B.V. Goodger

An enzyme linked immunosorbent assay to diagnose Babesia bovis infection in cattle.

The optimum gel filtration fraction from lysate of Babesia bovis infected erythrocytes was determined for use as an antigen in an ELISA to diagnose B. bovis infection in cattle. Of four enzyme labels tested, horseradish peroxidase was the most suitable. The assay is both sensitive and specific in detecting antibody for 2–4 years after a single infection. False positive reactions were obtained only with some sera from some Anaplasma marginale infected cattle.

The immune response of cattle to Babesia bovis (syn. B. argentina). Studies on the nature and specificity of protection

Abstract Studies of the immune response to Babesia bovis (syn. B. argentina ) in Bos taurus cattle, using the passive transfer of serum from immune animals, indicated that an effector mechanism was mediated by antibodies which reacted with the parasitized erythrocytes. During removal from the peripheral blood, the parasites did not show reduced viability on subinoculation into other non-infected animals, and thus were not dead or irreversibly damaged at this time. It was concluded that opsonization of infected erythrocytes was probably the basis of protection by the system. There was some evidence that minor variation of the protective antigen(s) occurred within strains of the parasite but this had little effect on the efficiency of the hosts immune response. However, there was no cross-protection between the antibodies against different strains. These interstrain differences in antibody specificity were reconciled with earlier observations that cross-immunity commonly occurs between different strains in infected animals. It was concluded that the mechanism of cross-immunity relied on priming of the hosts immune system by the protective antigen(s) of the strain so that a secondary response against the heterologous strain occurred soon after challenge.

Bovine babesiosis: The immunization of cattle with fractions of erythrocytes infected with Babesia bovis (syn B. argentina)

Soluble antigen which protected susceptible cattle against challenge with Babesia bovis was extracted from B. bovis-infected erythrocytes by sonic disintegration and separation of the soluble from the insoluble matter by ultracentrifugation. The material was then fractionated by the precipitation of fibrinogen-like proteins. The precipitate contained the babesial antigens that were located on the stroma of the infected erythrocytes. Antigen originally located on the parasite remained in solution. Both fractions conferred protection on splenectomized calves against challenge with B. bovis. However, the fraction containing the parasite antigens appeared to have more potential for development as a killed vaccine because it was not heavily contaminated with antigenic material from bovine erythrocytes.

Babesia argentina: Intraerythrocytic location of babesial antigen extracted from parasite suspensions

Abstract Goodger B.V. 1973. Babesia argentina : Intraerythrocytic location of babesial antigen extracted from parasite suspensions. International Journal for Parasitology 3 :387–391. A crude soluble antigen obtained from a mixture of Babesia argentina parasites and infected erythrocyte stromata has been partly purified and characterized by immunological procedures and its intraerythrocytic site demonstrated by fluorescent antibody techniques. This product contained at least two distinct antigens which were species specific. One had an electrophoretic mobility. similar to serum prealbumin, had little HA activity, and was found in or on the internal rim of the erythrocytic membrane. The other had an electrophoretic mobility similar to serum β 1 globulins, was highly active in HA tests, and was found in granules on the internal stromata of the infected erythrocytes. The evidence suggests that both antigens are produced either from the parasite and are associated closely with erythrocytic components or are produced by digestion of erythrocytic components and represent metabolites of the parasite. In either case the antigenic compounds detected are specific to B. argentina .

Cloning and characterisation of cDNA clones encoding two Babesia bovis proteins with homologous amino- and carboxy-terminal domains

A dextran sulphate protein (DSP) fraction derived from Babesia bovis has previously been shown to induce a protective immune response in cattle. A B. bovis cDNA library was screened with both the complete anti-DSP serum and a subfraction of the anti-DSP serum affinity purified on a native B. bovis protein of approx. 80 kDa. cDNA clones encoding two different B. bovis proteins were identified. The product of one gene, Bv80, has a single divergent copy of a sequence of 149 amino acids (approx. 30% amino acid identity) in both the amino- and carboxy-terminal domains. These domains are separated by an array of short variant repeat sequences rich in proline and glutamic acid. The product of the other gene, BvVAl (homologous to the previously described 225-kDa B. bovis protein)[19], is predicted to have a single divergent copy of a sequence of 170-171 amino acids (approx. 35% amino acid identity) in both the amino- and carboxy-terminal domains. These domains are also separated by an array of repeats. The 73-amino acid repeat unit of this array is composed of a number of variant derivatives of shorter repeat units. Detailed analysis of genomic clones flanking two alleles of the gene encoding BvVAl/225 kDa identified further members of a multi-gene family. This region of the genome of B. bovis has been subject to a large number of amplification processes.

