Immune responses of cattle to biochemically modified antigens from the midgut of the cattle tick, Boophilus microplus

Treatment of membrane antigens of the midgut (GM) of the cattle tick, Boophilus microplus with sodium metaperiodate (periodate), pronase and lipase significantly inhibited the reactivity of the GM with antibodies in the sera of 57 cattle vaccinated with GM. Treatment of GM with periodate only removed the correlation between antibody reactivity of sera and protection against infestation with ticks. A monoclonal antibody (MoAb QU13), which recognises protective antigens solubilized from GM (Lee + Opdebeeck 1991), did not react with GM treated with periodate. Cattle vaccinated with GM extracts were significantly protected against infestation with cattle ticks (P less than 0.05), whereas cattle vaccinated with either GM extracts treated with periodate or with antigens precipitated from GM extracts with MoAb QU13 and also treated with periodate, were not protected against infestation. These studies provide preliminary evidence that protective antigens in the tick midgut membrane either are carbohydrate or are dependent on carbohydrate for their specificity.     

Larval membrane antigens protect Hereford cattle against infestation with Boophilus microplus

Hereford cattle (Bos taurus) were immunized with antigens solubilized with Triton X-100 from larval membranes of the cattle tick (Boophilus microplus). Based on tick egg production compared to control cattle, vaccinated cattle were protected (78%) against challenge with 2 x 20,000 tick larvae. The soluble Triton X-100 extract of tick larval membranes was further purified by immunoaffinity chromatography, using immunoglobulin ligands (IgG1 and IgG2) from three immune steers, previously vaccinated with membrane antigens from the midgut of partly engorged adult female ticks. Cattle vaccinated with these purified antigens were protected in two separate experiments (80 and 89% respectively), against challenge with 2 x 20,000 larval ticks compared to control cattle. Whole larval membranes used as vaccines in cattle reduced the amount of eggs produced from ticks by 47% compared to control cattle, but this difference was not significant.     

Hereford cattle immunized and protected against Boophilus microplus with soluble and membrane-associated antigens from the midgut of ticks

Hereford cattle were immunized with membranes and soluble components extracted from the midgut of Boophilus microplus. Membrane vaccines protected cattle (91%) against challenge with 3 x 20,000 larval ticks administered at intervals of 7 days. Vaccines made from soluble antigens did not protect cattle. Antibody levels measured by enzyme-linked immunosorbent assay (ELISA) related to the levels of protection induced by vaccination.     

Effect of salivary gland extracts from the tick,Boophilus microplus,on leucocytes from Brahman and Hereford cattle

The effect of salivary gland extract (SGE) from Boophilus microplus on peripheral blood lymphocytes, neutrophils and monocytes from Brahman (Bos indicus) and Hereford (Bos taurus) cattle was investigated. SGE (8 micro g) significantly inhibited the proliferation response of lymphocytes to concanavalin A from both Brahman and Hereford cattle by 89% and 41%, respectively. The difference in inhibition between the two breeds was highly significant (P < 0.01), whilst at 1 micro g of SGE, significant inhibition of lymphocytes occurred only in Hereford cattle (34%). Flow cytometric analysis of monocytes and neutrophils showed that SGE (40 micro g) significantly reduced both the proportion of cells actively phagocytosing Escherichia coli labelled with fluorescein isothiocyanate (E. coli-FITC) and the uptake of E. coli-FITC in Brahman cattle. However, in Hereford cattle, a significant depression in uptake was only observed in neutrophils. The proportion of monocytes and neutrophils with oxidative activity was significantly suppressed in the presence of SGE in both breeds of cattle. These results indicate that peripheral blood leucocytes from different breeds of cattle respond differently to SGE.     

