Hostname: page-component-78c5997874-t5tsf Total loading time: 0 Render date: 2024-11-05T22:24:43.559Z Has data issue: false hasContentIssue false
  • English
  • Français

Specific antibody responses against Neospora caninum recombinant rNcGRA7, rNcSAG4, rNcBSR4 and rNcSRS9 proteins are correlated with virulence in mice

Published online by Cambridge University Press:  24 January 2013

ELENA JIMÉNEZ-RUIZ ,
GREGORI BECH-SÀBAT ,
GEMA ÁLVAREZ-GARCÍA ,
JAVIER REGIDOR-CERRILLO ,
LAURA HINOJAL-CAMPAÑA  and
LUIS M. ORTEGA-MORA
ELENA JIMÉNEZ-RUIZ
Affiliation:
SALUVET, Animal Health Department, Faculty of Veterinary Sciences, Complutense University of Madrid, Ciudad Universitaria s/n, 28040 Madrid, Spain
GREGORI BECH-SÀBAT
Affiliation:
SALUVET, Animal Health Department, Faculty of Veterinary Sciences, Complutense University of Madrid, Ciudad Universitaria s/n, 28040 Madrid, Spain
GEMA ÁLVAREZ-GARCÍA*
Affiliation:
SALUVET, Animal Health Department, Faculty of Veterinary Sciences, Complutense University of Madrid, Ciudad Universitaria s/n, 28040 Madrid, Spain
JAVIER REGIDOR-CERRILLO
Affiliation:
SALUVET, Animal Health Department, Faculty of Veterinary Sciences, Complutense University of Madrid, Ciudad Universitaria s/n, 28040 Madrid, Spain
LAURA HINOJAL-CAMPAÑA
Affiliation:
SALUVET, Animal Health Department, Faculty of Veterinary Sciences, Complutense University of Madrid, Ciudad Universitaria s/n, 28040 Madrid, Spain
LUIS M. ORTEGA-MORA
Affiliation:
SALUVET, Animal Health Department, Faculty of Veterinary Sciences, Complutense University of Madrid, Ciudad Universitaria s/n, 28040 Madrid, Spain
*
*Corresponding author: SALUVET, Animal Health Department, Faculty of Veterinary Sciences, Complutense University of Madrid, Ciudad Universitaria s/n. 28040-Madrid, Spain. Tel: +34 913944095. Fax: +34 913944098. E-mail: [email protected]
Get access Rights & Permissions [Opens in a new window]

Summary

The intraspecific diversity of Neospora caninum is a determinant for in vivo parasite virulence and in vitro parasite behaviour. The relationship between isolate virulence and specific antibody responses against key parasite proteins has not been well characterized. The response kinetics and the differences in specific anti-rNcGRA7, -rNcSAG4, -rNcBSR4 and -rNcSRS9 antibody levels were analysed by recombinant protein-based ELISA in groups of mice inoculated with 10 different N. caninum isolates that differ in their virulence. The majority of the virulence parameters analysed correlated with the specific antibody levels against the 4 recombinant proteins. The antibodies developed against the highly immunogenic protein NcGRA7 were significantly higher in mice inoculated with high virulence isolates than in those inoculated with low-to-moderate virulence isolates in both non-pregnant and pregnant mouse models. Moreover, these levels were correlated with the anti-N. caninum IgG1 and IgG2a responses and the in vitro tachyzoite yield at 56 h. The antibodies directed against the bradyzoite-specific proteins were not detected in a non-pregnant mouse model. However, some seropositive mice were found in groups inoculated with high virulence isolates in a pregnant mouse model. NcGRA7 and NcSAG4 are proteins clearly correlated with virulence, and to a lesser extent NcBSR4 and NcSRS9 proteins. Moreover, antibodies to bradyzoite-specific proteins appear to also be related to virulence in mice. Further analyses should be performed in order to verify the usefulness of these proteins as predictive markers for virulence in an experimental bovine model of neosporosis.

