scholarly journals Human Antibody Responses to VlsE Antigenic Variation Protein of Borrelia burgdorferi

1999 ◽  
Vol 37 (12) ◽  
pp. 3997-4004 ◽  
Author(s):  
M. B. Lawrenz ◽  
J. M. Hardham ◽  
R. T. Owens ◽  
J. Nowakowski ◽  
A. C. Steere ◽  
...  
Keyword(s):  
Lyme Disease ◽  
Borrelia Burgdorferi ◽  
Lupus Erythematosus ◽  
Antigenic Variation ◽  
Erythema Migrans ◽  
Disease Patient ◽  
Enzyme Linked Immunosorbent Assay ◽  
Human Antibody ◽  
Whole Cell ◽  
Cell Elisa

VlsE is a 35-kDa surface-exposed lipoprotein of Borrelia burgdorferi that was shown previously to undergo antigenic variation through segmental recombination of silent vlscassettes with vlsE during experimental mouse infections. Previous data had indicated that sera from North American Lyme disease patients and experimentally infected animals contained antibodies reactive with VlsE. In this study, sera from patients with Lyme disease, syphilis, and autoimmune conditions as well as from healthy controls were examined for reactivity with VlsE by Western blotting and enzyme-linked immunosorbent assay (ELISA). Strong Western blot reactivity to a recombinant VlsE cassette region protein was obtained consistently with Lyme disease sera. Although sera from Lyme disease patients also reacted with a band corresponding to VlsE in B. burgdorferi B31-5A3, interpretation was complicated by low levels of VlsE expression in in vitro-cultured B. burgdorferi and by the presence of comigrating bands. An ELISA using recombinant VlsE was compared with an ELISA using sonically disrupted B. burgdorferi as the antigen. For a total of 93 Lyme disease patient sera examined, the VlsE ELISA yielded sensitivities of 63% for culture-confirmed erythema migrans cases and 92% for later stages, as compared to 61 and 98%, respectively, for the “whole-cell” ELISA. The specificities of the two assays with healthy blood donor sera were comparable, but the VlsE ELISA was 90% specific with sera from syphilis patients, compared to 20% specificity for the whole-cell ELISA with this group. Neither assay showed reactivity with a panel of sera from 20 non-Lyme disease arthritis patients or 20 systemic lupus erythematosus patients. Our results indicate that VlsE may be useful in the immunodiagnosis of Lyme disease and may offer greater specificity than ELISAs using whole B. burgdorferi as the antigen.

scholarly journals Evaluation of Selected Borrelia burgdorferi lp54 Plasmid-Encoded Gene Products Expressed during Mammalian Infection as Antigens To Improve Serodiagnostic Testing for Early Lyme Disease

2015 ◽  
Vol 22 (11) ◽  
pp. 1176-1186 ◽  
Author(s):  
Zachary P. Weiner ◽  
Rebecca M. Crew ◽  
Kevin S. Brandt ◽  
Amy J. Ullmann ◽  
Martin E. Schriefer ◽  
...  
Keyword(s):  
Lyme Disease ◽  
Borrelia Burgdorferi ◽  
Recombinant Proteins ◽  
Erythema Migrans ◽  
Disease Patient ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Early Stages ◽  
Lyme Carditis ◽  
Mammalian Infection

ABSTRACTLaboratory testing for the diagnosis of Lyme disease is performed primarily by serologic assays and is accurate for detection beyond the acute stage of the infection. Serodiagnostic assays to detect the early stages of infection, however, are limited in their sensitivity, and improvement is warranted. We analyzed a series ofBorrelia burgdorferiproteins known to be induced within feeding ticks and/or during mammalian infection for their utility as serodiagnostic markers against a comprehensive panel of Lyme disease patient serum samples. The antigens were assayed for IgM and IgG reactivity in line immunoblots and separately by enzyme-linked immunosorbent assay (ELISA), with a focus on reactivity against early Lyme disease with erythema migrans (EM), early disseminated Lyme neuroborreliosis, and early Lyme carditis patient serum samples. By IgM immunoblotting, we found that recombinant proteins BBA65, BBA70, and BBA73 reacted with early Lyme EM samples at levels comparable to those of the OspC antigen used in the current IgM blotting criteria. Additionally, these proteins reacted with serum samples from patients with early neuroborreliosis and early carditis, suggesting value in detecting early stages of this disease progression. We also found serological reactivity against recombinant proteins BBA69 and BBA73 with early-Lyme-disease samples using IgG immunoblotting and ELISA. Significantly, some samples that had been scored negative by the Centers for Disease Control and Prevention-recommended 2-tiered testing algorithm demonstrated positive reactivity to one or more of the antigens by IgM/IgG immunoblot and ELISA. These results suggest that incorporating additionalin vivo-expressed antigens into the current IgM/IgG immunoblotting tier in a recombinant protein platform assay may improve the performance of early-Lyme-disease serologic testing.

