A first-tier rapid assay for the serodiagnosis of Borrelia burgdorferi infection.

BACKGROUND: The present recommendation for the serologic diagnosis of Lyme disease is a 2-tier process in which a serum sample with a positive or equivocal result by an enzyme-linked immunosorbent assay (ELISA) or immunofluorescent assay is then followed by supplemental testing by Western blot. Our laboratory has developed recombinant chimeric proteins composed of key Borrelia epitopes. These novel antigens are consistent and are easily standardized. METHODS: We adapted these recombinant proteins into a new immunochromatographic format that can be used as a highly sensitive and specific first-tier assay that can be used to replace the ELISA or immunofluorescent assay. RESULTS: This rapid test was equally sensitive (P>.05) and more specific (P<.05) than a frequently used commercial whole cell ELISA. The overall clinical accuracy achieved on agreement studies among 3 Lyme research laboratories on clinically defined serum panels was shown to be statistically equivalent to the commercial ELISA. The assay can detect anti-Borrelia burgdorferi antibodies in either serum or whole blood. CONCLUSION: This sensitive and specific rapid assay, which is suited for the physician's office, streamlines the 2-tier system by allowing the physician to determine if a Western blot is necessary at the time of the initial office visit.

Lyme disease: cause of a treatable peripheral neuropathy.

Peripheral nerve dysfunction was demonstrated in 36% of patients with late Lyme disease. Of 36 patients evaluated, 14 had prominent limb paresthesias. Thirteen of these had neurophysiologic evidence of peripheral neuropathy; neurologic examinations were normal in most. Repeat testing following treatment documented rapid improvement in 11 of 12. We conclude that this neuropathy, which is quite different from the infrequent peripheral nerve syndromes previously described in this illness, is commonly present in late Lyme disease. This neuropathy presents with intermittent paresthesias without significant deficits on clinical examination and is reversible with appropriate antibiotic treatment. Neurophysiologic testing provides a useful diagnostic tool and an important measure of response to treatment.

Lyme neuroborreliosis: central nervous system manifestations.

We evaluated 85 patients with serologic evidence of Borrelia burgdorferi infection. Manifestations included encephalopathy (41), neuropathy (27), meningitis (2), multiple sclerosis (MS) (6), and psychiatric disorders (3). We performed lumbar punctures in 53, brain MRI in 33, and evoked potentials (EPs) in 33. Only patients with an MS-like illness had abnormal EPs, elevated IgG index, and oligoclonal bands in the cerebrospinal fluid. Twelve of 18 patients with encephalopathy, meningitis, or focal CNS disease had evidence of intrathecal synthesis of anti-B burgdorferi antibody, compared with no patients with either MS-like or psychiatric illnesses, and only 2/24 patients with neuropathy. MRIs were abnormal in 7/17 patients with encephalopathy, 5/6 patients with an MS-like illness, and no others. We conclude that (1) intrathecal concentration of specific antibody is a useful marker of CNS B burgdorferi infection; (2) Lyme disease causes an encephalopathy, probably due to infection of the CNS; (3) MS patients with serum immunoreactivity against B burgdorferi lack evidence of CNS infection with this organism.

Cross-reactive antigenic domains of the flagellin protein of Borrelia burgdorferi.

The p41 flagellin of Borrelia burgdorferi is the most common antigen recognized by serum of patients with Lyme borreliosis. This antigen shares amino acid homology, particularly in the amino and carboxy termini, with periflagellar antigens found in other microorganisms including Treponema pallidum. We cloned and expressed the p41 open reading frame in Escherichia coli and expressed it both as TrpE fusion and full-length unfused proteins. Also, we generated deletion constructs of various portions of the gene. Sera from patients with late Lyme borreliosis and secondary syphilis were used to identify the recombinant proteins by immunoblot analysis. Sera from 26 patients with Lyme borreliosis, 20 with secondary syphilis and 10 controls were used to identify cross-reactive domains of the B. burgdorferi flagellin. The variable region (amino acids 131-234) of the protein was recognized by 59% (15/26) of patients with late Lyme borreliosis compared to 30% (6/20) of patients with secondary syphilis and no (0/10) control patients. It appears that cross-reactive epitopes between B. burgdorferi and T. pallidum extend to the variable region of the flagellin.

Treatment of Lyme borreliosis.

