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Life cycle of Ich. Infective theronts bore through the surface mucus and take up residence within the epithelium of susceptible fish. Theronts differentiate into feeding trophonts that grow and exit the host (as tomonts) within 4 to 7 days. Tomonts swim for a brief period and then adhere to an inert support where they secrete a gelatinous capsule. Tomonts divide within the capsule to form hundreds of tomites that differentiate into infective theronts within 18 to 24 hours at room temperature. Theronts that fail to infect fish die within 1 to 2 days.

Life cycle of Ich. Infective theronts bore through the surface mucus and take up residence within the epithelium of susceptible fish. Theronts differentiate into feeding trophonts that grow and exit the host (as tomonts) within 4 to 7 days. Tomonts swim for a brief period and then adhere to an inert support where they secrete a gelatinous capsule. Tomonts divide within the capsule to form hundreds of tomites that differentiate into infective theronts within 18 to 24 hours at room temperature. Theronts that fail to infect fish die within 1 to 2 days.

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Ichthyophthirius multifiliis, commonly known as Ich, is a highly pathogenic ciliate responsible for 'white spot', a disease causing significant economic losses to the global aquaculture industry. Options for disease control are extremely limited, and Ich's obligate parasitic lifestyle makes experimental studies challenging. Unlike most well-studied...

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... The increase in large-scale 'omics data from diverse members of the Ciliophora, (Aeschlimann et al., 2014, Coyne et al., 2011Maurer-Alcalá et al., 2018;Swart et al., 2013;Slabodnick et al., 2017;Wang et al., 2018;Xiong et al., 2015;Yan et al., 2019;Zheng et al., 2018) provides an opportunity to expand our understanding of the cell biology across the lineage. In this study, we have undertaken a comparative molecular evolutionary analysis across the ciliate lineage of SM proteins and Qa-SNAREs subfamilies, which together promote compartment specificity and are important components of vesicle fusion to the membrane. ...
Article
Protists in the phylum Ciliophora possess a complex membrane trafficking system, including osmoregulatory Contractile Vacuoles and specialized secretory organelles. Molecular cell biological investigations in Tetrahymena thermophila have identified components of the protein machinery associated with the secretory organelles, mucocysts. The Qa‐SNARE Syn7lp plays a role in mucocyst biogenesis as do subunits of the CORVET tethering complex (specifically Vps8). Indeed, Tetrahymena thermophila possesses expanded gene complements of several CORVET components, including Vps33 which is also a Sec1/Munc18 (SM) protein that binds Qa‐SNAREs. Moreover, the Qa‐SNAREs in Paramecium tetraurelia have been localized to various endomembrane organelles. Here, we use comparative genomics and phylogenetics to determine the evolutionary history of the SM and Qa‐SNARE proteins across the Ciliophora. We identify that the Last Ciliate Common Ancestor possessed the four SM proteins and six Qa‐SNAREs common to eukaryotes, including the uncommonly retained Syntaxin 17. We furthermore identify independent expansion of these protein families in several ciliate classes, including concurrent expansions of the SM protein‐Qa SNARE partners Sec1:SynPM inhe oligohymenophorean ciliates lineage, consistent with novel Contractile Vacuole specific innovations. Overall, these data are consistent with SM proteins and Qa‐SNAREs being a common set of components for endomembrane modulation in the ciliates.
... Ciliates are one of the most diverse, highly differentiated and ancient groups of microbial eukaryotes (Coyne et al., 2011). The characteristics of nuclear dimorphism (one large macronucleus and one small micronucleus) make ciliates considered to be an excellent model organism in the genetic investigation (Juranek and Lipps, 2007). ...
... 2.8.0) and Minimap (v. 2.17) sorftwares were used to retain the reads containing GC ratios less than 30% or accurately mapped to the genomes of closely homologous species were retained (Coyne et al., 2011). Then, the filtered nanopore data were assembled into contigs by Flye (v. ...
