Sensitivity and specificity of multiple Kato-Katz thick smears and a circulating cathodic antigen test for Schistosoma mansoni diagnosis pre- and post-repeated-praziquantel treatment

Abstract

Background: Two Kato-Katz thick smears (Kato-Katzs) from a single stool are currently recommended for diagnosing Schistosoma mansoni infections to map areas for intervention. This 'gold standard' has low sensitivity at low infection intensities. The urine point-of-care circulating cathodic antigen test (POC-CCA) is potentially more sensitive but how accurately they detect S. mansoni after repeated praziquantel treatments, their suitability for measuring drug efficacy and their correlation with egg counts remain to be fully understood. We compared the accuracies of one to six Kato-Katzs and one POC-CCA for the diagnosis of S. mansoni in primary-school children who have received zero to ten praziquantel treatments. We determined the impact each diagnostic approach may have on monitoring and evaluation (M&E) and drug-efficacy findings.

Method/principle findings: In a high S. mansoni endemic area of Uganda, three days of consecutive stool samples were collected from primary school-aged children (six - 12 years) at five time-points in year one: baseline, one-week-post-, four-weeks-post-, six-months-post-, and six-months-one-week-post-praziquantel and three time-points in years two and three: pre-, one-week-post- and four-weeks-post-praziquantel-treatment/retreatment (n = 1065). Two Kato-Katzs were performed on each stool. In parallel, one urine sample was collected and a single POC-CCA evaluated per child at each time-point in year one (n = 367). At baseline, diagnosis by two Kato-Katzs (sensitivity = 98.6%) or one POC-CCA (sensitivity = 91.7%, specificity = 75.0%) accurately predicted S. mansoni infections. However, one year later, a minimum of three Kato-Katzs, and two years later, five Kato-Katzs were required for accurate diagnosis (sensitivity >90%) and drug-efficacy evaluation. The POC-CCA was as sensitive as six Kato-Katzs four-weeks-post and six-months-post-treatment, if trace readings were classified as positive.

Conclusions/significance: Six Kato-Katzs (two/stool from three stools) and/or one POC-CCA are required for M&E or drug-efficacy studies. Although unable to measure egg reduction rates, one POC-CCA appears to be more sensitive than six Kato-Katzs at four-weeks-post-praziquantel (drug efficacy) and six-months-post-praziquantel (M&E).

Figure 1. Recruitment and inclusion of primary-school children and their samples in the study.

Final numbers of children who provided three stools for a total of six Kato-Katz thick smears (6KK) and one urine sample for a point-of-care circulating antigen test (POC-CCA) at each time-point. Final numbers and percentage female in parentheses. Totaling 1065 samples for Kato-Katz thick smears accuracy analyses and 367 for POC-CCA analyses over the whole study. Samples were collected pre-praziquantel treatment, one-week-post-praziquantel treatment (1 Wk-Post-Treat) and four-weeks-post-praziquantel treatment (4 Wks-Post-Treat) at four time points: Baseline, six months later, one year later and two years later.

Figure 2. Sensitivity and negative predictive values of one to five Kato-Katzs for S. mansoni diagnosis.

(A) Sensitivity and (B) Negative Predictive Values of one to five Kato-Katzs thick smears (1KK to 5KK) for S. mansoni diagnosis over 11 time-points using six Kato-Katzs as the ‘gold standard’. All individuals were treated with 40 mg/kg praziquantel after the start of the study (pre) and then again at six-months, one-year and two-years. All individuals providing stool samples with >100 eggs per gram (EPG) at one-week-post-praziquantel treatment and any infected children at all other time-points were re-treated with 40 mg/kg praziquantel. 95% confidence intervals are excluded for clarity, but can be seen in Table 2.

Figure 3. Effect of S. mansoni arithmetic mean infection intensity on sensitivity of one to five Kato-Katzs.

The sensitivity of one to five Kato-Katz thick smears (1KK to 5KK) for diagnosing S. mansoni infections at a range of community arithmetic mean S. mansoni infection intensities (measured as eggs per gram of stool (EPG) from six Kato-Katz thick smears). S. mansoni infection intensities were measured at three primary schools, at 11 time points each, ranging from pre-treatment to two-years-four-weeks-post-praziquantel treatment. Lines are best fit linear lines.

Figure 4. Effect of diagnostic technique and sampling effort on S. mansoni prevalence measures.

The effect of (A) number of Kato-Katz thick smears (Kato-Katzs), from one to six (1KK to 6KK) on the recorded prevalence of S. mansoni infection with multiple praziquantel treatments. All individuals were treated after sampling at pre, six-months, one-year and two-years. All individuals excreting more than 100 EPG at one-week and any infected children at all other time-points were re-treated. 95% confidence intervals are excluded for clarity. Difference in sensitivities between the current standard two Kato-Katzs and our ‘gold standard’ of six Kato-Katzs determined by the McNemar test is significant at all time-points except baseline. The effect of (B) diagnosis method using six Kato-Katzs or one point-of-care circulating cathodic antigen test (POC-CCA), with a trace counting as either a positive (POC-CCA-t+) or negative (POC-CCA-t−) on S. mansoni infection prevalence with 95% confidence intervals. Difference in sensitivities between the POC-CCA and our ‘gold standard’ of six Kato-Katzs determined by the McNemar test: ^* significant for POC-CCA-t− or ^† significant POC-CCA-t+.

Figure 5. S. mansoni intensity measures from one to three days of duplicate Kato-Katz thick smears.

Data shown over multiple rounds of praziquantel treatments with standard error bars.

Figure 6. Comparison of S. mansoni arithmetic mean intensity measured by six Kato-Katzs with POC-CCA band strength.

S. mansoni mean intensity in eggs per gram of stool (EPG) in comparison to the band strengths of a single urine point-of-care circulating cathodic antigen test (CCA) at five time-points, from (A) pre-treatment at baseline, (B) one-week-post-, (C) four-weeks-post-, (D) six-months-post- and (E) six-months-one-week-post-praziquantel treatment. Spearman's rank coefficients and significance values are given for each time point. Trace readings were included in the + category. NB y axes vary.