RESEARCH Open Access

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

Safety of the novel influenza viral vector Brucella abortus vaccine in pregnant heifers

ABSTRACT: The present study provides the first information about the safety of a new influenza viral vector vaccine expressing the Brucella ribosomal protein L7/L12 or Omp16 containing the adjuvant Montanide Gel01 in pregnant heifers. Immunization of pregnant heifers was conducted via the conjunctival (n=10) or subcutaneous (n=10) route using cross prime and booster vaccination schedules at an interval of 28 days. The vector vaccine was evaluated in comparison with positive control groups vaccinated with B. abortus S19 (n=10) or B. abortus RB51 (n=10) and a negative (PBS+Montanide Gel01; n=10) control group. Clinical studies, thermometry, assessment of local reactogenicity and observation of abortion showed that the vector vaccine via the conjunctival or subcutaneous route was completely safe for pregnant heifers compared to the commercial vaccines B. abortus S19 or B. abortus RB51. The only single adverse event was the formation of infiltration at the site of subcutaneous injection; this reaction was not observed for the conjunctival route

Safety of the novel vector vaccine against Brucella abortus based on recombinant influenza viruses expressing Brucella L7/L12 and OMP16 proteins, in cattle

This paper presents the results of a study of the safety of new vector vaccine against B. abortus based on recombinant influenza A subtype H5N1 or H1N1 (viral constructs vaccine formulation) viruses expressing Brucella ribosomal protein L7/L12 and Omp16,in cattle. To increase the effectiveness of the vaccine, adjuvants such as Montanide Gel01 or chitosan were included in its composition. Immunization of cattle (5 animals per group) with the viral constructs vaccine formulation only, or its combination with adjuvants Montanide Gel01 or chitosan, were conducted via the conjunctival method using cross prime (influenza virus subtype H5N1) and booster (influenza virus subtype H1N1) vaccination schedules. Vaccine candidates were evaluated in comparison with the positive (B. abortus S19) and negative (PBS) controls. Comprehensive studies involving thermometry and clinical examination, hematology and biochemical blood analysis, showed that all of the viral constructs vaccine formulation, as well as their combination with adjuvants, compared to the commercial bacterial vaccine B. abortus S19 were completely safe in cattle. Furthermore it is shown that the developed vaccines can effectively differentiate vaccinated animals from infected animals.</p

Improved influenza viral vector based Brucella abortus vaccine induces robust B and T-cell responses and protection against Brucella melitensis infection in pregnant sheep and goats.

We previously developed a potent candidate vaccine against bovine brucellosis caused by Brucella abortus using the influenza viral vector expressing Brucella Omp16 and L7/L12 proteins (Flu-BA). Our success in the Flu-BA vaccine trial in cattle and results of a pilot study in non-pregnant small ruminants prompted us in the current study to test its efficacy against B. melitensis infection in pregnant sheep and goats. In this study, we improved the Flu-BA vaccine formulation and immunization method to achieve maximum efficacy and safety. The Flu-BA vaccine formulation had two additional proteins Omp19 and SOD, and administered thrice with 20% Montanide Gel01 adjuvant, simultaneously by both subcutaneous and conjunctival routes at 21 days intervals in pregnant sheep and goats. At 42 days post-vaccination (DPV) we detected antigen-specific IgG antibodies predominantly of IgG2a isotype but also IgG1, and also detected a strong lymphocyte recall response with IFN-γ production. Importantly, our candidate vaccine prevented abortion in 66.7% and 77.8% of pregnant sheep and goats, respectively. Furthermore, complete protection (absence of live B. melitensis 16M) was observed in 55.6% and 66.7% of challenged sheep and goats, and 72.7% and 90.0% of their fetuses (lambs/yeanlings), respectively. The severity of B. melitensis 16M infection in vaccinated sheep and goats and their fetuses (index of infection and rates of Brucella colonization in tissues) was significantly lower than in control groups. None of the protection parameters after vaccination with Flu-BA vaccine were statistically inferior to protection seen with the commercial B. melitensis Rev.1 vaccine (protection against abortion and vaccination efficacy, alpha = 0.18-0.34, infection index, P = 0.37-0.77, Brucella colonization, P = 0.16 to P > 0.99). In conclusion, our improved Flu-BA vaccine formulation and delivery method were found safe and effective in protecting pregnant sheep and goats against adverse consequences of B. melitensis infection

Preclinical Evaluation of TB/FLU-04L—An Intranasal Influenza Vector-Based Boost Vaccine against Tuberculosis

