New Zealand White Rabbits Effectively Clear Borrelia burgdorferi B31 despite the Bacterium’s FunctionalvlsEAntigenic Variation System

Abstract

Borrelia burgdorferi is a tick-borne bacterium responsible for approximately 300,000 annual cases of Lyme disease (LD) in the United States, with increasing incidences in other parts of the world. The debilitating nature of LD is mainly attributed to the ability of B. burgdorferi to persist in patients for many years despite strong anti-Borrelia antibody responses. Antimicrobial treatment of persistent infection is challenging. Similar to infection of humans, B. burgdorferi establishes long-term infection in various experimental animal models except for New Zealand White (NZW) rabbits, which clear the spirochete within 4 to 12 weeks. LD spirochetes have a highly evolved antigenic variation vls system, on the lp28-1 plasmid, where gene conversion results in surface expression of the antigenically variable VlsE protein. VlsE is required for B. burgdorferi to establish persistent infection by continually evading otherwise potent antibodies. Since the clearance of B. burgdorferi is mediated by humoral immunity in NZW rabbits, the previously reported results that LD spirochetes lose lp28-1 during rabbit infection could potentially explain the failure of B. burgdorferi to persist. However, the present study unequivocally disproves that previous finding by demonstrating that LD spirochetes retain the vls system. However, despite the vls system being fully functional, the spirochete fails to evade anti-Borrelia antibodies of NZW rabbits. In addition to being protective against homologous and heterologous challenges, the rabbit antibodies significantly ameliorate LD-induced arthritis in persistently infected mice. Overall, the current data indicate that NZW rabbits develop a protective antibody repertoire, whose specificities, once defined, will identify potential candidates for a much-anticipated LD vaccine.

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

“…several surface proteins of B. burgdorferi have been identified. Interestingly, the newly identified epitopes are distinct from those recently delineated via the mouse repeated immunization assay (58). Importantly, when available, the published data cited below have empirically demonstrated that newly identified PA epitopes are surface exposed and that their respective proteins are considered to be strong LD vaccine candidates.…”

“…In contrast to the 50% protection rate achieved for mice via repeated immunization with the ΔVlsE mutant (59), the rabbit antibodies consistently prevented 100% of mice from host-adapted, VlsE-expressing spirochetes. Moreover, the rabbit antibodies significantly reduced the pathology of LD arthritis in persistently infected mice (58). Thus, in the present work, via subtractive RV (58), we have compared the antibody repertoires of nonprotective (day 14) and protective (day 28) rabbit sera with the goal of identifying the specificities of protective antibodies and, therefore, their respective surface targets.…”

“…Importantly, as shown by quantitative PCR, the NZW rabbits whose biopsy specimens from day 28 were still culture positive consistently harbored very few spirochetes, suggesting the final stage of skin infection clearance at about this time point (58). Moreover, only when they were passively immunized with day 28 rabbit sera did immunocompetent mice gain resistance to the host-adapted B. burgdorferi B31-A3 (B31) strain (58). Thus, by day 28 p.i., NZW rabbits develop a repertoire of highly protective antibodies, which are responsible for the clearance of VlsE-expressing spirochetes (58).…”

“…and 21 p.i., when rabbit skin biopsy specimens were consistently culture positive, the day 28 skin tissues of some NZW rabbits were already negative by culture. Importantly, as shown by quantitative PCR, the NZW rabbits whose biopsy specimens from day 28 were still culture positive consistently harbored very few spirochetes, suggesting the final stage of skin infection clearance at about this time point (58). Moreover, only when they were passively immunized with day 28 rabbit sera did immunocompetent mice gain resistance to the host-adapted B. burgdorferi B31-A3 (B31) strain (58).…”

“…Moreover, only when they were passively immunized with day 28 rabbit sera did immunocompetent mice gain resistance to the host-adapted B. burgdorferi B31-A3 (B31) strain (58). Thus, by day 28 p.i., NZW rabbits develop a repertoire of highly protective antibodies, which are responsible for the clearance of VlsE-expressing spirochetes (58). In order to identify the specificities of the protective antibodies and, consequently, map their targets, we analyzed the global antibody repertoires of immune sera collected from B31-infected rabbits at days 14 and 28 p.i., the time points at which the rabbit antibodies are not yet protective or are already protective, respectively.…”