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PubMed2005

Recombinant antigens for serodiagnosis of cysticercosis and echinococcosis.

Diagnosis of cysticercosis/echinococcosis is primarily based on imaging techniques. These imaging techniques are sometimes limited by the small size of visualized lesions and atypical images, which are difficult to be distinguished from abscesses or neoplasms. Therefore, efforts have been directed toward identification and characterization of specific antigens of parasites for development of serodiagnostic method that can detect specific antibody. For cysticercosis, glycoproteins of 10-26 kDa in cyst fluid of Taenia solium have been widely accepted for serodiagnosis purpose. The glycoproteins consist of a very closely related family of 8-kDa proteins. We identified four genes (designated Ag1, Ag1V1, Ag2 and Ag2V1) encoding the 7- and 10-kDa polypeptides. Based on the antigenicities of these clones, Ag1V1 and Ag2 were chosen as ELISA antigens and the Ag1V1/Ag2 chimeric protein was expressed. The Ag1V1/Ag2 chimeric protein showed the similar sensitivity and specificity as the native glycoproteins. For alveolar echinococcosis, the 65-kDa protein of Echinococcus multilocularis protoscolices and Em18 has been considered as serodiagnostic antigens. The sensitivity and specificity of Em18 are very compatible to those of the recombinant 65-kDa protein. Recently, we demonstrated that Em18 was the proteolytic product of the 65-kDa protein following the action by cysteine proteinases. From the information of N-terminal amino acid sequences, molecular size and isoelectric point of Em18, recombinant Em18 ((349)K to (508)K of the 65-kDa protein, RecEm18) was expressed and evaluated for serodiagnostic value. RecEm18 has the potential for use in the differential serodiagnosis of alveolar echinococcosis.

Animals↗

PubMed1998

Analysis of the antigenic properties of the L. infantum Hsp70: design of synthetic peptides for specific serodiagnosis of human leishmaniasis.

The 70-kDa heat shock protein (Hsp70) is a major target of the humoral immune response during Leishmania infections. In the present paper, it is shown that 84.6% of sera from mucocutaneous leishmaniasis (MCL) patients and 78.9% of sera from visceral leishmaniasis (VL) patients reacted with the L. infantum Hsp70. The mapping of the antigenic determinants indicated that this protein is highly rich in linear B-epitopes. As the complete protein cannot be used for specific serodiagnosis of VL because it is also recognized by the sera from Chagas' disease patients, a search for specific epitopes recognized by leishmaniasis patients was undertaken. A 50-mer synthetic peptide, located in the most divergent region of the protein, was found to be recognized by a high percentage of leishmaniasis sera and not recognized by chagasic sera. Such a tool would be particularly useful for serodiagnosis of leishmaniasis in geographical areas where mixed infections with Trypanosoma cruzi and Leishmania occurs.

Amino Acid Sequence↗

PubMed2003

Development of competitive ELISA for serodiagnosis on African horsesickness virus using baculovirus expressed VP7 and monoclonal antibody.

VP7, the sero-group common antigen, of African horsesickness virus (AHSV-4) was expressed in insect cells by recombinant baculovirus. To develop a specific diagnostic method, monoclonal antibody (Mab) against VP7 was prepared and investigated as diagnostic reagent with the baculovirus expressed VP7. However, the Mab against VP7 of AHSV cross-reacted with Chuzan virus by the indirect immunofluorescence assay (IFA), confirming the presence of conserved domain of VP7 among Orbiviruses. This study describes two types of ELISA; Mab linked indirect (I-ELISA) and competitive-ELISA (C-ELISA) using baculovirus expressed VP7 as an antigen. These ELISAs were compared for serodiagnosis of AHSV showing that C-ELISA was more specific than I-ELISA. The results indicated that C-ELISA is applicable to serodiagnosis of AHSV regardless of serotypes.

African Horse Sickness↗

PubMed1996

Serodiagnosis of tuberculosis: detection of mycobacterial antibodies and antigens.

