Identification and Preliminary Evaluation of a Novel Recombinant Protein for Serodiagnosis of Strongyloidiasis

Abstract

Strongyloides stercoralis is a human parasite that can cause a long-term infection. In immunosuppressed patients, strongyloidiasis may be fatal when there is overwhelming autoinfection resulting in the migration of large numbers of larvae through many organs. Definitive diagnosis is still a challenge, and a combination of symptoms, microscopic identification, and serology test results are often used to arrive at a clinical decision. However, intermittent larval excretion, low parasite burden, and occult infections are challenges with parasitological diagnosis of infection with S. stercoralis. Meanwhile, serologic tests using immunoglobulin G and parasite antigen extract have problems of cross-reactivity with other helminthic infections. Recombinant antigen-based serodiagnosis is a good alternative to overcome the laboratory diagnostic issues. Herein, we report on the isolation of cDNA clone encoding an antigen of potential diagnostic value identified from immunoscreening of a S. stercoralis cDNA library. The translated protein had highest similarity to Strongyloides ratti immunoglobulin-binding protein 1. The recombinant antigen produced, rSs1a, was assessed using western blot and enzyme-linked immunosorbent assay. The latter showed 96% diagnostic sensitivity and 93% specificity; thus, rSs1a has good potential for use in serodiagnosis of human strongyloidiasis.

Figure 1.

Representative images of IgG4-tertiary immunoscreening for evaluation of diagnostic sensitivity and specificity of clone Ss1a using individual positive and negative pre-adsorbed serum samples. NC = negative control serum; Neg = serum from group B; PC = positive control serum; Pos = serum from group A. This figure appears in color at www.ajtmh.org.

Figure 2.

SDS-PAGE analysis of purified rSs1a; M: Precision Plus Protein^™ Unstained Standard Marker (Bio-Rad). This figure appears in color at www.ajtmh.org.

Figure 3.

Representative image of western blot analysis of purified rSs1a: (A) probed with anti-His-HRP (His); (B) probed with positive serum samples from Group A (lane 1) and control serum samples from Group B: lanes 2–5: healthy individuals; lanes 6–12: mixed hookworm infection and trichuriasis; lane 13: hookworm; lane 14: ascariasis; lanes 15–17: trichuriasis; lane 18: toxocariasis; lanes 19–20: amoebiasis; lanes 21–22: schistosomiasis; lane 23: lymphatic filariasis; and M: Precision Plus Protein Unstained Standard Marker (Bio-Rad). This figure appears in color at www.ajtmh.org.

Figure 4.

Receiver operating characteristic curve of rSs1a IgG4-enzyme-linked immunosorbent assay for the detection of strongyloidiasis (area under the curve = 0.942, 95% confidence interval = 0.90–0.97, P < 0.0001). This figure appears in color at www.ajtmh.org.

Figure 5.

Scattergraph plot of optical density (OD) readings of IgG4-enzyme-linked immunosorbent assay (ELISA) using the positive and control serum samples. An OD reading of 0.165 was used as the cutoff value (COV) to discriminate between positive and negative results. This figure appears in color at www.ajtmh.org.