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R F Schell

Publications and source records attributed to R F Schell.

At least 37 records · Page 2Linked to original sources
PubMed1996

Detection of anti-Borrelia burgdorferi antibody responses with the borreliacidal antibody test, indirect fluorescent-antibody assay performed by flow cytometry, and western immunoblotting.

Borreliacidal antibodies participate in the resolution of Lyme disease by clearing Borrelia burgdorferi sensu lato from the host. Detection of borreliacidal antibodies is also valuable for determination of the specific serodiagnosis of Lyme disease. We show in this work that antibody detected by the borreliacidal antibody test did not correlate with antibody detected by the indirect fluorescent-antibody assay or Western immunoblotting. Detection of borreliacidal antibody decreased with elimination of the spirochete from the host in the presence or absence of therapy. By contrast, the antibody responses detected by the indirect fluorescent-antibody assay or Western immunoblotting remained elevated or continued to expand, respectively. This suggests that the borreliacidal antibody test is a prognostic indicator for clearance of the spirochete. Additional investigations with humans are needed to confirm the prognostic potential of the borreliacidal antibody test.

Animals↗
PubMed1996

Sensitivity and specificity of the borreliacidal-antibody test during early Lyme disease: a "gold standard"?

The serodiagnosis of early Lyme disease has been plagued with problems of sensitivity and specificity. We found that the flow-cytometric borreliacidal-antibody test had a sensitivity of 72% for the detection of patients with early Lyme disease. By contrast, the sensitivity of the enzyme immunofluorescence assay was 28%. The enhanced sensitivity of the borreliacidal-antibody test was due to the use of Borrelia burgdorferi 50772, which lacks OspA and OspB. When B. burgdorferi 297, which expresses both OspA and OspB, was used, the sensitivity of the borreliacidal-antibody test was 15%. Our results also showed that the borreliacidal-antibody test was specific. No borreliacidal activity was detected in normal sera or in sera from patients with mononucleosis, rheumatoid factor, or syphilis. These results demonstrate that the flow-cytometric borreliacidal-antibody test may be the laboratory "gold standard" for the serodiagnosis of Lyme disease.

Adolescent↗
PubMed1996

Macrophages exposed to Borrelia burgdorferi induce Lyme arthritis in hamsters.

The mechanism(s) by which Lyme arthritis is induced has not been elucidated. In this study, we showed that macrophages have a direct, effector role in the pathogenesis of Lyme arthritis. Severe destructive arthritis was induced in recipients of macrophages obtained from Borrelia burgdorferi-vaccinated and nonvaccinated hamsters exposed to Formalin-inactivated B. burgdorferi in vitro and then challenged with the Lyme spirochete. Swelling of the hind paws was detected within 8 h of infection, increased rapidly, and peaked at 21 h. This initial swelling decreased, and by day 4 only slight swelling was detected. Severe swelling of the hind paws was detected 8 days after infection and increased rapidly, with peak swelling occurring on day 11. Histopathologic examination affirmed that macrophages exposed to Formalin-inactivated spirochetes induced a severe destructive Lyme arthritis. The onset and severity of the severe destructive arthritis were dependent on the number of macrophages transferred. By contrast, macrophages not exposed to Formalin-inactivated B. burgdorferi failed to induce severe destructive arthritis in recipients after challenge with B. burgdorferi. Similarly, severe destructive arthritis was not detected in recipients of macrophages injected with spirochetal growth medium. Our results also showed that transferred macrophages could not protect hamsters from infection with B. burgdorferi, as spirochetes were readily recovered from their tissues when cultured. These findings demonstrate that macrophages exposed to B. burgdorferi are directly involved in the induction of Lyme arthritis.

Animals↗
PubMed1996

Rapid susceptibility testing for nontuberculosis mycobacteria using flow cytometry.

