Generation of Antigenic Variants via Gene Conversion: Evidence for Recombination Fitness Selection at the Locus Level in Anaplasma marginale

Abstract

Multiple bacterial and protozoal pathogens utilize gene conversion to generate antigenically variant surface proteins to evade immune clearance and establish persistent infection. Both the donor alleles that encode the variants following recombination into an expression site and the donor loci themselves are under evolutionary selection: the alleles that encode variants that are sufficiently antigenically unique yet retain growth fitness and the loci that allow efficient recombination. We examined allelic usage in generating Anaplasma marginale variants during in vivo infection in the mammalian reservoir host and identified preferential usage of specific alleles in the absence of immune selective pressure, consistent with certain individual alleles having a fitness advantage for in vivo growth. In contrast, the loci themselves appear to have been essentially equally selected for donor function in gene conversion with no significant effect of locus position relative to the expression site or origin of replication. This pattern of preferential allelic usage but lack of locus effect was observed independently for Msp2 and Msp3 variants, both generated by gene conversion. Furthermore, there was no locus effect observed when a single locus contained both msp2 and msp3 alleles in a tail-to-tail orientation flanked by a repeat. These experimental results support the hypothesis that predominance of specific variants reflects in vivo fitness as determined by the encoding allele, independent of locus structure and chromosomal position. Identification of highly fit variants provides targets for vaccines that will prevent the high-level bacteremia associated with acute disease.

Identification ofAnaplasma marginaleOuter Membrane Protein Antigens Conserved betweenA. marginaleSensu Stricto Strains and the LiveA. marginalesubsp.centraleVaccine

“…This preferential reactivity of the N terminus over the C terminus is not specific to live immunization nor to the immunoblot assay, as enzymelinked immunosorbent assay (ELISA) mapping following outer membrane immunization yielded the same conclusion (1,44). While Msp3 was not similarly mapped, the shared gene locus structure, HVR recombination mechanism, and pattern of conserved N-and C-terminal regions with a central, surfaceexposed HVR domain supports that lessons learned from Msp2 are applicable to Msp3 (7,9,18,26).…”

Anaplasma marginale Superinfection Attributable to Pathogen Strains with Distinct Genomic Backgrounds

“…The numbers of msp2 loci differ among strains, and there is genomic evidence of gene duplication and divergence (16)(17)(18). The significant increase in the number of msp2 variants in tropical versus temperate region, ϳ3.5ϫ (22), may arise from both an increase in the number of alleles per strain and the consequent increase in the number of segmental recombinants (15) and thus cannot be directly compared to the increase (ϳ2.2ϫ) in the number of omp alleles between tropical and temperate infections in the present study.…”

“…Alternatively, strain A may expand its allelic repertoire on the same genomic background to generate a closely related strain, A=. The latter possibility is supported by msp2 allelic gene duplication within a single strain and the presence of different numbers of msp2 loci and alleles among strains (16)(17)(18)(19). Either strain A= or B would generate the observed increase in the number and diversity of expressed msp2 alleles within the high-prevalence tropical regions (22).…”

“…A. marginale establishes persistent infection within an animal by antigenic variation in the immunodominant surface protein, major surface protein 2 (MSP2), which is generated via gene conversion using a genomic repertoire of distinct msp2 alleles (13)(14)(15)(16). Genomic comparison of multiple strains revealed that although A. marginale has a "closed core" genome with essentially the same gene content among all strains, the repertoire of msp2 alleles is distinct (17,18).…”

Anaplasma phagocytophilumAPH_1387 Is Expressed throughout Bacterial Intracellular Development and Localizes to the Pathogen-Occupied Vacuolar Membrane

“…Tandem repeat proteins of pathogenic bacteria have been implicated in hijacking host signaling pathways, adhesion, immune evasion, and other host-pathogen interactions (15,20,26,27,31,50,52,56). The genomes of A. phagocytophilum and E. chaffeensis each encode multiple acidic tandem repeat-carrying proteins.…”