The Transcriptional Regulator MucR, but Not Its Controlled Acid-Activated Chaperone HdeA, Is Essential for Virulence and Modulates Surface Architecture and Properties inPA.

Abstract

is a non-zoonotic bacterium causing contagious epididymitis and other genital lesions in rams and responsible for significant economic losses in sheep-breeding areas. It is a naturally rough (without O-chains in the lipopolysaccharide)species whose virulence mechanisms have been less explored than those of zoonotic smooth brucellae (bearing O-chains that mask other outer membrane molecules). Considering the rough nature of, the influence of surface components other than O-chains on its biological properties may be greater than in smoothspecies. Here we describe the construction and characterization of thedeletion mutant of virulentPA, which is defective in a transcriptional regulator, affecting surface properties and virulence in smooth brucellae. This mutant showed increased amounts of three proteins identified as HdeA (acid-activated chaperone), Omp25d (outer membrane protein undetectable in the parental strain), and BOV_A0299 (hypothetical protein of unknown function). This observation correlated with the enhanced transcription of the corresponding genes and constitutes the first report on this type of proteome alteration inΔmutants. The upstream regions of the three genes contained AT rich domains with T-A steps described as binding sites for MucR in the2308promoter (gene also upregulated inΔ), which suggests that, andexpression could be repressed by MucR through a direct binding to their promoter regions. Relative quantification of transcripts of several other genes selected according to the transcriptome of smooth brucellae Δmutants revealed not only similarities but also relevant differences among strains, such as those detected in flagellar andgenes. Periplasmic HdeA has been related to the resistance ofto acidic pH, conditions encountered byinside phagocytes, but the deletion ofinPA and the Δmutant did not modify any of the evaluated properties of these strains. ThePA Δand ΔΔmutants had defectivegrowth and altered surface properties and architecture, exemplified by detectable amounts of Omp25d. Moreover, they showed virulence attenuation but established persistent splenic infection in mice, which encourages their evaluation as specifical attenuated vaccines against.