The structure and function of platelet integrins.

Integrins are a ubiquitous family of non-covalently associated alpha/beta transmembrane heterodimers linking extracellular ligands to intracellular signaling pathways [1] [Cell, 2002; 110: 673]. Platelets contain five integrins, three beta1 integrins that mediate platelet adhesion to the matrix proteins collagen, fibronectin and laminin, and the beta3 integrins alphavbeta3 and alphaIIbbeta3 [2] [J Clin Invest, 2005; 115: 3363]. While there are only several hundred alphavbeta3 molecules per platelet, alphavbeta3 mediates platelet adhesion to osteopontin and vitronectin in vitro [3] [J Biol Chem, 1997; 272: 8137]; whether this occurs in vivo remains unknown. By contrast, the 80,000 alphaIIbbeta3 molecules on agonist-stimulated platelets bind fibrinogen, von Willebrand factor, and fibronectin, mediating platelet aggregation when the bound proteins crosslink adjacent platelets [2] [J Clin Invest, 2005; 115: 3363]. Although platelet integrins are poised to shift from resting to active conformations, tight regulation of their activity is essential to prevent the formation of intravascular thrombi. This review focuses on the structure and function of the intensively studied beta3 integrins, in particular alphaIIbbeta3, but reference will be made to other integrins where relevant.

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.

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.

Current aspects of Lyme disease and other Borrelia burgdorferi infections.

Lyme disease, acrodermatitis chronica atrophicans, and borrelial lymphocytoma are caused by species of the spirochete Borrelia burgdorferi. Lyme disease has emerged as the leading vector-borne infectious disease in the United States. This article presents a current review of these entities.

Bloodstream invasion in early Lyme disease: results from a prospective, controlled, blinded study using the polymerase chain reaction.

PURPOSE: The purposes of this study were to determine (1) the optimal techniques for and potential diagnostic usefulness of the polymerase chain reaction (PCR) in early Lyme disease, and (2) the true frequency and clinical correlates of PCR-documented blood-borne infection in the dissemination of Lyme disease. PATIENTS AND METHODS: We performed a prospective, controlled, blinded study of PCR, culture, and serology on fractionated blood samples from 105 patients; 76 with physician-diagnosed erythema migrans and 29 controls. Clinical characteristics of the patients were obtained with a standardized data entry form and correlated with results of the laboratory studies. RESULTS: Only 4 of the 76 (5.3%) patients with erythema migrans were culture positive; however, 14 of 76 (18.4%) had spirochetemia documented by PCR of their plasma. None of 29 controls were PCR or culture positive (P = 0.007, versus patients). PCR-documented spirochetemia correlated with clinical evidence of disseminated disease; 10 of 33 patients (30.3%) with systemic symptom(s) were PCR positive compared to 4 of 43 (9.3%) without such evidence (P = 0.02). PCR positivity was more frequent among patients with each of four specific symptoms: fever, arthralgia, myalgia, and headache (all P < 0.05). A higher total number of symptoms (median 2.5 in PCR-positive patients versus 0 in PCR-negative controls; P < 0.01) and the presence of multiple skin lesions (37.5% of patients with multiple, versus 13.3% of patients with single lesions [P = 0.04] were also correlated with PCR positivity. Patients with both systemic symptoms and multiple skin lesions had a 40% PCR-positivity rate; however, 4 of 42 (9.5%) asympatomatic patients with only single erythema migrans lesions were also PCR positive. In multivariate analysis using logistic regression, the number of systemic symptoms was the strongest independent predictor of PCR positivity (P = 0.004). CONCLUSIONS: PCR detection of Borrelia burgdorferi is at least three times more sensitive than culture for identifying spirochetemia in early Lyme disease and may be useful in rapid diagnosis. PCR positivity significantly correlates with clinical evidence of disease dissemination. Bloodstream invasion is an important and common mechanism for the dissemination of the Lyme disease spirochete.

Cultivation of Borrelia burgdorferi from human tick bite sites: a guide to the risk of infection.

