The transcriptome of the rumen ciliate Entodinium caudatum reveals some of its metabolic features

Abstract

BackgroundRumen ciliates play important roles in rumen function by digesting and fermenting feed and shaping the rumen microbiome. However, they remain poorly understood due to the lack of definitive direct evidence without influence by prokaryotes (including symbionts) in co-cultures or the rumen. In this study, we used RNA-Seq to characterize the transcriptome of Entodinium caudatum, the most predominant and representative rumen ciliate species.ResultsOf a large number of transcripts, > 12,000 were annotated to the curated genes in the NR, UniProt, and GO databases. Numerous CAZymes (including lysozyme and chitinase) and peptidases were represented in the transcriptome. This study revealed the ability of E. caudatum to depolymerize starch, hemicellulose, pectin, and the polysaccharides of the bacterial and fungal cell wall, and to degrade proteins. Many signaling pathways, including the ones that have been shown to function in E. caudatum, were represented by many transcripts. The transcriptome also revealed the expression of the genes involved in symbiosis, detoxification of reactive oxygen species, and the electron-transport chain. Overall, the transcriptomic evidence is consistent with some of the previous premises about E. caudatum. However, the identification of specific genes, such as those encoding lysozyme, peptidases, and other enzymes unique to rumen ciliates might be targeted to develop specific and effective inhibitors to improve nitrogen utilization efficiency by controlling the activity and growth of rumen ciliates. The transcriptomic data will also help the assembly and annotation in future genomic sequencing of E. caudatum.ConclusionAs the first transcriptome of a single species of rumen ciliates ever sequenced, it provides direct evidence for the substrate spectrum, fermentation pathways, ability to respond to various biotic and abiotic stimuli, and other physiological and ecological features of E. caudatum. The presence and expression of the genes involved in the lysis and degradation of microbial cells highlight the dependence of E. caudatum on engulfment of other rumen microbes for its survival and growth. These genes may be explored in future research to develop targeted control of Entodinium species in the rumen. The transcriptome can also facilitate future genomic studies of E. caudatum and other related rumen ciliates.

Extending Burk Dehority’s Perspectives on the Role of Ciliate Protozoa in the Rumen

“…En. caudatum still expresses hydrogenases (Wang et al, 2020), whereas hydrogenosome-linked hydrogenases in other protozoa might be more constitutively linked with fermentation pathways because some fermentative enzymes (especially if sensitive to O 2 ) also might be hydrogenosomal, as documented for Dasytricha and Isotricha (Williams and Coleman, 1992). Although counts of mixed protozoa were positively associated with methanogenesis (Guyader et al, 2014), a decrease in protozoal counts also was associated with a decrease in NDF digestibility and in dry matter intake.…”

“…The ciliate's cellular signaling mechanisms should be coordinated with its cell cycle (Firkins et al, 2007;Diaz et al, 2014), which is supported by transcriptomics profiling in En. caudatum (Wang et al, 2020). These same eukaryotic cell cycle controls should both increase growth and decrease growth rate, depending on stage of incubation.…”

“…In the transcriptome of En. caudatum, at least 25 different putative signal transduction pathways were recorded (Wang et al, 2020). Those transcripts relatively highly expressed included well-described eukaryotic pathways MAPK, Ras, calcium, cGMP-PKG, cAMP, FoxO, phosphatidylinositol, sphingolipid, TOR, PI3K-Akt, AMPK, Wnt, and Apelin.…”

“…Genomics and transcriptomics, empowered by NGS and TGS technologies, would be enabling approaches to help gain new insights into the fundamental biological features of rumen protozoa. Genomic (Park et al, 2018) and transcriptomic (Wang et al, 2020) studies of En. caudatum are providing starting points for future studies investigating roles of prokaryotes or interactions such as in defaunation studies (discussed previously).…”

“…For example, should we continue to study inhibiting agents without studying how the protozoal community adapts to them (and how they do it)? Indeed, Entodinium caudatum expressed eukaryotic stress response pathways when the inhibitors monensin and wortmannin were introduced (Wang et al, 2020). We questioned if protozoal autolysis in the rumen is less than that projected from cultures (Diaz et al, 2014).…”