The detection of IgM and IgG antibodies against Babesia bigemina in bovine sera using semi-defined antigens in enzyme immunoassays

Abstract

Soluble extracts prepared from Babesia bigenina merozoites were tested for anti-genicity in class-specific enzyme immunoassays currently being evaluated for the differential serodiagnosis of bovine babesiosis. Intact merozoites were harvested from erythrocytes from an experimentally-infected calf by controlled hypotonic lysis and differential ultracentrifugation. The merozoites were disrupted by ultrasonication and a crude soluble extract obtained by ultracentrifugation. Fractionation of the crude extract on calibrated Sephadex G-200 column consistently produced 4 fractions with molecular weights of 600, 40, 15 and 5 k (k = 10 3 daltoons). Only the 100 and 15 k fractions proved to be antigenic when reacted against bovine immune sera. These fractions were incorporated into IgM- and IgG-specific enzyme immunoassays and used to determine the kinetics of the host-antibody responses to infection. The use of semi-defined antigens allowed assay standardization and good reproducibility of the results. A calf infected with a cryopreserved stabilate of B. bigemina originating from adult Boophilus microplus ticks developed a mild transient fever from 6–4 days post-infection (d.p.i.) and low parasitaemia levels from 7–16 d.p.i. IgG-antibodies first appeared at 7 d.p.i.,peaked in intensity at 12 d.p.i. and then persisted at these levels until the end of the test period at 49 d.p.i. IgM-antibodies appeared at 7 d.p.i., peaked in intensity from 12–22 d.p.i., but then declined to low levels by 28 d.p.i.. The importance of this transitory IgM-antibody response in the serodiagnosis of acute B. bigemina infections remains to be determined in clinical and field situations.