Major Histocompatibility Complex Class II DR-Restricted Memory CD4 + T Lymphocytes Recognize Conserved Immunodominant Epitopes of Anaplasma marginale Major Surface Protein 1a

Abstract

ABSTRACT Native major surface protein 1 (MSP1) of Anaplasma marginale , composed of covalently associated MSP1a and MSP1b proteins, stimulates protective immunity in cattle against homologous and heterologous strain challenge. Protective immunity against pathogens in the family Anaplasmataceae involves both CD4 + T cells and neutralizing immunoglobulin G. Thus, an effective vaccine should contain both CD4 + T- and B-lymphocyte epitopes that will elicit strong memory responses upon infection with homologous and heterologous strains. Previous studies demonstrated that the predominant CD4 + T-cell response in MSP1 vaccinates is directed against the MSP1a subunit. The present study was designed to identify conserved CD4 + T-cell epitopes in MSP1a presented by a broadly represented subset of major histocompatibility complex (MHC) class II molecules that would be suitable for inclusion in a recombinant vaccine. Transmembrane protein prediction analysis of MSP1a from the Virginia strain revealed a large hydrophilic domain (HD), extending from amino acids (aa) 1 to 366, and a hydrophobic region extending from aa 367 to 593. The N terminus (aa 1 to 67) includes one 28-aa form A repeat and one 29-aa form B repeat, which each contain an antibody neutralization-sensitive epitope [Q(E)ASTSS]. In MSP1 vaccinates, recombinant MSP1a HD (aa 1 to 366) stimulated recall proliferative responses that were comparable to those against whole MSP1a excluding the repeat region (aa 68 to 593). Peptide mapping determined a minimum of five conserved epitopes in aa 151 to 359 that stimulated CD4 + T cells from cattle expressing DR-DQ haplotypes common in Holstein-Friesian breeds. Peptides representing three epitopes (aa 231 to 266, aa 270 to 279, and aa 290 to 319) were stimulatory for CD4 + T-cell clones and restricted by DR. A DQ-restricted CD4 + T-cell epitope, present in the N-terminal form B repeat (VSSQSDQASTSSQLG), was also mapped using T-cell clones from one vaccinate. Although form B repeat-specific T cells did not recognize the form A repeat peptide (VSSQS_EASTSSQLG), induction of T-cell anergy by this peptide was ruled out. The presence of multiple CD4 + T-cell epitopes in the MSP1a HD, in addition to the neutralization-sensitive epitope, supports the testing of this immunogen for induction of protective immunity against A. marginale challenge.