Keywords

Neospora caninumvirulenceBALB/c micerNcGRA7rNcSAG4rNcBSR4rNcSRS9antibody response
Type
Research Article
Information
Parasitology , Volume 140 , Issue 5 , April 2013 , pp. 569 - 579
Copyright
Copyright © Cambridge University Press 2013

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

Footnotes

†

Both authors contributed equally to this work.

References

REFERENCES

Aguado-Martínez, A., Álvarez-García, G., Fernández-García, A., Risco-Castillo, V., Arnaiz-Seco, I., Rebordosa-Trigueros, X., Navarro-Lozano, V. and Ortega-Mora, L. M. (2008). Usefulness of rNcGRA7- and rNcSAG4-based ELISA tests for distinguishing primo-infection, recrudescence, and chronic bovine neosporosis. Veterinary Parasitology 157, 182–195. doi: 10.1016/j.vetpar.2008.08.002.Google Scholar
Aguado-Martínez, A., Ortega-Mora, L. M., Álvarez-García, G., Rodríguez-Marco, S., Risco-Castillo, V., Marugán-Hernández, V. and Fernández-García, A. (2009 a). Stage-specific expression of Nc SAG4 as a marker of chronic Neospora caninum infection in a mouse model. Parasitology 136, 757–764. doi: 10.1017/S0031182009006076.CrossRefGoogle ScholarPubMed
Aguado-Martínez, A., Álvarez-García, G., Fernández-García, A., Risco-Castillo, V., Marugán-Hernández, V. and Ortega-Mora, L. M. (2009 b). Failure of a vaccine using immunogenic recombinant proteins rNcSAG4 and rNcGRA7 against neosporosis in mice. Vaccine 27, 7331–7338. doi: 10.1016/j.vaccine.2009.09.050.Google Scholar
Aguado-Martínez, A., Álvarez-García, G., Schares, G., Risco-Castillo, V., Fernández-García, A., Marugán-Hernández, V. and Ortega-Mora, L. M. (2010). Characterisation of NcGRA7 and NcSAG4 proteins: immunolocalisation and their role in the host cell invasion by Neospora caninum tachyzoites. Acta Parasitologica 55, 304–312. doi: 10.2478/s11686-010-0056-9.Google Scholar
Al-Qassab, S., Reichel, M. P., Ivens, A. and Ellis, J. T. (2009). Genetic diversity amongst isolates of Neospora caninum, and the development of a multiplex assay for the detection of distinct strains. Molecular and Cellular Probes 23, 132–139. doi: 10.1016/j.mcp.2009.01.006.CrossRefGoogle ScholarPubMed
Álvarez-García, G., Pitarch, A., Zaballos, A., Fernández-García, A., Gil, C., Gómez-Bautista, M., Aguado-Martínez, A. and Ortega-Mora, L. M. (2007). The NcGRA7 gene encodes the immunodominant 17 kDa antigen of Neospora caninum . Parasitology 134, 41–50. doi: 10.1017/S0031182006001284.Google Scholar
Bartley, P. M., Wright, S., Sales, J., Chianini, F., Buxton, D. and Innes, E. A. (2006). Long-term passage of tachyzoites in tissue culture can attenuate virulence of Neospora caninum in vivo . Parasitology 133, 421–432. doi: 0.1017/S0031182006000539.Google Scholar
Basso, W., Schares, S., Barwald, A., Herrmann, D. C., Conraths, F. J., Pantchev, N., Vrhovec, M. G. and Schares, G. (2009). Molecular comparison of Neospora caninum oocyst isolates from naturally infected dogs with cell culture-derived tachyzoites of the same isolates using nested polymerase chain reaction to amplify microsatellite markers. Veterinary Parasitology 160, 43–50. doi: 10.1016/j.vetpar.2008.10.085.Google Scholar
Caspe, S. G., Moore, D. P., Leunda, M. R., Cano, D. B., Lischinsky, L., Regidor-Cerrillo, J., Alvarez-Garcia, G., Echaide, I. G., Bacigalupe, D., Ortega Mora, L. M., Odeon, A. C. and Campero, C. M. (2012). The Neospora caninum-Spain 7 isolate induces placental damage, fetal death and abortion in cattle when inoculated in early gestation. Veterinary Parasitology 189(2–4), 171–811. doi: 10.1016/j.vetpar.2012.04.034.Google Scholar
