Investigating Field Efficacy and Safety of Conjunctival Brucella abortus S19 Vaccine in Cattle

Background: Vaccination is the most fundamental strategy in the control and eradication of brucellosis. Several vaccination programs with different vaccines have been carried out in many countries in which brucellosis continues to be a problem in livestock. One of the recommended vaccines against brucellosis in cattle is the live Brucella abortus S19 vaccine. The aim of this study is to evaluate the results of field safety and efficacy trials for the conjunctival Brucella abortus S19 vaccine prior to the mass vaccination program. Methods: In this study, 81 female cattle were vaccinated with a reduced dose of Brucella abortus S19 vaccine with the conjunctival route. The immune response after vaccination was investigated by suggested serological tests; namely, Rose Bengal Plate Test, Serum Agglutination Test and Complement Fixation Test. Result: No adverse effect was observed within the scope of safety. Isolation of vaccine strain was observed only in a milk sample of lactating animals. Excluding the diagnosis criteria of the serological tests, humoral immune response was observed in most of the animals by all the serological tests one month after vaccination. Antibody levels lasted approximately 4 months after vaccination. In conclusion, the results of this study demonstrated that besides vaccine-induced antibodies, the vaccine including changes in dose and administration way in this study did not cause any significant risks for the target animals.

Brucella abortus S19 GFP-tagged vaccine allows the serological identification of vaccinated cattle

Bovine brucellosis induces abortion in cows, produces important economic losses, and causes a widely distributed zoonosis. Its eradication was achieved in several countries after sustained vaccination with the live attenuated Brucella abortus S19 vaccine, in combination with the slaughtering of serologically positive animals. S19 induces antibodies against the smooth lipopolysaccharide (S-LPS), making difficult the differentiation of infected from vaccinated bovines. We developed an S19 strain constitutively expressing the green fluorescent protein (S19-GFP) coded in chromosome II. The S19-GFP displays similar biological characteristics and immunogenic and protective efficacies in mice to the parental S19 strain. S19-GFP can be distinguished from S19 and B. abortus field strains by fluorescence and multiplex PCR. Twenty-five heifers were vaccinated withS19-GFP (5×109 CFU) by the subcutaneous or conjunctival routes and some boosted with GFP seven weeks thereafter. Immunized animals were followed up for over three years and tested for anti-S-LPS antibodies by both the Rose Bengal test and a competitive ELISA. Anti-GFP antibodies were detected by an indirect ELISA and Western blotting. In most cases, anti-S-LPS antibodies preceded for several weeks those against GFP. The anti-GFP antibody response was higher in the GFP boosted than in the non-boosted animals. In all cases, the anti-GFP antibodies persisted longer, or at least as long, as those against S-LPS. The drawbacks and potential advantages of using the S19-GFP vaccine for identifying vaccinated animals in infected environments are discussed.

Use of Postbiotics as an Immunomodulatory of the Immune Response against Brucellosis in Nellore Calves Immunized with S 19 Vaccine

Background: Brucellosis is an important public health disease and a great problem in the cattle production. Objectives: The aim of this study was to evaluate the immunomodulatory efficiency of a commercial postbiotic in Nellore calves immunized with the Brucella abortus S19 vaccine. Methods: We used 40 calves negative for Brucella spp. organizated into four treatments during 15 days: InRum (Ingulbal Ruminant®); InPro (Ingulbal Protein®); RumPro (Ingulbal Ruminant® and Ingulbal Protein®); and Cont: control. Collections of whole blood and serum samples were performed at the beginning of the administration of postbiotic and at 15, 45, 75 and 105 days after start the experiment. At 15 days, all animals received the mandatory vaccine S19. In order to assess immunological parameters, the means of total white cells counts, total lymphocytes, monocytes and neutrophils, and total IgG antibodies were determined. Results: It found a significative increase (P<0.05) of white cells counts and serum IgG levels in the three treatment groups throughout the experiment. Additionally, we observe a tendency not significative in a greater number of lymphocytes, monocytes and neutrophils counts in the treat calves. Conclusion: Our results suggest that the supplementation with postbiotic is a promising alternative to modulate both the cellular and humoral immune response of S19 vaccine against bovine brucellosis.