... For the genome survey analysis, the number of 17-mer was 44,364,461,437, K_depth was estimated as 92.6, GC content and heterozygosity were about 26.86 and 0.5%, respectively. And the corrected genome size is estimated to be 45.67 Mb ( Supplementary Table S1), which is similar to the I. multifiliis genome size (Coyne et al., 2011). A common single-peak pattern was observed in the K-mer frequency distribution analysis, indicating that the genome may have a low level of heterozygosity and repetitive regions ( Figure 1B). ...
... By contrast, Hou (2008) did not observe this relationship from the six test species of dinoflagellates, which might be due to the small number of species used and the small difference in cell size among the test species of Karenia, Karlodinium, Symbiodinium, and Prorocentrum, as they all are small sized (<30 µm). In other marine protists, ciliates usually have relatively small macronuclear genome sizes (50-104 Mb) (Aury et al., 2006;Eisen et al., 2006;Coyne et al., 2011), but possess high copy numbers of rRNA gene (3385 to 705,287 rRNA genes per cell; Gong et al., 2013;Wang et al., 2017). For example, Tetrahymena thermophila has a 100 Mb of macronuclear genome and 9000 copies of rRNA gene per cell (Eisen et al., 2006). ...
Article
Dinoflagellates are an ecologically important group of protists in aquatic environment and have evolved many unusual and enigmatic genomic features such as immense genome sizes, high repeated genes, and a large portion of hydroxymethyluracil in DNA. Although previous studies have observed positive correlations between the large subunit (LSU) rRNA gene copy number and genome size of a variety of eukaryotic organisms (e.g. higher plants and animals), or between cell volume and LSU rRNA gene copy number, and/or between genome size and cell size, which suggests a possible co-evolution among these three features in different lineages of life, it remains an open question regarding the relationships among these three parameters in dinoflagellates. For the first time, we estimated the copy numbers of the LSU rRNA gene, the genome sizes, and cell volumes within a broad range of dinoflagellates (covering 15 species of 11 genera) using single-cell qPCR-based assay (determining LSU rRNA gene copy number), FlowCAM (cell volume measurement), and ultraviolet spectrophotometry (genome size estimation). The measured copy number of LSU rRNA gene ranged from 398 ± 184 (Prorocentrum minimum) to 152,078 ± 33,555 copies·cell-1 (Alexandrium pacificum), while the genome size and the cell volume ranged from 5.6 ± 0.2 (Karlodinium veneficum) to 853 ± 19.9 pg·cell-1 (Pseliodinium pirum), and from 1,070 ± 225 (Kar. veneficum) to 168,474 ± 124,180 μm3 (Ps. pirum), respectively. Together with the three parameters measured in literature, there are significant positive linear correlations between LSU rRNA gene copy numbers and genome sizes, cell volumes and LSU rRNA gene copy numbers, and between genome sizes and cell volumes via comparisons of multi-model regression analyses, suggesting a dependence of genome size and rRNA gene copy number on the cell volumes of dinoflagellates. Validation of the measurement methods was conducted via comparisons between reported data in the literature and that predicted using the linear equations we obtained, and between genome size measured by flow cytometry (FCM) and ultraviolet spectrophotometry (Nanodrop). These results provide insightful understandings of dinoflagellate evolution in terms of the relationships among genomes, gene copy number, and cell volume, and of rRNA gene-based studies in intra-populational and intra-individual genetic diversity, taxonomy, and diversity assessment in the environment of dinoflagellates. The results also provide a dataset useful for reads calibration in environmental metabarcoding studies of dinoflagellates and selection of candidate species for whole genome sequencing.
... Numerous proteases and lytic enzymes from parasites are wellknown virulence factors that can destroy host tissues, such as the extracellular matrix and interstitial tissues, through catabolism of host membranes and cytoadherence (64). Several studies have reported on the existence of multiple types of cysteine proteases in I. multifiliis according to gelatin-precast zymography and genomic information from the macro-nucleus (7,(64)(65)(66). These virulence factors can directly affect erythrocytes, thus inducing hemolysis and anemia, as shown in this study (decrease in Ht from 39 to 21% after I. multifiliis infection) as well as other studies (55,57,58). ...