We demonstrated previously that susceptibility testing of Mycobacterium tuberculosis could be accomplished within 24 h after the organisms were incubated with antituberculosis agents by using fluorescein diacetate (FDA) staining and flow cytometry. Continued studies have now shown that assay suspensions containing M. avium, M. fortuitum, M. gordonae, or M. marinum incubated with various concentrations of ciprofloxacin, clarithromycin, erythromycin, kanamycin, rifampin, tobramycin hydrolyzed less FDA than drug-free controls. Suspensions of treated and nontreated mycobacteria could be easily differentiated at 6 and 24 h after the initiation of the susceptibility assays by using FDA staining and flow cytometry. In addition, multiplication of the mycobacteria was not required to discern differences between drug-free suspensions of mycobacteria and those treated with antimycobacterial agents. The flow cytometric assay is simple, reproducible, and rapid.

Drug Resistance, Microbial↗
PubMed1995

Involvement of CD4+ T lymphocytes in induction of severe destructive Lyme arthritis in inbred LSH hamsters.

We determined that Borrelia burgdorferi-specific CD4+ T lymphocytes are responsible for the development of severe destructive Lyme arthritis and affect the production of borreliacidal antibody. Severe destructive Lyme arthritis was readily evoked in immunocompetent inbred LSH hamsters vaccinated with a whole-cell preparation of Formalin-inactivated B. burgdorferi sensu stricto isolate C-1-11 in adjuvant when challenged with B. burgdorferi sensu stricto isolate 297. When vaccinated hamsters were depleted of CD4+ T lymphocytes by the administration of monoclonal antibody GK1.5 and challenged, they failed to develop severe destructive arthritis. Similarly, nonvaccinated hamsters with or without the depletion of CD4+ T lymphocytes failed to develop severe destructive arthritis. In addition, depleting CD4+ T lymphocytes impaired the development of borreliacidal antibody in vaccinated and nonvaccinated hamsters challenged with the Lyme borreliosis spirochete. These findings show that CD4+ T lymphocytes are important for the recognition of arthritogenic and protective antigens of B. burgdorferi sensu lato isolates. Additional studies are needed to define the mechanisms responsible for the development of severe destructive Lyme arthritis and the production of borreliacidal antibody.

Animals↗
PubMed1995

Borrelia burgdorferi-specific T lymphocytes induce severe destructive Lyme arthritis.

This is the first documentation that Borrelia burgdorferi-specific T lymphocytes are involved in the pathogenesis of Lyme arthritis. We present direct evidence that T lymphocytes obtained from inbred LSH hamsters vaccinated with a whole-cell preparation of Formalin-inactivated B. burgdorferi sensu stricto isolate C-1-11 in adjuvant conferred on naive recipient hamsters the ability to develop severe destructive arthritis when challenged with B. burgdorferi sensu stricto isolates C-1-11 and 297. By contrast, recipients infused with normal T lymphocytes and challenged with B. burgdorferi sensu stricto isolates C-1-11 and 297 failed to develop severe destructive arthritis. The T lymphocytes transferred were obtained from the lymph nodes of vaccinated and nonvaccinated hamsters by depleting B lymphocytes by using monoclonal antibody 14-4-4s (< 1% B lymphocytes by flow cytometric analysis). The enriched T lymphocytes showed enhanced proliferation to stimulation with concanavalin A and failed to respond to lipopolysaccharide. Moreover, only the enriched T lymphocytes from vaccinated hamsters proliferated on exposure to a whole-cell preparation of B. burgdorferi sensu stricto isolate C-1-11 in the presence of mitomycin-treated syngeneic antigen-presenting cells. These results demonstrate that B. burgdorferi-specific T lymphocytes primed by vaccination with a whole-cell preparation of inactivated B. burgdorferi sensu stricto isolate C-1-11 in adjuvant are involved in the development of severe destructive arthritis. Additional experiments are needed to define the precise mechanism(s) responsible for the development of Lyme arthritis.

Animals↗
PubMed1995

Abilities of OspA proteins from different seroprotective groups of Borrelia burgdorferi to protect hamsters from infection.