Collantes-Fernandez, E., Arrighi, R. B., Alvarez-Garcia, G., Weidner, J. M., Regidor-Cerrillo, J., Boothroyd, J. C., Ortega-Mora, L. M. and Barragan, A. (2012). Infected dendritic cells facilitate systemic dissemination and transplacental passage of the obligate intracellular parasite Neospora caninum in mice. PloS One 7, e32123. doi: 10.1371/journal.pone.0032123.Google Scholar
Collantes-Fernández, E., López-Pérez, I., Álvarez-García, G. and Ortega-Mora, L. M. (2006). Temporal distribution and parasite load kinetics in blood and tissues during Neospora caninum infection in mice. Infection and Immunity 74, 2491–2494. doi: 10.1128/IAI.74.4.2491-2494.2006.CrossRefGoogle ScholarPubMed
Dubey, J. P. and Schares, G. (2011). Neosporosis in animals – the last five years. Veterinary Parasitology 180, 90–108. doi: 10.1016/j.vetpar.2011.05.031.CrossRefGoogle ScholarPubMed
Dubey, J. P., Buxton, D. and Wouda, W. (2006). Pathogenesis of bovine neosporosis. Journal of Comparative Pathology 134, 267–289. doi: 10.1016/j.jcpa.2005.11.004.Google Scholar
Dubey, J. P., Schares, G. and Ortega-Mora, L. M. (2007). Epidemiology and control of neosporosis and Neospora caninum . Clinical Microbiology Reviews 20, 323–367. doi: 10.1128/CMR.00031-06.Google Scholar
Fernández-García, A., Risco-Castillo, V., Zaballos, A., Álvarez-García, G. and Ortega-Mora, L. M. (2006). Identification and molecular cloning of the Neospora caninum SAG4 gene specifically expressed at bradyzoite stage. Molecular and Biochemical Parasitology 146, 89–97. doi: 10.1016/j.molbiopara.2005.08.019.CrossRefGoogle ScholarPubMed
Hemphill, A., Felleisen, R., Connolly, B., Gottstein, B., Hentrich, B. and Muller, N. (1997). Characterization of a cDNA-clone encoding Nc-p43, a major Neospora caninum tachyzoite surface protein. Parasitology 115, 581–590. doi: 10.1017/S0031182097001650.Google Scholar
Hemphill, A., Gajendran, N., Sonda, S., Fuchs, N., Gottstein, B., Hentrich, B. and Jenkins, M. (1998). Identification and characterisation of a dense granule-associated protein in Neospora caninum tachyzoites. International Journal for Parasitology 28, 429–438. doi: 10.1016/S0020-7519(97)00193-8.CrossRefGoogle ScholarPubMed
Hill, R. D., Gouffon, J. S., Saxton, A. M. and Su, C. (2012). Differential gene expression in mice infected with distinct Toxoplasma strains. Infection and Immunity 80, 968–974. doi: 10.1128/IAI.05421-11.Google Scholar
Jiménez-Ruiz, E., Álvarez-García, G., Aguado-Martínez, A., Salman, H., Irache, J. M., Marugán-Hernández, V. and Ortega-Mora, L. M. (2012). Low efficacy of NcGRA7, NcSAG4, NcBSR4 and NcSRS9 formulated in poly-epsilon-caprolactone against Neospora caninum infection in mice. Vaccine 30, 4983–4992. doi: 10.1016/j.vaccine.2012.05.033.Google Scholar
Kim, S. K. and Boothroyd, J. C. (2005). Stage-specific expression of surface antigens by Toxoplasma gondii as a mechanism to facilitate parasite persistence. Journal of Immunology 174, 8038–8048.Google Scholar
Kim, S. K., Karasov, A. and Boothroyd, J. C. (2007). Bradyzoite-specific surface antigen SRS9 plays a role in maintaining Toxoplasma gondii persistence in the brain and in host control of parasite replication in the intestine. Infection and Immunity 75, 1626–1634. doi: 10.1128/IAI.01862-06.Google Scholar
