RecombinantBrucella abortusProteins That Induce Proliferation and Gamma-Interferon Secretion by CD4+T Cells fromBrucella-Vaccinated Mice and Delayed-Type Hypersensitivity in Sensitized Guinea Pigs

Optimal protective immunity toBrucella abortusinfection is dependent on a coordinate interaction between different T-cell subsets which leads to an antigen-specific T-lymphocyte-mediated activation of macrophages, the main cellular reservoir for the bacterium. As an initial step in the identification of bacterial proteins that mediate cellular immunity, we have subcloned theB. abortus ssb, uvrA, GroES,andGroELgenes into the prokaryotic expression vector pMAL-c2 using PCR.Escherichia coliDH5α was transformed with the pMAL-ssb, pMAL-uvrA, pMAL-GroES, and pMAL-GroEL constructs separately, and gene expression was induced by isopropyl-β-D-thiogalactopyranoside. The resulting fusion proteins were purified by affinity chromatography and confirmed by Western blot analysis using an anti-maltose-binding protein antibody. Furthermore, we have examined the pattern of T helper (Th) cell response from vaccinated BALB/c mice afterin vitrostimulation with the recombinant (r) fusion proteins. In addition to T-cell proliferative responses, CD4+T cells were tested for interleukin-2 (IL-2), IL-4, and gamma interferon (IFN-γ) secretion. Primed CD4+T cells proliferated to the rUvrA, rGroES, and rGroEL, but not to rSsb. The cytokine profile of the proliferating cells was characteristic of a Th1 type, as we detected IL-2 and IFN-γ but not IL-4 in the T-cell culture supernatants. The recombinantB. abortusproteins were also screenedin vivoto their ability to elicit DTH reaction inBrucella-sensitized guinea pigs. Moreover, the results of this study suggest thatB. abortusrUvrA, rGroES, and rGroEL might be important sources of potentially protective molecules.