Killing of Brucella abortus by bovine serum (original) (raw)
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Veterinary Microbiology, 1991
Belzer, C.A., Tabatabai, L.B. and Deyoe, B.L., 1991. Differentiation by western blotting of immune responses of cattle vaccinated with Brucella abortus strain 19 or infected experimentally or naturally with virulent Brucella abortus. Vet. Microbiol., 27: 79-90. Brucella abortus strain 19 salt-extractable proteins fractionated by differential ammonium sulfate precipitation were used in a western blotting method to detect bovine immunoglobulin G antibodies to B. abortus. Sera from infected cattle and from cattle vaccinated with strain 19 and subsequently exposed to virulent B. abortus bound to a common group of antigens ranging in molecular weights from 31 000 to 45 000 daltons. Immunoglobulin G antibodies in sera from the latter group in addition also bound to antigens with molecular weights of 66 000 to 71 000 daltons. Some sera from cattle vaccinated when sexually mature reacted similar to those from infected cattle, while immunoglobulin G antibodies in sera from Brucella-free cattle and vaccinated calves did not bind to either group of antigens. In general, fractionation of the proteins by ammonium sulfate precipitation offered no advantage for detecting differences between groups of sera. Ammonium sulfate fraction 0 to 35% reacted with a larger number of sera from a naturally infected group than fraction 0 to 70%. Both fractions reacted equally well with sera from the other groups of cattle, while fractions 35 to 70% and 70 to 100% reacted poorly in this technique. The attractive feature of the blot is that sera from calfhoodvaccinated cattle did not react. INTRODUCTION Standard serologic tests for diagnosis of bovine brucellosis have been in use since 1940 (Manthei et al., 1956), but the most difficult task has been in distinguishing antibodies of infected from those of vaccinated animals. To date, most diagnostic tests for Brucella abortus rely on detecting a humoral immune response, although the most definitive diagnostic test is bacterial ~To whom correspondence should be addressed.
Journal of Veterinary Medicine, Series B, 2008
The comparison of serological responses in a sample of adult, vaccinated and ®eld-infected bovines with Brucella abortus is reported. Indirect enzyme immunoassay (EIA) titration curves and Western blotting tests for smooth-type lipopolysaccharide (S-LPS), rough-type LPS (R-LPS) and lipid A were performed. In the initial screening of sera, an overall prevalence of 20.5 % was found, which corresponds to a country with a high incidence of brucellosis. End-point EIA titres against LPS antigens from vaccinated and ®eld-infected cows were not signi®cantly different. However, the absorbance values in the titration curves were signi®cantly higher for S-LPS as compared with the other antigens. A high correlation coef®cient (r 0.933) was obtained when the titres to R-LPS versus lipid A were compared. Western blotting reactions of vaccinated and ®eld-infected animals were indistinguishable. S-LPS, R-LPS and lipid A epitopes were recognized in a heterogeneous manner. In general, the number of bovines that reacted against LPS was higher in the ®eld-infected group, with a stronger binding to S-LPS. Based on our observations, the vaccinated and ®eldinfected bovines are capable of producing similar antibody responses to the Brucella main outer surface antigen, LPS. It should be emphasized that the humoral response of cattle to Brucella LPS contains signi®cant amounts of antibodies to other antigenic moieties of this important surface molecule, which may contribute to the immunity to brucellosis.
PubMed, 1989
Purified O chain of Brucella abortus was passively attached to polystyrene to differentiate antibody responses of cattle vaccinated with B abortus strain 19 from those of naturally infected cattle. In the indirect assay, using O polysaccharide as antigen, a single serum dilution was used and mouse monoclonal antibody to bovine L chain conjugated with horseradish peroxidase was the detection reagent. Measurable antibody was not found in sera of vaccinated cattle, except for 3 sera from cattle that were persistently infected with strain 19. Sera from 25 cattle infected with pathogenic strains contained antibody on the basis of results of indirect enzyme immunoassay, using smooth lipopolysaccharide or O chain as antigens, or results of competitive enzyme immunoassay, using the O-chain antigen. Results in sera from calves with experimentally induced Yersinia enterocolitica serotype 0:9 infection or inoculated with a low dose of B abortus strain 2308 were comparable with those in sera of cattle that were vaccinated with strain 19. The data correlated with those from competitive enzyme immunoassay, using one serum dilution and horseradish peroxidase-conjugated mouse monoclonal antibody to smooth lipopolysaccharide. On the basis of results of the indirect enzyme immunoassay, all sera (except those samples obtained before inoculation) contained antibody to smooth lipopolysaccharide.
