Efficacy of different commercial and new inactivated vaccines against ovine enzootic abortion

The protective efficacy of two inactivated commercial (A, B) and two new inactivated vaccines (M7, QS) against ovine enzootic abortion was determined in two separate experiments in sheep. Vaccine A contained chlamydiae propagated in chicken embryos, adjuvated with Marcol 82, and vaccine B contained chlamydiae cultured in cell monolayers, adjuvated with aluminium hydroxide. For the preparation of the experimental vaccines, Chlamydophila abortus AB7 strain was cultured in McCoy cells and adjuvated with QS-21 (QS) or Montanide ISA 773 (M7). The ewes were vaccinated twice subcutaneously and challenged at 90 days of gestation. Protection was evaluated by clinical, bacteriological and serological examinations, and compared to two control groups: one of infected but not vaccinated ewes, and another of vaccinated but not infected ewes. The experimental vaccines induced considerably better protection than the two commercial ones. The new vaccine M7 especially showed no abortions, a good antibody response, the highest newborn lamb weights and the lowest level of C. abortus shedding at lambing.     

Role of polymorphonuclear neutrophils (PMNs) and NK cells in the protection conferred by different vaccines against Chlamydophila abortus infection

Ovine enzootic abortion (OEA) is caused by Chlamydophila abortus, an intracellular bacterium which acts by infecting the placenta, causing abortion in the last term of gestation. The main prevention strategy against OEA is the vaccination of flocks. An effective vaccine against C. abortus must induce a Th1-like specific immune response, which is characterized by the early production of IFN-gamma and the activation of CD8(+)T cells. Moreover, vaccine effectiveness could be modulated by the functioning of the innate immunity. The purpose of this study was to ascertain how polymorphonuclear neutrophils (PMNs) and NK cells might influence vaccine-induced protection. The live attenuated 1B vaccine and two inactivated experimental vaccines, adjuvated with aluminium hydroxide (AH) or QS-21 (QS), were used in PMN-depleted or NK cell-depleted mice. For PMN depletion, RB6-8C5 monoclonal antibody, which recognizes GR1(+) receptors (Robben, P.M., LaRegina, M., Kuziel, W.A., Sibley, L.D. 2005. Recruitment of Gr-1(+) monocytes is essential for control of acute toxoplasmosis. The Journal of Experimental Medicine 201, 1761-1769.) was used, while for NK cell-depletion the anti-asialo GM1 polyclonal antibody was used. The depletion of PMNs caused 100% mortality in non-vaccinated mice (NV) and 60% mortality in the AH-vaccinated mice by day 10 p.i., while both groups showed a significant increase in their bacterial burden in the liver by day 4 p.i. The depletion of NK cells caused mortality only in the NV group (50% by day 10 p.i.), although this group and the 1B vaccinated mice showed an increased bacterial burden in the liver at day 4 p.i. Our results suggest that the importance of PMNs in inactivated vaccines depends on the adjuvant chosen. The results also demonstrated that the importance of NK cells is greater in live vaccines than in inactivated vaccines.     

Les vaccins dans la prevention de la rhinotracheite infectieuse du chat adjuvant BCG gel d''alumine (preparation, controle, immunite)

Immunogenicity of different vaccines against the main viral diseases was compared: feline infectious rhinotracheitis, feline panleucopenia virus and calicivirus. These inactivated vaccines developed higher protective activity when adjuvants such as BCG or aluminium hydroxide were used. Animals vaccinated and boostered with an identical dose of these vaccines resisted the viral challenges. Neutralizing antibodies titers obtained with BCG adjuvated vaccines were twofold higher than those containing aluminium hydroxide.     

Protection of mice against Chlamydophila abortus infection with a bacteriophage-mediated DNA vaccine expressing the major outer membrane protein

A bacteriophage-delivered DNA vaccine against Chlamydophila abortus was constructed by cloning a eukaryotic cassette containing the ompA gene (which expresses the Major Outer Membrane Protein) into a bacteriophage lambda vector. Four groups, each of 20 BALB/c mice were inoculated separately with the phage vaccine, a conventional DNA vaccine based on the same ompA expression cassette, a live attenuated vaccine (strain 1B) or the empty phage vector. The phage and DNA vaccines and empty phage vector were administered intramuscularly on days 0, 14 and 28; the attenuated vaccine was given once on day 0. Half the animals in each group were challenged on day 42 by intraperitoneal injection of live C. abortus and sacrificed on day 49. Phage-vaccinated mice developed moderate antibody levels against C. abortus and yielded higher levels of IFN-γ and IL-2 compared with the attenuated live vaccine group. Clearance of chlamydiae from spleens was significantly better in the attenuated vaccine group compared with the phage vaccine group, while both groups were significantly superior to the DNA vaccine and control groups (p<0.01). Although levels of protection in the mouse model were lower in phage-vaccinated animals, than in 1B vaccinated animals, phage vaccines offer several other advantages, such as easier handling and safety, potentially cheaper production and no chance of reversion to virulence. Although these are preliminary results in a model system, it is possible that with further optimisation immunization with phage vaccines may provide a novel way to improve protection against C. abortus infection and trials in large animals are currently being initiated.     

