Antibody profiling of canine IgG responses to the OspC protein of the Lyme disease spirochetes supports a multivalent approach in vaccine and diagnostic assay development

Abstract

OspC performs essential functions during the enzootic cycle of the Lyme disease (LD) spirochetes. In this study, the specificity of antibody (Ab) responses to OspC was profiled to define the antigenic determinants during infection and after vaccination. Several OspC variants or 'types' were screened with serum from SNAP4Dx C6 positive dogs and with serum from rabbits hyperimmunized with OspC proteins. The OspC type-specific nature of the Ab response revealed that variable domains of OspC are immunodominant during infection and upon vaccination. To assess the potential of OspC to elicit Ab in the context of a bacterin vaccine, OspC production in strains cultivated in vitro was assessed. Immunoblot and indirect immunofluorescent antibody analyses demonstrated that production is low and that only a subset of cells actively produces OspC in vitro, raising questions about the potential of bacterin vaccines to stimulate significant anti-OspC Ab responses. The specificity of the OspC Ab response in experimentally infected mice over time was assessed to determine if domains shielded in the OspC homodimer become accessible and stimulate Ab production as infection progresses. The results demonstrate that the OspC Ab response remains focused on surface exposed variable regions of the protein throughout infection. In contrast to some earlier studies, it is concluded that conserved domains of OspC, including the C7 or C10 domain, do not elicit significant Ab responses during infection or upon vaccination. Collectively, the results indicate that OspC diversity must be considered in vaccine design and in the interpretation of diagnostic assays that employ OspC as a diagnostic antigen.

Analysis of the antigenic determinants of the OspC protein of the Lyme disease spirochetes: Evidence that the C10 motif is not immunodominant or required to elicit bactericidal antibody responses

“…The OspC specific IgG log titers elicited by each protein were similar (4.8 and 4.4 for OspC and OspCΔC10, respectively) and anti-OspC and anti-OspCΔC10 displayed equal reactivity with both proteins as assessed by immunoblotting (Figure 4). Since OspC is produced at low and variable levels during cultivation [39,50], to measure bactericidal Ab activity, a strain (B31-HE-OspC) that constitutively produces OspC [51] was used as the assay strain. Controls included B. burgdorferi B31(wild type strain; low level OspC production) and B. burgdorferi B31Δ ospC (negative control; no OspC production) [40].…”

“…PCR primers were designed with ligase independent cloning (LIC) tails to allow for annealing into linearized pET46-Ek/LIC vector as described by the supplier (Novagen). All methods pertaining to cloning, protein production and purification were exactly as decribed in an earlier study [39].…”

“…The rat-anti-OspC and rat-anti-OspCΔC10 antisera were used at dilutions of 1:1000. Serum from rabbits immunized with OspC proteins of different OspC types [39] were used at a dilution of 1:30000. Serum from infected dogs were obtained from North Carolina State University.…”

Sequence Variation of Rare Outer Membrane Protein β-Barrel Domains in Clinical Strains Provides Insights into the Evolution of Treponema pallidum subsp. pallidum , the Syphilis Spirochete

“… pallidum strains that are circulating in at-risk populations worldwide. Similar vaccine strategies have been proposed for Borrelia burgdorferi , the Lyme disease spirochete ( 87 – 89 ). The ability to use conserved extracellular loops (e.g., L2 in TprC/D/D2, L5 in TprD/D2, and L3 in BamA) would circumvent the difficulties associated with expression and purification of full-length OMPs on a mass scale ( 70 ).…”

Epitope mapping of Borrelia burgdorferi OspC protein in homodimeric fold

“…These results were confirmed in the experiments utilising nucleocapsid of hepatitis B virus displaying OspC protein in the native fold as confirmed by cryo‐electron microscopy . Nevertheless, the opposite approach based on immunization with synthetic peptides was implemented successfully as well …”