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In the present study inactivated human immunodeficiency virus type 1 (HIV-1) was conjugated to Brucella abortus and tested for immunogenicity in normal and anti-L3T4-treated BALB/c mice. HIV-BA was more immunogenic than uncoupled HIV in normal mice, since 6-fold less virus in HIV-BA preparations elicited higher titer responses than HIV-1 alone. Furthermore, the HIV-BA antibody response reached higher levels before the HIV-1 response. Immunoblot analysis showed that most of the HIV-1 antigens were recognized by antibodies induced by either HIV-1 or HIV-BA. Isotype analysis revealed that HIV-1 induced similar levels of IgG1 and IgG2a antibodies, whereas the IgG2a responses to HIV-BA were more pronounced than the IgG1 response. These different IgG subclass patterns suggest that conjugation of HIV-1 to BA changed the immunogenic nature of HIV-1. The requirement for helper T cells was examined by immunizing mice that were depleted of CD4+ T cells by in vivo anti-L3T4 treatment. Under these conditions the IgG responses to HIV-1 were completely eliminated. Although HIV-BA antibody responses were markedly reduced in anti-L3T4-treated mice, anti-HIV-1 antibodies, mainly of the IgG2a isotype, were produced. The antibodies generated by HIV-1 and HIV-BA immunization were also tested for their ability to inhibit syncytia formed by infecting CD4 + CEM cells with gp160 vaccinia. Sera from normal mice, immunized with either HIV-1 or HIV-BA were capable of inhibiting syncytia. In contrast, following anti-L3T4 treatment, only mice immunized with HIV-BA, but not HIV-1, produced antibodies capable of inhibiting syncytia.

In view of its unique ability to stimulate human B cells, we have considered using Brucella abortus (BA) as a carrier for human vaccines. Recently we showed that HIV-1 coupled to BA, but not unconjugated HIV-1, was able to stimulate murine responses even in the relative absence of CD4+ T cells. This result suggested that HIV-BA may be useful in boosting the immunity of individuals infected with HIV-1 and who have impaired CD4+ T cell function. In order to refine this carrier we purified lipopolysaccharide (LPS) from BA and examined its effects on immune responses. Similar to LPS from E. coli (LPS-EC), LPS-BA was capable of stimulating mouse B cells to proliferate. In addition, LPS-BA could activate mouse spleen cells to secrete antibodies in vitro. Isotype analysis revealed that IgM and all the IgG subclasses were elicited. When comparing these responses to those of LPS-EC, LPS-BA induced a greater percentage of IgG2a and LPS-EC evoked more IgG3. IgG2a is probably important in protection against murine viral infection. LPS-BA was haptenated with trinitrophenol TNP-LPS (BA) and tested for carrier effect. Similar to TNP-BA and TNP-LPS (EC), TNP-LPS (BA) triggered anti-TNP antibody of the IgM and all IgG subclasses. In contrast, TNP-ficoll induced mainly IgM and only small amounts of IgG3. These results suggest that LPS-BA, like intact BA, behaves as a T-independent type 1 carrier, and as such may be advantageous as a carrier for human vaccines development