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Ichthyophthirius multifiliis is a major pathogen that causes a high mortality rate in trout farms. However, systemic responses to the pathogen and its interactions with multiple organs during the course of infection have not been well described. In this study, dual-organ transcriptomic responses in the liver and head kidney and hemato-serological indexes were profiled under I. multifiliis infection and recovery to investigate systemic immuno-physiological characteristics. Several strategies for massive transcriptomic interpretation, such as differentially expressed genes (DEGs), Poisson linear discriminant (PLDA), and weighted gene co-expression network analysis (WGCNA) models were used to investigate the featured genes/pathways while minimizing the disadvantages of individual methods. During the course of infection, 6,097 and 2,931 DEGs were identified in the head kidney and liver, respectively. Markers of protein processing in the endoplasmic reticulum, oxidative phosphorylation, and the proteasome were highly expressed. Likewise, simultaneous ferroptosis and cellular reconstruction was observed, which is strongly linked to multiple organ dysfunction. In contrast, pathways relevant to cellular replication were up-regulated in only the head kidney, while endocytosis- and phagosome-related pathways were notably expressed in the liver. Moreover, interestingly, most immune-relevant pathways (e.g., leukocyte trans-endothelial migration, Fc gamma R-mediated phagocytosis) were highly activated in the liver, but the same pathways in the head kidney were down-regulated. These conflicting results from different organs suggest that interpretation of co-expression among organs is crucial for profiling of systemic responses during infection. The dual-organ transcriptomics approaches presented in this study will greatly contribute to our understanding of multi-organ interactions under I. multifiliis infection from a broader perspective.
... The massive retention of duplicates 39 ubiquitin carboxyl-terminal hydrolase family members and 15 members of the threonine proteases occurred due to extensive gene duplication events. This mirrors the fundamental function of the proteasome system in I. multifiliis regulating cell-cycle and stress responses [24]. ...
... I. multifiliis contains 9 of the 14 highly conserved core proteins associated with centriole and basal body biogenesis and function [24]. ...
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The skin mucus is the fish primary defense barrier protecting from infections via the skin epidermis. In a previous study, we have investigated the proteome of common carp (Cyprinus carpio) skin mucus at two different time points (1 and 9 days) post-exposure to Ichthyophthirius multifiliis. Applying a nano-LC ESI MS/MS technique, we have earlier revealed that the abundance of 44 skin mucus proteins has been differentially regulated including proteins associated with host immune responses and wound healing. Herein, in skin mucus samples, we identified six proteins of I. multifiliis associated with the skin mucus in common carp. Alpha and beta tubulins were detected in addition to the elongation factor alpha, 26S proteasome regulatory subunit, 26S protease regulatory subunit 6B, and heat shock protein 90. The identified proteins are likely involved in motility, virulence, and general stress during parasite growth and development after parasite attachment and invasion. Two KEGG pathways, phagosome and proteasome, were identified among these parasite proteins, mirroring the proteolytic and phagocytic activities of this parasite during host invasion, growth, and development, which represent a plausible host invasion strategy of this parasite. The results obtained from this study can support revealing molecular aspects of the interplay between carp and I. multifiliis and may help us understand the I. multifiliis invasion strategy at the skin mucus barrier. The data may advance the development of novel drugs, vaccines, and diagnostics suitable for the management and prevention of ichthyophthiriosis in fish.
... Ciliate mitogenomes are generally in size of 20-70 kb, with high A+T content (58.53% for N. ovalis~81.51% for I. multifiliis) and relatively large gene complements (20-30 protein-coding genes) [39,41,44]. Mitochondrial DNA has been proven extremely useful in phylogenetic analyses to determine ciliate relationships and species delimitation [45]. ...
