Brucellosis 2003 proceedings - PHIDIAS

Poster Session
study of B. abortus and cross-reacting bacteria was conducted to identify proteins,
which may help to distinguish Brucella spp. from V. cholerae, F. tularensis, X.
maltophilia, etc. B. abortus and several cross-reacting bacteria were lysed. Total
proteins were separated by isoelectric focusing in the first dimension using
immobilized pH gradient strips, followed by SDS-PAGE in the second dimension.
Proteins were visualized by silver staining and all gels were analyzed using 2D-Elite
ImageMaster Software. For Surface-enhanced Laser Desorption/Ionization Mass
Spectrometry (SELDI-MS) proteins were prepurified by using affinity ligands on
ProteinChips. Retained proteins were eluted by laser desorption and ionization and
their mass was determined by Time-of-Flight Mass Spectrometry.
Clear differences between B. abortus and cross-reacting bacteria were
detected both in 2D-gels and in SELDI spectra. Potential biomarkers may be isolated
this way. Although various differences in the proteome of the bacteria could be
identified, the immunological role of these proteins has to be examined in future
studies. Thus, proteomics may lead to medical advances in brucellosis, e.g. more
significant diagnostic tools and possible vaccine candidates.
108- IDENTIFICATION OF IMMUNOGENIC PROTEINS IN THE PROTEOME OF
Brucella melitensis AND Brucella abortus USING TWO-DIMENSIONAL
ELECTROPHORESIS AND IMMUNOBLOTTING.
Al Dahouk S. 1 , Tomaso H. 1 , Bartling C. 1 , Neubauer H. 1 (1) Institute of Microbiology, Federal Armed
Forces, Munich, Germany.
Human brucellosis is predominantly caused by three species of the genus
Brucella – B. melitensis, B. abortus, and B. suis. Clinical signs and symptoms of
brucellosis are unspecific, antibiotic therapy often fails and effective human vaccines
are still not available. Because of these characteristics Brucella was among the first
biological agents weaponized in the 1950ies. An effective human vaccine at least for
personnel with high occupational risks of infection would be desirable. The aim of our
study was to identify specific immunogenic proteins in the proteome of Brucella
melitensis and Brucella abortus. B. melitensis and B. abortus were lysed and total
proteins were separated by isoelectric focusing in the first dimension using
immobilized pH gradient strips, followed by SDS-PAGE in the second dimension.
Proteins were visualized by silver staining and all gels were analyzed using 2D-Elite
ImageMaster Software. A second gel was blotted on a nitrocellulose membrane and
the immunogenic proteins were identified using B. abortus or B. melitensis
hyperimmune serum.
Live-attenuated Brucella strains are used in veterinary medicine for
vaccination. As accidental inoculation may cause a severe human disease, antigenic
fractions of Brucella have been discussed for human uses, e.g. Brucella protective
antigen and phenol-insoluble fraction. However, neither the efficacy nor the duration
of protection could be clearly established. Using two-dimensional gel electrophoresis
we were able to demonstrate the whole proteomes of human pathogenic Brucella
spp. Immunoblotting revealed various immunogenic proteins in B. melitensis and B.
abortus. A single specific protein or a defined cocktail of several immunogenic
proteins may be the key for an adaequate human vaccine.
Brucellosis 2003 International Research Conference
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