Transcriptome-Wide Comparisons and Virulence Gene Polymorphisms of Host-Associated Genotypes of the Cnidarian Parasite Ceratonova shasta in Salmonids

Abstract

Ceratonova shasta is an important myxozoan pathogen affecting the health of salmonid fishes in the Pacific Northwest of North America. C. shasta exists as a complex of host-specific genotypes, some with low to moderate virulence, and one that causes a profound, lethal infection in susceptible hosts. High throughput sequencing methods are powerful tools for discovering the genetic basis of these host/virulence differences, but deep sequencing of myxozoans has been challenging due to extremely fast molecular evolution of this group, yielding strongly divergent sequences that are difficult to identify, and unavoidable host contamination. We designed and optimized different bioinformatic pipelines to address these challenges. We obtained a unique set of comprehensive, host-free myxozoan RNA-seq data from C. shasta genotypes of varying virulence from different salmonid hosts. Analyses of transcriptome-wide genetic distances and maximum likelihood multigene phylogenies elucidated the evolutionary relationship between lineages and demonstrated the limited resolution of the established Internal Transcribed Spacer marker for C. shasta genotype identification, as this marker fails to differentiate between biologically distinct genotype II lineages from coho salmon and rainbow trout. We further analyzed the datasets based on polymorphisms in two gene groups related to virulence: cell migration and proteolytic enzymes including their inhibitors. The developed SNP-calling pipeline identified polymorphisms between genotypes and demonstrated that variations in both motility and protease genes were associated with different levels of virulence of C. shasta in its salmonid hosts. The prospective use of proteolytic enzymes as promising candidates for targeted interventions against myxozoans in aquaculture is discussed. We developed host-free transcriptomes of a myxozoan model organism from strains that exhibited different degrees of virulence, as a unique source of data that will foster functional gene analyses and serve as a base for the development of potential therapeutics for efficient control of these parasites.

Genetic Diversity of Serine Protease Inhibitors in Myxozoan (Cnidaria, Myxozoa) Fish Parasites

“…This was in accordance with the findings of Irving et al [ 9 ] regarding the phylogenetic analysis of non-vertebrate serpins. The genetic differences among C. shasta serpins detected by Alama-Bermejo et al [ 32 ] were confirmed in our study. Although all three analyzed serpins were putatively intracellular (clade B), Cs-S3 were located far from Cs-S1 & Cs-S2 on the phylogenetic tree, potentially indicating an independent origin.…”

“…The function of myxozoan serpins was predicted based on translated protein sequences only. However, it is likely that serpins may be involved in anti-inflammatory and anti-coagulatory processes during host invasion [ 32 ]. Previous study concluded that the phylogenetic clustering according to serpin type is more prominent for vertebrates [ 9 ].…”

“…PRJNA485581) [ 29 ]; Ceratonova shasta (BioProject No. PRJNA241036) [ 32 ]; Sphaerospora molnari (BioProject No. PRJNA522909) [ 33 ]; Thelohanellus kitauei (BioProject No.…”

Evolutionary Analysis of Cystatins of Early-Emerging Metazoans Reveals a Novel Subtype in Parasitic Cnidarians

“…Recently, next-generation sequencing (NGS) data have become available for three of these parasite lineages, i.e., two Edwardsiella spp. [ 44 , 45 ], Polypodium hydriforme [ 46 , 47 ], and several species of the large group Myxozoa [ 47 , 48 , 49 , 50 , 51 , 52 , 53 , 54 ]. While Edwardsiella and Polypodium parasitize their hosts exclusively during the larval stage of their development [ 45 , 55 ], the microscopic myxozoans are an entirely parasitic group with complex life cycles [ 56 ].…”

Serine protease inhibitors of the whirling disease parasite Myxobolus cerebralis (Cnidaria, Myxozoa): Expression profiling and functional predictions

“…The serpins in the genome of T. kitauei extensively diversified compared to those of the free-living cnidarians or other myxozoans [29,30]. The recently elucidated myxozoan transcriptomes, such as those of Henneguya salminicola [31], Ceratonova shasta [32] or Sphaerospora molnari [33], have made it possible to identify new myxozoan serpin homologs, and to study their genetic diversity in detail [30]. Among 224 serpins from 71 species ranging from protists to vertebrates (including ten myxozoan species), seven M. cerebralis serpins (named Mc-S1 to Mc-S7) were distinguished, which formed four clusters on the phylogenetic tree.…”