SETTING: The diagnosis of tuberculosis is based primarily on identification of mycobacteria and on clinical evidence. Recently, serological studies have been widely used experimentally as a diagnostic approach. OBJECTIVE: The aim of our study was to optimize serodiagnosis of tuberculosis by detecting mycobacterial antigens and antibodies in sera from patients with lung tuberculosis, non-related diseases and healthy controls. DESIGN: Mycobacterium tuberculosis H37Rv was disintegrated by pressure. Cell walls were extracted with 3 M KCL and were subjected to gel filtration in Toyopearl gel. Immune sera were prepared by immunization of rabbits with cell wall material. Anti H37Rv antibodies were purified by affinity chromatography. The reagents obtained were used to detect serum antibodies and antigens (following immune complex dissociation) using ELISA. RESULTS: Using fraction 6 of cell wall extract, antibodies were detected in 72.2% of TB patients; there were no positive reactions in control subjects. By use of affinity-purified antibodies, antigens were detected in 77.1% of TB patients, 10% of patients with unrelated diseases and 6.7% of healthy controls. CONCLUSION: Effective serodiagnosis of tuberculosis can be achieved only by combining detection of both circulating antibodies and antigens using highly specific purified reagents and immune complex-dissociated sera.

Antibodies, Bacterial↗

PubMed1996

Purification and characterisation of isocitrate dehydrogenase and malate dehydrogenase from Mycobacterium tuberculosis and evaluation of their potential as suitable antigens for the serodiagnosis of tuberculosis.

SETTING: Enzymes from Mycobacterium tuberculosis are potent antigens and might thus be of interest in the serodiagnosis of tuberculosis. OBJECTIVE: The purpose of the study was to purify and characterize the two enzymes isocitrate dehydrogenase (IDH) and malate dehydrogenase (MDH) from, M. tuberculosis and to evaluate their potential in the serodiagnosis of tuberculosis. DESIGN: The two enzymes were analysed for specificity by electrophoresis and then purified by means of affinity chromatography using reactive dyes and ion exchange chromatography. The two isolated enzyme fractions were analysed by ELISA, using antisera against related organisms. They were then tested as antigens in ELISA together with sera from tuberculous patients and controls. RESULTS: The electrophoretical analyses showed that the two enzymes each differed markedly from the corresponding enzymes of other mycobacteria. The serological analyses, however, could not distinguish between either IDH or MDH from other mycobacteria, but organisms of other genera, such as Nocardia, gave much weaker responses. When IDH and MDH were tested with sera from tuberculous patients and controls the former gave clearly higher optical density values than the latter. CONCLUSION: The enzymes/antigens IDH and MDH may be of value in developing a serological test for tuberculosis. The latter fraction seemed particularly capable of discriminating patients from controls.

Animals↗

PubMed2003

Immune complex antigens as a tool in serodiagnosis of kala-azar.

The 63 kDa surface antigen of Leishmania promastigotes is one of the most important virulent factors in establishing the host parasite relationship. This glycoprotein is revealed by surface iodination study as well as by metabolic labeling and immunoblot methods. In search of this specific antigen for serodiagnosis, immune complexes (ICs) were isolated from kala-azar patient sera and analysed by SDS-PAGE and Western immunoblotting. The immunoblot of kala-azar IC with patient sera, anti-promastigote sera and anti gp63 sera detected the major antigen of 55 kDa. This recognition suggests that 55 kDa antigen and gp63 have common antigenic epitope(s). Normal IC did not react with anti gp63 sera indicating absence of this antigen in normal IC. To confirm the parasitic origin of the 55 kDa antigen of kala-azar IC, in vitro IC was formed with parasite antigen and acid dissociated kala-azar IC antibody. This indicated the antigenic similarity of the 55 kDa antigen and gp63 antigen of the parasite. This also suggested that the former antigen may have been processed from gp63. In summary, identification of parasite antigen (55 kDa) in IC of kala-azar patients' sera may be useful in developing a serodiagnostic assay of visceral leishmaniasis. Several other antigens are visualized in kala-azar IC when developed with patient sera. But specificity and efficacy of these antigens have not yet been evaluated in serodiagnosis of the disease.

