Clearance of bacteria from lymph nodes in sheep immunized with Brucella suis S2 vaccine is associated with M1 macrophage activation.

Ovine brucellosis is a global zoonotic disease of sheep caused by Brucella melitensis, which inflicts a significant burden on human and animal health. Brucella suis strain S2 (B. suis S2) is a smooth live attenuated vaccine for the prevention of ovine brucellosis in China. However, no previous studies have assessed the immunogenicity of B. suis S2 vaccine after oral immunization in sheep. Here, we attempted to evaluate the ovine immune response over the course of B. suis S2 immunization and to identify in vivo predictors for vaccine development. Body temperature, serum Brucella antibodies, serum cytokines (IL-12p70 and interferon [IFN]-γ), and bacterial load in the mandibular lymph nodes (LN), superficial cervical LN, superficial inguinal LN, and spleen were investigated to determine the safety and efficacy of the vaccine. The abnormal body temperature of sheep occurred within 8 days post-infection (dpi). Brucella suis S2 persisted for a short time (< 21 dpi) in the mandibular LN. The highest level of IL-12p70 was observed at 9 dpi, whereas serum IFN-γ levels peaked at 12 dpi. Transcriptome analysis and quantitative reverse transcription PCR were performed to determine gene expression profiles in the mandibular LN of sheep. Antigen processing and presentation pathway was the dominant pathway related to the dataset. Our studies suggest that the immune response in ovine LN resembled type 1 immunity with the secretion of IL-12p70 and IFN-γ after B.suis S2 immunization and the vaccine may eliminate Brucella via stimulation of M1 macrophages through the course of Th cells.

Immunization effect of lipopolysaccharide antigen in conjugation with PLGA nanoparticles as a nanovaccine against Brucella melitensis infection.

Brucella is an infectious disease with difficult treatment faced with drug resistance and recurrence of infection. Despite advances in the development of effective vaccines against brucellosis infections, there is still a need for more effective vaccine against brucellosis. In this study, we developed a nanovaccine for delivery of lipopolysaccharide Brucella melitensis antigen to the immune system using PLGA nanoparticles to prevent Brucella infection, which is associated with the stimulation of both humoral and cellular immune systems. In particular, we determined the rate of produced immunoglobulines and their functional effectiveness on the immune system by carring out opsonophagocytosis and challenge tests. According to the results, it was determined that PLGA improve the delivery of LPS antigen to the immune system to enhance the production of immunoglobulins levels and their efficiency to remove Brucella bacteria.

Knowledge, attitudes, and practices (KAP) related to brucellosis and factors affecting knowledge sharing on animal diseases: a cross-sectional survey in the dry zone of Sri Lanka.

Farmers' lack of knowledge is assumed to have affected the presence of brucellosis in Sri Lanka for decades. This study, carried out in the Ampara district in the dry zone of Sri Lanka, revealed that there is a significant knowledge gap for brucellosis compared to foot and mouth disease (FMD) (p < 0.001). Only 8.3% of farmers knew that brucellosis causes cattle abortions. Only 2.6% knew that it is zoonotic. The difference in knowledge of the symptoms and transmission of brucellosis and FMD was significant (p < 0.001). Farmers' attitudes and practices related to the spread of the disease were poor. Farmers' education and spoken language had a negative influence on knowledge. Young people and those with strong social relationships were efficient in knowledge sharing. It can be concluded that brucellosis knowledge, attitudes, and practices are poor; thus, there is a need for more attention in disease control policymaking. Backward farmer groups should be the focus in animal health extension programs.

Prediction of T cell epitopes of Brucella abortus and evaluation of their protective role in mice.

Brucellae are Gram-negative intracellular bacteria that cause an important zoonotic disease called brucellosis. The animal vaccines are available but have disadvantage of causing abortions in a proportion of pregnant animals. The animal vaccines are also pathogenic to humans. Recent trend in vaccine design has shifted to epitope-based vaccines that are safe and specific. In this study, efforts were made to identify MHC-I- and MHC-II-restricted T cell epitopes of Brucella abortus and evaluate their vaccine potential in mice. The peptides were designed using online available immunoinformatics tools, and five MHC-I- and one MHC-II-restricted T cell peptides were selected on the basis of their ability to produce interferon gamma (IFN-γ) in in vivo studies. The selected peptides were co-administered with poly DL-lactide-co-glycolide (PLG) microparticles and evaluated for immunogenicity and protection in BALB/c mice. Mice immunized with peptides either entrapped in PLG microparticles (EPLG-Pep) or adsorbed on PLG particles (APLG-Pep) showed significantly higher splenocyte proliferation and IFN-γ generation to all selected peptides than the mice immunized with corresponding irrelevant peptides formulated PLG microparticles or phosphate-buffered saline (PBS). A significant protection compared to PBS control was also observed in EPLG-Pep and APLG-Pep groups. A plasmid DNA vaccine construct (pVaxPep) for peptides encoding DNA sequences was generated and injected to mice by in vivo electroporation. Significant protection was observed (1.66 protection units) when compared with PBS and empty vector control group animals. Overall, the MHC-I and MHC-II peptides identified in this study are immunogenic and protective in mouse model and support the feasibility of peptide-based vaccine for brucellosis.

