title: Identification of a gene cluster for cell-surface genes of the SRS superfamily in Neospora caninum and characterization of the novel SRS9 gene.
abstract: Here we present the detection of a gene cluster for Neospora caninum surface genes, similar to the Toxoplasma gondii SRS9 locus, and the cloning and characterization of the NcSRS9 gene. PCR genome walking, using NcBSR4 gene as a framework, allows the identification, upstream NcBSR4, of 2 sequences homologous to the SRS5 and the Ubiquinol-cytochrome C reductase genes and, downstream NcBSR4, of an ORF of 1191 bp coding for a 396-amino acid polypeptide with 59% similarity to the TgSRS9 antigen. A putative 39-residue signal peptide was found at the NH2-terminus followed by a hydrophilic region, and a potential site for a glycosylphosphatidylinositol anchor at the COOH-terminus. A recombinant NcSRS9 protein was produced and was recognized on a Western blot by a low proportion of sera from a panel of naturally infected cows and calves. In addition, Western blot analysis using polyclonal anti-rNcSRS9 revealed stage-specific expression of NcSRS9 in bradyzoites but not in tachyzoites, and immunohistochemistry on brain from a congenitally infected calf showed NcSRS9 recognition in bradyzoites contained in tissue cysts. However, bradyzoite-specific expression of NcSRS9 could not be proven by immunofluorescence on bradyzoites obtained in vitro and RT-PCR analysis showed no significant variations of NcSRS9 transcripts during in vitro tachyzoite-bradyzoite switch, probably due to incomplete maturity of in vitro bradyzoites. Initial characterization of NcSRS9 in this study may lead to further studies for a better understanding of N. caninum persistence.
authors.affiliation: SALUVET, Animal Health Department, Faculty of Veterinary Sciences, Complutense University of Madrid, Ciudad Universitaria s/n, 28040 Madrid, Spain.
authors.forename: V
authors.initials: V
authors.lastname: Risco-Castillo
authors.affiliation: SALUVET, Animal Health Department, Faculty of Veterinary Sciences, Complutense University of Madrid, Ciudad Universitaria s/n, 28040 Madrid, Spain.
authors.forename: V
authors.initials: V
authors.lastname: Marugán-Hernández
authors.affiliation: SALUVET, Animal Health Department, Faculty of Veterinary Sciences, Complutense University of Madrid, Ciudad Universitaria s/n, 28040 Madrid, Spain.
authors.forename: A
authors.initials: A
authors.lastname: Fernández-García
authors.affiliation: SALUVET, Animal Health Department, Faculty of Veterinary Sciences, Complutense University of Madrid, Ciudad Universitaria s/n, 28040 Madrid, Spain.
authors.forename: A
authors.initials: A
authors.lastname: Aguado-Martínez
authors.affiliation: SALUVET, Animal Health Department, Faculty of Veterinary Sciences, Complutense University of Madrid, Ciudad Universitaria s/n, 28040 Madrid, Spain.
authors.forename: E
authors.initials: E
authors.lastname: Jiménez-Ruiz
authors.affiliation: SALUVET, Animal Health Department, Faculty of Veterinary Sciences, Complutense University of Madrid, Ciudad Universitaria s/n, 28040 Madrid, Spain.
authors.forename: S
authors.initials: S
authors.lastname: Rodríguez-Marco
authors.affiliation: SALUVET, Animal Health Department, Faculty of Veterinary Sciences, Complutense University of Madrid, Ciudad Universitaria s/n, 28040 Madrid, Spain.
authors.forename: G
authors.initials: G
authors.lastname: Alvarez-García
authors.affiliation: SALUVET, Animal Health Department, Faculty of Veterinary Sciences, Complutense University of Madrid, Ciudad Universitaria s/n, 28040 Madrid, Spain.
authors.forename: L M
authors.initials: LM
authors.lastname: Ortega-Mora
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000913:Antibodies, Protozoan / Q000276:immunology*; D000953:Antigens, Protozoan / Q000276:immunology*; D000954:Antigens, Surface / Q000276:immunology*; D001921:Brain / Q000469:parasitology; D002417:Cattle; D002874:Chromosome Mapping / Q000662:veterinary; D003048:Coccidiosis / Q000469:parasitology / Q000662:veterinary*; D005260:Female; D056426:Genetic Loci; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D005810:Multigene Family; D018688:Neospora / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D021382:Protein Sorting Signals / Q000235:genetics; D015800:Protozoan Proteins / Q000235:genetics* / Q000378:metabolism; D011817:Rabbits; D011994:Recombinant Proteins; D016415:Sequence Alignment / Q000662:veterinary; D026801:Synteny
chemical_list: D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan; D000954:Antigens, Surface; D008565:Membrane Proteins; D021382:Protein Sorting Signals; D015800:Protozoan Proteins; D011994:Recombinant Proteins