Vaccination of cattle against the Boophilus microplus using a mucin-like membrane glycoprotein

An antigen, BMA7, which induced partial immunity against tick infestation has been isolated from Boophilus microplus using two different protein fractionation protocols, accompanied by vaccination and parasite challenge trials. The antigen is a 63 kDa glycoprotein isolated from semi-engorged adult female ticks. Though significant, the induced immunity is less striking than that previously reported for antigen Bm86 from the same parasite. However, co-vaccination with Bm86 and BMA7 can enhance immunity over that seen with a commercial vaccine based on Bm86 alone. Limited peptide sequence information shows significant variation in the BMA7 protein occurs. The antigen has approximately 36 kDa of glycosylation, in both N-linked and O-linked oligosaccharides. There is evidence that both polypeptide and oligosaccharide are antigenic, but the chemical nature of the protective antigenic sites is not clear. There is little or no immunological response to the antigen during natural infestation with parasites, suggesting the antigen is 'concealed' and protective immunity dependent on artificial vaccination. The antigen has some similarities with the vertebrate mucins. It is widely distributed in tick tissues and membrane bound but its function is currently unknown .     

Comparative vaccination of cattle against Boophilus microplus with recombinant antigen Bm86 alone or in combination with recombinant Bm91

Cattle were vaccinated either with a single recombinant tick antigen, Bm86 or with a combination of two recombinant antigens, Bm86 and Bm91 from the tick Boophilus microplus . In three experiments, the responses of cattle to subsequent challenge with the tick were assessed. The addition of the Bm91 antigen enhanced the efficacy of the vaccination over that with Bm86 alone to a statistically significant degree. Moreover, co-vaccination with two antigens did not impair the response of cattle to the Bm86 antigen. Finally, responses of individual cattle to the two antigens were independent. All of these results may be relevant to the increase in efficacy expected from a dual antigen vaccine.     

Detection of Bm86 antigen in different strains of Boophilus microplus and effectiveness of immunization with recombinant Bm86

The control of tick populations by using conventional strategies poses several problems, including the appearance of organophosphate resistant strains, among others. The possibility of using alternative strategies such as vaccination with tick antigens has been suggested by several authors. One particular antigen (Bm86) has been described and shown to be able to induce a protective immunity against the cattle tick Boophilus microplus. In this paper we demonstrate by means of immunohistochemical staining that this antigen is conserved among several strains of this species. These results correlate with those showing that animals vaccinated with a preparation of recombinant Bm86 were protected against challenge with the four different strains tested, including one resistant to organophosphates. These results favour the immunization with recombinant Bm86 for the control of the cattle tick B. microplus.     

Peripheral cellular and humoral responses to infestation with the cattle tick Rhipicephalus microplus in Santa Gertrudis cattle

Resistance to cattle tick infestation in single‐host ticks is primarily manifested against the larval stage and results in the immature tick failing to attach successfully and obtain a meal. This study was conducted to identify immune responses that characterize the tick‐resistant phenotype in cattle. Thirty‐five tick‐naïve Santa Gertrudis heifers were used in this study, thirty of which were artificially infested for thirteen weeks with tick larvae while five animals remained at a tick‐free quarantine property to serve as a control group. Following thirteen weeks of tick infestation, the animals in this trial exhibited highly divergent tick‐resistant phenotypes. Blood samples collected throughout the trial were used to measure peripheral immune parameters: haematology, the percentage of cellular subsets comprising the peripheral blood mononuclear cell (PBMC) population, tick‐specific IgG₁ and IgG₂ antibody titres, IgG1 avidity for tick antigens and the ability of PBMC to recognize and proliferate in response to stimulation with tick antigens in vitro. The tick‐susceptible cattle developed significantly higher tick‐specific IgG₁ antibody titres compared to the tick‐resistant animals. These results suggest that the heightened antibody response either does not play a role in resistance or might contribute to increased susceptibility to infestation.     

Antibody responses to invariant antigens of Trypanosoma congolense in cattle of differing susceptibility to trypanosomiasis

Five trypanotolerant N'Dama (Bos taurus) and five susceptible Boran (Bos indicus) cattle were challenged by tsetse flies infected with Trypanosoma congolense IL 13-E3. These animals had experienced five previous infections with T. congolense, each terminated by drug therapy. Immunoblotting and ELISA were used to determine isotype and specificity of antibody responses to trypanosome invariant antigens. Both IgM and IgG1 were elicited, but the IgG1 responses were directed against a greater diversity of antigens. A 69 kD antigen was the major invariant antigen which elicited IgM antibodies in both breeds, but the N'Damas also responded with high levels of specific IgG1. Analysis of isotypic responses to whole trypanosome extract also revealed lower levels of IgG1 and higher levels of IgM in the Borans than in the N'Damas, suggesting that a dysfunction in the switch from IgM to IgG might occur in infected Boran cattle. A 33 kD antigen appeared to elicit only IgG1. Sera from all five N'Damas and the two Borans which were most resistant to the disease reacted with this antigen prior to and following re-infection. Furthermore, during the primary T. congolense infection in the same animals, anti-33 kD antibodies were detectable in all five trypanotolerant N'Damas, but in none of the five susceptible Borans. Thus, the presence of antibodies to the 33 kD antigen ofT. congolense appeared to be associated with a capacity to control the disease.     