Miller, C. M., Quinn, H. E., Windsor, P. A. and Ellis, J. T. (2002). Characterisation of the first Australian isolate of Neospora caninum from cattle. Australian Veterinary Journal 80, 620–625. doi: 10.1111/j.1751-0813.2002.tb10967.x.Google Scholar
Pardini, L., Maksimov, P., Herrmann, D. C., Bacigalupe, D., Rambeaud, M., Machuca, M., More, G., Basso, W., Schares, G. and Venturini, M. C. (2012). Evaluation of an in-house TgSAG1 (P30) IgG ELISA for diagnosis of naturally acquired Toxoplasma gondii infection in pigs. Veterinary Parasitology 189, 204–210. doi: 10.1016/j.vetpar.2012.4.014.CrossRefGoogle ScholarPubMed
Pedraza-Díaz, S., Marugán-Hernández, V., Collantes-Fernández, E., Regidor-Cerrillo, J., Rojo-Montejo, S., Gómez-Bautista, M. and Ortega-Mora, L. M. (2009). Microsatellite markers for the molecular characterization of Neospora caninum: application to clinical samples. Veterinary Parasitology 166, 38–46. doi: 10.1016/j.vetpar.2009.07.043.Google Scholar
Pereira García-Melo, D., Regidor-Cerrillo, J., Collantes-Fernández, E., Aguado-Martínez, A., Del Pozo, I., Minguijon, E., Gómez-Bautista, M., Aduriz, G. and Ortega-Mora, L. M. (2010). Pathogenic characterization in mice of Neospora caninum isolates obtained from asymptomatic calves. Parasitology 137, 1057–1068. doi: 10.1017/S0031182009991855.Google Scholar
Pérez-Zaballos, F. J., Ortega-Mora, L. M., Álvarez-García, G., Collantes-Fernández, E., Navarro-Lozano, V., García-Villada, L. and Costas, E. (2005). Adaptation of Neospora caninum isolates to cell-culture changes: an argument in favor of its clonal population structure. Journal of Parasitology 91, 507–510. doi: 10.1645/GE-381R1.Google Scholar
Quinn, H. E., Miller, C. M., Ryce, C., Windsor, P. A. and Ellis, J. T. (2002). Characterization of an outbred pregnant mouse model of Neospora caninum infection. Journal of Parasitology 88, 691–696. doi: 10.1645/0022-3395(2002)088[0691:COAOPM]2.0.CO;2.Google Scholar
Rettigner, C., De Meerschman, F., Focant, C., Vanderplasschen, A. and Losson, B. (2004). The vertical transmission following the reactivation of a Neospora caninum chronic infection does not seem to be due to an alteration of the systemic immune response in pregnant CBA/Ca mice. Parasitology 128, 149–160.Google Scholar
Regidor-Cerrillo, J., Pedraza-Díaz, S., Gómez-Bautista, M. and Ortega-Mora, L. M. (2006). Multilocus microsatellite analysis reveals extensive genetic diversity in Neospora caninum . Journal of Parasitology 92, 517–524. doi: 10.1645/GE-713R.1.Google Scholar
Regidor-Cerrillo, J., Gómez-Bautista, M., Del Pozo, I., Jiménez-Ruiz, E., Aduriz, G. and Ortega-Mora, L. M. (2010). Influence of Neospora caninum intra-specific variability in the outcome of infection in a pregnant BALB/c mouse model. Veterinary Research 41, 52. doi: 10.1051/vetres/2010024.CrossRefGoogle Scholar
Regidor-Cerrillo, J., Gómez-Bautista, M., Sodupe, I., Aduriz, G., Álvarez-García, G., Del Pozo, I. and Ortega-Mora, L. M. (2011). In vitro invasion efficiency and intracellular proliferation rate comprise virulence-related phenotypic traits of Neospora caninum . Veterinary Research 42, 41. doi: 10.1186/1297-9716-42-41.CrossRefGoogle ScholarPubMed