Effective, broad spectrum control of virulent bacterial infections using cationic DNA liposome complexes combined with bacterial antigens.

Protection against virulent pathogens that cause acute, fatal disease is often hampered by development of microbial resistance to traditional chemotherapeutics. Further, most successful pathogens possess an array of immune evasion strategies to avoid detection and elimination by the host. Development of novel, immunomodulatory prophylaxes that target the host immune system, rather than the invading microbe, could serve as effective alternatives to traditional chemotherapies. Here we describe the development and mechanism of a novel pan-anti-bacterial prophylaxis. Using cationic liposome non-coding DNA complexes (CLDC) mixed with crude F. tularensis membrane protein fractions (MPF), we demonstrate control of virulent F. tularensis infection in vitro and in vivo. CLDC+MPF inhibited bacterial replication in primary human and murine macrophages in vitro. Control of infection in macrophages was mediated by both reactive nitrogen species (RNS) and reactive oxygen species (ROS) in mouse cells, and ROS in human cells. Importantly, mice treated with CLDC+MPF 3 days prior to challenge survived lethal intranasal infection with virulent F. tularensis. Similarly to in vitro observations, in vivo protection was dependent on the presence of RNS and ROS. Lastly, CLDC+MPF was also effective at controlling infections with Yersinia pestis, Burkholderia pseudomallei and Brucella abortus. Thus, CLDC+MPF represents a novel prophylaxis to protect against multiple, highly virulent pathogens.

Escheriosomes entrapped DNA vaccine co-expressing Cu-Zn superoxide dismutase and IL-18 confers protection against Brucella abortus.

In the present study, we evaluated prophylactic prospective of liposome based DNA vaccine co-expressing Cu-Zn superoxide dismutase (SOD) along with interleukin-18 (IL-18) against experimental murine brucellosis. The immunization schedule involves liposome-mediated delivery of pVsod (encoding SOD of Brucella abortus) and pVIL18-sod (encoding IL-18 of mouse and SOD of B. abortus) DNA constructs. The data highlight potential of Escherichia coli lipid liposome (escheriosome) based DNA delivery vehicle to induce SOD specific humoral and cellular immune responses in the immunized mice. The co-expression of SOD along with IL-18 ensued in higher lymphoproliferative response and IFN-gamma production in comparison to the group of animals that were immunized with free form of SOD-DNA. Antibody response developed upon immunization with both DNA vaccines was of IgG2a type mainly. The results of the present study show that co-expression of IL-18 along with SOD polarized the antigen specific immune responses toward Th-1 direction, a desirable feature to control intracellular pathogens.

Protective efficacy of a canine brucellosis vaccine candidate based on live attenuated Salmonella expressing recombinant Brucella BCSP31, Omp3b and SOD proteins in Beagles.

The aim of this study was to establish a proof-of-concept of protective efficacy of Salmonella-based B. abortus vaccine candidate in Beagles. Group A Beagles (n=10) were subcutaneously (SC) inoculated with S. Typhimurium delivery strain containing pMMP65 (vector to deliver antigens) only as vector control. Group B Beagles (n=10) were SC vaccinated with the mixture of the three Salmonella delivery strains expressing the recombinant B. abortus BCSP31, Outer membrane protein 3b (Omp3b), and superoxide dismutase (SOD) proteins, respectively. No Salmonella delivery strains were isolated from all tissues tested. Serum IgG, interleukin-4, tumor necrosis factor-alpha, and interferon-gamma concentrations were significantly higher in group B than in group A. Following intraconjunctival challenge with B. abortus 544, among 5 group B Beagles, the challenge strain was isolated from mandibular, and retropharyngeal lymph nodes of three Beagles, and no isolates were observed from all tissues of two Beagle. However, the challenge strains were detected from spleen, uterus (except two Beagles), and mandibular, prescapular, retropharyngeal, and superficial inguinal lymph nodes of all group A Beagles. These results suggest that the mixture of three S. Typhimurium delivery strains be a good vaccine candidate against brucellosis by B. abortus in dogs. Further investigations are needed to improve the protective efficacy of the Salmonella-based B. abortus vaccine candidate and explore its practical application in dogs.