Heterogeneity of IgG antibody responses to cloned Onchocerca volvulus antigens in microfiladermia positive individuals from Esmeraldas Province, Ecuador

The prevalence of IgG antibodies to three recombinant O. volvulus antigens, OvMBP/10, OvMBP/11 and OvMBP/29 was determined in a group of 94 microfilaria positive (mf+) individuals resident in the hyperendemic onchocercal area of Esmeraldas Province, Ecuador. Clone OvMBP/11 was the antigen most frequently recognized by patients sera followed by OvMBP/10 and OvMBP/29. When a cocktail of the three recombinant antigens was used the proportion of positive sera increased to 100%. Antibody responses to the fusion partner maltose binding protein (MBP) were low in comparison with those to the cloned antigens and no correlation of responses between individual antigens was observed. The relative level of antibody response to each of the clones in the cocktail varied between individuals. The distribution of IgG responses to OvMBP/11 was bimodal and those to OvMBP/29 and OvMBP/10 were positively and negatively skewed, respectively. When the three recombinant antigens were used in combination this variation was minimized and the pattern of responses showed a normal distribution as was also seen to crude O. volvulus antigen. The cocktail of recombinants thus offers excellent diagnostic sensitivity in combination with the parasite specificity demonstrated previously.     

Isolation and characterization of salivary antigens from the female tick, Dermacentor andersoni

The salivary glands of ixodid ticks are complex organs which are known to contain the antigens responsible for tick resistance in animals. We have identified a large number of proteins from salivary gland extracts (SGE), at least some of which are immunologically recognized by tick resistant animals and which are therefore presumed to be secreted salivary components. During the 6 to 10 day feeding process, a number of these antigens alter in concentration according to individual kinetics, and some of these changes correlate with the kinetics of skin test reactivity of SGE obtained at different times throughout the feeding period. By use of immunoaffinity chromatography we have isolated large quantities of many of the salivary antigens (SGA) contained in SGE, and found that they contain several esterase activities. SGA stimulates both immediate and delayed skin reactions in tick resistant guinea-pigs, and these reactions are about 200-fold more intense, per unit protein, than those elicited by SGE. The skin reactions to SGA are basophil-mediated and have many features in common with the cutaneous basophil hypersensitivity reactions of tick resistant animals. The demonstrated antigenic complexity of the glands may have profound implications for attempts to develop anti-tick vaccines, as it may eventually be found that candidate vaccines will have to incorporate more than one tick antigen in order to be effective.     

Temporal changes in the humoral immune response of cattle during experimental infections with Onchocerca lienalis

In order to gain insights into the immune response in onchocerciasis during early infection, laboratory-reared calves were infected with 1000 Onchocerca lienalis infective larvae and examined serologically over a period of 508 days. Levels of serum antibodies measured by ELISA against adult worm extract revealed a multiphasic response, characterized by a broadly similar profile of peaks in individual animals arising at 15–30, 79 and >266 days after infection. Timings of these changes in responsiveness closely mirrored parasite development, coinciding with larval moults and with the onset of a patent infection. The levels of individual antibody isotypes directed against parasite antigens was strongly skewed. The dominant response was of IgG1, although limited reactivities were also found for IgG2 and IgM: No parasite-specific IgA antibodies were detected. Immuno-blots of adult worms extracts revealed a pattern of antigen recognition over time that matched the results obtained by ELISA. Again, the IgGl response was strongest, although certain lgG2 and IgM specificities were well represented. In general, there was a steady increase in the number of individual antigens recognized as the infection progressed, with a striking expansion of antibody specificities from day 79 following the fourth larval moult. Antibodies to a 16kDa component were a prominent feature of the response following development of a patent infection. These data reveal the strong influence of parasite biology on the development of the immune response in onchocerciasis.     

