Brucellosis remains one of the most prevalent zoonotic diseases worldwide, causing substantial economic losses and significant human morbidity. Despite decades of research, no licensed human vaccine against Brucella infection exists, and current veterinary vaccines exhibit considerable safety limitations including residual virulence, pregnancy complications, and diagnostic interference. Here we report the development of a multi-epitope messenger RNA (mRNA) vaccine candidate targeting Brucella melitensis, the most pathogenic species responsible for human brucellosis. Using an integrated immunoinformatics pipeline, we screened ten outer membrane proteins (OMPs) and identified Omp25, Omp31, and BP26 as the most immunodominant antigens. A fusion construct incorporating 24 HLA class I and 31 HLA class II predicted epitopes achieved 87.3% global HLA population coverage. Codon optimization improved the codon adaptation index from 0.55 to 0.93. Molecular dynamics simulations over 200 ns confirmed structural stability of the vaccine-TLR4 complex, with a binding free energy of -65.7 kcal/mol. In vitro assays demonstrated robust Th1-biased immune activation, with IFN-gamma reaching 125.6 pg/mL. In a BALB/c mouse challenge model, the mRNA vaccine conferred 80.0% protection, comparable to the live-attenuated S19 vaccine (92.3%) but without associated safety risks. These findings establish a promising platform for Brucella vaccine development warranting further evaluation in large animal models and human clinical trials.
Zhiheng Dong, Sha Li, Jiarong Guo et al.· Research in Veterinary Scien...· 0 citations
Brucellosis is one of the most severe Class B infectious diseases prevalent in the agricultural and pastoral areas of northern China. Current attenuated live vaccines (e.g., M5, S19) have defects such as residual virulence, causing abortion in pregnant animals, and the inability to differentiate between natural infection and vaccination (DIVA). Relying on the ABSL-3 laboratory of the Inner Mongolia Center for Disease Control and Prevention, this study established a chronic infection model in C57BL/6J mice using the virulent strain Brucella melitensis M16. Single-cell transcriptome sequencing (10x Genomics) was employed to map the heterogeneity of splenic immune cells. Whole-genome scanning and pangenomic analysis were performed on four strains with different virulence levels (M16, 544 A, M5, 104 M) using second-generation sequencing. Membrane/secreted proteins unique and conserved in virulent strains were screened as candidate antigens, prepared via prokaryotic expression systems, and their humoral and cellular immune levels were detected by indirect ELISA and flow cytometry. A stable chronic infection model was successfully constructed (bacterial load Log10 CFU > 4.5). Single-cell sequencing yielded 45,231 cells, annotated into 12 immune cell subpopulations, revealing significant expansion of Effector CD4 + T cells (P < 0.01) and high expression of the Ifng gene. Pangenomic analysis identified three candidate antigens (BMEI0021, BMEI1943, BMEI0367) that are 100% conserved in virulent strains but absent in the vaccine strain M5. Immunogenicity assays showed that BMEI1943 induced high levels of IgG2a subtype antibodies (titer Log2 13.45 ± 0.52) and IFN-γ + CD4 + T cell responses (frequency 15.80% ± 2.10%). Through multi-omics integration analysis, this study successfully identified a novel candidate antigen, BMEI1943, with strong Th1-type immunogenicity, providing an experimental basis for the development of safe and efficient subunit vaccines against brucellosis.
Zhiheng Dong, Sha Li, Jiarong Guo et al.· Brazilian Journal of Microbi...· 0 citations
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