Dominant Epitopes of the C6 Diagnostic Peptide ofBorrelia burgdorferiAre Largely Inaccessible to Antibody on the Parent VlsE Molecule

Abstract

Lyme borreliosis (LB) is a disease for which antibody-based detection assays are often required for diagnosis. The variable surface molecule VlsE and IR6, one of its invariable regions, are commonly targeted by the antibody response in infected individuals. A series of enzyme-linked immunosorbent assays was performed to comparatively examine the antibody responses of North American LB patients (n ‫؍‬ 37) to VlsE and invariable segments of this molecule. Both immunoglobulin M (IgM) and IgG responses to full-length VlsE and to peptides reproducing invariable regions 2, 4, and 6, as well as the invariable domains at the amino and carboxyl termini of VlsE, were assessed. The proportions and specificities of reactivity to the invariable segments were tested by using cognate peptides as competitors for VlsE binding by patient serum antibodies. IR6 epitopes (by the C6 peptide) were found to dominate the response to invariable segments. IR6 (C6)-specific antibodies were detected in 78% of the serum specimens, whereas <40% of patients generated antibodies that bound the N-or C-terminal domain and <12% of patients responded to either IR2 or IR4. Interestingly, 15 of 37 patients generated IgG antibodies that reacted with C6 but not with VlsE. Conversely, IgM responses were frequent for VlsE but not for invariable segments. A representative number of the serum specimens (n ‫؍‬ 8) that contained IgG antibodies reacting with both C6 and VlsE was assessed in competition experiments, using C6 as a competitor. Only half of these specimens contained IgG antibodies whose binding to VlsE could be inhibited >50% by competition with the added C6 peptide. The median percent inhibition was 45.5%. These findings indicate that IR6 epitopes are largely concealed from the VlsE molecular surface and that full-length VlsEbased diagnosis likely detects antibodies to conformational and/or variable region epitopes.

Dynamic Longitudinal Antibody Responses during Borrelia burgdorferi Infection and Antibiotic Treatment of Rhesus Macaques

“…This dose corresponded to Ͼ12 mg/kg of body weight/day to ensure that an effective blood level was achieved. Blood was collected at the following time points: 0, 2, 6, 10, 14,16,18,22,26,28,34,40, and 47 weeks p.i.…”

“…The C6 test, for instance, is more sensitive for human patients with early or late disseminated disease than for patients in the localized phase (5,16). While the C6 peptide is highly conserved, other ELISA and Western blotting tests utilize whole-cell lysates or recombinant proteins from one species/ strain/isolate, despite the enormous potential for antigenic variability that can preclude recognition by antibody.…”

Multiplex Immunoassay for Lyme Disease Using VlsE1-IgG and pepC10-IgM Antibodies: Improving Test Performance through Bioinformatics

“…Given the complex nature of the host immune response to B. burgdorferi infection, the use of multiple serologic assays has been proposed to enhance either test sensitivity or specificity (2,6,12,17,34,35). Tests derived from continuous data generate binary (positive or negative) results when a cutoff value is chosen to achieve a desired specificity (e.g., 99%).…”

ROLE OF VlsE/C6 ANTIGEN AS A MARKER FOR EARLY LYME BORRELIOSIS DIAGNOSIS AND MONITORING THE EFFECTIVENESS OF ITS TREATMENT

“…What was also surprising was that only 15 patients observed reactions with C6, but not with full recombinant VlsE. According to the authors, in the full VlsE molecule used in the diagnostic tests, the C6 epitope is well covered by variable regions, and it is the variable regions of the protein surface that are responsible for the positive results [17]. Attention has also been paid to the possibility of using C6 in diagnostics of early Lyme borreliosis.…”