Brucellosis 2003 proceedings - PHIDIAS

Short Oral Communications
Taxonomy & Genomics and proteomics
presence of two different urease clusters, conserved in the three main species. The
sequences are highly conserved, except for a shorter ureE gene in B. suis cluster 1.
We have isolated several mutants deficient in urease activity from a Tn5 mutant
library of the Brucella abortus strain 2308. All the mutants isolated carry a single
insertion of Tn5, suggesting that only one enzyme is responsible for urease activity.
To ascertain the contribution of each cluster to urease activity we have deleted by
homologous recombination most of the gene codifying for the main structural subunit,
ureC, in both clusters and in the three species, B. abortus, B. suis and B. melitensis.
Measurement of urease activity in the parental strain, as well as the single and
double mutants shows that in the three species tested, the only active cluster in
laboratory conditions is cluster 1, as deduced by the initial mutagenesis. However,
the activities differ greatly between strains. Given the similarities at the DNA level, we
hypothesize the different activities depend on the presence or activity of regulatory
proteins.
GO4- SYSTEMATIC DISRUPTION OF GENES CODING FOR TRANSCRIPTIONAL
REGULATORS IN Brucella melitensis 16M.
V. Haine , M. Dozot, A. Sinon, P. Lestrate, R-M. Delrue, A. Tibor, C. Lambert, J.J. Letesson and X. De
Bolle. Unité de Recherche en Biologie Moléculaire (URBM), Laboratoire d’Immunologie et de
Microbiologie, Facultés Universitaires Notre-Dame de la Paix, Namur, Belgique.
The genome sequence of three species of Brucella is now available (B.
melitensis, suis and abortus). That information opens the way of the post genomic
approach (transcriptome, proteome, interactome, etc …). We performed the
systematic disruption of a category of genes coding for transcriptional regulators, the
control of Brucella virulence genes being poorly documented. First, all B. melitensis
16M transcriptional regulators predicted were grouped into families based on
sequence comparisons (essentially the helix-turn-helix signatures). We predicted 180
transcriptional regulators divided into 18 families. We selected 10 families that
include regulators involved in virulence and/or symbiosis in other bacteria (AraC,
ArsR, Crp, DeoR, GntR, IclR, LysR, MerR and TetR families). We constructed
mutants for the ten families (94 regulators) by integrative disruption in B. melitensis
16M. The virulence of the mutants was tested in the BALB/c model of infection using
a method based on STM (signature-tagged mutagenesis). We already identified 7
transcriptional regulators playing a role in Brucella virulence. This library of mutants is
also very useful to study promoters of genes of interest in Brucella.
GO5- IDENTIFICATION OF A TRANSCRIPTIONAL REGULATOR INVOLVED IN
Brucella melitensis FLAGELLAR GENE EXPRESSION.
S. Léonard, A. Tibor, J. Ferooz, D. Fretin, C. Nijskens and J.-J. Letesson. Unité de Recherche en
Biologie Moléculaire (URBM), Laboratoire d’Immunologie et de Microbiologie, Facultés Universitaires
Notre-Dame de la Paix, Namur, Belgique.
Even though brucellae are described as non motile, the Brucella genome
contains three clusters of flagellar genes on the small chromosome. To identify an in
vitro culture condition which was appropriate for stimulating expression of flagellar
genes, we tested the effect of growth stage on flagellar gene expression by using a
Brucellosis 2003 International Research Conference
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