Veterinary World, 2021
Background and Aim: The present serodiagnosis of brucellosis in livestock is based on the whole cell or smooth lipopolysaccharide of the Brucella organism in which specificity is hampered by the cross-reactivity, especially with the antibodies against Yersinia enterocolitica O:9 organism. The problem can be addressed by screening for better immunodominant antigens. Hence, the present study was undertaken to screen protein antigens of Brucella abortus for their diagnostic potential in cattle brucellosis. Materials and Methods: Protein antigens of B. abortus (n=10) non-reactive to antibodies against Y. enterocolitica O:9 were selected, expressed in Escherichia coli, assessed the reactivity of expressed recombinant proteins by Western blot, standardized indirect-enzyme-linked immunosorbent assay (ELISA) for detecting Brucella antibodies in cattle serum, and comparative evaluation was done. Results: All the selected protein antigens were expressed and in the Western blot with Brucella antibodies positive cattle serum, six recombinant (Brucella protein 26 [BP26], Cu-Zn Superoxide dismutase [SodC], B. abortus I-1885, Serine protease, Bacterioferritin, and Brucella Lumazine Synthase [BLS]) proteins showed reaction whereas none of the proteins showed reactivity with Brucella negative cattle serum. ELISA has been done using known Brucella positive and negative cattle sera samples (n=113 each) in which the performance of recombinant proteins in diagnosing brucellosis was in the order of BP26 > BLS > SodC followed by rest of the proteins. BP26 based ELISA was found to be better with area under the curve as 0.953, and diagnostic sensitivity, diagnostic specificity, and Youden's index of 90.27%, 95.58%, and 0.8584, respectively, with the excellent agreement (k=0.85). Conclusion: BP26 could be a potential diagnostic antigen among the immunodominant proteins of B. abortus in ruling out Y. enterocolitica O:9 infection while diagnosing brucellosis in cattle herds.
Infection and immunity, 1992
Cellular immune responses are influential for protection against intracellular bacteria such as brucellae. Therefore, identification of Brucella abortus antigens that activate primed bovine lymphocytes is fundamental for discerning the breadth of cellular response in bovine brucellosis. Potentially antigenic components of B. abortus S19 were isolated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by nitrocellulose blotting. Specific one-dimensional blot segments induced proliferation of peripheral blood lymphocytes from all 25 of the vaccinated cattle tested and were defined as immunodominant. Individual proteins that stimulated lymphocyte proliferation were further characterized by two-dimensional cellular immunoblotting by two different approaches. Individual one-dimensional stimulatory blot segments were eluted, concentrated, and then subjected to two-dimensional cellular immunoblotting. Alternatively, entire two-dimensional gels containing all of the B. ab...
Identification of immunotolerance in the progeny of cows infected with Brucella abortus
African Journal of Microbiology Research, 2012
The progeny of cows infected with Brucella abortus could acquire the bacterium during their fetal stage and generate immune tolerance. In order to identify them, a clinical assay was conducted with two groups of seronegative calves. In Group I, seven bovine females received a standard S19 dose, and in Group II, eight males served as control. All animals were sampled seven times by official serological tests every 45 days. Also, complete blood with anticoagulant was collected and tested by the polymerase chain reaction (PCR) 45 and 135 days after vaccination. Results were analyzed by the test of proportions. In Group I, all the animals had their maximum antibodies title at the first sampling, but from the fourth sampling, their titles decreased until they became undetectable for the screening test. All animals in Group II remained negatives. Animals from both groups recorded negative with PCR. Significant differences (P<0.05) between groups were observed for seroconversion in the first two samplings. It was concluded that none of the tested animals were immunotolerant and that PCR may not be an appropriate method to demonstrate immunotolerence in some individuals.
Modulation of the Bovine Trophoblastic Innate Immune Response by Brucella abortus
Infection and Immunity, 2008
Brucella abortus and trophoblastic cells, which is essential for better understanding the pathogenesis of the Brucella-induced placentitis and abortion, a key event for transmission of the disease. The goal of this study was to evaluate the profile of gene expression by bovine trophoblastic cells during infection with B. abortus. Explants of chorioallantoic membranes were inoculated with B. abortus strain 2308. Microarray analysis was performed at 4 h after infection, and expression of cytokines and chemokines by trophoblastic cells was assessed by real-time reverse transcription-PCR at 6 and 12 h after inoculation. In addition, cytokine and chemokine expression in placentomes from experimentally infected cows was evaluated.
Characterization of Brucella abortus Soluble Antigen Employed in Immunoassay
Journal of Clinical Microbiology, 1980
A soluble antigen extract of Brucella abortus (BASA) has been prepared by the National Veterinary Services Laboratories and furnished to a number of workers who are examining antibody-mediated and cell-mediated immune responses of cattle infected with B. abortus. Three lots of BASA were examined. There were quantitative but not qualitative differences among lots by content of protein, total carbohydrate, hexose, fatty acid, and 2-keto-3-deoxyoctonic acid. The presence of smooth lipopolysaccharide was demonstrated by the presence of 2-keto-3-deoxyoctonic acid and lipid, by Limulus lysate gelation activity, and by formation of characteristic lipopolysaccharide precipitates in immunoelectrophoresis. A polysaccharide antigen as well as two nonsurface antigens, A2 and C, were also identified. BASA is a satisfactory antigen for use in the enzyme-linked immunosorbent assay since the smooth lipopolysaccharide component bound to polystyrene and functioned in the test. Normal murine spleen ce...