Development of a genetically engineered vaccine against feline leukemia virus infection.

A genetically engineered subunit vaccine against FeLV infection was developed. The protective immunogen in the vaccine was a purified recombinant protein containing the entire amino acid sequence of FeLV subgroup A gp70 envelope protein. The optimal adjuvant was determined to be a highly purified saponin, QS-21, derived from Quillaja saponaria Molina. A vaccine formulation containing the recombinant protein, QS-21, and aluminum hydroxide was tested in specific-pathogen-free kittens and was shown to induce neutralizing antibodies as well as appreciable antibody responses to native gp70 by enzyme immunoassay and protein (western) immunoblot analysis and of whole virus preparations.     

An improved Corynebacterium pseudotuberculosis vaccine for sheep

Extensive experiments in mice confirmed that the immunogenicity of a Corynebacterium pseudotuberculosis vaccine could not be significantly improved with the use of various adjuvants. Immunity against C. pseudotuberculosis likewise could not be enhanced by incorporating various immunostimulants into the vaccine or by the use of live vaccines. However, a combination of aluminium hydroxide gel and saponin as adjuvant did have a beneficial effect. This vaccine was tolerated better, and a smaller dose apparently protected sheep more effectively against intralymph node challenge than the currently available alum-precipitated vaccine.     

Ginseng extract in aluminium hydroxide adjuvanted vaccines improves the antibody response of pigs to porcine parvovirus and Erysipelothrix rhusiopathiae

Ginseng, the dry extract prepared from the Panax ginseng C.A. Mayer-root contain immunomodulators named ginsenosides, which in the pig enhance the antibody response to viral and bacterial antigens. The enhancing effect of ginseng was demonstrated vaccinating pigs against porcine parvovirus (PPV) and Erysipelothrix rhusiopathiae infections, using commercially available vaccines. The potency of the licensed, aluminium hydroxide adjuvanted; vaccines were compared with those supplemented with ginseng. The antibody response to PPV was measured by the haemagglutination inhibition (HI) test whereas the mouse potency test and ELISA evaluated the immune response to E. rhusiopathiae. Antibodies to the 64-66 kDa glycoprotein of the E. rhusiopathiae were demonstrated by immunoblotting. The qualitative antibody responses were evaluated by means of ELISA(s) using monoclonal antibodies to swine IgG1 and IgG2. The addition of 2mg ginseng per vaccine dose, potentiate the antibody response of the commercial vaccines without altering their safety. Significantly higher (P<0.001) antibody titres were achieved to both PPV and to E. rhusiopathiae by the supplementation with ginseng. Aluminium hydroxide adjuvanted vaccines favoured the production of IgG1 antibodies. Interestingly, the vaccines supplemented with ginseng favoured IgG2. The vaccines used in the evaluations varied in their immunogenic potency. However, after the addition of ginseng the less immunogenic vaccine proved to be as potent as the better one without ginseng. Thus, the use of ginseng as a co-adjuvant provides a simple, safe and cheap alternative for improving the potency of aluminium hydroxide adjuvanted vaccines.     

Factors affecting the immunogenicity of Pasteurella haemolytica in mice

An appreciable level of immunity from intraperitoneal infection with Pasteurella haemolytica was established in mice by using a vaccine prepared in a conventional bacteriological culture medium, with aluminium hydroxide gel as adjuvant. The level of immunity could not be elevated by using bacteria grown in tissue culture media, enriched brain heart infusion broth, the addition of serum to the media or by using bacteria that had been harvested in the logarithmic growth phase. Although various extracts of the bacteria elicited a distinct immunity, the immunogenicity of vaccines containing bacteria could not be enhanced by augmentation with those products. The potential application of the vaccine in cattle and sheep is discussed.     

