Isolation, identification and expression of a Fasciola hepatica cDNA encoding a 2.9-kDa recombinant protein for the diagnosis of ovine fasciolosis (original) (raw)

Abstract

A 400-bp Fasciola hepatica cDNA clone was isolated from an expression library by immunological screening using rat sera taken 2 weeks after experimental infection. The nucleotide sequence of the cDNA revealed the presence of an open reading frame of 78 bp which encoded a 25 amino acid polypeptide with a predicted molecular weight of 2.9 kDa. This polypeptide was expressed in bacteria as a GST-fusion protein and used for the production of specific antigen. The 2.9 kDa recombinant protein (APS) was evaluated against sera from experimentally infected sheep using an indirect ELISA, and the results were compared to those obtained using F. hepatica excretory/secretory products (ESP). The pattern of IgG was very similar both against the recombinant and the native proteins, increasing early following the infection. After treatment with triclabendazole, the IgG response against the APS seroreverted to negative values, whereas it remained elevated against the ESP. We conclude that this recombinant protein could be used in diagnostic assays for the identification of recently infected sheep.

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Acknowledgements

This work was supported, in part, by a research grant from the Xunta de Galicia (XUGA, Spain) to Dr. A. Paz-Silva, and by the Projects XUGA PGIDT00PXI26102PR (Xunta de Galicia, Spain), DCICYT AGL2001-2990 (Ministerio de Ciencia y Tecnología, Spain), NIH-NIGMS Grant No. SO6 GM08224, NCRR-RCMI G12 RR03051 (USA), and NSF-EPSCOR SPACS Program (USA), and complies with the current laws for Animal Health Research in Spain. We are indebted to Mrs. B. Valcárcel for preparing and editing the manuscript. The nucleotide sequence data reported in this paper have been submitted to GenBank database with accession no. AY676331.

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Authors and Affiliations

1. Departamento de Patología Animal, Parasitología y Enfermedades Parasitarias y Epidemiología, Facultad de Veterinaria, Universidad de Santiago de Compostela, 27071 , Lugo, Spain
   A. Paz-Silva, R. Sánchez-Andrade, M. Arias, P. Morrondo & P. Díez-Baños
2. Laboratory of Parasite Immunology and Pathology, School of Medicine, University of Puerto Rico, Suite 617-A, Medical Sciences Campus, 00936–5067, San Juan, Puerto Rico
   G. V. Hillyer
3. Department of Biochemistry, School of Medicine, University of Puerto Rico, Medical Sciences Campus, 00936–5067, San Juan, Puerto Rico
   J. R. Rodríguez-Medina

Authors

1. A. Paz-Silva
2. G. V. Hillyer
3. R. Sánchez-Andrade
4. J. R. Rodríguez-Medina
5. M. Arias
6. P. Morrondo
7. P. Díez-Baños

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Correspondence toA. Paz-Silva.

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Paz-Silva, A., Hillyer, G.V., Sánchez-Andrade, R. et al. Isolation, identification and expression of a Fasciola hepatica cDNA encoding a 2.9-kDa recombinant protein for the diagnosis of ovine fasciolosis.Parasitol Res 95, 129–135 (2005). https://doi.org/10.1007/s00436-004-1202-9

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• Received: 19 July 2004
• Accepted: 20 July 2004
• Published: 18 December 2004
• Issue date: January 2005
• DOI: https://doi.org/10.1007/s00436-004-1202-9

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