Comparison of Kato Katz, antibody-based ELISA and droplet digital PCR diagnosis of schistosomiasis japonica: Lessons learnt from a setting of low infection intensity

Abstract

Background: Zoonotic schistosomiasis in Asia, caused by Schistosoma japonicum, remains a major public health concern in China and the Philippines. The developing epidemiological and socio-economic picture of the disease in endemic areas necessitates the development of affordable and highly accurate field diagnostics as an important component in evaluating ongoing integrated control and elimination efforts.

Methods: Three diagnostic methods, namely Kato-Katz (KK) stool microscopy, ELISA and droplet digital (dd) PCR assays, were compared by detecting infection in a total of 412 participants from an area moderately endemic for schistosomiasis in the Philippines.

Results: This comprehensive comparison further defined the diagnostic performance and features for each assay. Compared with the ddPCR assay analysing DNA from faeces (F_ddPCR), which exhibited the highest sensitivity, the SjSAP4 + Sj23-LHD-ELISA had the best accuracy (67.2%) among all five ELISA assays assessed. Schistosomiasis prevalence determined by the SjSAP4 + Sj23-LHD-ELISA and ddPCRs was similar and was at least 2.5 times higher than obtained with the KK method. However, the agreement between these assays was low. In terms of cost and logistical convenience, the SjSAP4 + Sj23-LHD-ELISA represents a cost-effective assay with considerable diagnostic merits. In contrast, although the ddPCR assays exhibited a high level of diagnostic performance, the high cost and the need for specialized equipment presents a major obstacle in their application in screening campaigns.

Conclusion: The SjSAP4 + Sj23-LHD-ELISA represents a cost-effective tool for the diagnosis of schistosomiasis that could prove an important component in the monitoring of integrated control measures as elimination draws closer, whereas the ddPCR assays, in addition to their high sensitivity and specificity, are capable of quantifying infection intensity. However, the high cost of ddPCR hinders its wider application in screening programs, although it could be a valuable reference in the development and improvement of other diagnostic assays.

Fig 1. Flowchart showing the workflow for the detection of S. japonicum infection with different diagnostic methods in a cohort from an endemic area of Northern Samar, the Philippines.

Fecal samples were examined with the KK technique, serum samples were tested by ELISA, while both samples were also analyzed by ddPCR.

Fig 2. Classification of S. japonicum infection in the study cohort according to parasite load.

A) Infection status and intensity determined by the KK method. B) Parasite load (EPG) in the different age groups. Boxes represent the interquartile range of the data with lines across the boxes indicating the median values. The hash marks above and below the boxes indicate the 90th and 10th percentiles, respectively. No statistically significant difference was observed between age groups (Non-parametric Kruskal-Wallis test, p > 0.05).

Fig 3. Comparison of prevalence of S. japonicum infection by different diagnostic tests.

A) The prevalence of S. japonicum infection determined by the KK, SR_ddPCR, F_ddPCR and SjSAP4 + Sj23-LHD-ELISA for the total cohort by different age groups. B) Fold changes in the prevalence of S. japonicum infection determined by the SR_ddPCR, F_ddPCR and SjSAP4 + Sj23-LHD-ELISA vs the KK for the total cohort by each age group. Cut-off value for the SjSAP4 + Sj23-LHD-ELISA: 0.1503.