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Background  We generated novel, effective candidate vaccine against  Brucella abortus  based on recombinant influenza viruses expressing the  Brucella  ribosomal protein L7/L12 or outer membrane protein (Omp)-16 from the NS1 open reading frame. The main purpose of this work was to evaluate the safety, immunogenicity and protectiveness of vaccine candidate in laboratory animals.    Methods and Results  Four recombinant influenza A viral constructs of the subtypes Н5N1 or H1N1 expressing the  Brucella  proteins L7/L12 or Omp16 were obtained by a reverse genetics method: Flu-NS1-124-L7/L12-H5N1, Flu-NS1-124-Omp16-H5N1, Flu-NS1-124-L7/L12-H1N1 and Flu-NS1-124-Omp16-H1N1. Despite of substantial modification of NS1 gene, all constructs replicated well and were retain their  Brucella  inserts over five passages in embryonated chicken eggs (CE). Administration of the mono- or bivalent vaccine formulation via prime-boost intranasal ( i.n .), conjunctival ( c .) or subcutaneous ( s.c. ) immunization was safe in mice; no deaths, body weight loss or pathomorphological changes were observed over 56 days. Moreover, guinea pigs vaccinated  i.n.  with vaccine vectors did not shed the vaccine viruses through their upper respiratory tract after the prime and booster vaccination. These findings confirmed the replication-deficient phenotype of viral vectors. The highest antibody response to  Brucella  antigen was obtained with constructs expressing L7/L12 (ELISA, GMT 242.5-735.0); whereas the highest T-cell immune response- with construct expressing Omp16 (ELISPOT, 337 ± 52-651 ± 45 spots/4×10 5 cells), which was comparable ( P > 0.05 ) to the response induced by the commercial vaccine  B. abortus 19 . Interestingly,  c.  immunization appeared to be optimal for eliciting T-cell immune response. In guinea pigs, the highest protective efficacy after challenge with  B. abortus  544 was achieved with Omp16 expressing constructs in both monovalent or bivalent vaccine formulations; protective efficacy was comparable to those induced by a commercial live  B. abortus  19 vaccine.    Conclusion  Thus, influenza vectors expressing  Brucella  protective antigens can be developed as novel influenza vectored vaccine against  B. abortus  infection.

Influenza viral vectors expressing the Brucella OMP16 or L7/L12 proteins as vaccines against B. abortus infection