For the ecological niches where relapsing fever and Lyme disease causing spirochetes overlap (11
) and the serological cross-reactivity from infected patients occurs (37
), identifying antigens that are species specific is important. We demonstrate here that a protein, which we have designated Borrelia
immunogenic protein A (BipA), is surface localized and immunogenic in patients and mice infected with B. hermsii.
Also, with no orthologue of bipA
in B. burgdorferi
), lack of reactivity to rBipA from serum samples of patients infected with Lyme disease causing spirochetes was expected.
Interestingly, pooled sera from mice immunized with a GGI rBipA bound to the protein only in GGI isolates. Due to the regions of amino acid deletions in GGI isolates, the quaternary structure of the protein may differ between genomic groups. Thus, the mice used to produce antisera to rBipA may have generated antibodies against an epitope of the protein that is unique in GGI isolates. In addition, the amino acid alignments of BipA from GGI and GGII isolates indicated conservation at the C terminus of the protein. Therefore, mice infected with the GGII isolates YOR and MTW-4 may have generated antibodies to the C terminus of BipA, suggesting differential antigen processing of BipA in mice immunized with the recombinant protein and during natural infection.
Although GGI isolates displayed more nucleotide diversity in the bipA
locus, there was no evidence for the horizontal gene transfer of bipA
between GGI and GGII isolates, as shown for another plasmid-encoded gene, the variable tick protein (43
). The nucleotide diversity of the bipA
locus in GGI isolates implies a selective pressure on this gene. The surface localization and immunogenicity of BipA supports the hypothesis that immune pressure may be responsible for the observed genetic heterogeneity in GGI isolates. Furthermore, GGII isolates may represent a more recent derivative from GGI isolates, possibly explaining the lesser amount of nucleotide diversity of bipA
in GGII isolates.
Further analysis of the bipA
locus identified identical sequences from B. hermsii
isolates spread over a large geographical area, a finding similar to that when the 16S-23S ribosomal DNA noncoding intergenic spacer region (IGS) was sequenced in these isolates (53
). In addition, sequence analysis of B. hermsii
detected in a squirrel in Montana was identical to that of a spirochete found in a Northern spotted owl in Kittias County, WA (23
). These data further support a role of birds in spreading relapsing fever spirochetes in nature as previously suggested (23
), possibly explaining the distant locations of B. hermsii
with identical bipA
The current antigen used for serodiagnosis of infection by relapsing fever spirochetes in North American and Africa is recombinant glycerophosphodiester phosphodiesterase (rGlpQ) (41
). In addition, the factor H binding protein may be a promising antigen for diagnosing infection caused by B. hermsii
). Since BipA is unique to relapsing fever spirochetes with no identified homologues in other bacteria, and human patients produce high titers of antibody against rBipA, this protein also has potential as a diagnostic antigen. Also, with orthologues of glpQ
, and Klebsiella
species having ca. 45 to 50% amino acid identity to B. hermsii
GlpQ (data not shown), using rBipA with rGlpQ or the factor H binding protein in an enzyme-linked immunosorbent assay could improve serodiagnosis. Furthermore, given the amino acid heterogeneity of BipA between species of relapsing fever spirochetes, expressing bipA
orthologues as recombinant proteins may aid in determining the species causing infection.