The infectious process of Lyme disease can appear as chronic dermatologic, rheumatologic, or neurologic. To rationally select a treatment regimen, the physician must have an appreciation of the clinical manifestations of the illness and of the systemic nature of the infection. The authors discuss the proper treatment protocols for each stage in the progression of Lyme disease.

Immunodiagnosis of Lyme borreliosis.

The clinical definition of Lyme disease depends on the epidemiologic association of signs and symptoms with a measureable immune response to B. burgdorferi. The dependence on the demonstration of an immune response to B. burgdorferi has made the understanding of this systemic spirochetosis critical for the physician when making a diagnosis.

New chemotherapeutic approaches in the treatment of Lyme borreliosis.

1. It was demonstrated that while B. burgdorferi may be sensitive to relatively small concentrations of penicillin and ceftriaxone, the organism is killed slowly. This implies that, as in syphilis, prolonged blood levels of these drugs may be necessary in order to ensure cure. In contrast, the activity of tetracycline is more rapid in its action but is more dependent on drug concentration achieved. Unfortunately, the MIC and MBC for some strains are at or above the peak level achieved under optimal conditions. 2. Increasing the concentrations of penicillin or ceftriaxone above the MIC for the organism has little effect on the rate of killing. In contrast, the killing by tetracycline can be augmented by increasing concentrations of the drug. 3. Ceftriaxone is more active than penicillin, as measured by MIC, against the five strains of B. burgdorferi tested. 4. Ceftriaxone was efficacious in the treatment of Lyme borreliosis, which was recalcitrant to penicillin therapy. In a randomized trial comparing ceftriaxone to high-dose penicillin therapy, ceftriaxone was significantly more efficacious than penicillin in the treatment of the late complications of Lyme borreliosis.

Infection with multiple strains of Borrelia burgdorferi sensu stricto in patients with Lyme disease.

OBJECTIVE: To assess human skin biopsy specimens from erythema migrans lesions for the presence of infection with multiple strains of the Lyme disease spirochete, Borrelia burgdorferi. DESIGN: Skin biopsy specimens were obtained prospectively from patients with erythema migrans. To determine allelic differences and strain identification of B burgdorferi, the biopsy specimens were analyzed by cold single-strand conformation polymorphism of an amplified fragment of the outer surface protein C (ospC) gene. Further single-strand conformation polymorphism patterns of amplified ospC genes from culture isolates were compared with polymerase chain reaction products obtained directly from erythema migrans biopsy specimens. SETTING: A private dermatology office and a university medical center outpatient department. PATIENTS: Sixteen patients presenting with erythema migrans. RESULTS: Two of the 16 patients in this cohort were infected with 2 B burgdorferi sensu stricto strains, as evidenced by 2 ospC alleles in their skin biopsy results. CONCLUSION: This is the first documented description of the existence of more than a single strain of B burgdorferi sensu stricto in a human specimen.

Antibiotic treatment of Lyme borreliosis.

Unlike most bacterial infections, where diagnosis is by identification of the causal organism, diagnosis of infection by Borrelia burgdorferi (Lyme's borreliosis) relies mostly upon indirect techniques. This situation has some short-comings. As long as no technology permits a microbiological diagnosis of this infection, controversy will exist as to the clinical symptoms and the criteria for the cure of the disease. Despite the lack of consensus upon both the clinical definition and the treatment of Lyme's borreliosis, it is widely agreed that the affection is best understood if regarded as a progressive general infectious disease. Indeed, following a bite with local infection, there occurs a fairly rapid dissemination of the spirochaetes. In vivo therapeutic trials have shown the potential effectiveness of beta-lactams and tetracyclines, but no treatment is considered universally effective. Most of the first trials were empirical, as antibiograms were not used. Antibiotic concentrations reached with some oral therapies are too low for the protection of certain sites such as the central nervous system. In vitro studies conducted on various strains of B. burgdorferi both in the US and in Europe are very enlightening. Among the more perplexing results of some of these studies, it is worth noting the high resistance rate of some B. burgdorferi strains to penicillin, reported by Johnson et al. and by Preac Mursic et al. Therapy for Lyme's borreliosis is discussed in light of both the in vivo and in vitro studies.

Lymphocyte subsets in women on low dose oral contraceptives.

