BackgroundIn an effort to enhance accuracy of diagnosis of Schistosoma haematobium, this study explores day-to-day variability and diagnostic performance of real-time PCR for detection and quantification of Schistosoma DNA compared to other diagnostic tools in an endemic area before and after treatment.MethodologyPreviously collected urine samples (N = 390) from 114 preselected proven parasitological and/or clinical S. haematobium positive Kenyan schoolchildren were analyzed by a Schistosoma internal transcribed spacer-based real-time PCR after 14 years of storage. Pre-treatment day-to-day fluctuations of PCR and microscopy over three consecutive days were measured for 24 children using intra-class correlation coefficient. A combined ‘gold standard’ (PCR and/or microscopy positive) was used to measure sensitivity and negative predictive value (NPV) of several diagnostic tools at baseline, two and 18 months post-treatment with praziquantel.Principal FindingsAll 24 repeatedly tested children were PCR-positive over three days with little daily variation in median Ct-values, while 83.3% were found to be egg-positive for S. haematobium at day 1 and 75.0% at day 2 and 3 pre-treatment, signifying daily fluctuations in microscopy diagnosis. Of all 114 preselected schoolchildren, repeated microscopic measurements were required to detect 96.5% versus 100% of positive pre-treatment cases by single PCR. At two months post-treatment, microscopy and PCR detected 22.8% versus 69.3% positive children, respectively. Based on the ‘gold standard’, PCR showed high sensitivity (>92%) as compared to >31% sensitivity for microscopy, both pre- and post-treatment.Conclusions/SignificanceDetection and quantification of Schistosoma DNA in urine by real-time PCR was shown to be a powerful and specific diagnostic tool for detection of S. haematobium infections, with less day-to-day variation and higher sensitivity compared to microscopy. The superior performance of PCR before, and two and 18 months post-treatment provides a compelling argument for PCR as an accurate and reproducible tool for monitoring treatment efficacy.

SummaryWe evaluated the impact of praziquantel therapy (40 mg/kg body weight) on indicators of infection with Schistosoma haematobium by following a cohort of infected children from schools located 12 km apart in the Coast province of Kenya, at 0, 2, 4, 6, 12 and 18 months after treatment. Within this period, measurements of infection parameters pertaining to egg counts and haematuria (micro-, macro-and history) were evaluated at all time points. The initial prevalence of 100% dropped significantly 8 weeks after treatment with a similar trend in the intensity of infection. Microhaematuria followed the same trend as observed for egg counts while macrohaematuria remained low after treatment. Reinfection following successful therapy differed significantly between schools; in one school the children were reinfected immediately while those in the other remained uninfected despite similar starting prevalences, intensities of infection and cure rates. Transmission between the two areas looked homogeneous before treatment but when both groups were treated, contrasting transmission patterns became evident. In a regression model we evaluated factors that might be associated with reinfection, and after allowing for pretreatment infection level, age and sex, area (school) remained a highly significant predictor.

Abstract. A cohort of 117 school children infected with Schistosoma haematobium was followed-up after therapy with praziquantel (0, 2, 4, 6, 12, and 18 months) and various infection and morbidity parameters (egg counts, hematuria, soluble egg antigen [SEA] in urine, and ultrasonography-detectable pathology) were quantified. At the onset of the study, 97% of the children were positive for S. haematobium with a geometric mean egg count of 45.7 eggs/10 ml of urine. Eighty-one percent of the children were positive for SEA in urine with a geometric mean SEA concentration of 218.8 ng/ml of urine. Ninety-two percent and 56% of the children were microhematuria positive and macrohematuria positive, respectively. Two months after treatment, all infection and morbidity indicators had significantly decreased. Reinfection after treatment as determined by detection of eggs in urine was observed by four months post-treatment while the other parameters remained low. The clearance of SEA was slower than that of egg counts while pathology resolved at an even slower pace. Levels of SEA and egg output showed similar correlations with ultrasound detectable pathology; these correlations were better than the correlation between hematuria and pathology.

A panel of 17 monoclonal antibodies (MAbs) against Schistosoma soluble egg antigens (SEAs) was produced from BALB/c mice immunized with antigens secreted/excreted by Schistosoma mansoni eggs. In this study, we demonstrate that 16 MAbs were reactive with S. haematobium SEA in addition to S. mansoni SEA. The MAbs were tested as potential immunodiagnostic reagents in a homologous sandwich ELISA format to detect circulating soluble egg antigens (CSEAs) in serum and urine samples of S. mansoni-or S. haematobium-infected individuals. When samples of S. mansoni-infected individuals were tested, none of these MAbs performed as good as the previously described S. mansoni-specific 114-5B1-A and 114-4D12-A MAbs. However, 11 MAbs (of the IgM isotype) detected CSEA in urine samples of S. haematobium-infected individuals. Three MAbs, 290-2E6-A, 291-3D5-A, and 291-5D5-A, were selected for a pilot study with 47 urine samples of S. haematobium-infected individuals from Kenya. The CSEA levels detected with each of these ELISAs showed a significant correlation with urinary egg counts (Spearman rho Ͼ 0.37, P Ͻ 0.01) and with each other (Spearman rho Ͼ 0.74, P Ͻ 0.001). Based on the 92% specificity and 90% sensitivity of the assay, the ELISA using MAb 290-2E6-A was found to be the most promising assay for immunodiagnosis of S. haematobium infections.

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