Genetic control of immune response to a purified Schistosoma mansoni antigen. I. Effect of MHC class II antigens on the cellular, humoral and protective responses

The influence of the I region of the major histocompatibility complex (MHC) on T-dependent immune responses against a purified schistosome antigen (9B antigen) was investigated. H-2 congenic mice expressing both I-A and I-E antigens (I-E+) showed a higher in-vitro proliferation to 9B antigen as compared to the recombinant strains expressing only I-A (I-E-). These two strains of mice differed both qualitatively and quantitatively in the humoral responses elicited by the purified antigen. Furthermore, in-vivo protection experiments showed that mice which do not express I-E molecules can be partially protected against the disease by prior immunization with the 9B antigen in contrast to their I-E expressing counterparts. The possible role of the I-E molecule in the immune responses elicited during Schistosoma mansoni infection is discussed.     

Fasciola hepatica in the rat: immune responses associated with the development of resistance to infection

F. hepatica infections were established in rats and immune responses were monitored during primary and challenge infections. Antibody levels peaked at 3 weeks post-primary infection and at 6 days post-challenge infection. No significant correlation was found between antibody titre and number of flukes recovered at autopsy. Immunoblotting revealed a limited number of immunogenic polypeptides. When antibodies from these reactive bands were eluted and tested by IFA they all gave identical binding patterns: on juvenile fluke sections tegumental syncytium, tegumental cells and gut cells were labelled, while on adult sections the same antibodies labelled gut cells, reproductive tissue, excretory ducts and flame cells. This suggested that these tissues shared a common epitope or range of epitopes. A pronounced eosinophilia was observed throughout the infection period studied and infected liver sections showed massive cellular infiltration. Histochemical and immunocytochemical investigation of infected liver revealed the presence of large numbers of eosinophils, neutrophils, lymphocytes and phagocytes. The implications of these findings, to an understanding of concomitant immunity in the rat are discussed.     

Passive protection against Taenia saginata infection in cattle by a mouse monoclonal antibody reactive with the surface of the invasive oncosphere

The surface, excretory/secretory and intracellular compartments of Taenia saginata oncospheres were analysed by a combination of immunochemical techniques and by the use of selected hybridoma antibodies. The surface proteins of the oncospheres were directly iodinated by the lactoperoxidase technique and the intracellular and excretory/secretory components were labelled biosynthetically by culture in vitro with 35S-methionine. Analysis of radiolabelled proteins by SDS-PAGE revealed a restricted number of components in all of these three compartments. Mouse derived monoclonal antibodies directed against the oncospheral stage of this parasite demonstrated both stage specific and common determinants on the surfaces of the oncospheres and the metacestodes. One monoclonal (IgM) antibody, reactive with the oncosphere surface, which had a half life of 4.1 days when injected into calves, conferred protection against oral infection with T. saginata eggs. A monoclonal antibody reactive with a major secreted component did not confer passive protection.     

Protective immune response to Plasmodium chabaudi, developed by mice after drug controlled infection or vaccination with parasite extracts: analysis of stage specific antigens from the asexual blood cycle

Summary The protective immune response to asexual blood infection by Plasmodium chabaudi was studied in mice immunized either by drug controlled infection or by vaccination with preparations of merozoïtes or free parasites at different stages of development. Animals immunized by the first method developed a sterile immunity. The passive transfer of their serum protected naive recipients from the lethal development of the infection, but affected only moderately the initial course of the parastiaemia. Animals immunized with either ring, schizont or merozoi'te preparations exhibited a limited but significant resistance to infection: when challenged with 10⁶ parasites of the homologous strain they exhibited a reduced parasitaemia as compared to control mice, and in addition, 50% of them recovered from the infection. Immunochemical analysis of parasite antigens showed that a family of high molecular weight proteins synthesized essentially at the schizont stage and conserved in the merozoites are important immunogens. Quantitative rather than qualitative differences were observed in the pattern of parasite proteins immunoprecipitated by serum of animals exhibiting sterile immunity or moderate protective immunity. A schizont specific polypeptide of mol. wt 82 Kd which is found in the surface of the merozoite is preferentially immunoprecipited by serum from animals exhibiting sterile immunity.