The role of PlsC in Brucella melitensis virulence: impacts on membrane homeostasis, stress tolerance, and pathogenesis

Abstract

Brucella melitensis, a facultative intracellular pathogen, relies on membrane integrity and homeostasis to resist host defenses and establish infection. The plsC gene encodes 1-acyl-sn-glycerol-3-phosphate acyltransferase, a key enzyme in the glycerophospholipid pathway that catalyzes the synthesis of phosphatidic acid, an essential precursor for membrane lipid formation. However, its role in B. melitensis virulence remains poorly understood. Here, we constructed a plsC deletion mutant (ΔplsC) and a complemented strain (ΔplsC-Com) in B. melitensis strain M5 and characterized their phenotypes. Deletion of plsC impaired bacterial growth in nutrient-limited media, reduced tolerance to hydrogen peroxide and polymyxin B, and decreased lipid synthesis while increasing outer membrane permeability. Ultrastructural analysis revealed surface roughness, cytoplasmic voids, and nucleoid condensation in the mutant. Although ΔplsC retained normal adhesion and invasion capabilities in RAW264.7 macrophages and HeLa cells, its intracellular survival was specifically attenuated in macrophages at 48 h post-infection. In a mouse model, ΔplsC showed significantly reduced colonization of the spleen and liver and induced fewer and smaller liver granulomas as compared with the parental and complemented strains. These results demonstrate that PlsC is essential for maintaining membrane homeostasis and stress resistance in Brucella, which in turn supports its survival within professional phagocytes and full virulence in vivo. Our study suggests a critical link between phospholipid metabolism and Brucella pathogenicity.