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Genetic Bases of the Rifampin Resistance Phenotype in Brucella spp.

Dipartimento di Sanità Alimentare ed Animale, Istituto Superiore di Sanità, Rome, Italy

Received 1 April 2004/ Returned for modification 10 July 2004/ Accepted 1 August 2004

Rifampin is one of the most potent and broad-spectrum antibiotics against bacterial pathogens. Its bactericidal activity is due to its ability to bind to the ß subunit of the DNA-dependent RNA polymerase encoded by the rpoB gene. Mutations of the rpoB gene have been characterized in rifampin-resistant (Rif^r) strains of Escherichia coli and Mycobacterium tuberculosis. The genetic bases of Rif^r in Brucella spp. are still unknown. In the present study, the nucleotide sequences of the rpoB gene of the Rif^r vaccine strain Brucella abortus RB51 and of 20 Rif^r clones derived in our laboratory from two Brucella melitensis isolates were determined. These sequences were then compared to those of the respective rifampin-susceptible (Rif^s) parental strains and to the published B. melitensis strain 16M. All Rif^r strains carried one or more missense mutations mapping in two regions of the rpoB gene. These two "hot" regions were investigated in eight additional Rif^r Brucella laboratory mutants and in 20 reference Rif^s Brucella strains. rpoB mutations were found in all Rif^r mutants. In contrast, no missense mutations were found in any analyzed Rif^s strains. Our results represent the first from a study of the molecular characterization of rpoB mutations in resistant Brucella strains and provide an additional proof of the association of specific rpoB mutations with the development of the Rif^r phenotype in prokaryotes. In addition, because of the relationship between Rif^r and the attenuation of virulence in Brucella spp., studies of virulence in these mutants may provide useful information about the genetic basis of pathogenesis in Brucella.

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