Rapid latex agglutination assays for serodiagnosis of human

Abstract

Burkholderia pseudomallei is the etiologic agent of melioidosis, a significant community-acquired and severe disease in endemic regions. Diagnosis and treatment of human infections caused by this organism is challenging. In the absence of chemotherapeutic intervention, acute disease is frequently fatal. The objective of this project is to identify and characterize immunogens expressed by B. pseudomallei for developing rapid serodiagnostic tests. Recently, we have demonstrated that the O-polysaccharides (OPS) and capsular polysaccharides (CPS) are suited for this task. We evaluated the use of OPS- and CPS-based latex agglutination assays as a rapid approach for serodiagnosing human melioidosis. OPS and CPS antigens were isolated from B. pseudomallei Bp82, using a modified hot aqueous-phenol extraction procedure. Following purification by sequential rounds of ultracentrifugation and size-exclusion chromatography, the carbohydrate antigens were chemically activated with sodium periodate and covalently linked to aliphatic amine-derivatized latex beads via reductive amination to produce two distinct immunoassays. Utilizing rabbit antiserum samples raised against formalin fixed B. pseudomallei, B. thailandensis and B. cenocepacia isolates, the resulting OPS- and CPS-coated beads exhibited agglutination patterns consistent with those that would be predicted. Using 156 culture-confirmed human melioidosis patients and 143 healthy donor serum samples, the sensitivity and specificity of OPS-coated beads were 82.1% and 62.9% respectively, and of CPS-coated beads were 65.4 and 69.2% respectively. The performance of these assays was equal to or greater than those associated with indirect hemagglutination assays (sensitivity 68.0%, specificity 67.1%). Our rapid latex agglutination assays may be useful as stable, affordable, point-of-care tests for serodiagnosing human melioidosis.