Chlamydophila abortus infection in the mouse: a useful model of the ovine disease

Chlamydophila (C.) abortus is an obligate intracellular bacterium able to colonize the placenta of several species of mammals, which may induce abortion in the last third of pregnancy. The infection affects mainly small ruminants resulting in major economic losses in farming industries worldwide. Furthermore, its zoonotic risk has been reported in pregnant farmers or abattoir workers. Mouse models have been widely used to study both the pathology of the disease and the role of immune cells in controlling infection. Moreover, this animal experimental model has been considered a useful tool to evaluate new vaccine candidates and adjuvants that could prevent abortion and reduce fetal death. Future studies using these models will provide and reveal information about the precise mechanisms in the immune response against C. abortus and will increase the knowledge about poorly understood issues such as chlamydial persistence.     

Comparison between immune responses and resistance induced in BALB/c mice vaccinated with RB51 and Rev. 1 vaccines and challenged with Brucella melitensis bv. 3

BALB/c mice were immunized with live rough Brucella abortus RB51 or smooth Brucella melitensis Rev. 1 vaccines and challenged with a B. melitensis field strain. Protection was assessed by a variety of serological tests and recovery of vaccinal and challenge strains by culture. Mice vaccinated with RB51 gave negative results in the conventional serological tests prior to challenge, namely; standard tube agglutination test (SAT), Rose Bengal plate test (RBPT), buffered acidified plate antigen test (BAPAT), and mercaptoethanol test (MET). Sero-conversion took place to a whole-cell bacterial buffered RB51 antigen after vaccination and persisted for 7 weeks post-vaccination. Mice challenged with B. melitensis were assessed for bacterial load and immune response for 12 weeks after challenge. Protection units were showed that Rev. 1 vaccine was superior to RB51 vaccine in protection of mice against B. melitensis. However, RB51 vaccine has the advantage that it would not elicit antibodies to standard serological tests based on the LPS O antigen. RB51 vaccine could therefore be used for control of B. melitensis infection and avoid confusion in the use of standard sero-diagnostic tests.     

A bovine ephemeral fever vaccine incorporating adjuvant Quil A: a comparative study using adjuvants Quil A, aluminium hydroxide gel and dextran sulphate

Various vaccines containing the 919 strain of ephemeral fever virus were evaluated in experimental calves and in commercial cattle. The vaccine virus was mixed with one of the adjuvants, Quil A (a saponin derivative), aluminium hydroxide gel, dextran sulphate or combinations of these. The response of experimental calves was evaluated by measuring the production of neutralising antibodies and by resistance to challenge with virulent virus; the response of commercial cattle was judged only by the production of neutralising antibody. Twelve calves given two doses of vaccine containing Quil A produced neutralising antibodies to bovine ephemeral fever virus and all were resistant to challenge with virulent virus given 28 to 76 days after the second vaccination. The vaccine given in three of these calves also contained aluminium hydroxide gel. Six of eight unvaccinated control calves succumbed to experimental challenge. In commercial cattle (17 to 26 animals per group) the serological response after two doses of vaccine containing Quil A or Quil A and dextran sulphate was significantly better than that after vaccines containing only dextran sulphate or after vaccines containing combinations of aluminium hydroxide gel and Quil A. The adjuvant Quil A alone was tested in cattle and shown to produce a transient soft swelling at the injection site as well as a rise in rectal temperature of greater than 1 degree C one day after inoculation. At least 99.99 per cent of viral infectivity was destroyed when the vaccine was mixed with Quil A, suggesting that live virus may not be essential in the immunogenicity of the vaccine. This vaccine overcame two of the problems associated with previous attenuated vaccines tested in Australia; the necessity for adjuvant and virus to be mixed immediately before use and the large volume of the vaccine.     

New challenges for vaccination to prevent chlamydial abortion in sheep

Ovine enzootic abortion (OEA) is caused by the obligate intracellular Gram-negative bacterium Chlamydia abortus. OEA remains a common cause of infectious abortion in many sheep-rearing countries despite the existence of commercially available vaccines that protect against the disease. There are a number of confounding factors that influence the uptake and use of these vaccines, which includes an inability to discriminate between infected and vaccinated animals (DIVA) using conventional serological diagnostic techniques. This suggests that the immunity elicited by current vaccines is similar to that observed in convalescent, immune sheep that have experienced OEA. The existence of these vaccines provides an opportunity to understand how protection against OEA is elicited and also to understand why vaccines can occasionally appear to fail, as has been reported recently for OEA. Interferon-gamma (IFN-γ), the cytokine that classically defines Th1-type adaptive immunity, is a strong correlate of protection against OEA in sheep and has been shown to inhibit the growth of C. abortus in vitro. Humoral immunity to C. abortus is observed in both vaccinated and naturally infected sheep, but antibody responses tend to be used more as diagnostic markers than targets for strategic vaccine design. A future successful DIVA vaccine against OEA should aim to elicit the immunological correlate of protection (IFN-γ) concomitantly with an antibody profile that is distinct from that of the natural infection. Such an approach requires careful selection of protective components of C. abortus combined with an effective delivery system that elicits IFN-γ-producing CD4+ve memory T cells.     