Animals↗

PubMed2003

Identification and evaluation of LPS antigen for serodiagnosis of uveitis associated with leptospirosis.

Leptospirosis is a widespread zoonotic disease that affects all mammals in different parts of the world. Though there are many commercial kits available for the diagnosis of systemic leptospirosis, the nature of the antigen has not been described. Therefore, identification of a specific antigen is important. Since ocular involvement in leptospirosis has been reported, there is a need to identify and characterize the leptospiral antigen for diagnosis of uveitis associated with past leptospiral infection (leptospiral uveitis) and for confirming the clinical diagnosis. Seven-day-old culture of Leptospira biflexa serovar Patoc was used for preparing the antigen. The present study included serum samples from 81 patients with clinical criteria for leptospiral uveitis, 15 cataract controls and 15 non-leptospiral uveitis controls. Serum samples were assayed by ELISA using our antigenic preparation and by a microscopic agglutination test (MAT) using 19 serovars. The antigen prepared had 280 micro g LPS ml(-1) and no detectable amount of protein. Silver-staining of SDS-PAGE for protein and LPS, dot blot and Western blot analysis and proteinase K and periodate treatment showed that LPS (13-21 kDa and 28 kDa) in our preparation was the relevant antigen for serodiagnosis. IgG antibodies showed reactivity in both leptospiral uveitis patients and controls. However, on the basis of IgM response to LPS, 48 % of the leptospiral uveitis patients were significantly positive compared with controls; 58 % of leptospiral uveitis patients and none of the controls were positive for MAT. When MAT and IgM ELISA results were considered together, 77 % were significantly positive. LPS is identified as a candidate antigen for serodiagnosis of leptospiral uveitis and has sensitivity and specificity of 48 and 90 %, respectively, in ELISA for IgM antibodies. Confirmation of clinical diagnosis with a specific laboratory test would help to initiate the most appropriate treatment for leptospiral uveitis.

Adult↗

PubMed1998

Rapid serodiagnosis of Mycobacterium avium-intracellulare complex infection by ELISA with cord factor (trehalose 6, 6'-dimycolate), and serotyping using the glycopeptidolipid antigen.

Mycobacterium avium-intracellulare complex (MAC) is one of the most important opportunistic pathogens, particularly in patients with acquired immunodeficiency syndrome (AIDS). The aim of this study was to determine whether an enzyme-linked immunosorbent assay (ELISA) using trehalose 6,6'-dimycolate (TDM) as an antigen can be used for the rapid serodiagnosis of MAC infection. We also identified MAC serotypes by ELISA using serotype-specific glycopeptidolipid (GPL) antigen. To confirm our findings, the thin-layer chromatographic (TLC) behavior of serotype-specific GPL of the strains isolated from MAC-infected patients was also tested. Forty patients infected with MAC and 30 healthy controls were tested. Thirty-two of the 40 MAC-infected patients had higher titers of serum antibodies against MAC TDM than against MTB TDM, while all 30 healthy control sera were unreactive to MAC TDM and MTB TDM. Results of the GPL ELISA indicated that 20 of the 40 MAC-infected patients' sera were reactive against serotype 4 GPL, 3 against serotype 8 GPL, and 1 against serotype 16 GPL. A TLC analysis of the GPL of the 40 MAC isolates showed that 16 strains were of serotype 4, 5 of serotype 8, and 2 of serotype 16. Results of the GPL ELISA were in good accord with those of the TLC analysis for most patients. Our findings suggest that ELISA using TDM is useful for rapid serodiagnosis of MAC infection, and that complementary ELISA testing using serotype-specific GPL gives additional detailed information concerning MAC serotypes.