Oral contraceptive users have been reported to have a higher incidence of viral, bacterial and fungal infections. This study was undertaken to try to elucidate some of the mechanisms responsible for this increased susceptibility to infection. Peripheral blood lymphocytes were labelled with murine monoclonal antibodies directed against T cells and the various lymphocyte subsets: helper cells, suppressor/cytotoxic cells, or natural killer cells. The lymphocytes were then analyzed on a Coulter Electronics Epics V fluorescent activated cell sorter (FACS). A total of 27 control and 33 oral contraceptive users were studied. In comparison to the control group, there was no significant difference between the two groups in percentage or absolute numbers of total T cells, helper cells, suppressor/cytotoxic cells or natural killer cells. This study suggests that the increase in herpes virus, C. trachomonas, candida, and other infections in women taking oral contraceptives is not related to alterations in the numbers of T lymphocyte subsets or in levels of NK cells.

Overview of the clinical manifestations of Borrelia burgdorferi infection.

Lyme disease, caused by the spirochete Borrelia burgdorferi, has classically been divided into three stages: erythema migrans; neurological or cardiac involvement; and arthritis. Rather than defining a set disease pattern, however, one should, more logically, conceptualize a progressive infection that may be localized or disseminated, acute or chronic. Erythema migrans, the earliest and most easily recognized manifestation of B burgdorferi infection, is an expanding annular erythematous skin lesion with a central clearing that develops soon after the bite of an infected ixodes tick. Musculoskeletal manifestations are common, with approximately one-half of untreated individuals developing arthritis. Of these, only 10% have chronic arthritis. Invasion of the central nervous system occurs as the infection disseminates hematogenously, with encephalitis, myelitis and meningopolyneuritis being the most severe results. Acute cardiac involvement is recognized in up to 8% of adult patients, and less often in children. Early antibiotic treatment of the infection is highly effective.

Failure of tetracycline therapy in early Lyme disease.

We describe the clinical courses of 5 patients with Lyme disease who developed significant late complications, despite receiving tetracycline early in the course of their illness. All 5 patients had been treated for erythema chronicum migrans with a course of tetracycline that met or exceeded current recommendations. The late manifestations of Lyme disease included arthritis, cranial nerve palsy, peripheral neuropathy, chronic fatigue, and changes in mental function. Our findings suggest that the use of tetracycline at a dosage of 250 mg, 4 times a day for 10 days, as a treatment for early Lyme disease should be reconsidered. To determine optimal therapy for early Lyme disease, a study that compares an increased dosage of tetracycline with alternative treatments is indicated.

Complications of Lyme borreliosis.

Lyme borreliosis is the multisystem infectious disease caused by the spirochete Borrelia burgdorferi. Complications of this infection can involve many organ systems, especially the skin, joints, nervous system, and heart. These manifestations may be acute, or evolve slowly over months or years. Diagnosis is not always straightforward, and is currently hampered by lack of a specific serologic assay. This review discusses the syndromes associated with Lyme borreliosis and addresses issues of diagnosis and treatment.

Inhibition of sensitization of T-cells by alpha-fetoprotein.

A two-phase model of allograft immunity was studied. In the first phase, specific immune T-cells were generated by incubation of responder cells with mitomycin-C treated allogeneic stimulator cells of the same H-2 type as mouse mastocytoma tumor cells. In the second step, the ability of the sensitized cells to kill Cr51-labelled P-815 mastocytoma cells was assayed. Alpha-fetoprotein (AFP) was shown to inhibit the generation of immune cytotoxic T-cells at low concentrations (1-100 ng/ml) when added at the beginning of the first phase. When added at the end of the first phase or in the second (killing) phase, AFP was found to have no significant effect on cytotoxicity, indicating that it did not inhibit the killer T-cell once it was generated.

Antigen-specific T-cell anergy in progressive Mycobacterium marinum infection in humans.

The cellular immune responses of 12 patients with culture-proved Mycobacterium marinum infection involving deep tissues were studied. Of the 12 patients, 11 failed to respond to M. marinum antigens. All of the patients had, however, normal blastogenic responses to concanavalin A, phytohemagglutinin, pokeweed mitogen, and the antigens tetanus toxoid and Candida albicans. In contrast, 9 of 14 healthy fishermen responded to the M. marinum antigens with significant in-vitro blastogenesis. Infection of deep tissues with M. marinum is associated with antigen-specific T-cell anergy to M. marinum antigens.