Monophosphoryl lipid A-induced immune enhancement of Brucella abortus salt-extractable protein and lipopolysaccharide vaccines in BALB/c mice.

A study was conducted to determine the effect of monophosphoryl lipid A (MPL) and trehalose dimycolate (TDM) as adjuvants on the protective responses in BALB/c mice vaccinated with Brucella abortus salt-extractable protein (BCSP) or proteinase-K-treated B abortus lipopolysaccharide (PKLPS). Mice were vaccinated with different doses of BCSP or PKLPS given alone or in combination with MPL or TDM. Mice were challenge-exposed 4 weeks later with virulent B abortus strain 2308. Two weeks after challenge exposure, the number of B abortus colony-forming units (CFU) per spleen, spleen weights, and spleen cell interleukin 1 production were measured. Serum IgG and IgM concentrations specific for vaccinal immunogens were measured before and after challenge exposure with B abortus. Spleen weights and mean B abortus CFU per vaccine group were significantly lower in BCSP- and PKLPS-vaccinated mice, compared with those of nonvaccinated control mice. Monophosphoryl lipid A enhanced the suppression of splenic infection when given with the BCSP vaccine, but not when given with the PKLPS vaccine. Trehalose dimycolate had no effect on mean CFU when given with BCSP, but incorporation of TDM resulted in a significant increase in mean CFU when given with PKLPS. Spleen weights in BCSP- or PKLPS-vaccinated mice were not different when these vaccines were combined with MPL or TDM. Because of the wide variation in the results, we could not conclude that vaccination with BCSP or PKLPS alone, or in combination with MPL altered spleen cell interleukin-1 production in B abortus-infected mice.(ABSTRACT TRUNCATED AT 250 WORDS)     

I型鸭疫里默氏杆菌荚膜免疫的研究

用盐浸法提取了I型鸭疫里默氏杆菌（RA）的荚膜,并制成油佐剂疫苗。其免疫原性测定结果表明：用荚膜粗提物免疫雏鸭可得到很好的免疫保护效果（9／10）。同时进行了RA荚膜油佐剂疫苗与各常规佐剂疫苗的免疫效力比较,免疫保护效果依次为：英膜油乳剂苗、油乳剂灭活苗、蜂胶灭活苗、铝胶RA灭活苗、无佐剂RA灭活苗。结果显示,鸭疫里默氏杆菌英膜免疫的研究为进一步研制亚单位疫苗打下基础。     

A Salmonella abortusovis inactivated vaccine protects mice from abortion after challenge.

Two types of Salmonella abortusovis vaccines were prepared, one with aluminium hydroxide (vaccine A) and the other with water in oil (vaccine B) adjuvants. They were compared in a pregnant mouse model, aiming at protecting them from abortions after challenge with a virulent strain of S. abortusovis. The protection for vaccine A was from 74% to 77.6% and that for vaccine B from 71% to 79.6%. Abortions occurred 5-10 days post challenge and S. abortusovis was isolated from all aborted fetuses and from the liver and the spleen of their mothers at the end of the experiment (18 days post challenge). The presence of salmonella in the liver and the spleen of vaccinated non-pregnant but challenged mice was studied in a separate experiment. The bacterium was isolated from one out of 12 vaccinated mice 6 days post challenge as well as from the six controls.     

类M蛋白亚单位疫苗的佐剂筛选

将铝胶、ISA206和CpG三种佐剂分别与热盐酸提取的马链球菌兽疫亚种(Streptococcus equi subsp. zooepidemicus)ATCC35246株类M蛋白配合,制成疫苗接种20日龄的小鼠,并设不加佐剂的类M蛋白亚单位疫苗免疫对照组.二次免疫后用ATCC35246株活菌攻击,铝胶、ISA206、CpG和对照组的保护率分别为75%(12/16)、81.25%(13/16)、68.75%(11/16)和56.25%(9/16).试验结果表明ISA206的免疫增强作用高于其他两种佐剂,可望与类M蛋白亚单位疫苗配合,用于预防猪链球菌病.     