Adult↗

PubMed2005

Evaluation of Taenia solium and Taenia crassiceps cysticercal antigens for the serodiagnosis of neurocysticercosis.

We evaluated the usefulness of seven cysticercal antigen extracts, four from Taenia solium cysticerci (whole parasite-TsoW, membrane-TsoMe, vesicular fluid-TsoVF and scolex-TsoSc) and three from T. crassiceps cysticerci (whole parasite-TcraW, membrane-TcraMe and vesicular fluid-TcraVF), for serodiagnosis of neurocysticercosis with an enzyme-linked immunosorbent assay (ELISA). Cysticercus-specific IgG were screened in serum samples from 23 patients with neurocysticercosis, 32 patients with other infections and 48 healthy persons. The best results were obtained with the TsoVF-ELISA (91.3% sensitivity; 96.2% specificity) and TcraVF-ELISA (91.3% sensitivity; 95% specificity). The ELISA done with whole parasite and membrane extracts from cysts of T. solium and T. crassiceps and the scolex extract from T. solium cysts showed a low performance in terms of sensitivity, ranging from 47.8% to 73.9%. None of the antigen preparations from T. solium and T. crassiceps cysticerci used in this study showed outstanding performance for the serodiagnosis of neurocysticercosis. However, considering the results obtained with the seven antigen preparations, vesicular fluid from T. solium and T. crassiceps cysticerci may be useful for detecting specific antibodies in sera from patients with neurocysticercosis.

Animals↗

PubMed1996

IgG avidity in the serodiagnosis of acute Toxoplasma gondii infection: a multicenter study.

OBJECTIVE: To evaluate the validity of the IgG avidity test in the serodiagnosis of acute T. gondii infection; to verify the maturation of IgG avidity during the course of infection; to observe whether the kinetics of IgG maturation could be affected by antibiotic treatment. METHODS: Serial serum samples, collected in three Italian hospitals (Perugia, Treviso and Bologna), from untreated and antibiotic-treated patients with primary toxoplasmic infection, were assayed for IgG avidity, and IgM and IgA positivity. Single serum samples from patients at different stages of infection were assayed for IgG avidity and the results were correlated to the likely stage of infection. RESULTS: The IgG avidity value increased from 3.5% in the first month to 38.7% 1 year from the onset of infection. Antibiotic-treated patients showed significantly different values of IgG avidity at 2 and 4 months after the onset of infection. In single serum samples the IgG avidity values correlated with the likely stage of infection. CONCLUSIONS: The IgG avidity test was confirmed as a useful tool in the serodiagnosis of acute T. gondii infection and could be predictive of the stage of infection. Antibiotic treatment may affect the kinetics of the maturation of IgG avidity.

Journal Article↗

PubMed1997

Serodiagnosis of tuberculosis and leprosy by enzyme immunoassay.

OBJECTIVE: To evaluate the use of serodiagnosis for tuberculosis and leprosy using mycobacterial antigen 38 kDa, with kits from Omega laboratories, to detect IgG by enzyme immunoassay (EIA). METHODS: The study population consisted of 58 patients with evidence of tuberculous infection (culture of Mycobacterium tuberculosis complex or microscopic evidence), of whom 23 had pulmonary and 35 had extrapulmonary disease. There were six subjects who had recently been treated for tuberculosis, 11 patients on treatment for leprosy and 137 patients suspected of having tuberculosis on clinical or radiologic grounds (without laboratory evidence). A control group comprised 35 healthy individuals or patients suffering from diseases other than tuberculosis. RESULTS: The tests showed that there was a significant difference in antibody levels between the patients with active pulmonary disease, extrapulmonary tuberculosis and leprosy in comparison with the control group (p<0.001). The sensitivities of the two tests together for proven pulmonary tuberculosis were 100% and 95.7% at 1.0--1.5 and >1.6 EIA cut-off points respectively, while the specificities were 88.5% and 100% at the same cut-off points. The sensitivities for extrapulmonary tuberculosis were 71.4% and only 51.4% at 1.0--1.5 and >1.6 EIA cut-off points. The test was positive in 30 (21.9%) of the 137 suspected patients, while 43 (31.4%) had an equivocal result and the remaining 64 (47.7%) suspects were definitely negative. There was again a significant difference in positivity rates between suspects and the control group. CONCLUSIONS: Omega IgG test is useful in the serodiagnosis of active pulmonary tuberculosis and leprosy, but less sensitive in extrapulmonary disease, particularly in children. Equivocal results may only add to the evidence of tuberculosis in early or minimal disease.