Recombinant L7/L12 protein entrapping PLGA (poly lactide-co-glycolide) micro particles protect BALB/c mice against the virulent B. abortus 544 infection.

Brucella abortus is the etiologic agent of Brucellosis, a zoonotic infection affecting a wide range of animals. It is a highly infectious disease of pandemic potential reporting over 500,000 new human cases annually. Till date, there is no reported vaccine for humans and the available animal vaccines are unsafe, therefore a safe and effective subunit vaccine is highly sought for. In this study, we have evaluated rL7/L12 protein encapsulated in microparticles of PLGA (85:15), a biocompatible and biodegradable polymer approved by FDA for human use. In this work, BALB/c mice have been immunized with rL7/L12 entrapped in microparticles in a prime-boost regimen. Further, evaluation of the immunogenicity of the formulation showed that the IgG antibody titre reached a maxima of 2.2×10(5) (p value 0.0001 v/s control) after the injection of the booster dose. A mixed IgG isotype profile (IgG1/IgG2a) indicated the stimulation of both the cellular as well as humoral immunity which has increased parallely and gradually since the first immunization. High levels of IFN-γ, 815±55pg/ml were recorded depicting an optimal elicitation of the cellular wing of immunity leading to clearance of splenic bacteria upto 1.69 log units.

PLGA (85:15) nanoparticle based delivery of rL7/L12 ribosomal protein in mice protects against Brucella abortus 544 infection: A promising alternate to traditional adjuvants.

There is a compelling need for the development of suitable adjuvants for human use to enhance the efficacy of the upcoming vaccines for the prevention of life threatening infections. In the current study, we have tried to explore the immunogenic potential of nanoparticles (NPs) made of PLGA (poly lactic-co-glycolic acid), a biodegradable and biocompatible polymer approved by FDA for human use after entrapping rL7/L12 protein, an immunodominant antigen of Brucella. Adjuvant properties were exhibited by the formulation as it elicited high IgG antibody titers just after first immunization which increased significantly after the booster administration. A good elicitation of the Th1 cytokines especially IFN-γ was recorded. Amongst the IgG antibody subclasses, IgG1 remained the predominant subclass to be elicited in mice serum after immunization; however IgG1/2a ratio showed a mixed profile of Th1/Th2 response. Lymphocyte proliferation assay as a marker of amplification in cellular immunity demonstrated that the splenocytes of the immunized mice had a high proliferation index with reference to the control, revealing that L7/L12 entrapping PLGA nanoparticles are potent inducer of inflammatory cell response indispensable to combat Brucella infection. Enumeration of splenic CFU after 14 days of infection with Brucella abortus 544 showed a significant reduction in log CFU of splenic bacteria in the vaccinated mice as compared to the control group. Therefore it is evident that PLGA nano formulations delivering the entrapped vaccine candidate in mice elicit specific humoral as well as cellular responses specific to the entrapped Brucella antigen. So there is much promise in this approach and this work by highlighting the adjuvant properties of the PLGA nanospheres will accelerate the development of improved vaccines safe for human as well as veterinary use.

Protective effect of Brucella outer membrane complex-bearing liposomes against experimental murine brucellosis.

Liposomes of stable multilamellar type, which previously demonstrated great efficiency in antibiotic transport, were used in this study as transport vehicles of antigenic extracts of Brucella melitensis (HS: complex of lipopolysaccharide/phospholipids/outer membrane proteins). The incorporation of HS into positively charged liposomes produced a protective effect against experimental murine brucellosis when they were administered 1 day before or 2 days after infection, as the number of colony-forming units in the spleen was reduced in relation to the untreated control group (P < 0.01). On the other hand, the use of HS-free or bound in liposomes with negative net charge did not produce a significant effect. Moreover, the incorporation of HS into cationic liposomes eliminated the toxicity of the lipopolysaccharide.

Polyester microparticles as a vaccine delivery system for brucellosis: influence of the polymer on release, phagocytosis and toxicity.