scholarly journals Impact of Clinical Variables on Borrelia burgdorferi-Specific Antibody Seropositivity in Acute-Phase Sera from Patients in North America with Culture-Confirmed Early Lyme Disease

2008 ◽  
Vol 15 (10) ◽  
pp. 1519-1522 ◽  
Author(s):  
Gary P. Wormser ◽  
John Nowakowski ◽  
Robert B. Nadelman ◽  
Paul Visintainer ◽  
Andrew Levin ◽  
...  
Keyword(s):  
Lyme Disease ◽  
Borrelia Burgdorferi ◽  
Acute Phase ◽  
Erythema Migrans ◽  
Skin Lesions ◽  
Enzyme Linked Immunosorbent Assay ◽  
Clinical Variables ◽  
Gender And Age ◽  
Serologic Assay ◽  
Single Lesion

ABSTRACT Erythema migrans, the most common manifestation of Lyme disease, has been associated with highly variable rates of seropositivity for antibodies to Borrelia burgdorferi. Differences in the sensitivities of serologic assays for the detection of these antibodies, however, may not be the only or even the primary explanation for this observation. We investigated the impacts of four clinical variables on seropositivity—the duration of erythema migrans, the presence of single versus multiple skin lesions, and the gender and age of the patient. In this analysis, three different serologic tests were performed on acute-phase sera from 175 untreated patients with culture-confirmed erythema migrans: the C6 single-peptide enzyme-linked immunosorbent assay (ELISA), a commercially available ELISA in which a whole-cell sonicate of B. burgdorferi was the antigen, and a two-tier procedure. Irrespective of the serologic test performed, the results showed that seropositivity rates increased with the duration of the erythema migrans for patients with single lesions (P < 0.001) but not for those with multiple skin lesions. The variability in seropositivity rates was greatest for the two-tier testing strategy, with a >6-fold-higher rate of seropositivity among patients with a single lesion of 22- to 30-day duration than among those whose skin lesion was of 1- to 7-day duration (85.7 versus 14.1%; P < 0.001). Rates of seropositivity by each of the testing methods were also significantly higher for patients with multiple skin lesions than for those with single lesions (P < 0.001). In contrast, seropositivity rates were not affected by either the gender or the age of the patient. Thus, in patients with erythema migrans, certain clinical variables such as the duration and number of skin lesions had a profound impact on seropositivity rates, irrespective of the serologic assay performed.

scholarly journals Outer Surface Protein C Peptide Derived from Borrelia burgdorferi Sensu Stricto as a Target for Serodiagnosis of Early Lyme Disease

2013 ◽  
Vol 20 (4) ◽  
pp. 474-481 ◽  
Author(s):  
Paul M. Arnaboldi ◽  
Rudra Seedarnee ◽  
Mariya Sambir ◽  
Steven M. Callister ◽  
Josephine A. Imparato ◽  
...  
Keyword(s):  
Lyme Disease ◽  
Borrelia Burgdorferi ◽  
Bacterial Species ◽  
Erythema Migrans ◽  
Early Infection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Attractive Alternative ◽  
Serum Samples ◽  
Peptide Epitope ◽  
Content Type

ABSTRACTCurrent serodiagnostic assays for Lyme disease are inadequate at detecting early infection due to poor sensitivity and nonspecificity that arise from the use of whole bacteria or bacterial proteins as assay targets; both targets contain epitopes that are cross-reactive with epitopes found in antigens of other bacterial species. Tests utilizing peptides that contain individual epitopes highly specific forBorrelia burgdorferias diagnostic targets are an attractive alternative to current assays. Using an overlapping peptide library, we mapped linear epitopes in OspC, a critical virulence factor ofB. burgdorferirequired for mammalian infection, and confirmed the results by enzyme-linked immunosorbent assay (ELISA). We identified a highly conserved 20-amino-acid peptide epitope, OspC1. Via ELISA, OspC1 detected specific IgM and/or IgG in 60 of 98 serum samples (62.1%) obtained from patients with erythema migrans (early Lyme disease) at the time of their initial presentation. By comparison, the commercially available OspC peptide PepC10 detected antibody in only 48 of 98 serum samples (49.0%). In addition, OspC1 generated fewer false-positive results among negative healthy and diseased (rheumatoid arthritis and positive Rapid Plasma Reagin [RPR+] test result) control populations than did PepC10. Both highly specific and more sensitive than currently available OspC peptides, OspC1 could have value as a component of a multipeptide Lyme disease serological assay with significantly improved capabilities for the diagnosis of early infection.