Journal Article↗

PubMed2003

Improved serodiagnosis of early Lyme borreliosis: immunoblot with local Borrelia afzelii strain.

To improve the serodiagnosis of Lyme borreliosis (LB) the performances of four tests were evaluated. An indirect immunofluorescent assay based on Borrelia burgdorferi s.s., enzyme-linked immunosorbent assay (ELISA) based on local isolates of Borrelia afzelii and B. burgdorferi s.s., and immunoblot (IB) of B. afzelii were prepared. The serum panels contained 214 serum samples: control group (n=120) and patients at different stages of LB (n=94). The specificity of IB was 96%, of in-house ELISA 93%, and of IFA 89%. In early LB the sensitivity of IFA was 36%, ELISA 67%, and IB 93%. In late-stage LB the sensitivity was: 72% for IFA, 80% for ELISA, and 94% for IB. Comparison of in-house and Behring ELISA showed that the sensitivity of the serological assay could be increased when the test was based on local Borrelia strains. IgM and IgG antibodies from sera of patients with early and late LB most frequently demonstrated reactivity to OspC. The other significant proteins in early LB were: p39, p41 in IgM IB, and p83/100, p39, Osp17 in IgG IB; in late LB: p39, p41 in IgM IB, and p83/100, Osp17, p21 and p43 in IgG IB. Using IB based on local B. afzelii isolates improves the serodiagnosis of early LB in our geographical region.

Adult↗

PubMed1990

[Serodiagnosis of scrub typhus by dot-blot method].

Microimmunofluorescence (IF) and immunoperoxidase tests are generally used for the serodiagnosis of scrub typhus. While these tests give satisfactory results in the hands of experienced personnel, they can be troublesome for inexperienced technicians. To develop a simpler diagnostic method, dot-blot assay was examined in this study. Six antigenically distinctive strains of Gilliam, Karp, Kato, Shimokoshi, Kawasaki and Kuroki of Rickettsia tsutsugamushi and a strain of Rickettsia sibirica were dotted on a nitrocellulose sheet by a dot-blot instrument. The sheets were treated with patient sera, followed by peroxidase-conjugated anti-human immunoglobulin-antibody and then with the substrate of the enzyme, and the color development on the dots was compared by naked eye observation. By this procedure, anti-rickettsial antibody-positive patient sera showed clear color development on at least one, usually several dots, while the very faint color was observed by the treatment with antibody-negative sera. On the other hand, it was ascertained that the antigens on nitrocellulose sheets were stable for at least 4 months at room temperature. Therefore the diagnostic kits were prepared, and the practical application of this procedure for diagnosis of scrub typhus were tested in Shizuoka and Miyazaki Prefectural Pabulic Health Laboratories. The results indicated a very good comparability between the dot-blot assay and IF-tests, and this dot-blot method was ascertained as a simple and useful method for the scrub typhus serodiagnosis.

Adolescent↗

PubMed1997

Cloning and characterization of two recombinant Neospora protein fragments and their use in serodiagnosis of bovine neosporosis.

