21 research outputs found
Expression of the Tick-Associated Vtp Protein of Borrelia hermsii in a Murine Model of Relapsing Fever.
No full textBorrelia hermsii, a spirochete and cause of relapsing fever, is notable for its immune evasion by multiphasic antigenic variation within its vertebrate host. This is based on a diverse repertoire of surface antigen genes, only one of which is expressed at a time. Another major surface protein, the Variable Tick Protein (Vtp), is expressed in the tick vector and is invariable at its genetic locus. Given the limited immune systems of ticks, the finding of considerable diversity among the Vtp proteins of different strains of B. hermsii was unexpected. We investigated one explanation for this diversity of Vtp proteins, namely expression of the protein in mammals and a consequent elicitation of a specific immune response. Mice were infected with B. hermsii of either the HS1 or CC1 strain, which have antigenically distinctive Vtp proteins but otherwise have similar repertoires of the variable surface antigens. Subsequently collected sera were examined for antibody reactivities against Vtp and other antigens using Western blot analysis, dot blot, and protein microarray. Week-6 sera of infected mice contained antibodies that were largely specific for the Vtp of the infecting strain and were not attributable to antibody cross-reactivities. The antibody responses of the mice infected with different strains were otherwise similar. Further evidence of in vivo expression of the vtp gene was from enumeration of cDNA sequence reads that mapped to a set of selected B. hermsii genes. This measure of transcription of the infecting strain's vtp gene was ~10% of that for the abundantly-expressed, serotype-defining variable antigen gene but similar to that of genes known for in vivo expression. The findings of Vtp expression in a vertebrate host and elicitation of a specific anti-Vtp antibody response support the view that balancing selection by host adaptive immunity accounts in part for the observed diversity of Vtp proteins
Correction: Expression of the Tick-Associated Vtp Protein of Borrelia hermsii in a Murine Model of Relapsing Fever
No full textProtein array reactivities of sera from mice infected with HS1 or CC1 of <i>B</i>. <i>hermsii</i>.
No full textProtein array reactivities of sera from mice infected with HS1 or CC1 of B. hermsii.</p
Unrooted neighbor-joining distance phylogram of codon-aligned partial nucleotide sequences of selected <i>vsp</i>, <i>vtp</i>, and <i>ospC</i> genes of strains CC1, HS1, LPO, and Owl of <i>B</i>. <i>hermsii</i>, strain LB-2001 of <i>B</i>. <i>miyamotoi</i> (Bm), strain Oz of <i>B</i>. <i>turicatae</i> (Bt), or strain B31 of <i>B</i>. <i>burgdorferi</i> (Bb).
No full textThe accession numbers are given in Table 1, and the sequences for the alignment are given in S1 File. The bar indicates the nucleotide distance.</p
Western blot (WB) analyses of antisera to whole cells of <i>B</i>. <i>hermsii</i>.
No full textPanel A. Western blot analysis of whole cell lysates of in vitro cultures of strain HS1 B. hermsii isolates Vtp+ wildtype, vtpFS83 mutant, and vtp::kan knock-out. Blots were incubated with polyclonal mouse antisera to either the mutant or knock-out. Bound antibodies were detected with alkaline phosphatase-labeled anti-mouse immunoglobulin. An immunoreactive band in the WT lysate that was of the same size as Vtp is indicated by an arrow. A band corresponding to the Alp protein in the mutant lysate is indicated an asterisk. Molecular weight standards (MWS) in kilodaltons are shown on the left. Panel B. By Western blot analysis antibodies in the antiserum to vtpFS83 cells bound to purified recombinant Vtp (rVtp) of strain HS1 but not rVtp of strain CC1. Binding of antibodies in the antiserum to vtp::kan knock-out cells to either Vtp was not detected. Molecular weight standards (MWS) in kilodaltons are shown on the left.</p
RNA-Seq with selected genes of <i>B</i>. <i>hermsii</i> CC1 in blood of mice.