Regidor-Cerrillo, J., Álvarez-García, G., Pastor-Fernández, I., Marugán-Hernández, V., Gómez-Bautista, M. and Ortega-Mora, L. M. (2012). Proteome expression changes among virulent and attenuated Neospora caninum isolates. Journal of Proteomics 75, 2306–2318. doi: 10.1016/j.jprot.2012.01.039.CrossRefGoogle ScholarPubMed
Risco-Castillo, V., Fernández-García, A., Zaballos, A., Aguado-Martínez, A., Hemphill, A., Rodríguez-Bertos, A., Álvarez-García, G. and Ortega-Mora, L. M. (2007). Molecular characterisation of BSR4, a novel bradyzoite-specific gene from Neospora caninum . International Journal for Parasitology 37, 887–896. doi: 10.1016/j.ijpara.2007.02.003.CrossRefGoogle ScholarPubMed
Risco-Castillo, V., Marugán-Hernández, V., Fernadez-García, A., Aguado-Martínez, A., Jiménez-Ruiz, E., Rodriguez-Marco, S., Álvarez-García, G. and Ortega-Mora, L. M. (2011). Identification of a gene cluster for cell-surface genes of the SRS superfamily in Neospora caninum and characterization of the novel SRS9 gene. Parasitology 138, 1832–1842. doi: 10.1017/S0031182011001351.CrossRefGoogle ScholarPubMed
Rojo-Montejo, S., Collantes-Fernández, E., Blanco-Murcia, J., Rodríguez-Bertos, A., Risco-Castillo, V. and Ortega-Mora, L. M. (2009). Experimental infection with a low virulence isolate of Neospora caninum at 70 days gestation in cattle did not result in foetopathy. Veterinary Research 40, 49. doi: 10.1051/vetres/2009032.Google Scholar
Rojo-Montejo, S., Collantes-Fernandez, E., Rodriguez-Marcos, S., Perez-Zaballos, F., Lopez-Perez, I., Prenafeta, A. and Ortega-Mora, L. M. (2011). Comparative efficacy of immunization with inactivated whole tachyzoites versus a tachyzoite-bradyzoite mixture against neosporosis in mice. Parasitology 138, 1372–1383. doi: 10.1017/S0031182011001156.CrossRefGoogle ScholarPubMed
Saeij, J. P., Boyle, J. P., Coller, S., Taylor, S., Sibley, L. D., Brooke-Powell, E. T., Ajioka, J. W. and Boothroyd, J. C. (2006). Polymorphic secreted kinases are key virulence factors in toxoplasmosis. Science 314, 1780–1783. doi: 10.1126/science.1133690.CrossRefGoogle ScholarPubMed
Schock, A., Innes, E. A., Yamane, I., Latham, S. M. and Wastling, J. M. (2001). Genetic and biological diversity among isolates of Neospora caninum . Parasitology 123, 13–23. doi: 10.1017/S003118200100796X.CrossRefGoogle ScholarPubMed
Sousa, S., Ajzenberg, D., Vilanova, M., Costa, J. and Darde, M. L. (2008). Use of GRA6-derived synthetic polymorphic peptides in an immunoenzymatic assay to serotype Toxoplasma gondii in human serum samples collected from three continents. Clinical and Vaccine Immunology 15, 1380–1386. doi: 10.1128/CVI.00186-08.Google Scholar
Walsh, C. P., Vemulapalli, R., Sriranganathan, N., Zajac, A. M., Jenkins, M. C. and Lindsay, D. S. (2001). Molecular comparison of the dense granule proteins GRA6 and GRA7 of Neospora hughesi and Neospora caninum . International Journal for Parasitology 31, 253–258. doi: 10.1016/S0020-7519(00)00169-7.Google Scholar
Zhao, J. C., Zhao, Z. D., Wang, W. and Gao, X. M. (2005). Prokaryotic expression, refolding, and purification of fragment 450-650 of the spike protein of SARS-coronavirus. Protein Expression and Purification 39, 169–174. doi: 10.1016/j.pep.2004.10.004.Google Scholar
Supplementary material: File

Jiménez-Ruiz Supplementary Material

Appendix

Download Jiménez-Ruiz Supplementary Material(File)
File 17.8 KB
Supplementary material: File

Jiménez-Ruiz Supplementary Material

Appendix

Download Jiménez-Ruiz Supplementary Material(File)
File 13.8 KB