Identification of , and , Three Genes Involved in the Remodeling of Cell Envelope.

The brucellae are facultative intracellular bacteria that cause a worldwide extended zoonosis. One of the pathogenicity mechanisms of these bacteria is their ability to avoid rapid recognition by innate immunity because of a reduction of the pathogen-associated molecular pattern (PAMP) of the lipopolysaccharide (LPS), free-lipids, and other envelope molecules. We investigated the homologs of , and , three genes that in some pathogens encode enzymes that mask the LPS PAMP by upsetting the core-lipid A charge/hydrophobic balance. , which encodes a putative ethanolamine transferase, carries a frame-shift in but not in other spp. and phylogenetic neighbors like the opportunistic pathogen Consistent with the genomic evidence, a mutant lacked lipid A-linked ethanolamine and displayed increased sensitivity to polymyxin B (a surrogate of innate immunity bactericidal peptides), while carrying displayed increased resistance. encodes a putative phosphatase acting on lipid A or on a free-lipid that is highly conserved in all brucellae and Although we found no evidence of lipid A dephosphorylation, a mutant showed increased polymyxin B sensitivity, suggesting the existence of a hitherto unidentified free-lipid involved in bactericidal peptide resistance. Gene putatively encoding an acyl hydroxylase carries a frame-shift in all brucellae except and is intact in . Free-lipid analysis revealed that corresponded to , the gene coding for the ornithine lipid (OL) acyl hydroxylase active in and , while carrying the of and synthesized hydroxylated OLs. Interestingly, mutants in , or were not attenuated in dendritic cells or mice. This lack of an obvious effect on virulence together with the presence of the intact homolog genes in and but not in other brucellae suggests that LptA, LpxE, or OL β-hydroxylase do not significantly alter the PAMP properties of LPS and free-lipids and are therefore not positively selected during the adaptation to intracellular life.