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ABSTRACT: The gram-negative bacteria of the genus Brucella are facultative intracellular parasites that cause brucellosis, a worldwide-distributed zoonotic disease that represents a serious problem for animal and human health. There is no human-to-human contagion and, since there is no human vaccine, animal vaccination is essential to control brucellosis. However, current vaccines (all developed empirically) do not provide 100% protection and are infectious in humans. Attempts to generate new vaccines by obtaining mutants lacking the lipopolysaccharide O-polysaccharide, in purine metabolism or in Brucella type IV secretion system have not been successful. Here we propose a new approach to develop brucellosis vaccines based on the concept that Brucella surface molecules evade efficient detection by innate immunity, thus delaying protective Th1 responses and opening a time window to reach sheltered intracellular compartments. We showed recently that a branch of the core oligosaccharide section of Brucella lipopolysaccharide hampers recognition by TLR4-MD2. Mutation of glycosyltransferase WadC, involved in the synthesis of this branch, results in a lipopolysaccharide that, while keeping the O-polysaccharide essential for optimal protection, shows a truncated core, is more efficiently recognized by MD2 and triggers an increased cytokine response. In keeping with this, the wadC mutant is attenuated in dendritic cells and mice. In the mouse model of brucellosis vaccines, the B. abortus wadC mutant conferred protection similar to that provided by S19, the best cattle vaccine available. The properties of the wadC mutant provide the proof of concept for this new approach and open the way for more effective brucellosis vaccines.
Microbial Pathogenesis 12/2012; · 1.94 Impact Factor
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ABSTRACT: The brucellae are facultative intracellular pathogens of mammals that are transmitted by contact with infected animals or contaminated materials. Several major lipidic components of the brucella cell envelope are imperfectly recognized by innate immunity, thus contributing to virulence. These components carry large proportions of acyl chains of lactobacillic acid, a long chain cyclopropane fatty acid (CFA). CFAs result from addition of a methylene group to unsaturated acyl chains and contribute to resistance to acidity, dryness and high osmolarity in many bacteria and to virulence in mycobacteria. We examined the role of lactobacillic acid in Brucella abortus virulence by creating a mutant in ORF BAB1_0476, the putative CFA synthase gene. The mutant did not incorporate [(14)C]methyl groups into lipids, lacked CFAs and synthesized the unsaturated precursors, proving that BAB1_0476 actually encodes a CFA synthase. BAB1_0476 promoter-luxAB fusion studies showed that CFA synthase expression was promoted by acid pH and high osmolarity. The mutant was not attenuated in macrophages or mice, strongly suggesting that CFAs are not essential for B. abortus intracellular life. However, when the mutant was tested under high osmolarity on agar and acid pH, two conditions likely to occur on contaminated materials and fomites, they showed reduced ability to grow or survive. Since CFA synthesis entails high ATP expenses and brucellae produce large proportions of lactobacillic acyl chains, we speculate that the CFA synthase has been conserved because it is useful for survival extracellularly, thus facilitating persistence in contaminated materials and transmission to new hosts.
Microbiology 01/2012; 158(Pt 4):1037-44. · 3.06 Impact Factor
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Leyre Palacios-Chaves,
Raquel Conde-Álvarez, Yolanda Gil-Ramírez,
Amaia Zúñiga-Ripa,
Elías Barquero-Calvo,
Carlos Chacón-Díaz,
Esteban Chaves-Olarte,
Vilma Arce-Gorvel,
Jean-Pierre Gorvel,
Edgardo Moreno,
María-Jesús de Miguel,
María-Jesús Grilló,
Ignacio Moriyón,
Maite Iriarte
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ABSTRACT: The brucellae are α-Proteobacteria facultative intracellular parasites that cause an important zoonosis. These bacteria escape early detection by innate immunity, an ability associated to the absence of marked pathogen-associated molecular patterns in the cell envelope lipopolysaccharide, lipoproteins and flagellin. We show here that, in contrast to the outer membrane ornithine lipids (OL) of other Gram negative bacteria, Brucella abortus OL lack a marked pathogen-associated molecular pattern activity. We identified two OL genes (olsB and olsA) and by generating the corresponding mutants found that olsB deficient B. abortus did not synthesize OL or their lyso-OL precursors. Liposomes constructed with B. abortus OL did not trigger IL-6 or TNF-α release by macrophages whereas those constructed with Bordetella pertussis OL and the olsB mutant lipids as carriers were highly active. The OL deficiency in the olsB mutant did not promote proinflammatory responses or generated attenuation in mice. In addition, OL deficiency did not increase sensitivity to polymyxins, normal serum or complement consumption, or alter the permeability to antibiotics and dyes. Taken together, these observations indicate that OL have become dispensable in the extant brucellae and are consistent within the trend observed in α-Proteobacteria animal pathogens to reduce and eventually eliminate the envelope components susceptible of recognition by innate immunity.
PLoS ONE 01/2011; 6(1):e16030. · 4.09 Impact Factor
