94%). For the sewage sludge of Oran (Algeria), the results obtained with OFIFJ were compared to those obtained with the cladodes juice of the same plant OFIC, and also with a cationic polymer (Superfloc 8396). For an optimum dosage, it showed that OFIFJ has a flocculation activity as same as the cladodes juice OFIC for sludge conditioning and gives better results in terms of turbidity (dosage of 22.4 g/kg DM: 3.7 NTU for OFIC, dosage of 8.36: 3.63 NTU for OFIFJ. Dryness was enhanced from 14.91 to 22.93% (OFIC 16 g/kg DM) and to 24.48% (OFIF 20.9 g/kg DM) but for TTF, we found the opposite. In fact, this plant showed to be an available, biodegradable, and non-toxic flocculant. For kaolin synthetic sludge (30%), the optimum dosages of those conditioners were found to be 0.066 g kg-1 for OFIC, comparing between vacuum filtration and filtration compression; turbidity was enhanced for both techniques, contrary to dryness. Concerning the Oran city sewage sludge, both turbidity and dryness were optimized. Same thing for the France sewage sludge, all the studied parameters were enhanced with the two studied bioflocculants.

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.

Bio-waste mediated synthesis of zirconium nanoparticle fuel: Energy management strategy for performance evaluation in a diesel engine.

The impact of biosynthesized zirconium nanoparticles originated from biological waste, blended in diesel fuel processed through bio-refining strategy and its combustion, emissions, and overall diesel engine performance towards safety has been examined. Different weight fractions of zirconia nanoparticles were combined with crude diesel at 10, 20, and 30 mg/L values. According to the engine tests, Zirconia (20 nm) added to pure diesel at a concentration of 30 parts per million incremented thermal efficiencies by 4.9% compared to regular diesel fuel. The average reduction in specific fuel consumption for clean diesel fuel when the engine was operating at full power was 2.9%, 3.9%, and 4.9%. Diesel smoke, hydrocarbon, CO, and NOx emissions were reduced by 13%, 20%, 25%, and 29%, respectively, when nano additives were used at a concentration of 30 ppm.Nanoparticles enhance fuel stability, overcome detonation difficulties, and avoid fouling spark plugs. The pressure within cylinder, the temperature, and the rate at which heat is released was improved when alumina nanoparticles were appended to diesel fuel. However, both the length of the combustion and further delay in ignition were cut down. The ideal concentration of zirconia nanoparticles for improving combustion, efficiency, and emissions along with safety attainment in an internal combustion engine is recorded at 30 ppm.

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.

Comparative analysis of single-cell genome sequencing techniques toward the characterization of germline and somatic genomes in ciliated protists.

Ciliated protists contain both germline micronucleus (MIC) and somatic macronucleus (MAC) in a single cytoplasm. Programmed genome rearrangements occur in ciliates during sexual processes, and the extent of rearrangements varies dramatically among species, which lead to significant differences in genomic architectures. However, genomic sequences remain largely unknown for most ciliates due to the difficulty in culturing and in separating the germline from the somatic genome in a single cell. Single-cell whole genome amplification (WGA) has emerged as a powerful technology to characterize the genomic heterogeneity at the single-cell level. In this study, we compared two single-cell WGA, multiple displacement amplification (MDA) and multiple annealing and looping-based amplification cycles (MALBAC) in characterizing the germline and somatic genomes in ciliates with different genomic architectures. Our results showed that: 1) MALBAC exhibits strong amplification bias towards MAC genome while MDA shows bias towards MIC genome of ciliates with extensively fragmented MAC genome; 2) both MDA and MALBAC could amplify MAC genome more efficiently in ciliates with moderately fragmented MAC genome. Moreover, we found that more sample replicates could help to obtain more genomic data. Our work provides a reference for selecting the appropriate method to characterize germline and somatic genomes of ciliates.

Integrative Studies on Three Epibiotic Epistylis Species (Protozoa, Ciliophora, Peritrichia) in Lake Weishan Wetland, Northern China, Including the Establishment of a New Species.

Three epibiotic Epistylis species, i.e., Epistylis weishanensis sp. nov., Epistylis daphniae Fauré-Fremiet, 1905, and Epistylis pygmaeum (Ehrenberg, 1838) Foissner et al., 1999, were investigated based on their living morphology, infraciliature, and small subunit (SSU) rDNA sequence data. Epistylis weishanensis sp. nov. is characterized by its double-layered peristomial lip, contractile vacuole located on the dorsal wall of the infundibulum, infundibular polykinety 3 (P3) composed of three equal-length rows that terminate above infundibular polykinety 1 (P1), 50-62 silverlines between the peristome and the trochal band, and about 30 silverlines between the trochal band and the scopula. Based on previous and newly obtained data for E. daphniae and E. pygmaeum, improved diagnoses and redescriptions are provided including, for the first time, data on their infraciliature. Phylogenetic analyses reveal that all three species do not group within the major clade of Epistylis, supporting the assertion that the genus Epistylis should be an assemblage of morphospecies and therefore needs to be revised.

Taxonomy and molecular phylogeny of two poorly known ciliate genera, Balantidion and Acropisthium (Protista: Ciliophora: Litostomatea), including a new species of Balantidion.

The class Litostomatea Small & Lynn, 1981 is a morphologically diverse ciliate group including hundreds of free-living and endocommensal species. The genera Acropisthium Perty, 1852 and Balantidion Eberhard, 1862 previously consisted of one free-living freshwater species each. Here, we not only highlight additional morphological features of the two type species, but also investigate a new species, Balantidion foissneri sp. nov., isolated from a river flowing through Lake Weishan, China, based on complementary methods, i.e., living morphology, stained preparations, and 18S rRNA gene sequence data. Balantidion foissneri sp. nov. can be distinguished from the type species, B. pellucidum Eberhard, 1862, by the body size (115-170 × 50-80 µm vs. 70-100 × 25-45 µm in B. pellucidum), oral bulge (distinct vs. indistinct), extrusome shape (filiform vs. rod-shaped), and the number of somatic kineties (46-60 vs. 25-40). In Balantidion species, pre-encystment trophonts show similarly-shaped polymorphic cytoplasmic lepidosomes destined to adorn the outer surface of the resting cyst. Based on the current knowledge, assignment of Balantidion to the family Acropisthiidae Foissner & Foissner, 1988 is proposed. In addition, phylogenetic analyses based on molecular data show that the two Balantidion species form a fully-supported clade to which Acropisthium mutabile has a sister relationship.