Identification of Surface Epitopes Associated with Protection against Highly Immune-Evasive VlsE-Expressing Lyme Disease Spirochetes

Abstract

The tick-borne pathogen is responsible for approximately 300,000 Lyme disease (LD) cases per year in the United States. Recent increases in the number of LD cases, in addition to the spread of the tick vector and a lack of a vaccine, highlight an urgent need for designing and developing an efficacious LD vaccine. Identification of protective epitopes that could be used to develop a second-generation (subunit) vaccine is therefore imperative. Despite the antigenicity of several lipoproteins and integral outer membrane proteins (OMPs) on the surface, the spirochetes successfully evade antibodies primarily due to the VlsE-mediated antigenic variation. VlsE is thought to sterically block antibody access to protective epitopes of However, it is highly unlikely that VlsE shields the entire surface epitome. Thus, identification of subdominant epitope targets that induce protection when they are made dominant is necessary to generate an efficacious vaccine. Toward the identification, we repeatedly immunized immunocompetent mice with live-attenuated VlsE-deleted and then challenged the animals with the VlsE-expressing (host-adapted) wild type. Passive immunization and Western blotting data suggested that the protection of 50% of repeatedly immunized animals against the highly immune-evasive was antibody mediated. Comparison of serum antibody repertoires identified in protected and nonprotected animals permitted the identification of several putative epitopes significantly associated with the protection. Most linear putative epitopes were conserved between the main pathogenic genospecies and found within known subdominant regions of OMPs. Currently, we are performing immunization studies to test whether the identified protection-associated epitopes are protective for mice.

Delineating Surface Epitopes of Lyme Disease Pathogen Targeted by Highly Protective Antibodies of New Zealand White Rabbits

“…Peptide residues are rendered in uppercase if they are part of a BLASTP hit alignment and lowercase otherwise. Peptide identifiers (in parentheses) are prefixed with either "p" for protection-associated peptides in mice from previous work (59) or "q" for peptides uniquely recognized by day 28 rabbit sera. Residues of A3 marked with asterisks are critical for LA-2 binding and complement-dependent bactericidal activity (140).…”

“…Three individual serum samples from day 14 and three individual serum samples from day 28 were analyzed via RPPDL/NGS, the approach that was successfully applied to identify antibody repertoires in B. burgdorferi-infected mice (59,63). As a background control, preimmune sera collected from the three NZW rabbits prior to their challenge were pooled in equal volumes and analyzed as a single sample.…”

“…Identification of protection-associated surface epitopes of B. burgdorferi. To examine whether antibodies of the day 28 repertoire represented by the 309 peptides were specific to B31 surface proteins, the previously developed in silico algorithm was applied (59). Specifically, the 309 peptides were first mapped to the entire proteome of B. burgdorferi B31 (a total of 1,391 proteins) (64).…”

“…Comparison of protection-associated sequences with previously identified conformational surface epitopes of B. burgdorferi. In addition to linear epitopes, we attempted to map conformational surface epitopes associated with the PA peptides uniquely recognized by the rabbit day 28 sera vis-à-vis the PA peptides that were previously identified in mice (59). The analysis involved a total of five conformational epitopes within outer surface proteins A and B of B. burgdorferi B31 (OspA and OspB, respectively) ( Fig.…”

“…As before (59), aligned peptides were also analyzed in relation to three protective sequences ( Fig. 3), from OspA, OspB, and OspC, which had previously been shown to elicit antipeptide antibodies with complement-dependent (anti-OspA and anti-OspC) or complement-independent (anti-OspB) bactericidal activity (65)(66)(67).…”