Recombinant Flagellin A Proteins from
 Borrelia burgdorferi
 Sensu Stricto,
 B
 .
 afzelii
 , and
 B. garinii
 in Serodiagnosis of Lyme Borreliosis

Panelius, Jaana; Lahdenne, Pekka; Saxén, Harri; Heikkilä, Tero T.; Seppälä, Ilkka

doi:10.1128/jcm.39.11.4013-4019.2001

Cited by 19 publications

(20 citation statements)

References 39 publications

(17 reference statements)

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“…In an early exposure to borreliae, the expected IgM isotype antibodies could be detected only infrequently. The reason for this is unknown, but the low sensitivity of IgM serology has also been associated with most of the other new recombinant borrelial antigens tested (17,20,21,24). Hence, the present results imply that assessments for IgG antibodies, although not IgM antibodies, to BBK32 proteins may afford a major improvement in the serodiagnosis of early LB.…”

Section: Discussionmentioning

confidence: 74%

Recombinant BBK32 Protein in Serodiagnosis of Early and Late Lyme Borreliosis

et al. 2002

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Borrelial protein BBK32 was evaluated as an antigen in the serodiagnosis of early and disseminated Lyme borreliosis (LB). bbk32 was cloned and sequenced from eight isolates of the three pathogenic Borrelia species. The identities between the amino acid sequences of the BBK32 proteins from Borrelia burgdorferi sensu stricto, B. garinii, and B. afzelii isolates were 71 to 100%. By immunoglobulin G (IgG) Western blotting (WB) or enzyme-linked immunosorbent assay (ELISA), up to 74 and 100% of acute-and convalescent-phase samples, respectively, from 23 patients with erythema migrans (EM) were positive for recombinant BBK32 protein from B. afzelii. In the serology of disseminated LB, the three variant BBK32 antigens cross-reacted. In total, 14 of 14 samples from patients with neuroborreliosis and 15 of 15 samples from patients with Lyme arthritis were positive. The specificities of the IgG ELISA with the variant BBK32 antigens for EM and disseminated borreliosis were 81 to 92% and 89 to 95%, respectively. Our findings indicate that the BBK32 proteins are promising serodiagnostic antigens for the detection of early and disseminated LB but that variant BBK32 proteins may be needed either in parallel or in combination with an immunoassay for LB to cover all the relevant borrelial species that cause the disease.Lyme borreliosis (LB) is a tick-transmitted spirochetal infectious disease, caused by Borrelia burgdorferi, which is characterized by multistage skin, joint, neurologic, and cardiac manifestations (29). The diagnosis of LB is based on clinical evaluation of the patients, but serologic assays, most frequently, enzyme-linked immunosorbent assay (ELISA) and Western blotting (WB), are often used to provide supporting evidence of infection with B. burgdorferi. The antigens predominantly used in the present routine LB serodiagnostic tests are borrelial flagellin protein or whole-cell lysate (WCL) of the microbes cultured in vitro. In different laboratories, the performances of serologic assays with these antigens in terms of their specificities and sensitivities are highly variable (26). Especially in LB patients with early manifestations, e.g., erythema migrans (EM) and facial palsy, antibody responses to the antigens used at present may be weak or delayed (22). In Europe, evaluation of the serologic responses is further complicated by the existence of three species of B. burgdorferi sensu lato as causes of human LB: B. burgdorferi sensu stricto, B. afzelii, and B. garinii (6). One of the main reasons for these problems is the antigenic diversity due to variations in the sequences and expression of immunogenic proteins in these different borrelial species (26,27).The expression of borrelial proteins also varies at different stages in the life cycle of Borrelia in ticks and in the mammalian hosts. Several genes, e.g., bbk32, bbk50, vls, and ospE and ospF homologs (4,8,10,12,14,25,(30)(31)(32)(33)36), have been shown to be selectively expressed in vivo. Moreover, bbk32 expression is detectable in spirochetes during tick f...

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Section: Discussionmentioning

confidence: 74%

Recombinant BBK32 Protein in Serodiagnosis of Early and Late Lyme Borreliosis

et al. 2002

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“…BdrT, also called BdrF2, has been reported to be upregulated in "host-adapted" B. burgdorferi and to be specifically expressed during early infection in mice (75,97). This study revealed that several flagellar apparatus proteins besides FlaB flagellin (BB0147), the FlgE hook protein (BB0283), and the FlaA protein (BB0668) (42,51,69,77) elicit antibody responses during infection. Brinkmann et al found that FlgE of T. pallidum was frequently bound by antibodies in sera from patients with syphilis (11).…”