BACKGROUND: The risk of acquiring Lyme disease has been evaluated by xenodiagnostic procedures with laboratory strains of Borrelia burgdorferi and laboratory-reared Ixodes ticks, or by clinical trials in which diagnosis was based on clinical findings, culture, or serologic tests. OBJECTIVE: Our purpose was to determine the risk of infection from tick bites in a natural setting in which wild strains of B. burgdorferi were involved, by a biopsy culture technique. METHODS: Skin biopsy specimens were obtained from Ixodes scapularis tick bite sites, processed, and examined for the presence of B. burgdorferi. RESULTS: B. burgdorferi was cultivated from only 2 of 48 skin biopsy specimens. In both instances duration of tick attachment was approximately 24 hours. CONCLUSION: In a hyperendemic region for Lyme disease the risk of infection after a deer tick bite appears to be low, particularly if the tick has been attached for less than 24 hours.

Laboratory tests for Lyme disease.

The laboratory diagnosis of Lyme disease has been problematic because of the lack of sufficient sensitivity and specificity of the many tests used as well as the lack of interlaboratory standardization in the performance of these tests. Although helpful, the laboratory tests can only serve to support the clinical findings in making the diagnosis of active Lyme disease, the exception being a positive culture of Borrelia Burgdorferi.

Cultivation of Borrelia burgdorferi from the blood of two patients with erythema migrans lesions lacking extracutaneous signs and symptoms of Lyme disease.

BACKGROUND: We recently demonstrated that cultivation of Borrelia burgdorferi from skin biopsy specimens obtained from erythema migrans lesions was an efficacious procedure to confirm the diagnosis of Lyme disease. OBJECTIVE: Our purpose was to investigate the efficacy of our microbiologic technique on blood samples obtained from patients with Lyme disease and erythema migrans. METHODS: Whole blood samples were obtained from 52 patients with erythema migrans and early localized or early disseminated Lyme disease and placed into polystyrene tubes containing 6 ml of modified Barbour-Stoenner-Kelly medium, processed, and examined for B. burgdorferi by dark-field microscopy. RESULTS: B. burgdorferi was cultured from the blood of two of our 52 patients (4%). Clinically, both of these patients were considered to have early localized Lyme disease. CONCLUSION: The culture of B. burgdorferi from the blood of patients with early Lyme disease does not appear to be an efficacious procedure to confirm the diagnosis of Lyme disease. However, the demonstration of spirochetemia in patients with erythema migrans without any extracutaneous evidence of disseminated illness does have therapeutic significance.

Lyme disease.

Years before the spirochetal etiology of Lyme disease was determined, the effectiveness of antibiotic treatment for erythema chronicum migrans had been established. Revisions in antibiotic treatment have evolved in concert with a growing understanding of the pathogenesis of Lyme disease. Current treatment recommendations are discussed.

Lyme disease.

Lyme disease is a multisystem disorder caused by the spirochete Borrelia burgdorferi. It is transmitted to human and animal hosts primarily by ticks of the Ixodes ricinis complex. Recognition of its characteristic skin and eye manifestations facilitates diagnosis and treatment.

Failure of Borrelia burgdorferi to survive in the skin of patients with antibiotic-treated Lyme disease.

BACKGROUND: Borrelia burgdorferi has been cultivated from clinically normal skin (previous erythema migrans sites) after antibiotic therapy for Lyme disease. OBJECTIVE: We investigated the possibility of similar findings in 13 of our patients with antibiotic-treated Lyme disease from whom B. burgdorferi was cultivated from their erythema migrans lesions before antibiotic therapy. METHODS: After treatment with doxycycline or a combination of amoxicillin and probenecid, skin biopsy specimens were obtained from clinically normal skin adjacent to the previous biopsy sites and cultured. RESULTS: B. burgdorferi was not cultivated from these posttreatment biopsy sites. CONCLUSION: The failure of B. burgdorferi to survive in the former erythema migrans sites of our antibiotic-treated patients, as well as their favorable clinical response, supports the use of doxycycline or combined amoxicillin and probenecid in the treatment of early Lyme disease but does not preclude the survival of the organism in other tissues.

Cultivation of Borrelia burgdorferi from erythema migrans lesions and perilesional skin.

Skin biopsy specimens from the peripheral aspect of erythema migrans lesions (site 1) and from clinically normal perilesional areas (site 2) were compared as sources of Borrelia burgdorferi. This spirochete was isolated from the skin of 18 of 21 (86%) patients with untreated early Lyme disease at one or both biopsy sites. Site 1 specimens were superior to site 2 specimens for the isolation of B. burgdorferi. Site 1 specimens from 18 (86%) patients were culture positive, and site 2 specimens from 12 (57%) patients were culture positive. For patients whose site 2 specimens were culture positive, site 1 specimens were also found to be culture positive. B. burgdorferi was isolated from two patients with atypical lesions and from two patients with erythema migrans lesions that were less than 5 cm in diameter. This study demonstrates that the cultivation of B. burgdorferi from skin biopsy specimens from cutaneous lesions thought to be erythema migrans can be an efficacious procedure for confirming the diagnosis of Lyme disease and that the spirochete is present in clinically normal appearing perilesional skin.

