Blastomyces dermatitidis Antigen Detection by Quantitative Enzyme Immunoassay

Abstract

The second-generation MVista Blastomyces antigen enzyme immunoassay was not quantitative; therefore, specimens obtained previously were tested in the same assay as new specimens to assess the change in antigen levels. Furthermore, the sensitivity in serum had not been fully evaluated. The purpose of this study was to evaluate a quantitative Blastomyces antigen assay and detection of antigen in serum. Calibrators containing known concentrations of Blastomyces galactomannan were used to quantify antigen in urine and serum from patients with proven blastomycosis and from controls. Paired current and previously obtained urine specimens were tested to determine if quantification eliminated the need for concurrent testing to assess change in antigen. Pretreatment of serum with EDTA at 104°C was evaluated to determine if dissociation of immune complexes improved detection of antigenemia. Antigenuria was detected in 89.9% of patients with culture-or histopathology-proven blastomycosis. Specificity was 99.0% in patients with nonfungal infections and healthy subjects, but cross-reactions occurred in 95.6% of patients with histoplasmosis. Change in antigen level categorized as increase, no change, or decrease based on antigen units determined in the same assay agreed closely with the category of change in ng/ml determined from different assays. Pretreatment increased the sensitivity of detection of antigenemia from 35.7% to 57.1%. Quantification eliminated the need for concurrent testing of current and previously obtained specimens for assessment of changes in antigen concentration. Pretreatment increased the sensitivity for detection of antigenemia. Differentiation of histoplasmosis and blastomycosis is not possible by antigen detection.A ntigen detection is a useful method for diagnosis of blastomycosis. The sensitivity has been reported to be over 90% and specificity has been reported to be 100% in healthy subjects, but cross-reactions occur in most patients with histoplasmosis (6). In a recent report of 59 patients with proven blastomycosis (4), culture was positive in 86%, histopathology in 81%, antigen in 74%, cytopathology in 38%, and immunodiffusion for antibody in 32% (4). Antigen detection was considered to be a "reliable method to make an accurate and rapid diagnosis of blastomycosis, particularly when a large burden of disease is present, and when cytological analysis is performed at less-experienced centers" (4). Blastomyces antigen detection was also reported to be useful for diagnosis of blastomycosis in solid organ transplant patients (7).Because of interassay variability, specimens obtained earlier have been tested simultaneously with current specimens to assess the change in antigen levels. In a Histoplasma antigen assay, quantification of galactomannan antigen eliminated the need to test the previously obtained specimen with the current specimen to determine change in antigen level (5). In a recent study, quantification and improved detection of antigen in serum following EDTA treatment at 104°C in the t...

Serum and UrineBlastomycesAntigen Concentrations as Markers of Clinical Remission in Dogs Treated for Systemic Blastomycosis

“…Similar to prior reports in people and dogs,7, 8, 9, 10 we found urine Blastomyces antigen to be highly sensitive for the diagnosis of blastomycosis (100% sensitivity in this population). The sensitivity for serum antigen test was also quite high (about 90%), which is similar to previous reports (87%) 8.…”

“…In the absence of organism identification, diagnosis has been accomplished historically through a combination of clinical signs, radiographic findings, and serum antibodies 2, 4, 5, 6. Recently an antigen test has been developed, which detects a cell wall galactomannan, and has been found to have a high degree of both sensitivity and specificity for the diagnosis of blastomycosis in dogs and humans 7, 8, 9, 10. Although both urine and serum can be assayed to evaluate the presence of Blastomyces antigen, urine antigen measurement appears to have an increased sensitivity relative to serum antigen for the diagnosis of blastomycosis in dogs 8…”

Antigen Detection in Canine Blastomycosis: Comparison of Different Antibody-Antigen Combinations in Two Competitive ELISAs

“…In another human subject study, antigenuria was detected in approximately 90% of patients with proven blastomycosis. Specificity was 99% in healthy subjects or patients with nonfungal infections, but cross-reactions occurred in approximately 96% of patients with histoplasmosis [16]. An earlier study on antigen detection in dogs with blastomycosis revealed a sensitivity value of 94% in urine specimens and a value of 87% in serum specimens [7].…”

“…The primary objective of this present study was to compare different antibody-antigen combinations for the detection of antigen in urine specimens from dogs with diagnosed blastomycosis. Several recent studies have focused on antigen detection in blastomycosis in humans and dogs [7,15,16]. In one study, detectable antigenuria was achieved in approximately 85% of the subjects with newly diagnosed blastomycosis, but B. dermatitidis antigen was also detected in 15 patients with histoplasmosis [15].…”

“…In recent years other investigators have been approaching the problem of immunodiagnosis and have produced encouraging results with antigen detection assays as second-generation assays have been developed [7,[14][15][16][17][18]. Our laboratory has been focusing on the use of the competitive enzyme-linked immunosorbent assay (ELISA) in B. dermatitidis antigen detection studies in urine specimens from dogs with diagnosed blastomycosis [11].…”