When mice are intravenously inoculated with a virulent Brucella abortus strain at day 12 to 14 of pregnancy and killed three to five days later, colonization of placentae, fetuses and spleens can be estimated by the frequency and level (bacterial count) of infection and by linkage between individual placental and paired fetal infections. This linkage indicates the placental barrier effect, defined as the number of non-infected fetuses linked to 100 colonized placentae. Immune mice serum raised against two Brucella fractions injected one day before challenge (1) restricted the placental colonization (the dose required to infect 50 per cent placentae was increased by 50 to 70 times compared to controls), (2) decreased the level of splenic and placental infection, and (3) increased the barrier effect so that most fetuses were protected even when linked to a heavily infected placenta. Immune (B + T) spleen cells from mice vaccinated with a Brucella cell-wall fraction transferred to recipients seven to eight days before mating, that is, 22 days before challenge (1) restricted the frequency of placental and fetal colonization, (2) decreased the level of splenic, placental and fetal infections, and (3) increased the barrier effect. However, separated B- and T-cells were less active, in particular on the level of fetal infection. In contrast with serum, the cells did not decrease infection of the fetuses linked to heavily infected placentae.

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Brucellosis and immunity: humoral and cellular components in mice.

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Vaccines, vaccination in brucellosis.

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Immunity conferred upon mice by anti-LPS monoclonal antibodies in murine brucellosis.

An immune serum infused into mice prior to intravenous virulent Brucella abortus challenge may reduce initial colonization of the spleen on day 7 post-challenge and increase bacteriolysis in the spleen and liver, as evidenced on day 21. Three monoclonal antibodies (IgG1, IgG3 and IgG2a) directed toward the lipopolysaccharide-A epitope (LPS-A) of B. abortus were studied in this model and compared with three previously studied polyclonal immune sera. The three monoclonals restricted spleen infection on day 7 post-challenge and spleen and liver infections on day 21. These early and late effects were similar to those obtained with the two polyclonal sera of high anti-LPS-A titres, namely, serum from mice either infected by B. abortus or vaccinated with a cell-wall B. abortus fraction. In contrast, serum from mice vaccinated with LPS-M from B. melitensis, which had a high anti-LPS-M but a low anti-LPS-A titre, had lower activity, evidenced at day 7 only. Hence, immune serum protection was mediated by antibodies directed toward the LPS-dominant epitope of the challenge strain.

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Immunogenic activity of a cell wall fraction extracted from Brucella abortus in guinea-pigs.

A cell wall fraction (F8) extracted by boiling sodium dodecylsulfate at 4 % from Brucella abortus 99S was used with oil adjuvant to vaccinate groups of ten guinea-pigs, at doses equivalent to 1 X 10(9) and 1 X 10(10) bacteria, once or twice at 3 month intervals. H38 vaccine, a total cell vaccine from formalized B. melitensis 53 H38, was used as a reference, at doses 3 X 10(8) and 3 X 10(9) bacteria. These doses were chosen since they have about the same vaccinal activity in mice being respectively equal to 10 and 100 mice optimal dose (MOD). One extra-group of guinea-pigs received two injections of 100 microgram of smooth-lipopolysaccharide (LPS-S) of B. melitensis 16M, in adjuvant. Control group received the adjuvant only. Guinea-pigs were challenged 3 months after the last vaccination with 5,000 colony-forming units of B. abortus 544, and autopsied 40 days later. The spleen and 8 lymph nodes were cultured: a guinea-pig is considered as protected if no Brucella was found in any sample. Protection afforded by the two vaccines is dose-dependent. H38 vaccine gives a better protection (infected 24 %) than F8 (46 %) since a higher dose is needed to obtain the same level of protection: i. e., 100 MOD of F8 is about equal to 10 MOD of H38 (35 and 37 % respectively). Contrary to what was previously shown in mice, recall does not improve the immunity and LPS-S does not vaccinate at all.

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Antagonism between two immunogens extracted from Brucella (cell wall peptidoglycan and lipopolysaccharide fractions) and inactivity of the brucellin allergen in immunization of the mouse.

Three fractions extracted from Brucella possessing immunogenic activity were tested either as unique antigens or in combinations using various dose levels. Immunity in the mouse was tested one month after vaccination by the intraperitoneal administration of a virulent strain of B. abortus and the spleen count of Brucella 15 days after challenge. The brucellin protein fraction, the cutaneous hypersensitivity reagent did not immunize the mouse and did not interfere with the immunization produced by the PG (cell wall PG extracted with SDS) and LPS fractions. The PG fractions, extracted either from B. abortus (PG-F8) or from B. melitensis (PG-F7), conferred good dose-dependent immunity used on their own. The LPS fraction (B. melitensis) on its own also conferred good immunity but the dose-response curve was less regular. Immunity induced by either fraction was not affected by the presence of an oil adjuvant. Various dose-ratio combinations of PG and LPS were studied with respect to their interactions in three successive factorial experiments. The interactions were consistently significant and largely negative. According to the dose of each fraction and the ratio, the immunity was either equivalent to that produced by the most active fraction in the combination (no additive effect) or less (antagonism). The antagonism between two vaccinal fractions extracted from the same bacteria pleads in favour of purified vaccines.

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