The irradiation ofBabesia bovis

Babesia bovis parasites attenuated by 35 krads γ irradiation and parasites not exposed to irradiation, were injected into intact 2-year-old Hereford steers. All five animals receiving non-irradiated blood died but the five animals which received irradiated blood were only mildly affected.Highly significant differences were observed in changes to plasma fibrinogen, serum fibrinogen-like proteins, packed cell volume, partial thromboplastin time, prothrombin time, blood kinins, and plasma kininogen levels in the control animals but non-significant changes in these parameters occurred in the group receiving irradiated blood. Significant changes in the antiplasmin, α2M, and the antithrombin levels occurred in control cattle but not in the group receiving irradiated blood. Parasite multiplication rates and maximum parasitaemias were similar in both groups. Irradiation reduced the dose of living parasites from 1×108 to 2.5×103, but this was not the reason for the mild reactions. It was concluded that irradiation had selected an avirulent parasite population.Babesia bovis parasites attenuated by 35 krads gamma irradiation and parasites not exposed to irradiation, were injected into intact 2-year-old Hereford steers. All five animals receiving non-irradiated blood died but the five animals which received irradiated blood were only mildly affected. Highly significant differences were observed in changes to plasma fibrinogen, serum fibrinogen-like proteins, packed cell volume, partial thromboplastin time, prothrombin time, blood kinins, and plasma kininogen levels in the control animals but non-significant changes in these parameters occurred in the group receiving iradiated blood. Significant changes in the antiplasmin alpha 2M, and the antithrombin levels occurred in control cattle but not in the group receiving irradiated blood. Parasite multiplications rates and maximum parasitaemias were similar in both groups. Irradiation reduced the dose of living parasites from 1 x 10(8) to 2.5 x 10(3), but this was not the reason for the mild reactions. It was concluded that irradiation had selected an avirulent parasite population.

Proteinases in the lysate of bovine erythrocytes infected with Babesia bovis: initial vaccination studies.

Abstract The lysate of erythrocytes infected with Babesia bovis was tested for proteinases using an electrophoretic method in which substrate was included in the acrylamide matrix. Two babesial proteinases, which seemingly exist in both free and complexed forms, were detected. One of the proteinases was prepared by chromatography and preparative electrophoresis and used to vaccinate four splenectomized calves. The latter, along with a group of control splenectromized calves, were challenged with a strain of B. Bovis from which the proteinase was obtained. All the control calves died whereas only one of the vaccinates died. The protection was evident as a suppression of parasitaemia.

AcuteBabesia bovis infection: A study of the vascular lesions in kidney and lung

SummaryA histological study of kidney and lung from cattle acutely infected with the intra-erythrocytic protozoan parasiteBabesia bovis was undertaken. Samples of the same tissue were examined using haematoxylin-eosin, Lendrums acid picro-Mallory, and phosphotungstic acid-haematoxylin fibrin stains, direct fluorescent antibody techniques using rabbit anti-B. bovis cryofibrinogen and rabbit anti-bovine fibrinogen conjugates, and electron microscopy. Although positive fibrin reactions were observed with histochemical stains, examination of the same material by the electron microscope and fluorescent reagents failed to confirm its presence. It was concluded that the fibrin reactions were given by sludged erythrocytes packed tightly together in capillaries and coated with cryofibrinogen complexes.

Babesia bovis: the effect of acute inflammation and isoantibody production in the detection of babesial antigens.

Immunoblots ofBabesia bovis antigen contain dominant antigens which react not only with antisera toB. bovis but with sera from naive calves recovering from an acute inflammatory reaction. It seems likely these antigens are from the host rather than the parasite.

Babesia bovis (argentina): studies on the composition and location of antigen associated with infected erythrocytes.

Abstract Specific antisera against Babesia bovis (= argentina) antigens were prepared in rabbits by inoculation of precipitates from an extract of infected erythrocytes and absorption of the antisera with normal bovine components. Of three babesial antigens detected, one appeared to contain a modified stromal component. The antisera, when conjugated with fluoroscein isothiocyanate, stained aggregated infected erythrocytes in the microcirculation and located antigen in glomeruli and blood vessel endothelium. It was suggested that a babesial enzyme-fibrinogen complex contributes to the pathological changes of infection such as sludging and adherence of erythrocytes to blood vessel walls.