Research Article: Development of Ss-NIE-1 Recombinant Antigen Based Assays for Immunodiagnosis of Strongyloidiasis

Date Published: April 10, 2015

Publisher: Public Library of Science

Author(s): Lisa N. Rascoe, Courtney Price, Sun Hee Shin, Isabel McAuliffe, Jeffrey W. Priest, Sukwan Handali, Aysegul Taylan Ozkan.

Abstract: Strongyloides stercoralis is a widely distributed parasite that infects 30 to 100 million people worldwide. In the United States strongyloidiasis is recognized as an important infection in immigrants and refugees. Public health and commercial reference laboratories need a simple and reliable method for diagnosis of strongyloidiasis to identify and treat cases and to prevent transmission. The recognized laboratory test of choice for diagnosis of strongyloidiasis is detection of disease specific antibodies, most commonly using a crude parasite extract for detection of IgG antibodies. Recently, a luciferase tagged recombinant protein of S. stercoralis, Ss-NIE-1, has been used in a luciferase immunoprecipitation system (LIPS) to detect IgG and IgG4 specific antibodies. To promote wider adoption of immunoassays for strongyloidiasis, we used the Ss-NIE-1 recombinant antigen without the luciferase tag and developed ELISA and fluorescent bead (Luminex) assays to detect S. stercoralis specific IgG4. We evaluated the assays using well-characterized sera from persons with or without presumed strongyloidiasis. The sensitivity and specificity of Ss-NIE-1 IgG4 ELISA were 95% and 93%, respectively. For the IgG4 Luminex assay, the sensitivity and specificity were 93% and 95%, respectively. Specific IgG4 antibody decreased after treatment in a manner that was similar to the decrease of specific IgG measured in the crude IgG ELISA. The sensitivities of the Ss-NIE-1 IgG4 ELISA and Luminex assays were comparable to the crude IgG ELISA but with improved specificities. However, the Ss-NIE-1 based assays are not dependent on native parasite materials and can be performed using widely available laboratory equipment. In conclusion, these newly developed Ss-NIE-1 based immunoassays can be readily adopted by public health and commercial reference laboratories for routine screening and clinical diagnosis of S. stercoralis infection in refugees and immigrants in the United States.

Partial Text: Strongyloides stercoralis, an intestinal nematode that migrates through the skin and lung, is a widely distributed disease that infects 30 to 100 million people worldwide [1]. Unlike other helminthic parasites, S. stercoralis can complete its entire life cycle within a single human host through autoinfection and can cause an asymptomatic chronic infection that may go undetected for decades in immunocompetent hosts [2, 3]. In the United States, S. stercoralis causes more deaths than any other soil-transmitted helminth, with mortality rates as high as 87% in cases of hyper-infection in immunocompromised hosts [3].

Strongyloidiasis is an increasingly important health problem in the US among immigrants and refugees. Patients with occult strongyloidiasis are at risk of disease if they become immunosuppressed, and organ donors with unrecognized S. stercoralis infection pose a risk to recipients if their infected organs are transplanted [2]. Identification of the parasite in stool specimens is insensitive, and, because parasitological examination requires collection of multiple stool specimens over 3 days, serological testing would be preferred if available. We elected to develop novel immunodiagnostic assays to meet this need. We employed a well described recombinant protein with proven performance as an immunodiagnostic antigen, the Ss-NIE-1 protein [6–8]. Based on the potential importance of IgG4 antibody responses in filarial infections, we decided to develop methods that detect S. stercoralis-specific IgG4 antibodies. The Ss-NIE-1 IgG4 Luminex bead assay achieved a sensitivity and a specificity comparable to those reported for other strongyloidiasis assays such as the crude antigen ELISA, and 26-kDA ELISA, and Ss-NIE-1 ELISAs and the Ss-NIE-1 LIPS [6, 7, 9, 10, 12, 18]. Compared to the CDC S. stercoralis crude antigen ELISA, the Ss-NIE-1 IgG4 ELISA and the Ss-NIE-1 IgG4 Luminex assay achieved similar sensitivity without compromising specificity. The possible factors contributing to improved specificity could be the use of recombinant antigen, assay optimization, or detection of IgG4 versus IgG. During Ss-NIE-1 ELISA optimization, we found that non-specific antibody binding in the normal human sera could be decreased by adding a pre-blocking step with 10 mM nickel chloride in PBS/0.3% Tween/5% milk. The decrease in background noise allows the assay to have a higher specificity (93%).



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