Skip to main content
NCBI home page
Journal of Clinical Microbiology logoLink to Journal of Clinical Microbiology
. 1994 May;32(5):1159–1165. doi: 10.1128/jcm.32.5.1159-1165.1994

Evaluation of serological tests for diagnosis of Brucella melitensis infection of goats.

E Díaz-Aparicio 1, C Marín 1, B Alonso-Urmeneta 1, V Aragón 1, S Pérez-Ortiz 1, M Pardo 1, J M Blasco 1, R Díaz 1, I Moriyón 1
PMCID: PMC263630  PMID: 8051240

Abstract

Five serological assays were evaluated for the diagnosis of brucellosis in goats: the rose bengal test (RBT), complement fixation test (CFT), radial immunodiffusion (RID) with Brucella and Yersinia enterocolitica O:9 polysaccharides, counterimmunoelectrophoresis (CIEP) with cytosol, and enzyme-linked immunosorbent assay (ELISA) with polyclonal and protein G conjugates and smooth lipopolysaccharide (S-LPS), native hapten polysaccharide (NH), or cytosol antigens. For optimal sensitivity, RBT had to be used with sera-antigen at a 3:1 dilution. In the RID test, Brucella melitensis biotype 1 NH could not be replaced by Brucella abortus biotype 1 or Y. enterocolitica 0:9 polysaccharides. In the ELISA, S-LPS and NH gave similar results and the protein G conjugate increased the specificity. With the sera from 55 B. melitensis culture-positive goats, the sensitivity was 100% for RBT, CFT (titer > or = 4), and ELISA with S-LPS or NH; 94% for RID; and 93% for CIEP. All tests were negative (100% specific) when testing the sera from 127 brucella-free goats. Larger discrepancies among the results of the serological tests were obtained with sera from goats of areas where brucellosis is endemic. When the sera of 20 young goats vaccinated subcutaneously (10(9) CFU of B. melitensis Rev 1) and bled 6 months later were examined, the specificities were as follows: NH ELISA, 60%; CFT and S-LPS ELISA, 75%; RBT, 80%; CIEP, 90%; and RID, 94%. With the sera from 10 young goats vaccinated conjunctivally (10(9) CFU of B. melitensis Rev 1) all tests were 100% specific 4 months after vaccination. The proportion of goats giving a positive reaction after vaccination decreased faster in RID than in other tests.

