Antigenemia immunoassay for point of care dianostic melioidosis

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Status of laboratory diagnosis of melioidosis Currently, culture from multiple body sites is the gold standard for laboratory diagnosis. Culture requires experienced personnel and takes 3-4 days. Levels of bacteremia are very low (~ i CFU/ml). A point-of-care immunoassay for diagnosis of melioidosis could greatly impact patient outcome because a high percentage of patients with acute septicemia die within 24-48 h of admission, and the antibiotics used for empiric treatment of septicemia are not effective for B. pseudomallei. Assays for antibody have limited value due to an absence of antibody in the early stages of acute sepsis and a high level of background antibodies in endemic regions (6). Several studies reported PCR for detection of B. pseudomallei, but the reported lower limit of detection tends to fall above the range for the viable bacteria count in blood during human disease (118). Potential antigen targets for immunodiagnosis of melioidosis Detection of extracellular polysaccharides has been successful for a variety of bacterial infections. There are three candidate antigens for B. pseudomallei: the exopolysaccharide (EPS), the capsular polysaccharide (CPS), and the lipopolysaccharide (LPS). EPS is an unbranched polymer of a repeating tetrasaccharide: [-3)-p-D-Galp2Ac-(i-4)-a-D-Galp-(i-3)-p-D-Galp-(i-5)-|3-Kdo-(2-] (115). CPS is an unbranched homopolymer of [-3)-2-O-acetyl-6-deoxy-p-D-mannoheptopyranose-( i-] (120). LPS is an unbranched polymer of disaccharide repeating units having the basic structure: [-3-)-|3-D-glucopyranose-(i-3)-6-deoxy-a-L-talopyranose-(i-] (120). Melioidosis patients make antibodies to EPS, CPS and LPS, indicating that these antigens are produced in vivo. To date, no B. pseudomallei proteins have been identified as candidates for antigen-targeted diagnosis of melioidosis. The proposed InMAD process has the potential to identify B. pseudomallei antigens shed in vivo and may serve as a rapid broad discovery platform for many microbial threats.