scholarly journals Epitope Length, Genospecies Dependency, and Serum Panel Effect in the IR6 Enzyme-Linked Immunosorbent Assay for Detection of Antibodies to Borrelia burgdorferi

2007 ◽  
Vol 14 (7) ◽  
pp. 875-879 ◽  
Author(s):  
Maria J. C. Gomes-Solecki ◽  
Luciana Meirelles ◽  
John Glass ◽  
Raymond J. Dattwyler
Keyword(s):  
Lyme Disease ◽  
Borrelia Burgdorferi ◽  
Immunosorbent Assay ◽  
Erythema Migrans ◽  
The United States ◽  
Enzyme Linked Immunosorbent Assay ◽  
Diagnostic Assay ◽  
Assay Sensitivity ◽  
Selection Of ◽  
Antigenic Epitopes

ABSTRACTIn the absence of erythema migrans, the basis for diagnosis of Lyme disease is the demonstration of an antibody response againstBorrelia burgdorferiin an appropriate clinical setting. The C6 enzyme-linked immunosorbent assay, based on the IR6 region of VlsE, has become widely used in both the United States and Europe. We mapped the antigenic epitopes of IR6 to a shorter sequence that is equivalent in sensitivity and specificity to the full-length IR6 25-residue peptide. In addition, we observed significant differences in sensitivity between serum panels (60 to 100%), indicating that the selection of the serum panels can shape the apparent overall sensitivity of the assay. Contrary to prior reports, the assay sensitivity is greater when the IR6 peptide is derived from the sequence of the same infectingBorreliagenospecies. Using our North American panels and the two panels obtained from European Lyme disease patients, we determined that the IR6 assay that is based on a single genospecies ofBorreliaspp. is not optimal for use as a universal diagnostic assay for Lyme disease.

scholarly journals Recombinant Chimeric Borrelia Proteins for Diagnosis of Lyme Disease

2000 ◽  
Vol 38 (7) ◽  
pp. 2530-2535 ◽  
Author(s):  
Maria J. C. Gomes-Solecki ◽  
John J. Dunn ◽  
Benjamin J. Luft ◽  
Jonathan Castillo ◽  
Daniel E. Dykhuizen ◽  
...  
Keyword(s):  
Lyme Disease ◽  
Borrelia Burgdorferi ◽  
Sensitivity And Specificity ◽  
Immunosorbent Assay ◽  
Enzyme Linked Immunosorbent Assay ◽  
Chimeric Proteins ◽  
Whole Cell ◽  
Assay Optimization

Current serologic Lyme disease tests use whole borrelia cells as the source of antigen. These assays are difficult to standardize and to optimize for sensitivity and specificity. To help solve these problems, we constructed a library of recombinant chimeric proteins composed of portions of key antigens of Borrelia burgdorferi. These proteins were then used to develop an enzyme-linked immunosorbent assay. We compared our assay with the most sensitive of three whole-cell borrelia assays. We found that the recombinant assay could detect antibodies significantly better from early Lyme disease sera (P < 0.05), and had the same sensitivity for late Lyme disease sera, as the most sensitive whole-cell borrelia assay. On potentially cross-reactive sera, the recombinant assay was more specific, but not significantly so, than the best whole-cell borrelia assay. Optimization of the recombinant assay offers the potential for a significant improvement in both sensitivity and specificity.

scholarly journals Enhanced Detection of Host Response Antibodies to Borrelia burgdorferi Using Immuno-PCR

2013 ◽  
Vol 20 (3) ◽  
pp. 350-357 ◽  
Author(s):  
Micah D. Halpern ◽  
Sunny Jain ◽  
Mollie W. Jewett
Keyword(s):  
Lyme Disease ◽  
Borrelia Burgdorferi ◽  
Host Response ◽  
Magnetic Beads ◽  
Disease Patient ◽  
Rank Correlation ◽  
Protein Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Content Type

