Natural Antibody Contributes to Host Defense against an Attenuated Brucella abortus virB Mutant

Brucella abortus is an intracellular pathogen that persists within phagocytic cells of the reticuloendothelial system. To identify in vivo interactions between B. abortus and the host that lead to persistent infection, we studied the persistence of B. abortus and an isogenic virB mutant deficient in the VirB type IV secretion system (T4SS) in knockout mice. In contrast to control mice, mice lacking B cells (Igh6^−/−) were permissive for infection with the attenuated virB mutant. To determine the basis for this phenotype, we characterized immune functions of Igh6^−/− mice in the context of B. abortus infection. Igh6^−/− mice had greater numbers of extracellular bacteria in the spleen and increased early expression of proinflammatory cytokines during B. abortus infection. Further, a virB mutant, despite its wild-type level of survival, failed to elicit microgranuloma formation in the spleens of Igh6^−/− mice, suggesting a requirement for the T4SS to elicit this pathological change. Passive transfer of immunoglobulin G from naïve mice restored the ability of Igh6^−/− mice to control the persistence of the virB mutant by a complement-independent mechanism. Further, adoptive transfer of CD11b⁺ cells from C57BL/6 mice to Igh6^−/− mice restored the ability of the knockout mice to limit the replication of the virB mutant in the spleen, suggesting that the Igh6^−/− mutation affects phagocyte function and that phagocyte function can be restored by natural antibody.Human brucellosis is a febrile disease resulting from the transmission of Brucella abortus, B. suis, B. melitensis, or B. canis from its respective zoonotic reservoir in cattle, swine, goats and sheep, or dogs (47). These pathogens are endemic in many areas of the world, including Central and South America, the Mediterranean, and Central Asia, and are responsible for an estimated 500,000 new brucellosis cases each year (1). In human brucellosis, as well as in the zoonotic reservoir species, bacteria may persist for long periods of time in the reticuloendothelial system (7). This aspect of infection can be modeled in the mouse, which has been used to identify and characterize the virulence factors involved in the systemic persistence of Brucella spp. (2, 22).One essential virulence factor of the human pathogenic Brucella species is the type IV secretion system (T4SS) encoded by the virB locus on chromosome II (18, 29, 40). The T4SS of Brucella spp., similar to those of other bacterial pathogens, mediates the translocation of proteins into host cells; however, the functions of two newly identified B. abortus T4SS substrates, VceA and VceC, is not yet known (12, 25, 39, 45, 46). In cultured macrophages and dendritic cells (DC), the T4SS is essential for the intracellular replication and persistence of B. abortus (13, 36, 40). The T4SS mediates exclusion of late endosomal/lysosomal markers from the Brucella-containing vacuole and targeting of B. abortus to exit sites of the endoplasmic reticulum, where replication occurs (5, 6, 12, 41), suggesting that T4SS effectors are involved in this function. After intraperitoneal (i.p.) inoculation of mice, the T4SS is required not for the initial systemic dissemination of B. abortus but rather for persistence in the reticuloendothelial system (31, 34). In order to better understand the interactions between the host and B. abortus that lead to the persistence of wild-type (WT) strains and the eventual clearance of virB mutants, we examined the immune mechanisms required for clearance of the virB mutant. Unexpectedly, mice lacking B cells (Igh6^−/−) were permissive for the splenic persistence of the virB mutant, while the persistence of WT B. abortus was not increased. When cultured ex vivo, macrophages from Igh6^−/− mice behaved identically to macrophages from control mice in their ability to control the intracellular replication of the virB mutant while permitting the replication of WT B. abortus (34).In this study, we attempted to pinpoint the defect in Igh6^−/− mice that renders them permissive for persistent infection by the virB mutant. Our results show that nonspecific antibody can reverse the defect of these mice in controlling virB mutant replication without affecting WT B. abortus. These results suggest that the T4SS mediates the evasion of a natural antibody-dependent immune clearance function by B. abortus during persistence in vivo.