Abstract
BackgroundThe genome of the apicomplexan parasite Cystoisospora suis (syn. Isospora suis) has recently been sequenced and annotated, opening the possibility for the identification of novel therapeutic targets against cystoisosporosis. It was previously proposed that a 42 kDa uncharacterized merozoite protein, encoded by gene CSUI_005805, might be a relevant vaccine candidate due to its high immunogenic score, high expression level and species-specificity as determined in silico.MethodsThe 1170 bp coding sequence of the CSUI_005805 gene was PCR amplified and cloned into the bacterial expression vector pQE-31. The specificity of the expressed recombinant protein was evaluated in an immunoblot, and relative levels of expression in different developmental stages and subcellular localization were determined by quantitative real-time PCR and indirect immunofluorescence assay, respectively.ResultsThe CSUI_005805 gene encoded for a 389 amino acid protein containing a histidine-rich region. Quantitative RT-PCR showed that CSUI_005805 was differentially expressed during the early development of C. suis in vitro, with higher transcript levels in merozoites compared to sporozoites. The recombinant protein was specifically recognized by sera from chicken immunized with recombinant CSUI_005805 protein and sera from piglets experimentally infected with C. suis, all of which suggested that despite prokaryotic expression, the recombinant CSUI_005805 protein maintained antigenic determinants and could elicit an immune response in the host. Immunofluorescence labelling and confocal microscopy revealed localization primarily at the surface of the parasite.ConclusionsThe results suggest that CSUI_005805 is highly expressed in merozoites and might thus be critical for their survival and establishment inside host cells. Owing to its specificity, localization and expression pattern, CSUI_005805 could be exploited as an attractive candidate for alternative control strategies against C. suis such as vaccines.
Highlights
The genome of the apicomplexan parasite Cystoisospora suis
An alternative approach would be the introduction of subunit or recombinant vaccines, which demands a systematic search for antigenic proteins to find appropriate vaccine candidates for testing
Cloning and sequence analysis The CSUI_005805 gene included a 302 bp 5′-untranslated region (5′-Untranslated region (UTR)) before the ATG initiation codon and a 1170 bp coding sequence terminating with the TAG stop codon (Fig. 1), followed by a 44 bp 3′-UTR
Summary
The genome of the apicomplexan parasite Cystoisospora suis Isospora suis), an enteric protozoan parasite of swine, is a member of the phylum Apicomplexa and the causative agent of neonatal porcine coccidiosis (cystoisosporosis). It is distributed worldwide with high prevalence rates in intensive pig breeding facilities regardless of the farm management system [1, 2]. Virulent vaccines in large amounts, are impractical for the use in swine as even low infection doses can lead to disease in very young piglets [18], and attenuated lines have not yet been introduced for C. suis. An alternative approach would be the introduction of subunit or recombinant vaccines, which demands a systematic search for antigenic proteins to find appropriate vaccine candidates for testing
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