A. Cloeckaert et al., O-POLYSACCHARIDE EPITOPIC HETEROGENEITY AT THE SURFACE OF BRUCELLA SPP. STUDIED BY ENZYME-LINKED-IMMUNOSORBENT-ASSAY AND FLOW-CYTOMETRY, Clinical and diagnostic laboratory immunology (Print), 5(6), 1998, pp. 862-870
Smooth Brucella strains are classified into three serotypes, i.e., A()M(-), A(-)M(+), and A(+)M(+), according to slide agglutination with A
and M monospecific polyclonal sera. The epitopes involved have been l
ocated on the O-polysaccharide (O-PS) moiety of the smooth lipopolysac
charide (S-LPS), which represents the most exposed antigenic structure
on the surface of Brucella spp. By use of monoclonal antibodies (MAbs
) a number of epitope specificities on the O-PS have been reported: A,
M, and epitopes shared by both A and M dominant strains, which have b
een named common (C) epitopes. The latter have been further subdivided
, according to relative MAb binding in enzyme-linked immunosorbent ass
ays (ELISA) to A- and M-dominant Brucella strains and to cross-reactin
g Yersinia enterocolitica O:9, into five epitopic specificities: C (M>
A), C (M=A), C/Y (M>A), C/Y (M=A), and C/Y (A>M). In the present study
, we studied the occurrence of these epitopes at the surface of repres
entatives of all Brucella species and biovars including the live vacci
ne strains by analyzing the levels of MAb binding to whole Brucella ce
lls in ELISA and flow cytometry assays. In ELISA, the level of MAb bin
ding correlated well with the previously defined epitope specificity a
nd the serotype defined by polyclonal sera for each Brucella species,
biovar, or strain. However, MAbs to the C (M=A) and C (M>A) epitopes s
howed insignificant binding to B. suis biovar 2 strains and bound at l
ower titers to B. suis biovar 3 and B. neotomae than to the other Bruc
ella strains. Some of the flow cytometry results were contradictory to
those obtained by ELISA. In fact, it appeared by bow cytometry that a
ll O-PS epitopes, including the A and M epitopes, are shared to differ
ent degrees by Brucella spp. which nevertheless show a high degree of
O-PS heterogeneity according to MAb binding intensities. The subdivisi
on of MAb specificities and Brucella serotypes was therefore less evid
ent by Bow cytometry than by ELISA. Whereas in ELISA the MAb specific
for the A epitope showed insignificant binding to Y. enterocolitica O:
9, this MAb bound strongly to Y. enterocolitica O:9 in flow cytometry.
One of the two MAbs specific to the C (M=A) epitope also bound at a l
ow but significant level to B. suis biovar 2 strains. However, as in E
LISA the MAb specific for the C (M>A) epitope did not bind at all to B
. suis biovar 2 strains in flow cytometry. Flow cytometry provided new
information regarding specificity of the MAbs and may further explain
some aspects of the capacity of passive protection of some MAbs again
st smooth Brucella infection in mice, As shown in the present study th
e occurrence of Brucella strains apparently completely devoid of one s
pecific C O-PS epitope (e,g., B. suis biovar 2 devoid of the C [M>A] e
pitope) offers the possibility of obtaining vaccine strains devoid of
a diagnostic O-PS epitope, which could further help to resolve the pro
blem of discriminating infected from vaccinated animals that remains a
major goal in brucellosis research.