Brucellosis 2003 proceedings - PHIDIAS

Poster Session
68- DOES OXYGEN TENSION MODULATE GENE EXPRESSION DURING
GROWTH OF Brucella INSIDE ITS REPLICATIVE NICHE?.
S. Loisel, S. Köhler, J.-P. Liautard and V. Jubier-Maurin. INSERM U-431, Université Montpellier II,
34095 Montpellier, France.
Our previous study of the nik gene cluster of Brucella suis has shown that its
intracellularly induced promoter is also activated under in vitro microaerobic
conditions. On the other hand, a cydB mutant of B. abortus lacking the cytochrome
bd oxidase of high affinity for oxygen, was found highly attenuated in the mouse
model of infection. The complete genome sequence has revealed that Brucella
possesses a locus potentially encoding another high oxygen affinity oxidase, very
homologous to the cbb3 –type terminal oxidase of Rhizobium meliloti. Moreover, all
the necessary genes for a complete anaerobic respiratory system were discovered ,
which could allow Brucella to use nitrate, for instance, as terminal electron acceptor.
More recently, our work identified among the whole set of mutants representing the
virulome of B. suis, two attenuated strains showing miniTn5 insertion in cydD, part of
the operon encoding the cytochrome bd oxidase, and in caiB, encoding one of the
enzymes required for the carnitine metabolism during anaerobiosis. We conclude
that this environmental condition was a characteristic of the brucellosome,
necessitating bacteria adaptation. We decided to study how regulation by low
oxygen tension can influence expression of genes implicated in a better adaptation
of B. suis to the phagosomal environmental conditions. Plasmids were constructed in
which B. suis promoters were cloned upstream the gfp reporter gene to analyze their
regulation. Expression of the fixK gene, a putative oxygen sensor, and of its potential
target narK, first gene of the Nar operon encoding the nitrate reductase, were
examined in the wild type B. suis strain and in the fixK mutant under aerobic,
microaerobic and anaerobic conditions. Results were compared to the level obtained
in intracellular expression. Unexpected findings indicated that these two promoters
were expressed under normal oxygenation. Activation of the narK gene was actually
shown to be dependent of fixK.
69- INDUCTION OF ENHANCED CYTOTOXIC LYMPHOCYTE ACTIVITY BY
Brucella abortus RB51 OVEREXPRESSING CU/ZN SUPEROXIDE DISMUTASE
(SOD) AND LEAKING SOD.
Y. Hea 1 , R. Vemulapalli 2 , N. Sriranganathan 3 , S. Boyle 3 and G. G. Schurig 3 . (1) Virginia Bioinformatics
Institute, Virginia Tech, Virginia, USA. (2) Department of Veterinary Pathobiology, School of Veterinary
Medicine, Purdue University, Indiana, USA. (3) Center for Molecular Medicine and Infectious
Diseases, VA-MD Regional College of Veterinary Medicine, Virginia Tech, Virginia, USA.
Brucella abortus is a Gram negative, facultative intracellular pathogen of
several mammals, including humans. Cell-mediated immunity (CMI) is critical for
protection against brucellosis. B. abortus strain RB51 is currently being used as the
official live vaccine against bovine brucellosis in the US and several other countries.
We have previously reported that overexpression of Brucella Cu/Zn superoxide
dismutase (SOD) in a recombinant strain RB51 (strain RB51SOD) significantly
increased its vaccine efficacy against virulent B. abortus challenge in a mouse
model. Here, we describe that strain RB51SOD and another recombinant of strain
RB51 which overexpresses homologous SOD and simultaneously expresses
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Brucellosis 2003 International Research Conference