The ability of vaccination with recombinant OspA from six seroprotective groups of Borrelia burgdorferi sensu lato to induce protection against infection with homologous and other Lyme spirochetes was examined in hamsters. Antisera generated against the OspA proteins of B. burgdorferi sensu stricto S-1-10 and C-1-11 (seroprotective groups 1 and 2, respectively), Borrelia afzelii BV1 (seroprotective group 4), and Borrelia garinii LV4 (seroprotective group 5) were able to kill the homologous spirochete in vitro but not other isolates. Surprisingly, antisera against B. afzelii PKo (seroprotective group 6) and B. burgdorferi sensu lato LV5 (seroprotective group 3) OspA proteins were unable to kill the homologous organism, although LV5 OspA antisera killed the heterologous isolates S-1-10 and LV4. In vivo vaccination studies supported the in vitro findings, confirming that vaccination with a single OspA protein does not provide complete protection against challenge with all Lyme disease spirochetes. In addition, OspA antibodies from some isolates may not protect against the homologous isolate. The induction of protective antibodies against other B. burgdorferi proteins may be necessary to insure a comprehensive Lyme disease vaccine.

Amino Acid Sequence↗
PubMed1995

Rapid susceptibility testing of Mycobacterium tuberculosis (H37Ra) by flow cytometry.

The resurgence of tuberculosis has caused considerable effort to be focused on the development of rapid methods for determining the susceptibility of Mycobacterium tuberculosis to antimycobacterial agents. We demonstrated that susceptibility testing of M. tuberculosis can be accomplished rapidly by using flow cytometry. Results of tests were available within 24 h after M. tuberculosis organisms were incubated with ethambutol, isoniazid, rifampin, or streptomycin. The method was based on the ability of viable M. tuberculosis organisms to hydrolyze fluorescein diacetate (FDA) and the detection of fluorescent mycobacteria by flow cytometric analysis. The assay system also did not require multiplication of the mycobacteria. In contrast, M. tuberculosis organisms exposed to antimycobacterial agents hydrolyzed significantly less FDA. The use of flow cytometry and FDA staining shows considerable promise as a rapid method for obtaining susceptibility test results.

Antitubercular Agents↗
PubMed1995

Immune response in the hamster: definition of a novel IgG not expressed in all hamster strains.

A new IgG isotype is described in serum from Syrian hamsters. This 7S-IgG is called IgG3 and was isolated from IgG1 and IgG2 because of its great affinity for protein A. The unique antigenic determinants of IgG3 were identified with a specific rabbit antisera. IgG3 is the least expressed IgG subclass in Syrian hamsters, but serum levels increase more than 10-fold after immunization or infection. Although found in all tested outbred strains, IgG3 is expressed in only some of the commercially available inbred strains of Syrian hamsters. Five inbred hamster strains were examined, and in three strains (CB, LHC and MHA) IgG3 was not detected in normal serum or in immune serum, indicating serum levels at least 100-fold less than other normal inbred/outbred hamsters. The results of breeding experiments suggests a single gene defect is responsible for this non-expression of IgG3. Immunodeficiency was not associated with this IgG3 deficiency. Selective deficiencies of immunoglobulin classes/subclasses in experimental animals are rare. The evolution of a similar IgG3 deficiency in these three hamster strains during inbreeding suggests a novel and efficient mechanism for regulation of IgG3 synthesis in the Syrian hamster.

Animals↗
PubMed1994

Detection of borreliacidal antibodies by flow cytometry. An accurate, highly specific serodiagnostic test for Lyme disease.

BACKGROUND: Borreliacidal antibodies can be detected in serum samples from patients with early or late Lyme disease symptoms. When these serum samples are incubated with Borrelia burgdorferi and complement, spirochetes are rapidly killed. Detection of these antibodies can be used as a serodiagnostic test. METHODS: Individual serum samples containing IgM or IgG borreliacidal antibodies were Used to develop a method for detection using flow cytometry. An additional 10 case-defined Lyme disease serum samples and 10 normal serum samples were used to confirm appropriate flow cytometric parameters. To determine specificity, 157 normal serum samples and 104 potential cross-reactive serum samples were tested for borreliacidal activity and antibodies to B burgdorferi using indirect fluorescent antibody or enzyme immunoassay. RESULTS: Flow cytometry can be used to detect borreliacidal activity within 16 to 24 hours after incubation of B burgdorferi organisms. Lyme disease serum, and complement. Significant borreliacidal activity was detected in all Lyme disease serum samples. The percentages of positive normal serum samples were comparable (6% to 10%) using all three assays. In addition, the indirect fluorescent antibody and enzyme immunoassay identified 41 (39%) and 47 (45%) potential cross-reactive serum samples as positive, respectively. In contrast, significant borreliacidal activity was not detected in any potential cross-reactive serum samples. CONCLUSION: Detection of borreliacidal antibody, unlike indirect fluorescent antibody and enzyme immunoassay, is an accurate, highly specific serodiagnostic test for detection of Lyme disease.

