Oral Immunization of Mice with Gamma-Irradiated <i>Brucella neotomae</i> Induces Protection against Intraperitoneal and Intranasal Challenge with Virulent <i>B. abortus</i> 2308

<div><p><i>Brucella</i> spp. are Gram-negative, facultative intracellular coccobacilli that cause one of the most frequently encountered zoonosis worldwide. Humans naturally acquire infection through consumption of contaminated dairy and meat products and through direct exposure to aborted animal tissues and fluids. No vaccine against brucellosis is available for use in humans. In this study, we tested the ability of orally inoculated gamma-irradiated <i>B. neotomae</i> and <i>B. abortus</i> RB51 in a prime-boost immunization approach to induce antigen-specific humoral and cell mediated immunity and protection against challenge with virulent <i>B. abortus</i> 2308. Heterologous prime-boost vaccination with <i>B. abortus</i> RB51 and <i>B. neotomae</i> and homologous prime-boost vaccination of mice with <i>B. neotomae</i> led to the production of serum and mucosal antibodies specific to the smooth LPS. The elicited serum antibodies included the isotypes of IgM, IgG1, IgG2a, IgG2b and IgG3. All oral vaccination regimens induced antigen-specific CD4⁺ and CD8⁺ T cells capable of secreting IFN-γ and TNF-α. Upon intra-peritoneal challenge, mice vaccinated with <i>B. neotomae</i> showed the highest level of resistance against virulent <i>B. abortus</i> 2308 colonization in spleen and liver. Experiments with different doses of <i>B. neotomae</i> showed that all tested doses of 10⁹, 10¹⁰ and 10¹¹ CFU-equivalent conferred significant protection against the intra-peritoneal challenge. However, a dose of 10¹¹ CFU-equivalent of <i>B. neotomae</i> was required for affording protection against intranasal challenge as shown by the reduced bacterial colonization in spleens and lungs. Taken together, these results demonstrate the feasibility of using gamma-irradiated <i>B. neotomae</i> as an effective and safe oral vaccine to induce protection against respiratory and systemic infections with virulent <i>Brucella</i>.</p>