Tuberculosis is a major global threat to human health. Since the widely used BCG vaccine is poorly effective in adults, there is a demand for the development of a new type of boost tuberculosis vaccine. We designed a novel intranasal tuberculosis vaccine candidate, TB/FLU-04L, which is based on an attenuated influenza A virus vector encoding two mycobacterium antigens, Ag85A and ESAT-6. As tuberculosis is an airborne disease, the ability to induce mucosal immunity is one of the potential advantages of influenza vectors. Sequences of ESAT-6 and Ag85A antigens were inserted into the NS1 open reading frame of the influenza A virus to replace the deleted carboxyl part of the NS1 protein. The vector expressing chimeric NS1 protein appeared to be genetically stable and replication-deficient in mice and non-human primates. Intranasal immunization of C57BL/6 mice or cynomolgus macaques with the TB/FLU-04L vaccine candidate induced Mtb-specific Th1 immune response. Single TB/FLU-04L immunization in mice showed commensurate levels of protection in comparison to BCG and significantly increased the protective effect of BCG when applied in a “prime-boost” scheme. Our findings show that intranasal immunization with the TB/FLU-04L vaccine, which carries two mycobacterium antigens, is safe, and induces a protective immune response against virulent M. tuberculosis.</i

Improved influenza viral vector based <i>Brucella abortus</i> vaccine induces robust B and T-cell responses and protection against <i>Brucella melitensis</i> infection in pregnant sheep and goats - Fig 4

<p>Lymphocyte stimulation index (A, C) and levels of IFN-γ (B, D) in the supernatants of PBMCs of pregnant sheep and goats at 42 and 63 days post-vaccination (DPV). Pregnant sheep and goats in the group I (Flu-BA_Omp19-SOD) were immunized twice concurrently via the subcutaneous and conjunctival routes at an interval of 21 days with vaccines generated from the influenza viral vectors (IVV) subtypes H5N1 (prime vaccination) and H1N1 (booster vaccination). The vaccination of animals of group II (Flu-BA_Omp19-SOD_TV) was carried out in the same way as in group I, but only the vaccine generated from IVV subtypes H5N1 was used, which was administered three times at 21 days intervals. Sheep and goats in the negative control group (IV) were vaccinated with 20% Montanide Gel01 adjuvant in PBS three times at 21 days intervals. All data are presented as mean ± standard error; * <i>P = 0</i>.<i>02 to P<0</i>.<i>0001</i> vs. appropriate control group; † <i>P = 0</i>.<i>02–0</i>.<i>01</i> vs. Day 42 DPV. Statistical analysis was performed using two-way ANOVA followed by Tukey's multiple comparisons test. <i>P</i> values < 0.05 were considered significant. NI—not investigated.</p

Colonization and incidence of recovery of <i>B</i>. <i>melitensis</i> in tissues after challenge with <i>B</i>. <i>melitensis</i> 16M.

<p>Colonization and incidence of recovery of <i>B</i>. <i>melitensis</i> in tissues after challenge with <i>B</i>. <i>melitensis</i> 16M.</p

Index of infection for sheep and goats challenged with <i>B</i>. <i>melitensis</i> 16M at 113–120 days post-vaccination.

<p>Pregnant sheep and goats in the group I (Flu-BA_Omp19-SOD) were immunized twice concurrently via the subcutaneous and conjunctival routes of administration at an interval of 21 days with vaccines generated from the influenza viral vectors (IVV) subtypes H5N1 (prime vaccination) and H1N1 (booster vaccination). The vaccination of animals of group II (Flu-BA_Omp19-SOD_TV) was carried out in the same way as in group I, but only the vaccine generated from IVV subtypes H5N1 was used, which was administered three times at 21 days intervals. Sheep and goats in the negative control group (IV) were vaccinated with 20% Montanide Gel01 adjuvant in PBS three times at 21 days intervals. Animals in the positive control group (III) were immunized once subcutaneously in the axillary region (right side) with commercial vaccine <i>B</i>. <i>melitensis</i> Rev.1 according to the manufacturer's instructions. Challenge with the virulent strain <i>B</i>. <i>melitensis</i> 16M was performed via the subcutaneous route (10⁶ CFU/animal). The index of infection is the number of animals from which <i>Brucella</i> was isolated from the organs and lymph nodes. The data presented as mean ± standard error; * <i>P <0</i>.<i>0001</i> vs. appropriate control group IV. Statistical analysis was performed using two-way ANOVA followed by Tukey's multiple comparisons test. <i>P</i> values < 0.05 were considered significant.</p