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Determination and comparisons of complete mitochondrial genomes (mitogenomes) are important to understand the origin and evolution of mitochondria. Mitogenomes of unicellular protists are particularly informative in this regard because they are gene-rich and display high structural diversity. Ciliates are a highly diverse assemblage of protists and their mitogenomes (linear structure with high A+T content in general) were amongst the first from protists to be characterized and have provided important insights into mitogenome evolution. Here, we report novel mitogenome sequences from three representatives (Strombidium sp., Strombidium cf. sulcatum, and Halteria grandinella) in two dominant ciliate lineages. Comparative and phylogenetic analyses of newly sequenced and previously published ciliate mitogenomes were performed and revealed a number of important insights. We found that the mitogenomes of these three species are linear molecules capped with telomeric repeats that differ greatly among known species. The genomes studied here are highly syntenic, but larger in size and more gene-rich than those of other groups. They also all share an AT-rich tandem repeat region which may serve as the replication origin and modulate initiation of bidirectional transcription. More generally we identified a split version of ccmf, a cytochrome c maturation-related gene that might be a derived character uniting taxa in the subclasses Hypotrichia and Euplotia. Finally, our mitogenome comparisons and phylogenetic analyses support to reclassify Halteria grandinella from the subclass Oligotrichia to the subclass Hypotrichia. These results add to the growing literature on the unique features of ciliate mitogenomes, shedding light on the diversity and evolution of their linear molecular architecture.
... Similar to free-living ciliates, symbiotic rumen ciliates also have nuclear dimorphism [14]. The MAC genomes of about 10 ciliate species, mostly model ciliates, have been sequenced, including T. thermophila [15][16][17], P. tetraurelia [18], P. caudatum [19], Ichthyophthirius multifiliis [20], and O. trifallax [21]. The MAC genomes and subsequent genomic studies have provided new knowledge that was otherwise impossible to gain, including several unique genetic features such as single-gene nanochromosomes, tiny introns [22], and extensive genome fragmentation [21]. ...
Article
Entodinium caudatum is an anaerobic binucleated ciliate representing the most dominant protozoal species in the rumen. However, its biological features are largely unknown due to the inability to establish an axenic culture. In this study, we primally sequenced its macronucleus (MAC) genome to aid the understanding of its metabolism, physiology, ecology. We isolated the MAC of E. caudatum strain MZG-1 and sequenced the MAC genome using Illumina MiSeq, MinION, and PacBio RSII systems. De novo assembly of the MiSeq sequence reads followed with subsequent scaffolding with MinION and PacBio reads resulted in a draft MAC genome about 117 Mbp. A large number of carbohydrate-active enzymes were likely acquired through horizontal gene transfer. About 8.74% of the E. caudatum predicted proteome was predicted as proteases. The MAC genome of E. caudatum will help better understand its important roles in rumen carbohydrate metabolism, and interaction with other members of the rumen microbiome.
... Genome sequencing and analysis of the transcriptionally active macronucleus of I. multifiliis has revealed several gene classes functioning in behavior, cellular functions, and host immunogenicity, including protein kinases, membrane transporters, proteases, and surface antigens, providing avenues for selecting vaccine and drug targets [164]. These gene families are identified as lead vaccine targets in many parasites of medical and veterinary importance. ...
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Globally, parasites are increasingly being recognized as catastrophic agents in both aquaculture sector and in the wild aquatic habitats leading to an estimated annual loss between 1.05 billion and 9.58 billion USD. The currently available therapeutic and control measures are accompanied by many limitations. Hence, vaccines are recommended as the "only green and effective solution" to address these concerns and protect fish from pathogens. However, vaccine development warrants a better understanding of host-parasite interaction and parasite biology. Currently, only one commercial parasite vaccine is available against the ectoparasite sea lice. Additionally, only a few trials have reported potential vaccine candidates against endoparasites. Transcriptome, genome, and proteomic data at present are available only for a limited number of aquatic parasites. Omics-based interventions can be significant in the identification of suitable vaccine candidates, finally leading to the development of multivalent vaccines for significant protection against parasitic infections in fish. The present review highlights the progress in the immunobiology of pathogenic parasites and the prospects of vaccine development. Finally, an approach for developing a multivalent vaccine for parasitic diseases is presented. Data sources to prepare this review included Pub-med, google scholar, official reports, and websites.
