Ciliates are fundamental components of microzooplankton, with important ecological roles. However, ciliate communities are particularly difficult to monitor using conventional morphological approaches. New molecular tools, such as DNA metabarcoding, can facilitate the study of these communities. This study used high-throughput sequencing to examine the diversity and seasonal dynamics of ciliate communities in four estuarine ecosystems in the South China Sea from June 2019 to March 2020. The amplification of the V4 region of 18S rDNA using ciliate-specific primers identified a total of 1645 amplicon sequence variants (ASVs), corresponding to 13 ciliate classes, 97 families, and 157 genera. The dominant species across all four sampling stations were spirotrichs (including choreotrichs, oligotrichs, and stichotrichs), oligohymenophorean scuticociliates, litostomateans Didinium, and prostomateans Cryptocaryon. Significant differences in ciliate diversity and community composition in the four stations were mainly due to differences in rare, rather than abundant, ASVs. Analysis of the ciliate communities and seasonal patterns in their composition revealed that variations in habitat and environmental conditions have a greater effect than seasonal changes on community composition.

Background: Human babesiosis, caused by parasites of the genus Babesia, is an emerging and re-emerging tick-borne disease that is mainly transmitted by tick bites and infected blood transfusion. Babesia duncani has caused majority of human babesiosis in Canada; however, limited data are available to correlate its genomic information and biological features. Results: We generated a B. duncani reference genome using Oxford Nanopore Technology (ONT) and Illumina sequencing technology and uncovered its biological features and phylogenetic relationship with other Apicomplexa parasites. Phylogenetic analyses revealed that B. duncani form a clade distinct from B. microti, Babesia spp. infective to bovine and ovine species, and Theileria spp. infective to bovines. We identified the largest species-specific gene family that could be applied as diagnostic markers for this pathogen. In addition, two gene families show signals of significant expansion and several genes that present signatures of positive selection in B. duncani, suggesting their possible roles in the capability of this parasite to infect humans or tick vectors. Conclusions: Using ONT sequencing and Illumina sequencing technologies, we provide the first B. duncani reference genome and confirm that B. duncani forms a phylogenetically distinct clade from other Piroplasm parasites. Comparative genomic analyses show that two gene families are significantly expanded in B. duncani and may play important roles in host cell invasion and virulence of B. duncani. Our study provides basic information for further exploring B. duncani features, such as host-parasite and tick-parasite interactions.

Scuticociliatosis is an invasive external or systemic infection caused by ciliated protozoa, mainly those within the subclass Scuticociliatia (scuticociliates). Many scuticociliates are fish pathogens, including Miamiensis avidus, Philasterides dicentrarchi, Pseudocohnilembus persalinus, and Uronema marinum. Our previous study showed that hemolysis-related genes derived from bacteria through horizontal gene transfer (HGT) may contribute to virulence in P. persalinus. Hemorrhagic lesions are a common feature of scuticociliatosis, but it is not known whether other scuticociliates also have bacteria-derived hemolysis-related genes. In this study, we constructed a high-quality macronuclear genome of another typical pathogenic scuticociliate, U. marinum. A total of 105 HGT genes were identified in this species, of which 35 were homologs of hemolysis-related genes (including hemolysin-like genes) that had previously been identified in P. persalinus. Sequencing of an additional five species from four scuticociliate families showed that bacteria-derived hemolysis-related genes (especially hemolysin-like genes) are widely distributed in scuticociliates. Based on these findings, we suggest that hemolysin-like genes may have originated before the divergence of scuticociliates.

Microbial communities are essential components of aquatic ecosystems and are widely employed for the detection, protection, and restoration of water ecosystems. The polyurethane foam unit (PFU) method, an effective and widely used environmental monitoring technique, has been improved with the eDNA-PFU method, offering efficiency, rapidity, and standardization advantages. This research aimed to explore the colonization process of microbial communities within PFUs using eDNA-PFU technology. To achieve this, we conducted ten-day monitoring and sequencing of microbial communities within PFUs in a stable and controlled artificial aquatic ecosystem, comparing them with water environmental samples (eDNA samples). Results showed 1065 genera in eDNA-PFU and 1059 in eDNA, with eDNA-PFU detecting 99.95% of eDNA-identified species. Additionally, the diversity indices of bacteria and eukaryotes in both methods showed similar trends over time in the colonization process; however, relative abundance differed. We further analyzed the colonization dynamics of microbes in eDNA-PFU and identified four clusters with varying colonization speeds. Notably, we found differences in colonization rates between bacteria and eukaryotes. Furthermore, the Molecular Ecological Networks (MEN) showed that the network in eDNA-PFU was more modular, forming a unique microbial community differentiated from the aquatic environment. In conclusion, this study, using eDNA-PFU, comprehensively explored microbial colonization and interrelationships in a controlled mesocosm system, providing foundational data and reference standards for its application in aquatic ecosystem monitoring and beyond.

Balantidium ctenopharyngodoni is identified as the sole ciliate species that exclusively resides within the hindgut of grass carp with high prevalence and intensity. In this study, the successful cultivation of B. ctenopharyngodoni enabled us to collect enough cells for genome sequencing. Consequently, we acquired a high-quality genome assembly spanning 68.66 Mb, encompassing a total of 22,334 nanochromosomes. Furthermore, we predicted 29,348 protein-coding genes, and 95.5% of them was supported by the RNA-seq data. The trend of GC content in the subtelomeric regions of single-gene chromosomes was similar to other ciliates containing nanochromosomes. A large number of genes encoding carbohydrate-binding modules with affinities for starch and peptidoglycans was identified. The identification of mitochondrion-related organelles (MROs) within genome indicates its well-suited adaptation to the anaerobic conditions in the hindgut environment. In summary, our results will offer resources for understanding the genetic basis and molecular adaptations of balantidia to hindgut of herbivorous fish.