Microparticles, containing an antigenic complex from Brucella ovis (HS), were evaluated for vaccine purposes against brucellosis. They were prepared by the double emulsion solvent evaporation method using two different polyesters, poly-lactide-co-glycolide acid (75:25; RG 756) and poly-epsilon-caprolactone. The encapsulation efficiency and release of HS from the microparticles, their capacity to be phagocytosed and also their toxicity on murine monocytes J774.2 were investigated. Both polymers lead to smooth and spherical sub-5 microm particles, with approximately 30% of the antigen initial dose encapsulated. SDS-PAGE and immunoblot of extracted antigens confirmed that the apparent molecular weight and antigenicity remained unaltered after the encapsulation procedure. However, the in vitro release of the antigens differed among them. The release profile for PLGA microparticles was continuous, whereas PEC ones released the antigens in a triphasic release pattern. Phagocytosis was clearly influenced by the hydrophobicity of the polymer, increasing in the case of PEC microparticles. Toxicity assay showed that both types of microparticles induced similar levels of mitochondrial damage. In conclusion, HS-PEC microparticles could be used as an effective vaccine against brucellosis, as the antigen is released in boosters and they are greatly phagocytosed by macrophages.

Development of microparticles prepared by spray-drying as a vaccine delivery system against brucellosis.

The antigenic extract Hot Saline from Brucella ovis was microencapsulated by the spray-drying technique with different polyesters (poly-lactide-co-glycolide RG502H [PLGA], and blends with poly- epsilon -caprolactone [PEC]) in order to obtain microparticles smaller than 5 microm. Microparticles were tested for encapsulation efficiency, release studies, acidification of the in vitro release medium, and in vitro J744-macrophage experiments (phagocytosis and toxicity of the preparations) to determine the optimal formulation for vaccination purposes. Formulation containing no PCL showed the highest encapsulation efficiency, although the differences were not significant. The in vitro release kinetics were characterized by a high burst effect after 1 h of incubation, followed by a slow and continuous release. For the formulation based on PLGA, the pH of the medium during release dropped from 7.4 to 3.5 while the presence of PEC attenuated the pH drop. All formulations showed light toxicity by the MTT assay, but differences were observed in terms of phagocytosis, as particles prepared with PEC showed the higher uptake by J744-macrophages and cell respiratory burst, determined by oxygen peroxide release. All these characteristics suggest that the microparticulated antigenic formulation containing the higher ratio of PEC is susceptible to be used in animal vaccination studies.

Influence of the co-encapsulation of different excipients on the properties of polyester microparticle-based vaccine against brucellosis.

This work evaluates the influence of different pharmaceutical auxiliaries (Pluronic F68, polyvinylpyrrolidone [PVP] or Tween 20), when mixed with an antigenic extract from Brucella ovis (hot saline; HS), on the characteristics of the resulting poly(epsilon-caprolactone) (PEC) and poly(lactide-co-glycolide) (PLGA) microparticles. In all cases, PEC microparticles were smaller than PLGA ones. Concerning the HS loading, PLGA microparticles were highly dependent on the type of the excipient used, whereas all the PEC formulations displayed similar encapsulation efficiencies. For both types of microparticles, the presence of PVP induced a burst release effect. On the contrary, the use of Tween 20 or Pluronic F68 dramatically modified this profile. For PLGA-Tween 20 and PEC-Pluronic F68 microparticles, the HS was released in a pulsatil way during the first 7 days followed by a continuous release for at least 3 weeks. The antigenicity of the HS components was kept in all cases. Phagocytosis by murine monocytes showed a clear difference based just on the hydrophobicity of the polymer, being PEC microparticles better engulfed. Cell activation quantified by the release of H2O2 did not showed major differences between batches, however, microparticles of PEC and Pluronic F68 induced the highest nitric oxide production. Together, these results confirm the advantageous qualities of the "HS-PEC-Pluronic F68 microparticles" as favorable candidate for vaccine purposes against brucellosis.

Efficacy of calfhood vaccination with Brucella abortus strain RB51 in protecting bison against brucellosis.