ABSTRACTLyme disease is the fastest-growing zoonotic disease in North America. Current methods for detection ofBorrelia burgdorferiinfection are challenged by analysis subjectivity and standardization of antigen source. In the present study, we developed an immuno-PCR (iPCR)-based approach employing recombinantin vivo-expressedB. burgdorferiantigens for objective detection of a host immune response toB. burgdorferiinfection. iPCR is a liquid-phase protein detection method that combines the sensitivity of PCR with the specificity and versatility of immunoassay-based protocols. Use of magnetic beads coated with intact spirochetes provided effective antigen presentation and allowed detection of host-generated antibodies in experimentally infected mice at day 11 postinoculation, whereas host-generated antibodies were detected at day 14 by enzyme-linked immunosorbent assay (ELISA) and day 21 by immunoblotting. Furthermore, magnetic beads coated with recombinantB. burgdorferi in vivo-expressed antigen OspC or BmpA demonstrated positive detection of host-generated antibodies in mice at day 7 postinoculation with markedly increased iPCR signals above the background, with the quantification cycle (Cq) value for each sample minus the mean backgroundCqplus 3 standard deviations (ΔCq) being 4 to 10, whereas ΔCqwas 2.5 for intact spirochete-coated beads. iPCR demonstrated a strong correlation (Spearman rank correlation = 0.895,P< 0.0001) with a commercial ELISA for detection of host antibodies in human Lyme disease patient sera using theB. burgdorferiVlsE C6 peptide. In addition, iPCR showed potential applicability for direct detection of spirochetes in blood. The results presented here indicate that our iPCR assay has the potential to provide an objective format that can be used for sensitive detection of multiple host response antibodies and isotypes toB. burgdorferiinfection.

scholarly journals Development of a Multiantigen Panel for Improved Detection of Borrelia burgdorferi Infection in Early Lyme Disease

2015 ◽  
Vol 53 (12) ◽  
pp. 3834-3841 ◽  
Author(s):  
Lauren J. Lahey ◽  
Michael W. Panas ◽  
Rong Mao ◽  
Michelle Delanoy ◽  
John J. Flanagan ◽  
...  
Keyword(s):  
Lyme Disease ◽  
Borrelia Burgdorferi ◽  
Early Stage ◽  
Erythema Migrans ◽  
Disease Patient ◽  
High Specificity ◽  
The United States ◽  
Surface Proteins ◽  
Current Standard ◽  
Content Type

The current standard for laboratory diagnosis of Lyme disease in the United States is serologic detection of antibodies againstBorrelia burgdorferi. The Centers for Disease Control and Prevention recommends a two-tiered testing algorithm; however, this scheme has limited sensitivity for detecting early Lyme disease. Thus, there is a need to improve diagnostics for Lyme disease at the early stage, when antibiotic treatment is highly efficacious. We examined novel and established antigen markers to develop a multiplex panel that identifies early infection using the combined sensitivity of multiple markers while simultaneously maintaining high specificity by requiring positive results for two markers to designate a positive test. Ten markers were selected from our initial analysis of 62B. burgdorferisurface proteins and synthetic peptides by assessing binding of IgG and IgM to each in a training set of Lyme disease patient samples and controls. In a validation set, this 10-antigen panel identified a higher proportion of early-Lyme-disease patients as positive at the baseline or posttreatment visit than two-tiered testing (87.5% and 67.5%, respectively;P< 0.05). Equivalent specificities of 100% were observed in 26 healthy controls. Upon further analysis, positivity on the novel 10-antigen panel was associated with longer illness duration and multiple erythema migrans. The improved sensitivity and comparable specificity of our 10-antigen panel compared to two-tiered testing in detecting earlyB. burgdorferiinfection indicates that multiplex analysis, featuring the next generation of markers, could advance diagnostic technology to better aid clinicians in diagnosing and treating early Lyme disease.

scholarly journals The Borrelia burgdorferi 37-Kilodalton Immunoblot Band (P37) Used in Serodiagnosis of Early Lyme Disease Is the flaA Gene Product

1999 ◽  
Vol 37 (3) ◽  
pp. 548-552 ◽  
Author(s):  
Robert D. Gilmore ◽  
Rendi L. Murphree ◽  
Angela M. James ◽  
Sarah A. Sullivan ◽  
Barbara J. B. Johnson
Keyword(s):  
Lyme Disease ◽  
Borrelia Burgdorferi ◽  
Genomic Library ◽  
Disease Patient ◽  
Humoral Response ◽  
Immunoglobulin M ◽  
Leader Peptide ◽  
Serum Samples ◽  
Present Generation ◽  
Periplasmic Flagella