Antibodies, Bacterial↗
PubMed1994

Lyme disease: laboratory diagnosis and serologic testing.

Although identified less than 20 years ago, Lyme disease has proved to be the most common tick-borne disease in the United States: some 10,000 cases were reported in 1992. In some cases the disease may be transitory and of little consequence but in others it may become chronic and severely disabling. Accurate diagnosis is, therefore, of great importance but, as this article shows, laboratory testing techniques still need improvement.

Antibodies, Bacterial↗
PubMed1994

Seroprotective groups of Lyme borreliosis spirochetes from North America and Europe.

Five distinct seroprotective groups among North American and European isolates of Borrelia burgdorferi sensu stricto, Borrelia garinii, and Borrelia afzelii have been identified using the in vitro borreliacidal assay. The predominant North American seroprotective group comprised isolate 297 and B. burgdorferi isolates from California, Illinois, Wisconsin, New York, and Texas. A second group was represented by isolate C-1-11. The majority of European isolates belonged to a seroprotective group composed of B. garinii. Another European group contained isolates classified genetically as genospecies group VS461 (B. afzelii). A fifth group, represented by isolate LV5, could kill both North American and European isolates of B. burgdorferi sensu lato. These results suggest that combinations of immunogenic protective proteins of spirochetes will be necessary to provide a comprehensive vaccine.

Animals↗
PubMed1994

Detection of borreliacidal antibody by using acridine orange and flow cytometry.

Borreliacidal antibody has been shown to be important for the serodiagnosis of Lyme disease and determination of immune status. Our results show that borreliacidal antibody can be rapidly and accurately detected by flow cytometry. Acridine orange was added to normal and immune sera containing Borrelia burgdorferi organisms in the presence and absence of complement prior to data acquisition by flow cytometry. The flow cytometric parameters of side scatter and detection of acridine orange fluorescence were used to determine events per minute (number of labeled spirochetes), percent shift in fluorescence (number of dead spirochetes), and mean channel fluorescence (intensity of fluorescence-labeled spirochetes) of acridine orange-labeled spirochetes. Borreliacidal antibody was detected as early as 4 h, with optimal detection 16 to 24 h after incubation of B. burgdorferi organisms with immune serum and complement. Our results also showed that complement was necessary for detection of borreliacidal antibody. Flow cytometry with acridine orange-labeled spirochetes provides a rapid means for detection of borreliacidal antibody.

Acridine Orange↗
PubMed1994

Differentiation of borreliacidal activity caused by immune serum or antimicrobial agents by flow cytometry.

We demonstrated that borreliacidal activity caused by immune serum and complement can easily be differentiated by flow cytometry from killing activity caused by antimicrobial agents that are commonly used for the treatment of Lyme disease. Assay suspensions containing normal or immune serum were incubated with Borrelia burgdorferi in the presence or absence of ceftriaxone, doxycycline, penicillin, and phosphomycin for 2, 8, 16, and 24 h. Samples containing killing activity were identified by using flow cytometry and acridine orange. In 30 min, the effects of immune serum and complement were easily distinguished from the killing of spirochetes by antimicrobial agents by adding fluorescein isothiocyanate-conjugated goat anti-hamster immunoglobulin. This simple procedure greatly enhanced the usefulness of the borreliacidal assay by eliminating a major source of false-positive reactions.