... Only the species for which 5′ UTR annotation information was available for more than 25% of the protein-coding genes were considered in the analyses. Among all the 479 species meet this criteria, Ichthyophthirius multifiliis was excluded since UAA and UAG are reassigned to encode glutamine in this species 134 , which would interfere with the uORF and NTE prediction. ...
Article
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Upstream open reading frames (uORFs) play widespread regulatory functions in modulating mRNA translation in eukaryotes, but the principles underlying the genomic distribution and evolution of uORFs remain poorly understood. Here, we analyze ~17 million putative canonical uORFs in 478 eukaryotic species that span most of the extant taxa of eukaryotes. We demonstrate how positive and purifying selection, coupled with differences in effective population size (Ne), has shaped the contents of uORFs in eukaryotes. Besides, gene expression level is important in influencing uORF occurrences across genes in a species. Our analyses suggest that most uORFs might play regulatory roles rather than encode functional peptides. We also show that the Kozak sequence context of uORFs has evolved across eukaryotic clades, and that noncanonical uORFs tend to have weaker suppressive effects than canonical uORFs in translation regulation. This study provides insights into the driving forces underlying uORF evolution in eukaryotes.
... This tendency is particularly apparent in intracellular parasites such as microsporidia and apicomplexans, in which reductions in metabolic pathways evolved together with dramatic reductions in the genome (Heinz et al. 2012;Woo et al. 2015;Mathur et al. 2019). However, other parasites, such as parasitic kinetoplastids and ciliates, seem to have maintained most of their canonical physiological functions, and reduction primarily manifests as losses of functionally redundant paralogs (Coyne et al. 2011;Jackson et al. 2016). In addition to gene loss, parasite genomes are shaped by innovations, that is, gains of truly novel parasite-specific functions. ...
... However, although genomes of most important parasites have been sequenced, the genomes of their free-living neighbors are rarely studied, rendering comparative genomic studies impossible. Moreover, those free-living lineages that have been sampled primarily include organisms thriving in aerobic environments, for example, free-living kinetoplastids related to trypanosomatids (Jackson et al. 2016), chrompodellids (Janou skovec et al. 2015) related to apicomplexans (Woo et al. 2015;Mathur et al. 2019), and free-living ciliates related to Ichthyophthirius multifiliis (Coyne et al. 2011). Therefore, there exists an unsampled diversity of anaerobic free-living organisms that could provide novel comparative insights into the genome evolution of their parasitic relatives. ...
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The transition of free-living organisms to parasitic organisms is a mysterious process that occurs in all major eukaryotic lineages. Parasites display seemingly unique features associated with their pathogenicity; however, it is important to distinguish ancestral preconditions to parasitism from truly new parasite-specific functions. Here, we sequenced the genome and transcriptome of anaerobic free-living M. balamuthi and performed phylogenomic analysis of four related members of the Archamoebae, including Entamoeba histolytica, an important intestinal pathogen of humans. We aimed to trace gene histories throughout the adaptation of the aerobic ancestor of Archamoebae to anaerobiosis and throughout the transition from a free-living to a parasitic lifestyle. These events were associated with massive gene losses that, in parasitic lineages, resulted in a reduction in structural features, complete losses of some metabolic pathways, and a reduction in metabolic complexity. By reconstructing the features of the common ancestor of Archamoebae, we estimated preconditions for the evolution of parasitism in this lineage. The ancestor could apparently form chitinous cysts, possessed proteolytic enzyme machinery, compartmentalized the sulfate activation pathway in mitochondrion-related organelles, and possessed the components for anaerobic energy metabolism. After the split of Entamoebidae, this lineage gained genes encoding surface membrane proteins that are involved in host-parasite interactions. In contrast, gene gains identified in the M. balamuthi lineage were predominantly associated with polysaccharide catabolic processes. A phylogenetic analysis of acquired genes suggested an essential role of lateral gene transfer in parasite evolution (Entamoeba) and in adaptation to anaerobic aquatic sediments (Mastigamoeba).