RNA Seq analysis of the Eimeria tenella gametocyte transcriptome reveals clues about the molecular basis for sexual reproduction and oocyst biogenesis

Abstract

BackgroundThe protozoan Eimeria tenella is a common parasite of chickens, causing avian coccidiosis, a disease of on-going concern to agricultural industries. The high prevalence of E. tenella can be attributed to the resilient oocyst stage, which is transmitted between hosts in the environment. As in related Coccidia, development of the eimerian oocyst appears to be dependent on completion of the parasite’s sexual cycle. RNA Seq transcriptome profiling offers insights into the mechanisms governing the biology of E. tenella sexual stages (gametocytes) and the potential to identify targets for blocking parasite transmission.ResultsComparisons between the sequenced transcriptomes of E. tenella gametocytes and two asexual developmental stages, merozoites and sporozoites, revealed upregulated gametocyte transcription of 863 genes. Many of these genes code for proteins involved in coccidian sexual biology, such as oocyst wall biosynthesis and fertilisation, and some of these were characterised in more depth. Thus, macrogametocyte-specific expression and localisation was confirmed for two proteins destined for incorporation into the oocyst wall, as well as for a subtilisin protease and an oxidoreductase. Homologues of an oocyst wall protein and oxidoreductase were found in the related coccidian, Toxoplasma gondii, and shown to be macrogametocyte-specific. In addition, a microgametocyte gamete fusion protein, EtHAP2, was discovered.ConclusionsThe need for novel vaccine candidates capable of controlling coccidiosis is rising and this panel of gametocyte targets represents an invaluable resource for development of future strategies to interrupt parasite transmission, not just in Eimeria but in other Coccidia, including Toxoplasma, where transmission blocking is a relatively unexplored strategy.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1298-6) contains supplementary material, which is available to authorized users.

Distribution and Processing of Eimeria nieschulziOWP13, a New Protein of the COWP Family

“…By immunocytochemistry, the OWP6 of Eimeria tenella has been detected in WFBI but not in the oocyst wall (polyclonal serum) (Walker et al. ).…”

“…Potential candidates were published by Walker et al. (), but are not studied yet. As known from yeast, other enzymes can also be responsible for oxidative cross‐linking of tyrosine residues (Briza et al.…”

“…; Walker et al. ). BLAST analysis suggests at least one more formerly uncharacterized typical OWP in an Eimeria species.…”

Eimeria genomics: Where are we now and where are we going?

“…Exogenous and extracellular endogenous life cycle forms including the oocysts (unsporulated, sporulating and sporulated), sporozoites and merozoites have been most commonly sampled for transcriptomic analyses as a consequence of their availability, first as ESTs/ORESTES and more recently by Illumina-based RNAseq (Fig. 1) (Abrahamsen et al, 1993;Wan et al, 1999;Miska et al, 2008;Kim et al, 2010;Schwarz et al, 2010;Aarthi et al, 2011;Dong et al, 2011;Tuda et al, 2011;Amiruddin et al, 2012;Novaes et al, 2012;Heitlinger et al, 2014;Reid et al, 2014;Walker et al, 2015). As described for genome sequencing, progress from Sanger to NGS technologies has provided opportunities to expand transcriptome sequencing to additional E. tenella life cycle stages and species such as E. falciformis.…”

“…The requirement for microgram quantities of total RNA for library preparation restricted transcriptomic sequencing to only the most accessible lifecycle stages, hindering understanding of much of the eimerian life cycle's complexity. These problems have been overcome for some species and life cycle stages, such as E. tenella gametocytes, where specific culture and recovery protocols have allowed complementation of previous genomic analyses through development of the first RNAseq data (Katrib et al, 2012;Walker et al, 2015). Life cycle stages including trophozoites and developing schizonts are yet to be sampled, but complementary techniques including laser dissection from in vitro or in vivo cultured samples, ex vivo explant culture and fluorescence-activated cell sorting (FACS) of parasites transfected to express fluorescent reporter genes (Clark et al, 2008;Yan et al, 2009) offer good opportunities to access these stages.…”