Acridine Orange↗
PubMed1994

Development of destructive arthritis in vaccinated hamsters challenged with Borrelia burgdorferi.

We present the first direct evidence that adverse effects, particularly severe destructive arthritis, can develop in vaccinated hamsters after challenge with Borrelia burgdorferi sensu lato isolates. Hamsters were vaccinated with a whole-cell preparation of Formalin-inactivated B. burgdorferi sensu stricto isolate C-1-11 in adjuvant. A severe destructive arthritis was readily evoked in vaccinated hamsters challenged with the homologous B. burgdorferi sensu stricto isolate C-1-11 before high levels of protective borreliacidal antibody developed. Once high levels of C-1-11 borreliacidal antibody developed, hamsters were protected from homologous challenge and development of arthritis. Vaccinated hamsters, however, still developed severe destructive arthritis when challenged with other isolates of the three genomic groups of B. burgdorferi sensu lato (B. burgdorferi sensu stricto isolate 297, Borrelia garinii isolate LV4, and Borrelia afzelii isolate BV1) despite high levels of C-1-11 specific borreliacidal antibody. Vaccines that contained whole spirochetes in adjuvant induced destructive arthritis, but this effect was not dependent on the isolate of B. burgdorferi sensu lato or the type of adjuvant. These studies demonstrate that caution is necessary when employing whole spirochetes in adjuvant for vaccination to prevent Lyme borreliosis. Additional studies are needed to identify the antigen(s) responsible for the induction and activation of arthritis and to define the immune mechanisms involved.

Adjuvants, Immunologic↗
PubMed1994

Ability of canine Lyme disease vaccine to protect hamsters against infection with several isolates of Borrelia burgdorferi.

We used flow cytometry to determine levels of borreliacidal antibodies in hamsters after vaccination with a commercially available canine Lyme disease vaccine. In addition, we evaluated the ability of vaccinated hamsters to resist infection with several isolates of Borrelia burgdorferi. Borreliacidal antibodies could be detected 1 week after a primary vaccination, peaked at weeks 3 to 5, and then rapidly declined. One week after a booster vaccination, borreliacidal activity was detected at a dilution of 1:10,240, and it decreased fourfold by week 10 after the booster vaccination. Vaccinated hamsters were protected against infection with < or = 10(6) B. burgdorferi 297 organisms during the peak borreliacidal response (5 weeks after primary vaccination or 2 weeks after booster vaccination). However, hamsters were not fully protected from development of Lyme arthritis when the titer of borreliacidal antibodies was < 1:5,120. In addition, no significant borreliacidal activity was induced against B. burgdorferi C-1-11, LV4, or BV1, which belong to three other seroprotective groups. These studies demonstrate that vaccination with the canine Lyme disease vaccine induces protective antibodies against B. burgdorferi 297. However, significant levels of borreliacidal antibodies are not produced until 5 weeks after vaccination, and protection is short-lived. In addition, no borreliacidal activity was induced against other isolates of B. burgdorferi. Because of this, the incorporation of multiple isolates or protein subunits may be necessary to increase the effectiveness of future vaccines.

Animals↗
PubMed1993

Characterization of the borreliacidal antibody response to Borrelia burgdorferi in humans: a serodiagnostic test.

An in vitro assay was used to characterize the borreliacidal activity of sera from Lyme disease patients. The mean percentage of killing was 23% with sera from patients with a single erythema migrans lesion, 42% from patients with multiple lesions, 58% from patients with Lyme arthritis of short duration, and 83% from patients with Lyme arthritis of long duration. Borreliacidal activity was abrogated when Lyme disease serum was treated with anti-human IgM or IgG1. In addition, human sera from Lyme arthritis patients containing borreliacidal antibody prevented the induction of Lyme arthritis in irradiated hamsters challenged with the Lyme spirochete. Removal of outer surface protein A antibodies from late Lyme disease sera caused reductions in the borreliacidal antibody titer. The results demonstrate an important role for borreliacidal antibody against infection with B. burgdorferi in humans and confirm that detection of borreliacidal antibody in human sera can be a specific serodiagnostic test for Lyme disease.

Animals↗