The 37-kDa protein (P37) of Borrelia burgdorferi is an antigen that elicits an early immunoglobulin M (IgM) antibody response in Lyme disease patients. The P37 gene was cloned from aB. burgdorferi genomic library by screening with antibody from a Lyme disease patient who had developed a prominent humoral response to the P37 antigen. DNA sequence analysis of this clone revealed the identity of P37 to be FlaA, an outer sheath protein of the periplasmic flagella. Recombinant P37 expression was accomplished inEscherichia coli by using a gene construct with the leader peptide deleted and fused to a 38-kDa E. coli protein. The recombinant antigen was reactive in IgM immunoblots using serum samples from patients clinically diagnosed with early Lyme disease that had been scored positive for B. burgdorferi anti-P37 reactivity. Lyme disease patient samples serologically negative for theB. burgdorferi P37 protein did not react with the recombinant. Recombinant P37 may be a useful component of a set of defined antigens for the serodiagnosis of early Lyme disease. This protein can be utilized as a marker in diagnostic immunoblots, aiding in the standardization of the present generation of IgM serologic tests.

scholarly journals Serologic Evaluation of Patients from Missouri with Erythema Migrans-Like Skin Lesions with the C6 Lyme Test

2006 ◽  
Vol 13 (10) ◽  
pp. 1170-1171 ◽  
Author(s):  
Mario T. Philipp ◽  
Edwin Masters ◽  
Gary P. Wormser ◽  
Wayne Hogrefe ◽  
Dale Martin
Keyword(s):  
New York ◽  
Lyme Disease ◽  
Erythema Migrans ◽  
New York State ◽  
Skin Lesions ◽  
Etiologic Agent ◽  
Acute Stage ◽  
Enzyme Linked Immunosorbent Assay ◽  
Convalescent Phase ◽  
South Central

ABSTRACT Southern tick-associated rash illness (STARI), also known as Masters disease, affects people predominantly in the Southeast and South Central United States. These patients exhibit skin lesions that resemble erythema migrans (EM), the characteristic skin lesion in early Lyme disease. The etiology of STARI remains unknown, and no serologic test is available to aid in its diagnosis. The C6 Lyme enzyme-linked immunosorbent assay was used to evaluate coded serum specimens from patients with STARI at two laboratory sites. The specimens tested at one site consisted of acute- and convalescent-phase samples that were obtained from nine STARI patients from Missouri and from one patient with documented Borrelia lonestari infection who acquired this infection in either North Carolina or Maryland. All of these samples were C6 negative. Seventy acute- or convalescent-phase specimens from 63 STARI patients from Missouri were C6 tested at the second site. All but one of these STARI specimens were also negative. In contrast, of nine acute- and nine convalescent-phase serum specimens obtained from culture-confirmed Lyme disease patients with EM from New York state, seven were C6 positive at the acute stage, and eight were positive at convalescence. The C6 test is negative in patients with STARI, providing further evidence that B. burgdorferi is not the etiologic agent of this disease.

Comparative evaluation of three different ELISA methods for the diagnosis of early culture-confirmed Lyme disease in Italy

2005 ◽  
Vol 54 (4) ◽  
pp. 361-367 ◽  
Author(s):  
Antonella Marangoni ◽  
Monica Sparacino ◽  
Francesca Cavrini ◽  
Elisa Storni ◽  
Valeria Mondardini ◽  
...  
Keyword(s):  
Immune Response ◽  
Lyme Disease ◽  
Borrelia Burgdorferi ◽  
False Positive ◽  
Comparative Evaluation ◽  
Endemic Area ◽  
Blood Donors ◽  
Erythema Migrans ◽  
Serum Samples ◽  
Positive Reactivity

In this study the raising and development of the immune response to Borrelia burgdorferi infection in 45 Italian patients suffering from culture-confirmed Lyme borreliosis erythema migrans was investigated. A total of 95 serially collected serum samples were tested by using three different commercial ELISAs: recomWell Borrelia (Mikrogen), Enzygnost Borreliosis (DADE Behring) and Quick ELISA C6 Borrelia (Immunetics). The sensitivities of the ELISAs were as follows: Enzygnost Borreliosis IgM, 70.5 %; Quick ELISA C6 Borrelia, 62.1 %; recomWell Borrelia IgM, 55.7 %; recomWell Borrelia IgG, 57.9 %; and Enzygnost Borreliosis IgG, 36.8 %. In order to compare the specificity values of the three ELISAs, a panel of sera obtained from blood donors (210 samples coming from a non-endemic area and 24 samples from an endemic area) was tested, as well as sera from patients suffering from some of the most common biological conditions that could result in false-positive reactivity in Lyme disease serology (n = 40). RecomWell Borrelia IgG and recomWell Borrelia IgM were the most specific (97.1 % and 98.9 %, respectively), followed by Quick ELISA C6 Borrelia (96.7 %). Enzygnost Borreliosis IgG and IgM achieved 90.1 % and 92.3 % specificity, respectively. Sera that gave discrepant results when tested by the three ELISAs were further analysed by Western blotting.

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