Reactivity of Antibodies from Syphilis Patients to a Protein Array Representing the Treponema pallidum Proteome

Department of Molecular Virology and Microbiology, Baylor College of Medicine, One Baylor Plaza, Houston, Texas 77030, USA.
Journal of Clinical Microbiology (Impact Factor: 3.99). 04/2006; 44(3):888-91. DOI: 10.1128/JCM.44.3.888-891.2006
Source: PubMed


To identify antigens important in the human immune response to syphilis, the serum antibody reactivity of syphilitic patients
was examined with 908 of the 1,039 proteins in the proteome of Treponema pallidum subsp. pallidum using a protein array enzyme-linked immunosorbent assay. Thirty-four proteins exhibited significant reactivity when assayed
with human sera from patients in the early latent stage of syphilis. A subset of antigens identified were further scrutinized
for antibody reactivity at primary, secondary, and latent disease stages, and the results demonstrate that the humoral immune
response to individual T. pallidum proteins develops at different rates during the time course of infection.

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    • "A number of years ago, we predicted that rare OMPs would be poorly immunogenic because they are nonlipidated and expressed at extremely low levels (Radolf, 1995). Serologic studies using full-length TP0326 disproved this idea (Van Voorhis et al., 2003; McKevitt et al., 2005; Brinkman et al., 2006), suggesting that the spirochete employs an alternative stratagem to harness the Bam pathway while avoiding the deleterious consequences of the antibody responses TP0326 elicits. Immunoblot analysis with the POTRA and β-barrel constructs revealed what appears to be the spirochete's solution to this dilemma and an ostensibly striking dichotomy in the antibody responses to TP0326 elicited in humans and rabbits. "
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    ABSTRACT: Definitive identification of Treponema pallidum rare outer membrane proteins (OMPs) has long eluded researchers. TP0326, the sole protein in T. pallidum with sequence homology to a Gram-negative OMP, belongs to the BamA family of proteins essential for OM biogenesis. Structural modelling predicted that five polypeptide transport-associated (POTRA) domains comprise the N-terminus of TP0326, while the C-terminus forms an 18-stranded amphipathic β-barrel. Circular dichroism, heat modifiability by SDS-PAGE, Triton X-114 phase partitioning and liposome incorporation supported these topological predictions and confirmed that the β-barrel is responsible for the native protein's amphiphilicity. Expression analyses revealed that native TP0326 is expressed at low abundance, while a protease-surface accessibility assay confirmed surface exposure. Size-exclusion chromatography and blue native polyacrylamide gel electrophoresis revealed a modular Bam complex in T. pallidum larger than that of Escherichia coli. Non-orthologous ancillary factors and self-association of TP0326 via its β-barrel may both contribute to the Bam complex. T. pallidum-infected rabbits mount a vigorous antibody response to both POTRA and β-barrel portions of TP0326, whereas humans with secondary syphilis respond predominantly to POTRA. The syphilis spirochaete appears to have devised a stratagem for harnessing the Bam pathway while satisfying its need to limit surface antigenicity.
    Full-text · Article · Jun 2011 · Molecular Microbiology
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    • "It may be explained that the discordant results due to the different antigens used in different kits. Some studies described that antigens showed different antibody reactivity at different syphilitic stages [9]. TPPA kit is manufactured using whole pathogenic T. pallidum strain, while CLIA kit used several recombinant antigens of T. pallidum. "
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    ABSTRACT: To assess the sensitivity, specificity, and feasibility of a new chemiluminescence immunoassay (CLIA) in the diagnosis of syphilis. At first, a retrospective study was conducted, using 135 documented cases of syphilis and 30 potentially interfering samples and 80 normal sera. A prospective study was also performed by testing 2, 071 unselected samples for routine screening for syphilis. CLIA was compared with a nontreponemal test (TRUST) and a treponemal test (TPPA). There was an agreement of 100% between CLIA and TPPA in the respective study. The percentage of agreement among the 245 sera tested was 100.0%. Compared with TPPA, the specificity of CLIA was 99.9% (1817/1819), the sensitivity of CLIA was 100.0% (244/244) in the prospective study. CLIA showed 99.5% agreement with TPPA by testing 2, 071 unselected samples. And CLIA seemed to be more sensitive than TPPA in detecting the samples of primary syphilis. CLIA is easy to perform and the indicator results are objective and unequivocal. It may be suitable for large-scale screening as a treponemal test substituted for TPPA.
    Full-text · Article · Feb 2010 · European journal of medical research
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    • "TP0326 was predicted to be a virulence factor [26] and was experimentally verified to be an antigen [27]. It is of interest that the most variable region of the genome represents TP0136 (and sequence upstream of this gene) which encodes a protein that is antigenic in both rabbit and human infections [27,28] and was found to serve as fibronectin and laminin binding protein [29]. "
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    ABSTRACT: Syphilis spirochete Treponema pallidum ssp. pallidum remains the enigmatic pathogen, since no virulence factors have been identified and the pathogenesis of the disease is poorly understood. Increasing rates of new syphilis cases per year have been observed recently. The genome of the SS14 strain was sequenced to high accuracy by an oligonucleotide array strategy requiring hybridization to only three arrays (Comparative Genome Sequencing, CGS). Gaps in the resulting sequence were filled with targeted dideoxy-terminators (DDT) sequencing and the sequence was confirmed by whole genome fingerprinting (WGF). When compared to the Nichols strain, 327 single nucleotide substitutions (224 transitions, 103 transversions), 14 deletions, and 18 insertions were found. On the proteome level, the highest frequency of amino acid-altering substitution polymorphisms was in novel genes, while the lowest was in housekeeping genes, as expected by their evolutionary conservation. Evidence was also found for hypervariable regions and multiple regions showing intrastrain heterogeneity in the T. pallidum chromosome. The observed genetic changes do not have influence on the ability of Treponema pallidum to cause syphilitic infection, since both SS14 and Nichols are virulent in rabbit. However, this is the first assessment of the degree of variation between the two syphilis pathogens and paves the way for phylogenetic studies of this fascinating organism.
    Full-text · Article · Feb 2008 · BMC Microbiology
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