Section: Discussionmentioning

confidence: 98%

A Genome-Wide Proteome Array Reveals a Limited Set of Immunogens in Natural Infections of Humans and White-Footed Mice withBorrelia burgdorferi

et al. 2008

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Humans and other animals with Lyme borreliosis produce antibodies to a number of components of the agent Borrelia burgdorferi, but a full accounting of the immunogens during natural infections has not been achieved. Employing a protein array produced in vitro from 1,292 DNA fragments representing ϳ80% of the genome, we compared the antibody reactivities of sera from patients with early or later Lyme borreliosis to the antibody reactivities of sera from controls. Overall, ϳ15% of the open reading frame (ORF) products ( Infectious disease research has advanced rapidly with the accumulation of whole-genome sequences of pathogens and the subsequent use of genome-wide DNA microarrays to study gene expression. Equipped with arrays in different formats, investigators have identified different genes in a variety of pathogens that are more highly expressed in host animals or under in vitro conditions mimicking the in vivo environment. With few exceptions (27), these array studies have been performed with experimental animals, usually rodents, and in laboratory settings. Less is known about the proteins that are expressed during natural infections of humans or other host animals. Detection of a specific antibody during the course of infection is indirect evidence of in vivo expression by the pathogen. But the use of this approach to study large numbers of proteins has been largely limited to one-dimensional and two-dimensional gel electrophoresis of whole cells having an in vitro origin, followed by identification of the more abundant antigens by partial amino acid sequencing of reactive bands or spots and then searches of the databases (22,23,38,44,52,60,66).An alternative to using the pathogen itself as the source of the proteins is to produce recombinant polypeptides based on the deduced open reading frames (ORFs) of the pathogen's genome and to determine whether these polypeptides are antibody targets (11,61). A potential shortcoming of using this approach with cells, such as Escherichia coli or yeast (Saccharomyces cerevisiae) cells, is that some foreign proteins may not be expressed or the quantity may be insufficient. Felgner and coinvestigators increased the success rate for expression and decreased the cost with a high-throughput, cell-free, coupled transcription-translation system (26,29,30). Hundreds to thousands of individual recombinant proteins are printed on chips, which are then used to capture antibodies present in serum from infected individuals and other animals. The amount of captured antibody is quantified using a labeled secondary antibody. Whole-proteome microarrays have been employed in studies of experimental Francisella tularensis infections in laboratory mice (35,57) and of immune responses of humans to immunization with live vaccinia virus (26,29,31). McKevitt et al. (61) and Brinkmann et al. (11) used the enzyme-linked immunosorbent assay (ELISA) format to study the binding of antibodies of experimentally infected rabbits and people with syphilis to a nearly complete representation of the OR...

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“…The internal portion of the flagellin molecule, containing the variable, genus-specific immunodominant domain, is less cross-reactive with antigens of other bacteria than the whole protein (101,179,274). The flagellar outer sheath protein FlaA, with a molecular size of 37 kDa, is another immunodominant antigen, especially in early disease (7,84,104,246).…”

Section: Immunologic Diagnosis Of B Burgdorferi Sensu Lato Infectionmentioning

confidence: 99%

Diagnosis of Lyme Borreliosis

et al. 2005

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A large amount of knowledge has been acquired since the original descriptions of Lyme borreliosis (LB) and of its causative agent, Borrelia burgdorferi sensu stricto. The complexity of the organism and the variations in the clinical manifestations of LB caused by the different B. burgdorferi sensu lato species were not then anticipated. Considerable improvement has been achieved in detection of B. burgdorferi sensu lato by culture, particularly of blood specimens during early stages of disease. Culturing plasma and increasing the volume of material cultured have accomplished this. Further improvements might be obtained if molecular methods are used for detection of growth in culture and if culture methods are automated. Unfortunately, culture is insensitive in extracutaneous manifestations of LB. PCR and culture have high sensitivity on skin samples of patients with EM whose diagnosis is based mostly on clinical recognition of the lesion. PCR on material obtained from extracutaneous sites is in general of low sensitivity, with the exception of synovial fluid. PCR on synovial fluid has shown a sensitivity of up to >90% (when using four different primer sets) in patients with untreated or partially treated Lyme arthritis, making it a helpful confirmatory test in these patients. Currently, the best use of PCR is for confirmation of the clinical diagnosis of suspected Lyme arthritis in patients who are IgG immunoblot positive. PCR should not be used as the sole laboratory modality to support a clinical diagnosis of extracutaneous LB. PCR positivity in seronegative patients suspected of having late manifestations of LB most likely represents a false-positive result. Because of difficulties in direct methods of detection, laboratory tests currently in use are mainly those detecting antibodies to B. burgdorferi sensu lato. Tests used to detect antibodies to B. burgdorferi sensu lato have evolved from the initial formats as more knowledge on the immunodominant antigens has been collected. The recommendation for two-tier testing was an attempt to standardize testing and improve specificity in the United States. First-tier assays using whole-cell sonicates of B. burgdorferi sensu lato need to be standardized in terms of antigen composition and detection threshold of specific immunoglobulin classes. The search for improved serologic tests has stimulated the development of recombinant protein antigens and the synthesis of specific peptides from immunodominant antigens. The use of these materials alone or in combination as the source of antigen in a single-tier immunoassay may someday replace the currently recommended two-tier testing strategy. Evaluation of these assays is currently being done, and there is evidence that certain of these antigens may be broadly cross-reactive with the B. burgdorferi sensu lato species causing LB in Europe

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Recombinant Flagellin A Proteins from Borrelia burgdorferi Sensu Stricto, B . afzelii , and B. garinii in Serodiagnosis of Lyme Borreliosis

Cited by 19 publications

References 39 publications

Recombinant BBK32 Protein in Serodiagnosis of Early and Late Lyme Borreliosis

Recombinant BBK32 Protein in Serodiagnosis of Early and Late Lyme Borreliosis

A Genome-Wide Proteome Array Reveals a Limited Set of Immunogens in Natural Infections of Humans and White-Footed Mice withBorrelia burgdorferi

Diagnosis of Lyme Borreliosis

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