Temporal Analysis of the Antigenic Composition of Borrelia burgdorferi during Infection in Rabbit Skin

Abstract

The numbers of host-adapted Borrelia burgdorferi (HAB) organisms in rabbit skin were assessed by real-time PCR over the first 3 weeks of infection. Maximal numbers were found at day 11, while spirochete numbers decreased by more than 30-fold by day 21. The antigenic composition of HAB in skin biopsy samples was determined by use of a procedure termed hydrophobic antigen tissue Triton extraction. Immune sera from rabbits, sera from chronically infected mice, and monospecific antiserum to the antigenic variation protein, VlsE, were used to probe parallel two-dimensional immunoblots representing each time point. Individual proteins were identified using either specific antisera or by matching protein spots to mass spectrometryidentified protein spots from in vitro-cultivated Borrelia. There were significant changes in the relative expression of a variety of known and previously unrecognized HAB antigens during the 21-day period. OspC and the outer membrane proteins OspA and OspB were prominent at the earliest time point, day 5, when the antigenic variation protein VlsE was barely detected. OspA and OspB were not detected after day 5. OspC was not detected after day 9. VlsE was the most prominent antigen from day 7 through day 21. BmpA, ErpN, ErpP, LA7, OppA-2, DbpA, and an unidentified 15-kDa protein were also detected from day 7 through day 21. Immunoblot analysis using monospecific anti-VlsE revealed the presence of prominent distinct VlsE lower forms in HAB at days 9, 11, and 14; however, these lower forms were no longer detected at day 21. This marked diminution in VlsE lower forms paralleled the clearance of the spirochete from skin.

Verification and dissection of the ospC operator by using flaB promoter as a reporter in Borrelia burgdorferi

“…Previous studies showed that OspC antibodies administered to immunodeficient mice or the specific humoral response induced during infection of immunocompetent mice constitute an overwhelming force to reduce ospC expression to a baseline level in all tissues [17,18,22,23]. A recent study also showed that lack of the operator allows the ospC promoter to drive constitutive gene expression in all tissues and, as a result, completely diminishes the ability of B. burgdorferi to evade specific humoral immunity [34].…”

“…OspC is not only a strong immunogen but also an effective target of protective immunity; its expression induces a robust humoral response that imposes tremendous pressure on the pathogen [20,21]. To cause persistent infection, B. burgdorferi must down-regulate OspC [17,18,22,23]. If B. burgdorferi fails to repress OspC expression or undergo escape mutations on the ospC gene, infection would be cleared [20].…”

Biology of Infection with Borrelia burgdorferi

“…These experiments demonstrated that the bacterial gene expression profile changes, not only between hosts, but also at different times within a single host. Finally, one group has corroborated some of the gene expression patterns identified in these studies by directly examining the proteins made by bacteria growing in SCID mouse and rabbit skin, taking advantage of the larger number of bacteria found in these models [102,103]. Using these methods, researchers have identified genes whose products are, or are likely to be, produced in mice or ticks, some of which have been tested for their contribution to bacterial survival in the corresponding host (Fig.…”

“…OspC production begins in feeding ticks (or immediately after needle inoculation, Fig. 1) and lasts for the first couple of weeks of mammalian infection [49,51,103,[121][122][123]. Once the bacteria are established in a host, OspC production is not required for persistence [114].…”

“…1) [103]. Although its function is unknown, this lipoprotein has an elaborate system for variation [22].…”