Serodiagnostics of schistosomiasis using keyhole limpet hemocyanin (KLH) as antigen

Abstract: The diagnostics of invasive helminth infections is often indirect and based on antibody detection in serum specimens from infected individuals. This is the case in schistosomiasis (bilharzia) where no excreted eggs can be found in light, often asymptomatic, infection. Travellers, especially adventure tourists, and immigrants to endemic areas run a high risk of becoming infected. If the diagnosis is missed, infected individuals may carry intravascular worms for more than 30 years and thereby run a risk for tissue injury due to egg deposition. Early during infection antibodies against gut associated schistosome antigens (GAA) are detected by an indirect immunofluorescent assay where sections of adult worms are used as antigen. We observed a similar, previously unrecognised staining pattern in a significant proportion of GAApositive sera, which subsequently was shown to be due to antibodies, which may cross-react with keyhole limpet hemocyanin (KLH). This cross-reacting glycoprotein, from the mollusk (Megathura crenulata), has been introduced as diagnostic antigen for the detection of antibodies against schistosomes. However,. in the literature the limited experience using KLH in diagnostics of early infection shows discrepant results. The aim of this study was to understand why antibodies, reacting with KLH, are produced during schistosomiasis and to evaluate the diagnostic potential of such antibodies. This was done by localisation of KLH-cross-reactive epitopes in the different life stages of the parasite and by partial identification by immunoblotting of electrophoretically separated schistosome egg components. We found that shared antigens are expressed in different life stages of the parasite: at the surface and in the secretions of the cercaria, at the schistosomula surface, in the excretory/secretory (E/S) ducts of the adult worms, in reproductive tissues, at the surface of the miracidia, in and around eggs in granulomas, and in Kupffer cells in the liver. These observations explain why an immune response against KLH-cross-reactive schistosome components is readily induced in infected individuals. However, when we evaluated a simple dotELISA for the detection of anti-KLH antibodies, it had poor specificity and the sensitivity varied when different patient groups were tested. A significant finding was that trichinellosis patients had anti-KLH antibodies and also reacted with schistosome ducts. We conclude that KLH is a potentially useful target antigen for serodi agnostics. However, multiple antigenic epitopes in KLH may explain reported discrepant results and specificity and sensitivity problems.