Accuracy of a dual path platform (DPP) assay for the rapid point-of-care diagnosis of human leptospirosis

Abstract

Background: Diagnosis of leptospirosis by the gold standard serologic assay, the microscopic agglutination test (MAT), requires paired sera and is not widely available. We developed a rapid assay using immunodominant Leptospira immunoglobulin-like (Lig) proteins in a Dual Path Platform (DPP). This study aimed to evaluate the assay's diagnostic performance in the setting of urban transmission.

Methodology: We determined test sensitivity using 446 acute and convalescent sera from MAT-confirmed case-patients with severe or mild leptospirosis in Brazil. We assessed test specificity using 677 sera from the following groups: healthy residents of a Brazilian slum with endemic transmission, febrile outpatients from the same slum, healthy blood donors, and patients with dengue, hepatitis A, and syphilis. Three operators independently interpreted visual results without knowing specimen status.

Results: The overall sensitivity for paired sera was 100% and 73% for severe and mild disease, respectively. In the acute phase, the assay achieved a sensitivity of 85% and 64% for severe and mild leptospirosis, respectively. Within seven days of illness onset, the assay achieved a sensitivity of 77% for severe disease and 60% for mild leptospirosis. Sensitivity of the DPP assay was similar to that for IgM-ELISA and increased with both duration of symptoms (chi-square regression P = 0.002) and agglutinating titer (Spearman ρ = 0.24, P<0.001). Specificity was ≥93% for dengue, hepatitis A, syphilis, febrile outpatients, and blood donors, while it was 86% for healthy slum residents. Inter-operator agreement ranged from very good to excellent (kappa: 0.82-0.94) and test-to-test reproducibility was also high (kappa: 0.89).

Conclusions: The DPP assay performed acceptably well for diagnosis of severe acute clinical leptospirosis and can be easily implemented in hospitals and health posts where leptospirosis is a major public health problem. However, test accuracy may need improvement for mild disease and early stage leptospirosis, particularly in regions with high transmission.

Figure 1. DPP, IgM-ELISA, and 1∶100 MAT sensitivity and mean MAT titer for acute-phase sera of human leptospirosis.

Acute-phase sera from (A) 282 MAT-confirmed severe leptospirosis case-patients (259 from Salvador and 23 from Recife, Brazil) and from (B) 28 MAT-confirmed mild leptospirosis case-patients from Salvador. 95% confidence intervals calculated for point estimates of sensitivity and the geometric mean (± geometric standard deviation) calculated for reciprocal MAT titers. DPP = Dual Path Platform; MAT = microagglutination test; ELISA = enzyme-linked immunosorbant assay; ND = not determined because no case-patients were identified within the corresponding time interval.

Figure 2. Representative non-reactive, strongly reactive, and weakly reactive DPP assay results.

Demonstration of (A) non-reactive, (B) strongly reactive, and (C) weakly reactive visual interpretations for the DPP assay. Coloration of the test line (“T” on rapid test cartridge) indicates the presence of anti-rLig antibodies in the biological sample while coloration of the control line (“C” on rapid test cartridge) indicates the presence of non-specific antibodies and denotes a valid test result. DPP = Dual Path Platform.