No full textRNA-Seq with selected genes of B. hermsii CC1 in blood of mice.</p
Western blot analysis with anti-Vtp monoclonal antibody of aqueous (Aq) or detergent (Det) fractions of Triton X-114 extractions of cells <i>B</i>. <i>hermsii</i> HS1 wildtype, <i>vtp</i>FS83 mutant, and <i>vtp</i>::kan knock-out.
No full textBands that correspond to the predicted migrations of unlipidated preprotein and lipidated processed Vtp are indicated by an asterisk and arrows, respectively. Molecular weight standards (MWS) in kilodaltons are shown on the left.</p
Antibodies to Vtp in sera from infected mice with different serotypes of <i>B</i>. <i>hermsii</i>.
No full textPanel A. Dot blot analysis of binding of antibodies in pooled sera from mice infected for 6 weeks with either serotype 7 or serotype 19 to recombinant Vtp proteins from either strain HS1 or CC1 of B. hermsii. Purified proteins at a concentration of 20 μg/ml were spotted onto nitrocellulose membranes in duplicate and then incubated with sera. Panel B. Western blot analysis of pooled week-6 sera from infected mice to recombinant Vtp proteins at concentrations of 50, 5, or 0.1 μg/ml. Molecular weight standards in kilodaltons are shown on the left.</p
Pathogen and Host Response Dynamics in a Mouse Model of Borrelia hermsii Relapsing Fever
No full textMost Borrelia species that cause tick-borne relapsing fever utilize rodents as their natural reservoirs, and for decades laboratory-bred rodents have served as informative experimental models for the disease. However, while there has much progress in understanding the pathogenetic mechanisms, including antigenic variation, of the pathogen, the host side of the equation has been neglected. Using different approaches, we studied, in immunocompetent inbred mice, the dynamics of infection with and host responses to North American relapsing fever agent B. hermsii. The spirochete’s generation time in blood of infected mice was between 4–5 h and, after a delay, was matched in rate by the increase of specific agglutinating antibodies in response to the infection. After initiating serotype cells were cleared by antibodies, the surviving spirochetes were a different serotype and, as a population, grew more slowly. The retardation was attributable to the host response and not an inherently slower growth rate. The innate responses at infection peak and immediate aftermath were characterized by elevations of both pro-inflammatory and anti-inflammatory cytokines and chemokines. Immunodeficient mice had higher spirochete burdens and severe anemia, which was accounted for by aggregation of erythrocytes by spirochetes and their partially reversible sequestration in greatly enlarged spleens and elsewhere
Fibronectin-Binding Protein of Borrelia hermsii Expressed in the Blood of Mice with Relapsing Fever
No full textTo identify and characterize surface proteins expressed by the relapsing fever (RF) agent Borrelia hermsii in the blood of infected mice, we used a cell-free filtrate of their blood to immunize congenic naive mice. The resultant antiserum was used for Western blotting of cell lysates, and gel slices corresponding to reactive bands were subjected to liquid chromatography-tandem mass spectrometry, followed by a search of the proteome database with the peptides. One of the immunogens was identified as the BHA007 protein, which is encoded by a 174-kb linear plasmid. BHA007 had sequence features of lipoproteins, was surface exposed by the criteria of in situ protease susceptibility and agglutination of Vtp(-) cells by anti-BHA007 antibodies, and was not essential for in vitro growth. BHA007 elicited antibodies during experimental infection of mice, but immunization with recombinant protein did not confer protection against needle-delivered infection. Open reading frames (ORFs) orthologous to BHA007 were found on large plasmids of other RF species, including the coding sequences for the CihC proteins of Borrelia duttonii and B. recurrentis, but not in Lyme disease Borrelia species. Recombinant BHA007 bound both human and bovine fibronectin with Kd (dissociation constant) values of 22 and 33 nM, respectively, and bound to C4-binding protein with less affinity. The distant homology of BHA007 and its orthologs to BBK32 proteins of Lyme disease species, as well as to previously described BBK32-like proteins in relapsing fever species, indicates that BHA007 is a member of a large family of multifunctional proteins in Borrelia species that bind to fibronectin as well as other host proteins