Bovine neosporosis causes fetal abortion and/or congenital neurologic disease in cattle. For the serodiagnosis of this parasitic disease, two immunodominant clones from a bovine Neospora lambda gt11 library were identified, characterized, and expressed as recombinant proteins for the development of an enzyme-linked immunosorbent assay (ELISA). These two clones, designated N54 and N57, were 29 and 20 kDa, respectively, when expressed as histidine fusion proteins from the pRSET expression vector. Antibodies to recombinant protein N54 recognized five major bands from a Neospora tachyzoite lysate with molecular masses of 97, 87, 77, 67, and 64 kDa. Antibodies to recombinant protein N57 recognized four primary bands with molecular masses of 34, 31, 30, and 28 kDa. When a defined "gold standard" panel of bovine sera from confirmed Neospora-positive and Neospora-negative cattle were characterized by immunoblotting, 57 of the 60 Neospora-positive serum samples recognized proteins with the molecular masses of the N54 heptuplet. Binding to the N57 quadruplet was more variable. The same gold standard panel was used to evaluate and compare an N54-based ELISA, an N57-based ELISA, and a whole-tachyzoite lysate-based ELISA. The sensitivities and specificities were 95 and 96% (N54 ELISA), 82 and 93% (N57 ELISA), and 74 and 93% (lysate ELISA). Thus, compared to the whole-tachyzoite lysate-based ELISA, both recombinant-protein-based ELISAs had higher sensitivities and higher or the same specificities and can be used to replace the whole-tachyzoite lysate ELISA for the serodiagnosis of bovine neosporosis.

Animals↗

PubMed2001

Serodiagnosis of human cysticercosis by using antigens from vesicular fluid of Taenia crassiceps cysticerci.

Neurocysticercosis (NC), caused by the presence of Taenia solium metacestodes in tissues, is a severe parasitic infection of the central nervous system with universal distribution. To determine the efficiency of enzyme-linked immunosorbent assay (ELISA) and immunoblot with antigens of T. crassiceps vesicular fluid (Tcra) compared to standard techniques (indirect immunofluorescence test [IFT] and complement fixation test [CFT]) using T. solium cysticerci (Tso) for the serodiagnosis of NC, we studied serum samples from 24 patients with NC, 30 supposedly healthy individuals, 76 blood bank donors, 45 individuals with other non-NC parasitoses, and 97 samples from individuals screened for cysticercosis serology (SC). The sensitivity observed was 100% for ELISA-Tso and ELISA-Tcra, 91.7% for the IFT, and 87.5% for the CFT. The specificity was 90% for ELISA-Tso, 96.7% for ELISA-Tcra, 50% for IFT, and 63.3% for CFT. The efficiency was highest for ELISA-Tcra, followed by ELISA-Tso, IFT, and CFT. Of the 23 samples from SC group, which were reactive to ELISA-Tso and/or ELISA-Tcra, only 3 were positive to immunblot-Tcra (specific peptides of 14- and 18-kDa) and to glycoprotein peptides purified from Tcra antigen (gp-Tcra), showing the low predictive value of ELISA for screening. None of the samples from the remaining groups showed specific reactivity in immunoblot-Tcra. These results demonstrate that ELISA-Tcra can be used as a screening method for the serodiagnosis of NC and support the need for specific tests for confirmation of the results. The immunoblot can be used as a confirmatory test both with Tcra and gp-Tcra, with the latter having an advantage in terms of visualization of the results.

Animals↗

PubMed2002

Usefulness of hydatid cyst fluid of Echinococcus granulosus developed in mice with secondary infection for serodiagnosis of cystic Echinococcosis in humans.