Influence of the Th2 immune response established by Nippostrongylus brasiliensis infection on the protection offered by different vaccines against Chlamydophila abortus infection

Chlamydophila abortus is the aetiological agent of enzootic abortion in small ruminants in which it infects the placenta to cause abortion during the last trimester of gestation. In a mouse model, a Th1 immune response involving IFN-gamma production and CD8+ T cells is necessary for the infection to be resolved. The authors previously demonstrated that infection with Nippostrongylus brasiliensis, a rodent gastrointestinal nematode extensively used in experimental models to induce Th2 responses, alters the specific immune response against C. abortus infection, increasing bacterial multiplication in liver and reducing specific IFN-gamma production. The aim of the present work was to clarify whether a Th2 immune response has any influence on the success of vaccination using both inactivated and attenuated vaccines. The results showed that the Th2 response established prior to vaccination did not influence the induction of protection offered by the vaccines. However, the effectiveness of this protective response can be altered, depending on the adjuvant employed in the inactivated vaccines, when the Th2 response is established after vaccination, just before challenge with C. abortus.     

Protective activity to murine experimental brucellosis conferred by monoclonal antibody ISS/32 anti-B. abortus.

The protective properties of the monoclonal antibody ISS/32 anti-B. abortus were estimated by splenic infection with B. abortus 544. Five groups of Balb/c mice were used: two groups, previously vaccinated with a 45/20 antigen and a-LPS antigen, were challenged after 30 days intravenously by inoculation of 2.10(5) cells of B. abortus 544, one group was challenged with the same dose of B. abortus 544 preincubated with MAb-ISS/32 and another one with B. abortus 544 incubated with negative serum; the fifth group infected with B. abortus 544 only served as control. The results, expressed as an index of splenic infection, show significant protective properties of monoclonal antibody ISS/32. The infection index in the MAb-ISS/32 group of mice was a bit lower than in B. abortus 45/20 vaccine group.     

Protective adaptive immunity to Chlamydophila abortus infection and control of ovine enzootic abortion (OEA)

Ovine enzootic abortion (OEA) remains a major problem in sheep-rearing countries despite the availability of protective vaccines. The causative agent, Chlamydophila abortus, is a Gram-negative bacterium that can induce a persistent, subclinical infection in non-pregnant sheep. The development of a new safe, effective and practical vaccine requires a detailed understanding of host-pathogen interactions and the identification of clear correlates of protection. Since disease (abortion) is only observed during pregnancy, the nature of host immunity to C. abortus and the specialised immunological features that permit maternal acceptance of the semi-allogeneic fetus are central to the pathogenesis of OEA. We review the current literature on persistence of C. abortus, host immunity to infection and mechanisms of abortion. We identify the key outstanding questions surrounding OEA and discuss the current knowledge gaps with a view to developing improved control strategies.     

Diagnostic characterization of a feral swine herd enzootically infected with Brucella.

Eighty feral swine were trapped from a herd that had been documented to be seropositive for Brucella and which had been used for Brucella abortus RB51 vaccine trials on a 7,100-hectare tract of land in South Carolina. The animals were euthanized and complete necropsies were performed. Samples were taken for histopathology, Brucella culture, and Brucella serology. Brucella was cultured from 62 (77.5%) animals. Brucella suis was isolated from 55 animals (68.8%), and all isolates were biovar 1. Brucella abortus was isolated from 28 animals (35.0%), and isolates included field strain biovar 1 (21 animals; 26.3%), vaccine strain Brucella abortus S19 (8 animals, 10.0%), and vaccine strain Brucella abortus RB51 (6 animals, 7.5%). Males were significantly more likely to be culture positive than females (92.9% vs. 60.6%). Thirty-nine animals (48.8%) were seropositive. Males also had a significantly higher seropositivity rate than females (61.9% vs. 34.2%). The relative sensitivity rates were significantly higher for the standard tube test (44.6%) and fluorescence polarization assay (42.6%) than the card agglutination test (13.1%). Lesions consistent with Brucella infection were commonly found in the animals surveyed and included inflammatory lesions of the lymph nodes, liver, kidney, and male reproductive organs, which ranged from lymphoplasmacytic to pyogranulomatous with necrosis. This is the first report of an apparent enzootic Brucella abortus infection in a feral swine herd suggesting that feral swine may serve as a reservoir of infection for Brucella abortus as well as Brucella suis for domestic livestock.