In the studies reported here, protection induced by calfhood vaccination of bison with 1.2-6.1 x 10(10)CFU of Brucella abortus strain RB51 (SRB51) against a virulent strain of B. abortus was evaluated. Non-vaccinated and SRB51-vaccinated bison were intraconjunctivally challenged during midgestation with 3 x 10(7)CFU of virulent B. abortus strain 2308 (S2308). Maternal and fetal tissues were obtained within 24hour after abortion or parturition. Incidence of abortion was greater (P<0.05) in non-vaccinated as compared to SRB51-vaccinated bison (62% and 15%, respectively), with abortions occurring between 5 and 8 weeks after experimental challenge. Calves from bison vaccinated with SRB51 had a reduced (P<0.05) prevalence of fetal infection with S2308 as compared to calves from non-vaccinated bison (19% and 62%, respectively). Although the ability to recover the 2308 challenge strain from maternal tissues did not differ (P>0.05) between nonvaccinates and vaccinates (100% and 78%, respectively), calfhood vaccination with SRB51 reduced (P<0.05) recovery of S2308 from uterine or mammary gland tissues. In bison which did not abort, S2308 was routinely recovered in low numbers from maternal lymphatic tissues; particularly the parotid, bronchial, supramammary, and mandibular lymph nodes. The RB51 vaccine strain was not recovered at any time from maternal or fetal samples obtained at necropsy. Histological lesions associated with Brucella-induced abortions were suppurative placentitis, fetal broncho-interstitial pneumonia, and fetal histiocytic splenitis. The results of our studies suggest that calfhood vaccination of bison with SRB51 is efficacious in protecting against intramammary, intrauterine, and fetal infection following exposure to a virulent strain of B. abortus during pregnancy. As brucellosis is transmitted horizontally through fluids associated with the birth or abortion of an infected fetus, or vertically to the calf through the ingestion of milk containing B. abortus, our data suggest that calfhood vaccination with SRB51 will be beneficial in preventing transmission of brucellosis in bison.

Use of S-[2,3-bispalmitoyiloxy-(2R)-propyl]-R-cysteinyl-amido-monomethoxy polyethylene glycol as an adjuvant improved protective immunity associated with a DNA vaccine encoding Cu,Zn superoxide dismutase of Brucella abortus in mice.

This study was conducted to evaluate the immunogenicity and protective efficacy of a DNA vaccine encoding Brucella abortus Cu,Zn superoxide dismutase (SOD) using the Toll-like receptor 2/6 agonist S-[2,3-bispalmitoyiloxy-(2R)-propyl]-R-cysteinyl-amido-monomethoxy polyethylene glycol (BPPcysMPEG) as an adjuvant. Intranasal coadministration of BPPcysMPEG with a plasmid carrying the SOD-encoding gene (pcDNA-SOD) into BALB/c mice elicited antigen-specific humoral and cellular immune responses. Humoral responses were characterized by the stimulation of IgG2a and IgG1 and by the presence of SOD-specific secretory IgA in nasal and bronchoalveolar lavage fluids. Furthermore, T-cell proliferative responses and increased production of gamma interferon were also observed upon splenocyte restimulation with recombinant SOD. Cytotoxic responses were also stimulated, as demonstrated by the lysis of RB51-SOD-infected J774.A1 macrophages by cells recovered from immunized mice. The pcDNA-SOD/BPPcysMPEG formulation induced improved protection against challenge with the virulent strain B. abortus 2308 in BALB/c mice over that provided by pcDNA-SOD, suggesting the potential of this vaccination strategy against Brucella infection.

Pluronic P85 enhances the efficacy of outer membrane vesicles as a subunit vaccine against Brucella melitensis challenge in mice.

Brucellosis is the most common zoonotic disease worldwide, and there is no vaccine for human use. Brucella melitensis Rev1, a live attenuated strain, is the commercial vaccine for small ruminants to prevent B. melitensis infections but has been associated with abortions in animals. Moreover, strain Rev1 is known to cause disease in humans and cannot be used for human vaccination. Outer membrane vesicles (OMVs) obtained from B. melitensis have been shown to provide protection similar to strain Rev1 in mice against B. melitensis challenge. In the present work, we tested the efficacy of Pluronic P85 as an adjuvant to enhance the efficacy of Brucella OMVs as a vaccine. P85 enhanced the in vitro secretion of TNF-α by macrophages induced with OMVs and P85. Further, P85 enhanced the protection provided by OMVs against B. melitensis challenge. This enhanced protection was associated with higher total IgG antibody production but not increased IFN-γ or IL-4 cytokine levels. Moreover, P85 alone provided significantly better clearance of B. melitensis compared to saline-vaccinated mice. Further studies are warranted to find the mechanism of action of P85 that provides nonspecific protection and enhances the efficacy of OMVs as a vaccine against B. melitensis.

Epidemiological and clinical aspects of human brucellosis in eastern Anatolia.