The aim of this work was to assess the usefulness of hydatid cyst fluid (HCF) of Echinococcus granulosus, obtained from mice experimentally infected with hydatid cyst tissue homogenates, for the serodiagnosis of cystic echinococcosis (CE) in humans. The sensitivity and specificity of HCF obtained from mice for the detection of immunoglobulin G (IgG) antibodies in the sera of CE patients were compared with those of HCF from sheep and/or from a human CE patient by using immunoblotting (IB) and an enzyme-linked immunosorbent assay (ELISA). HCFs obtained from three different host species all were highly useful for immunoblotting, and sera from 19 (95%) of 20 CE patients equally recognized the antigen B subunit (approximately 8 kDa). HCF from mice showed a cross-reaction with 9 of 20 alveolar echinococcosis (AE) sera (45%), whereas HCFs from two other host species cross-reacted with 14 of the AE sera (70%). Although 2 (10%) of 20 sera from neurocysticercosis (NCC) patients were false positive with HCF from both sheep and humans, none of these sera showed a positive reaction with HCF from mouse origin. ELISAs with HCFs from both mouse and sheep origins detected all 20 CE and AE sera; however, these ELISAs showed 45% (9 of 20) and 60% (12 of 20) false-positive reactions with 20 NCC sera, respectively. The presence of nonspecific human IgG in HCF obtained from a CE patient prevented us from applying it to the ELISA. HCF of E. granulosus, obtained from laboratory mice with a secondary infection with hydatid cyst tissue homogenates, appears to be highly useful for the serodiagnosis of CE in humans and may be useful in domestic animals.

Animals↗

PubMed1991

Serodiagnosis of parasitic diseases.

In this review on serodiagnosis of parasitic diseases, antibody detection, antigen detection, use of monoclonal antibodies in parasitic serodiagnosis, molecular biological technology, and skin tests are discussed. The focus at the Centers for Disease Control on developing improved antigens, a truly quantitative FAST-enzyme-linked immunosorbent assay, and the very specific immunoblot assays for antibody detection is highlighted. The last two assays are suitable for field studies. Identification of patient response in terms of immunoglobulin class or immunoglobulin G subclass isotypes or both is discussed. Immunoglobulin isotypes may asist in defining the stage of some diseases. In other instances, use of a particular anti-isotype conjugate may increase the specificity of the assay. Monoclonal antibodies have played important roles in antigen purification and identification, in competitive antibody assays with increased sensitivity and specificity, and in assays for antigen detection in serum, body fluids, or excreta. Molecular biological technology has allowed significant advances in the production of defined parasitic serodiagnostic antigens.

Animals↗

PubMed1994

Humoral immune response to outer surface protein C of Borrelia burgdorferi in Lyme disease: role of the immunoglobulin M response in the serodiagnosis of early infection.

We determined the humoral immune response to outer surface protein C (OspC) of Borrelia burgdorferi in patients with early or late manifestations of Lyme disease and investigated the use of this antigen in the serodiagnosis of early infection. The ospC gene from the low-passage human isolate 297, a North American B. burgdorferi strain, was used to make a recombinant maltose-binding protein (MBP)-OspC fusion protein for serologic tests. This gene showed 84 to 85% nucleotide sequence identity and 76 to 79% amino acid identity with ospC of B. burgdorferi B31 and 2591. The antibody responses to MBP-OspC were determined in serial sera from 15 patients with Lyme disease who were monitored for 4 to 12 years of illness, in single-serum samples from 189 patients with early or late manifestations of the disorder, and in serum samples from 106 control patients. Early in the infection, patients with erythema migrans or meningitis commonly had weak to strong immunoglobulin M (IgM) responses to OspC and sometimes weak to moderate IgG responses. Months to years later, weak to strong IgG reactivity with this protein was often apparent in patients with arthritis, but this response was weak or absent in patients with chronic neuroborreliosis. When acute- and convalescent-phase serum samples from patients with erythema migrans were tested for reactivity against MBP-OspC, the sensitivity of the IgM test was 73% and the specificity was 98%, with either enzyme-linked immunosorbent assay (ELISA) or Western blotting. We conclude that the majority of patients with Lyme disease have a prominent IgM response to OspC early in the illness, which is often followed by a prominent IgG response in patients with arthritis. For the serodiagnosis of early infection, the sensitivity and specificity of IgM ELISA and Western blotting were comparable or slightly improved when MBP-OspC was used as the antigen compared with tests in which spirochetal lysates were used.

Amino Acid Sequence↗