Clinical implications of delayed growth of the Lyme borreliosis spirochete, Borrelia burgdorferi.

Lyme borreliosis, a spirochetal infection caused by Borrelia burgdorferi, may become clinically active after a period of latency in the host. Active cases of Lyme disease may show clinical relapse following antibiotic therapy. The latency and relapse phenomena suggest that the Lyme disease spirochete is capable of survival in the host for prolonged periods of time. We studied 63 patients with erythema migrans, the pathognomonic cutaneous lesion of Lyme borreliosis, and examined in vitro cultures of biopsies from the active edge of the erythematous patch. Sixteen biopsies yielded spirochetes after prolonged incubations of up to 10.5 months, suggesting that Borrelia burgdorferi may be very slow to divide in certain situations. Some patients with Lyme borreliosis may require more than the currently recommended two to three week course of antibiotic therapy to eradicate strains of the spirochete which grow slowly.

Clinical and microbiologic findings in six patients with erythema migrans of Lyme disease.

The clinical course of six patients with erythema migrans of Lyme disease was viewed in the context of antibiotic susceptibility studies of their own Borrelia burgdorferi isolates. An initial poor response by two patients to antibiotic therapy suggested the possibility of B. burgdorferi resistance to these agents. Minimum bactericidal concentration determinations of eight antimicrobial agents against these isolates did not support this suggestion.

Cutaneous manifestations of Lyme borreliosis.

Erythema migrans is the distinctive cutaneous marker of Lyme borreliosis. The clinical picture is variable but at some point in its evolution, erythema migrans presents as a red, centrifugally expanding, annular plaque. Erythema migrans may appear as a solitary lesion or in multiplicity. It may be accompanied by extra cutaneous signs and symptoms as fever, headache, musculoskeletal discomfort, and regional lymphadenopathy. The diagnosis of erythema migrans is based primarily on clinical findings because serologic tests to detect elevated antibody levels to Borrelia burgdorferi are frequently negative during the first few weeks of the illness. Identification of Borrelia burgdorferi from skin biopsy specimens obtained from erythema migrans lesions microbiologically or histopathologically will confirm the clinical diagnosis of erythema migrans.

Dermatologic manifestations of Lyme disease.

Erythema migrams (EM), the distinctive cutaneous lesion of Lyme disease, has a variable clinical appearance, but at some point presents as a centrifugally expanding, usually erythematous, annular patch. Of 237 patients with this condition, 201 (85%) were examined initially from May through September. Thirty-four (14%) remembered having been bitten by a deer tick. The median interval from the bite to the appearance of EM was 9 days (range, 1-36 days). Forty-one (17%) of the patients had multiple EM lesions. Of the 237 patients, 128 (54%) manifested major extracutaneous signs and symptoms. Although EM also has a variable histologic picture, the presence of a deep and superficial perivascular and interstitial lymphohistiocytic infiltrate containing plasma cells is diagnostic. Spirochetes can be demonstrated with Warthin-Starry staining in approximately 40% of the biopsy specimens. Concomitant cutaneous lesions appeared on some patients before and during antibiotic therapy. Nine patients with serologic evidence of Borrelia burgdorferi infection had cutaneous lesions other than EM, including granuloma annulare (three), erythema nodosum (two), papular urticaria (two), Henoch-Schönlein-like purpura (one), and morphea (one). Whether these entities are cutaneous markers of Lyme disease or are coincidental findings is yet to be determined.

Use of an autologous antigen in the serologic testing of patients with erythema migrans of Lyme disease.

We attempted to detect an early rise in antibody titers to Borrelia burgdorferi in the serum of patients with erythema migrans of Lyme disease by utilizing B. burgdorferi isolates obtained from patients' own skin lesions instead of the B31 reference strain. B. burgdorferi was isolated from nine of 23 skin biopsy specimens submitted for culture. Elevated antibody titers were not detected in any of the 23 acute serum samples by immunofluorescence assay. The antigens derived from patient isolates were no more effective than the reference strain in detecting antibodies in patients with early Lyme disease.