BACKGROUND: We investigated the seroprevalence of human brucellosis and risk factors in a village in Eastern Anatolia, Turkey, where a brucellosis outbreak among cattle had recently occurred. PATIENTS AND METHODS: All 501 inhabitants enrolled were screened with the Rose Bengal Test and were asked to fill out a questionnaire to determine the signs and symptoms of brucellosis and the risk factors. Patients' laboratory findings and clinical responses were also evaluated. RESULTS: The Rose Bengal Test was positive in 44 persons. In comparison of age groups, fever (20.2%), arthralgia (19.2%) and weight loss (8.4%) were frequently seen in children (p<0.05). Hepatomegaly (17.9%) was also prevalent in the age group of 0 to 14 years (p<0.05). The consumption of dairy products, a family history of brucellosis, and raising livestock were significantly related to seropositivity (p<0.05). Rifampicin plus doxycyline was the most common regimen administered in these cases (54.5%) and also had the highest relapse rate (p<0.05). CONCLUSION: Brucellosis remains an important public-health problem in the rural areas of Turkey. Appropriate public health measures and education must be pursued to eradicate human brucellosis.

Co-immunization with interlukin-18 enhances the protective efficacy of liposomes encapsulated recombinant Cu-Zn superoxide dismutase protein against Brucella abortus.

Brucellosis is a worldwide zoonotic disease caused by Brucella abortus and a number of closely related species. Brucellosis has severe impact on the health and economic prosperity of the developing countries due to the persistent nature of infection and unavailability of effective control measures. The Cu-Zn superoxide dismuatse (SOD) protein of Brucella have been extensively studied as a major antigen involved in bacterial evading mechanism of host defence. Being a critical pro-inflammatory cytokine interleukin-18 (IL-18) plays key role in induction of immune mediated protection against intracellular pathogens. In the present study, we aimed to investigate the immunogenic potential of fusogenic liposomes (escheriosomes) encapsulated recombinant Cu-Zn SOD (rSOD) protein alone or in combination with recombinant IL-18 (rIL-18). Escheriosomes encapsulated rSOD mediated immune responses were further increased upon co-immunization with rIL-18. Furthermore, immunization with escheriosomes encapsulated rSOD alone or in combination with rIL-18, increased resistance in mice against challenge with B. abortus 544.

Evaluation of particulate acellular vaccines against Brucella ovis infection in rams.

The attenuated Brucella melitensis Rev 1 vaccine, used against brucellosis infection, interferes with serological diagnosis tests, may induce abortions in pregnant animals, and may infect humans. In order to overcome these drawbacks, we developed acellular vaccines based on a Brucella ovis antigenic complex (HS) containing outer membrane proteins and R-LPS entrapped in poly(anhydride) conventional and mannosylated nanoparticles (NP-HS and MAN-NP-HS) or in poly(epsilon-caprolactone) microparticles (HS-PEC) as antigen delivery systems and immunoadjuvants. Brucellosis free rams were vaccinated subcutaneously with a single dose of particles containing 3mg of HS, and challenged 6 months thereafter. Protection was evaluated by clinical, bacteriological and serological examinations, in comparison with non-vaccinated control rams. HS-PEC vaccine induced protection (7 out of 13 animals were infected) equivalent to that induced by the reference Rev 1 vaccine (8/14). In contrast, animals immunized with NP-HS were not protected, showing similar results to that obtained in the control unvaccinated rams. Furthermore HS-PEC vaccine did not interfere against B. melitensis serodiagnostic tests. In summary, HS-PEC microparticles could be used as a safe and effective vaccine against brucellosis in rams.

Immunopotentiation of live brucellosis vaccine by adjuvants.

In a series of studies in SPF and conventional guinea pigs, various adjuvants (larifan, polyoxidonium-PO, natrium thiosulphate-NT, TNF-ß and Ribi adjuvant system-RAS) were evaluated for their ability to enhance immune responses to the live brucellosis vaccine, Brucella abortus strain 82-PS (penicillin-sensitive). Combining adjuvants with S82-PS increased synthesis of antibodies against rough (R) and smooth (S) Brucella antigens. Dynamics and levels of antibodies differed dependent upon the adjuvant. Adjuvants enhanced cell-mediated responses to S82-PS, and phagocytosis by macrophages. Humoral and cellular immune responses stimulated by the adjuvants correlated with increased vaccine protection against experimental challenge. The highest protection was demonstrated by combining TNF-ß or PO with S82-PS. Our data demonstrates the potential of adjuvants to improve immunogenic properties of live brucellosis vaccines.