Evaluation of lipopolysaccharides and polysaccharides of different epitopic structures in the indirect enzyme-linked immunosorbent assay for diagnosis of brucellosis in small ruminants and cattle

Abstract

Brucella abortus and Brucella melitensis have surface lipopolysaccharides and polysaccharides carrying B. melitensis-type (M) and B. abortus-type (A) epitopes as well as common (C) epitopes present in all smooth Brucella biotypes. Crude lipopolysaccharides, hydrolytic O polysaccharides, and native hapten polysaccharides of MC or AC specificity were evaluated in indirect enzyme-linked immunosorbent assays with polyclonal, monoclonal, or protein G conjugates by using sera from cattle, sheep, and goats infected with AC, MC, or AMC Brucella biotypes. Regardless of the antigen, the levels of antibodies were lower in goats than in sheep and highest in cattle. The diagnostic performance of the assay was not affected by the absence of lipid A-core epitopes, the presence of contaminating outer membrane proteins, the AC or MC epitopic structure of the absorbed antigen, or the conjugate used. Moreover, with sera from cattle vaccinated with B. abortus S19 (AC) or from sheep and goats vaccinated with B. melitensis Rev 1 (MC), AC and MC antigens showed similar levels of reactivity. The results show that antibodies to the C epitopes largely dominate in infection, and this is consistent with the existence of multiple overlapping C epitopes (V. Weynants, D. Gilson, A. Cloeckaert, A. Tibor, P. A. Denoel, F. Godfroid, J. N. Limet, and J.-J. Letesson, Infect. Immun. 65:1939-1943, 1997) rather than with one or two C epitopes. It is concluded that, by adaptation to the corresponding antibody levels, brucellosis in cattle, sheep, and goats can be diagnosed by immunosorbent assay with a single combination of conjugate and antigen.

FIG. 1

Reactivities of sera from goats infected with B. melitensis biotype 1 (MC) (continuous lines) and Brucella-free goats (dotted lines) in the ELISA with MC crLPS (•), MC PS (▴), MC NH (▾), AC crLPS (○), AC PS (▵), and AC NH (▿) (for clarity, error bars are not shown).

FIG. 2

Distribution of the results of the iELISA obtained with sera from Brucella-free cattle and cattle infected with B. abortus biotype 1 and B. melitensis biotype 3 and crLPSs, PSs, and NHs of AC or MC specificity.

FIG. 3

Distribution of the results of the iELISA obtained with sera from Brucella-free sheep and sheep infected with B. melitensis biotype 1 and B. melitensis biotype 3 and crLPSs, PSs, and NHs of AC or MC specificity.