Full text

PDF
1159

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Alonso-Urmeneta B., Moriyón I., Díaz R., Blasco J. M. Enzyme-linked immunosorbent assay with Brucella native hapten polysaccharide and smooth lipopolysaccharide. J Clin Microbiol. 1988 Dec;26(12):2642–2646. doi: 10.1128/jcm.26.12.2642-2646.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bell L. M., Hayles L. B., Chanda A. B. Evidence of reservior hosts of Brucella mellitensis. Med J Zambia. 1976 Dec;10(6):152–153. [PubMed] [Google Scholar]
  3. Björck L., Kronvall G. Purification and some properties of streptococcal protein G, a novel IgG-binding reagent. J Immunol. 1984 Aug;133(2):969–974. [PubMed] [Google Scholar]
  4. Cherwonogrodzky J. W., Nielsen K. H. Brucella abortus 1119-3 O-chain polysaccharide to differentiate sera from B. abortus S-19-vaccinated and field-strain-infected cattle by agar gel immunodiffusion. J Clin Microbiol. 1988 Jun;26(6):1120–1123. doi: 10.1128/jcm.26.6.1120-1123.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Diaz R., Garatea P., Jones L. M., Moriyon I. Radial immunodiffusion test with a Brucella polysaccharide antigen for differentiating infected from vaccinated cattle. J Clin Microbiol. 1979 Jul;10(1):37–41. doi: 10.1128/jcm.10.1.37-41.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Diaz R., Toyos J., Salvó M. D., Pardo M. L. A simple method for the extraction of polysaccharide B from Brucella cells for use in the radial immunodiffusion test diagnosis of bovine brucellosis. Ann Rech Vet. 1981;12(1):35–39. [PubMed] [Google Scholar]
  7. Douglas J. T., Palmer D. A. Use of monoclonal antibodies to identify the distribution of A and M epitopes on smooth Brucella species. J Clin Microbiol. 1988 Jul;26(7):1353–1356. doi: 10.1128/jcm.26.7.1353-1356.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Dubray G. Progrès récents sur la biochimie et les propriétés biologiques des antigènes de Brucella. Dev Biol Stand. 1984;56:131–150. [PubMed] [Google Scholar]
  9. Dáz R., Maravi-Poma E., Rivero A. Comparison of counter-immunoelectrophoresis with other serological tests in the diagnosis of human brucellosis. Bull World Health Organ. 1976;53(4):417–424. [PMC free article] [PubMed] [Google Scholar]
  10. Díaz-Aparicio E., Aragón V., Marín C., Alonso B., Font M., Moreno E., Pérez-Ortiz S., Blasco J. M., Díaz R., Moriyón I. Comparative analysis of Brucella serotype A and M and Yersinia enterocolitica O:9 polysaccharides for serological diagnosis of brucellosis in cattle, sheep, and goats. J Clin Microbiol. 1993 Dec;31(12):3136–3141. doi: 10.1128/jcm.31.12.3136-3141.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Falade S. A comparison of three serological tests in the diagnosis of caprine brucellosis. Res Vet Sci. 1978 May;24(3):376–377. [PubMed] [Google Scholar]
  12. Fensterbank R., Verger J. M., Grayon M. Conjunctival vaccination of young goats with Brucella melitensis strain Rev 1. Ann Rech Vet. 1987;18(4):397–403. [PubMed] [Google Scholar]
  13. Hurvell B. Serological cross-reactions between different Brucella species and Yersinia enterocolitica. Immunodiffusion and immunoelectrophoresis. Acta Vet Scand. 1972;13(4):472–483. doi: 10.1186/BF03547153. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Jiménez de Bagüs M. P., Marín C. M., Blasco J. M., Moriyón I., Gamazo C. An ELISA with Brucella lipopolysaccharide antigen for the diagnosis of B. melitensis infection in sheep and for the evaluation of serological responses following subcutaneous or conjunctival B. melitensis strain Rev 1 vaccination. Vet Microbiol. 1992 Feb;30(2-3):233–241. doi: 10.1016/0378-1135(92)90117-c. [DOI] [PubMed] [Google Scholar]
  15. Jones L. M., Berman D. T., Moreno E., Deyoe B. L., Gilsdorf M. J., Huber J. D., Nicoletti P. Evaluation of a radial immunodiffusion test with polysaccharide B antigen for diagnosis of bovine brucellosis. J Clin Microbiol. 1980 Dec;12(6):753–760. doi: 10.1128/jcm.12.6.753-760.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Jones L. M., García-Carrillo C., Alton G. G. Brucella melitensis Rev. 1 and Brucella abortus 45-20 vaccines in goats: serologic tests. Am J Vet Res. 1973 Feb;34(2):199–202. [PubMed] [Google Scholar]
  17. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  18. Leong D., Diaz R., Milner K., Rudbach J., Wilson J. B. Some structural and biological properties of Brucella endotoxin. Infect Immun. 1970 Feb;1(2):174–182. doi: 10.1128/iai.1.2.174-182.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. León A. P., Guerrero J. Inmunización de cabras contra la brucellosis con la cepa viva atenuada de Brucella melitensis 899B. Rev Invest Salud Publica. 1968 Oct-Dec;28(4):277–308. [PubMed] [Google Scholar]
  20. Lord V. R., Cherwonogrodzky J. W. Evaluation of polysaccharide, lipopolysaccharide, and beta-glucan antigens in gel immunodiffusion tests for brucellosis in cattle. Am J Vet Res. 1992 Mar;53(3):389–391. [PubMed] [Google Scholar]
  21. Markwell M. A., Haas S. M., Bieber L. L., Tolbert N. E. A modification of the Lowry procedure to simplify protein determination in membrane and lipoprotein samples. Anal Biochem. 1978 Jun 15;87(1):206–210. doi: 10.1016/0003-2697(78)90586-9. [DOI] [PubMed] [Google Scholar]
  22. Merril C. R., Switzer R. C., Van Keuren M. L. Trace polypeptides in cellular extracts and human body fluids detected by two-dimensional electrophoresis and a highly sensitive silver stain. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4335–4339. doi: 10.1073/pnas.76.9.4335. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Moreno E., Mayer H., Moriyon I. Characterization of a native polysaccharide hapten from Brucella melitensis. Infect Immun. 1987 Nov;55(11):2850–2853. doi: 10.1128/iai.55.11.2850-2853.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Moriyon I., Berman D. T. Effects of nonionic, ionic, and dipolar ionic detergents and EDTA on the Brucella cell envelope. J Bacteriol. 1982 Nov;152(2):822–828. doi: 10.1128/jb.152.2.822-828.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Nielsen K. H., Wright P. F., Kelly W. A., Cherwonogrodzky J. H. A review of enzyme immunoassay for detection of antibody to Brucella abortus in cattle. Vet Immunol Immunopathol. 1988 Jun;18(4):331–347. doi: 10.1016/0165-2427(88)90160-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Plommet M., Fensterbank R. La vaccination antibrucellique administrée par voie conjonctivale. Dev Biol Stand. 1984;56:681–687. [PubMed] [Google Scholar]
  27. Tsai C. M., Frasch C. E. A sensitive silver stain for detecting lipopolysaccharides in polyacrylamide gels. Anal Biochem. 1982 Jan 1;119(1):115–119. doi: 10.1016/0003-2697(82)90673-x. [DOI] [PubMed] [Google Scholar]
  28. Verstreate D. R., Creasy M. T., Caveney N. T., Baldwin C. L., Blab M. W., Winter A. J. Outer membrane proteins of Brucella abortus: isolation and characterization. Infect Immun. 1982 Mar;35(3):979–989. doi: 10.1128/iai.35.3.979-989.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. WARREN L. The thiobarbituric acid assay of sialic acids. J Biol Chem. 1959 Aug;234(8):1971–1975. [PubMed] [Google Scholar]
  30. Waghela S., Wandera J. G., Wagner G. G. Comparison of four serological tests in the diagnosis of caprine brucellosis. Res Vet Sci. 1980 Mar;28(2):168–171. [PubMed] [Google Scholar]
  31. Zundel E., Verger J. M., Grayon M., Michel R. Conjunctival vaccination of pregnant ewes and goats with Brucella melitensis Rev 1 vaccine: safety and serological responses. Ann Rech Vet. 1992;23(2):177–188. [PubMed] [Google Scholar]

Articles from Journal of Clinical Microbiology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES