RESUMO
Leucine aminopeptidase (LAP) and cathepsin L1 (CL1) are important enzymes for the pathogenesis and physiology of Fasciola hepatica. These enzymes were analysed in silico to design a chimeric protein containing the most antigenic sequences of LAP (GenBank; AAV59016.1; amino acids 192-281) and CL1 (GenBank CAC12806.1; amino acids 173-309). The cloned 681-bp chimeric fragment (rFhLAP-CL1) contains 270 bp from LAP and 411 bp from CL1, comprising three epitopes, DGRVVHLKY (amino acids 54-62) from LAP, VTGYYTVHSGSEVELKNLV (amino acids 119-137) and YQSQTCLPF (amino acids 161-169) from CL1. The ~25 kDa rFhLAP-CL1 chimeric protein was expressed from the pET15b plasmid in the Rosetta (DE3) Escherichia coli strain. The chimeric protein rFhLAP-CL1, which showed antigenic and immunogenic properties, was recognized in Western blot assays using F. hepatica-positive bovine sera, and induced strong, specific antibody responses following immunization in rabbits. The newly generated chimeric protein may be used as a diagnostic tool for detection of antibodies against F. hepatica in bovine sera and as an immunogen to induce protection against bovine fasciolosis.
Assuntos
Catepsina L/genética , Fasciola hepatica/genética , Fasciolíase/veterinária , Proteínas de Helminto/genética , Leucil Aminopeptidase/genética , Fígado/enzimologia , Animais , Catepsina L/análise , Catepsina L/imunologia , Bovinos , Doenças dos Bovinos/parasitologia , Epitopos/análise , Epitopos/genética , Epitopos/imunologia , Fasciola hepatica/imunologia , Fasciolíase/parasitologia , Expressão Gênica , Proteínas de Helminto/análise , Proteínas de Helminto/imunologia , Imunização , Leucil Aminopeptidase/análise , Leucil Aminopeptidase/imunologia , Coelhos , Proteínas Recombinantes de Fusão/análise , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologiaRESUMO
The aim of this study was to describe bovine neosporosis in dairy cattle from the Sierra region, Ecuador. A case-control study was performed on 841 dairy cattle from 5 dairy herds. The overall seroprevalence was 23.4% having significant association between abortion and seropositivity (p < .05). Additionally, 46 fetuses were recovered from a local slaughterhouse to evaluate the frequency of vertical transmission. Seventeen and 3 fetuses were positive by PCR and had compatible histopathological lesions, respectively. N. caninum infection must be considered as a relevant cause of reproductive losses in Ecuador.
Assuntos
Aborto Animal/epidemiologia , Doenças dos Bovinos/epidemiologia , Coccidiose/veterinária , Neospora/isolamento & purificação , Aborto Animal/parasitologia , Animais , Bovinos , Doenças dos Bovinos/parasitologia , Coccidiose/epidemiologia , Coccidiose/parasitologia , Indústria de Laticínios , Equador/epidemiologia , Feminino , Prevalência , Estudos SoroepidemiológicosRESUMO
Bovine tuberculosis is a major problem in many countries; hence, new and better diagnostic tools are urgently needed. In this work, we have tested ESAT6, CFP10, PE13, PE5, MPB70, TB10.4, and TB27.4 for their potentials as diagnostic markers in field animals from Northern Ireland, Mexico, and Argentina, regions with low, medium, and high prevalences of bovine tuberculosis, respectively. At all three sites, ESAT6 and CFP10 were superior diagnostic antigens, while their combination performed even better at the two sites where the combination was tested, providing the best coverage for the detection of diseased populations. The high sensitivity in the skin test reactor groups, combined with the high specificity in the tuberculosis-free groups, indicated that a diagnosis could correctly be made for 85% of the infected animals, based on their responses to these two antigens. Furthermore, TB10.4, PE13, and PE5 have the potential to supplement ESAT6 and CFP10 in a future five-component diagnostic cocktail.