Efficacy of Brucella abortus S19 and RB51 vaccine strains: a systematic review and meta-analysis

This systematic review and meta-analysis aimed to recalculate the efficacy of these two vaccine strains, and to discuss the main variables associated with controlled trials to evaluate bovine brucellosis vaccines efficacy. The most used vaccine strain was S19, at the dose of 10 colony forming units (CFU), followed by the vaccine strain RB51 at 10 CFU. The most used challenge strain was B. abortus 2308, at the dose of 10 CFU by intraconjunctival route. For the meta-analysis, trials were grouped according to the vaccine strain and dose to recalculate protection against abortion (four groups) or infection (five groups), using pooled risk ratio (RR) and vaccine efficacy (VE). For protection against abortion (n = 15 trials), S19 vaccine at 10 CFU exhibited the highest protection rate (RR = 0.25, 95% CI: 0.12 to 0.52; VE = 75.09%, 95% CI: 48.08 – 88.05), followed by RB51 10 (RR = 0.31, 95% CI: 0.16 to 0.61; VE = 69.25%, 95% CI: 39.48 – 84.38). For protection against infection (n = 23 trials), only two subgroups exhibited significant protection: S19 at 10 CFU (RR = 0.28, 95% CI: 0.14 to 0.55; VE = 72.03%, 95% CI: 57.70 – 81.50) and RB51 at 10 CFU dose (RR = 0.43, 95% CI: 0.22 to 0.84; VE = 57.05%, 95% CI: 30.90 – 73.30). In conclusion, our results suggest that the dose of 10 CFU for S19 and 10 CFU for RB51 are the most suitable for the prevention of abortion and infection caused by B. abortus.

Single-tube duplex-PCR for specific detection and differentiation of Brucella abortus S19 vaccine strain from other Brucella spp

A simple single-tube duplex-PCR assay was optimized for rapid and sensitive detection and differentiation of Brucella abortus (B.abortus) S19 vaccine strain from other Brucella spp.(B.abortus 544,B abortus S99,B.melitensis 16M,B.suis).This assay was optimized using two primer pairs that were designed, one targeting genus specific multicopy IS711 and another eryC for the development of the duplex-PCR assay. Specificity of the assay was assessed using DNA templates from various Brucella species (B.abortus, B.melitensis, B.suis) and non Brucella bacteria like Staphylococcus aureus, Proteus vulgaris, Pasteurella multocida, Pseudomonas aeruginosa, Citrobacter freundii, E. coli, Salmonella enteritidis, Campylobacter jejuni and Listeria monocytogenes. The assay was also evaluated on spiked milk samples with known B.abortus cultures. The assay was able to detect up to 2.47 x 103CFU of B. abortus S99 organism in spiked milk sample. The assay was further validated in 53 clinical specimens (aborted foetal stomach content). Results confirm that Single tube duplex-PCR assay was useful for early and rapid detection and differentiation of B. abortus vaccine strain S19 from other Brucella spp in milk as well as in clinical specimen.

Stability of the B. abortus S19 vaccine strain with a eukaryotic expression plasmid encoding the G glycoprotein from the rabies virus

Brucella abortus S19 is an intracellular vaccine strain against bovine brucellosis. Rabies is a lethal disease in cattle. Plasmids encoding the G glycoprotein from the rabies virus induce a protective immune response in different animal species. A vector called pBBR4-CMV-Ggp-SV40+, which encodes the G gene, regulated by the cytomegalovirus eukaryotic expression promoter, and which can be used to transform the B. abortus S19 vaccine strain, was constructed. The stability of the transformant strain was tested both in vitro and in vivo. In the in vitro assays, B. abortus S19 pBBR4-CMV-Ggp-SV40+ was grown for 5 sequential passages, and for the in vivo assays, female BALB/c mice were infected. Colony-forming unit counting and plasmid identification were performed in each passage and in the spleens at 7 days post-infection. To test the plasmid stability in the strain, all parameters were determined with and without antibiotic. The bacterial concentration was lower with antibiotic than without it, but the bacterial growth was more homogeneous. The plasmid was identified in antibiotic- and non-antibiotic-treated isolated colonies under both in vitro and in vivo conditions. The plasmid construct was also transfected into BHK-21 cells, which express the G glycoprotein. The B. abortus S19 pBBR4-CMV-Ggp-SV40+ strain showed stability, representing a suitable candidate vector for developing a bivalent vaccine against brucellosis and rabies. This is the first time that a Brucella species has been transformed with a eukaryotic expression plasmid.