Characterisation of novel peptides as candidates for diagnosis of schistosomiasis

BACKGROUND: Schistosomiasis is one of the most prevalent parasitic diseases worldwide: affecting around 250 million people, with 90% of cases occurring in sub-Saharan Africa. Elimination as a public health problem is ambitious given the current lack of sensitive and specific diagnostics. There is urgent need for the development of point-of-care tools which are rapid, affordable, and easy to use. The current study evaluates and characterises the reactivity of Schistosoma linear B-cell peptide epitopes via serological assays to assess their potential as diagnostic biomarkers. METHODOLOGY: In-silico analyses were carried out to identify protein family, sequence conservation and potential cross-reactivity of five selected peptides (4,5,9,11,15). The peptides were subsequently evaluated for immunoreactivity via indirect enzyme-linked immunosorbent assays (ELISAs). IgM and IgG responses were measured in serum samples from an endemic population in Zimbabwe with known infection status determined by microscopy. Sensitivity and specificity were assessed by comparing serological outcomes with microscopy as the reference standard. PRINCIPLE FINDINGS: Peptides 4,5,9, and 11 were localised to the parasite tegument and 4,5, and 15 were observable across all parasite life cycle stages. Accessibility and continual expression make peptides accessible to the host immune response. All peptides were relatively conserved across Schistosoma species and exhibited little concern of cross-reactivity outside the genus. IgG responses displayed a reversed control trend, and therefore all subsequent analyses focused on IgM. All peptides produced a measurable IgM response with detectable responses across a range of infection intensities and age-groups. Peptide 15 IgM assays displayed the highest reactivity and a corresponding sensitivity of 78.85%. However, sensitivity was at the cost of reduced specificity. Peptides 15 and 5 had the highest sensitivities (78.85% and 71.15% respectively), but the lowest specificities (33.33% and 42.59%). Contrastingly, peptide 4 had moderate sensitivity (50%) and the highest specificity (57.4%). Receiver operating characteristic (ROC) and area under the curve (AUC) analysis indicated that none of the peptides had of strong diagnostic discrimination (all AUC localised around 0.5). Infection intensity (measured as egg burden) had no significant effect on IgM absorbance levels for any peptide-ELISA (all p > 0.3). Age did however affect the absorbance of peptides 4, 9 and 11 (type II ANOVA: p = 0.047, p = 0.050, and p = 0.001 respectively). For peptide 11, the responses in the 0-5-year group were significantly lower than those in the 6-10 (estimate = -1.88, p = 0.004), 11–15 (estimate = −1.73, p = 0.009), and 16–30 (estimate = −1.87, p = 0.003) (Tukey-adjusted pairwise comparisons). For peptide 9, a significant difference was observed between the 0-5- and 11-15-year groups (estimate = -1.68, p = 0.047). CONCLUSION: Due to their immunogenic and sequence profiles, these peptides remain promising candidates for Schistosoma specific antibody detection. Future studies should prioritise assay repetition and expanding sample size to improve assay validation and optimisation. Overall, the creation and improvement of diagnostic methods is an urgent step towards the elimination of schistosomiasis of as a public health problem.