Sequence Variation and Immunologic Cross-Reactivity among Babesia bovis Merozoite Surface Antigen 1 Proteins from Vaccine Strains and Vaccine Breakthrough Isolates

Abstract

The Babesia bovis merozoite surface antigen 1 (MSA-1) is an immunodominant membrane glycoprotein that is the target of invasion-blocking antibodies. While antigenic variation has been demonstrated in MSA-1 among strains from distinct geographical areas, the extent of sequence variation within a region where it is endemic and the effect of variation on immunologic cross-reactivity have not been assessed. In this study, sequencing of MSA-1 from two Australian B. bovis vaccine strains and 14 breakthrough isolates from vaccinated animals demonstrated low sequence identity in the extracellular region of the molecule, ranging from 19.8 to 46.7% between the T vaccine strain and eight T vaccine breakthrough isolates, and from 18.7 to 99% between the K vaccine strain and six K vaccine breakthrough isolates. Although MSA-1 amino acid sequence varied substantially among strains, overall predicted regions of hydrophilicity and hydrophobicity in the extracellular domain were conserved in all strains examined, suggesting a conserved functional role for MSA-1 despite sequence polymorphism. Importantly, the antigenic variation created by sequence differences resulted in a lack of immunologic cross-reactivity among outbreak strains using sera from animals infected with the B. bovis vaccine strains. Additionally, sera from cattle hyperinfected with the Mexico strain of B. bovis and shown to be clinically immune did not cross-react with MSA-1 from any other isolate tested. The results indicate that isolates of B. bovis capable of evading vaccine-induced immunity contain an msa-1 gene that is significantly different from the msa-1 of the vaccine strain, and that the difference can result in a complete lack of cross-reactivity between MSA-1 from vaccine and breakthrough strains in immunized animals.

Merozoite Surface Antigen 2 Proteins of Babesia bovis Vaccine Breakthrough Isolates Contain a Unique Hypervariable Region Composed of Degenerate Repeats

“…bovis vaccine breakthrough isolates are genetically and antigenically distinct from vaccine strains (12,13). These differences are, in part, due to the MSA-1 proteins, which in these breakthrough isolates have recently been shown to encode strain-specific epitopes not shared with vaccine strains (12).…”

“…However, the composite genetic and antigenic changes that result in vaccine breakthrough are not defined. LeRoith et al recently showed that significant MSA-1 variation was present in every vaccine breakthrough isolate examined (12). Based on studies using American strains (defined by geographic origins), MSA-2 has been postulated to be less divergent despite apparent intragenic sites of genetic exchange (10).…”

Anaplasma marginaleType IV Secretion System Proteins VirB2, VirB7, VirB11, and VirD4 Are Immunogenic Components of a Protective Bacterial Membrane Vaccine

“…Recombinant VirB2 and VirD4F2 were purified once using a ProBond system, under denaturing conditions, and dialyzed in 10 mM Tris buffer at pH 7.8 containing 0.1% Triton X detergent (Bio-Rad) using Slide-A-Lyzer dialysis cassettes with a 10-kDa molecular mass cutoff, purified by immunoaffinity chromatography using an anti-FLAG agarose matrix (Sigma-Aldrich), and dialyzed against PBS. VirB9-1, VirB9-2, VirB10, and a negative-control protein from M07 strain Babesia bovis, merozoite surface antigen 1 (MSA1), and T4SS protein VirB7 were also expressed and purified for this study as previously described (43,46), using gene-specific primers (Table 1). AM306 (putative virb7), was amplified from A. marginale St. Maries strain genomic DNA without a C-terminal FLAG epitope.…”

Genome-Based Vaccinology Applied to Bovine Babesiosis

“…It could be due to the fact that msa-1 gene has an important allelic variation in strains from the nearby geographical regions. This variation suggests that the antibodies generated could not have a cross-reaction between different strains [34,35].…”