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PLoS Negl Trop Dis. 2012;6(12):e1946. doi: 10.1371/journal.pntd.0001946. Epub 2012 Dec 13.

Leishmaniasis direct agglutination test: using pictorials as training materials to reduce inter-reader variability and improve accuracy.

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  • 1Royal Tropical Institute Koninklijk Instituut voor de Tropen, Amsterdam, The Netherlands.



The Direct Agglutination Test (DAT) has a high diagnostic accuracy and remains, in some geographical areas, part of the diagnostic algorithm for Visceral Leishmaniasis (VL). However, subjective interpretation of results introduces potential for inter-reader variation. We report an assessment of inter-laboratory agreement and propose a pictorial-based approach to standardize reading of the DAT.


In preparation for a comparative evaluation of immunochromatographic diagnostics for VL, a proficiency panel of 15 well-characterized sera, DAT-antigen from a single batch and common protocol was sent to nine laboratories in Latin-America, East-Africa and Asia. Agreement (i.e., equal titre or within 1 titer) with the reading by the reference laboratory was computed. Due to significant inter-laboratory disagreement on-site refresher training was provided to all technicians performing DAT. Photos of training plates were made, and end-titres agreed upon by experienced users of DAT within the Visceral-Leishmaniasis Laboratory-Network (VL-LN).


Pre-training, concordance in DAT results with reference laboratories was only 50%, although agreement on negative sera was high (94%). After refresher training concordance increased to 84%; agreement on negative controls increased to 98%. Variance in readings significantly decreased after training from 3.3 titres to an average of 1.0 titre (two-sample Wilcoxon rank-sum (Mann-Whitney) test (z = -3,624 and p = 0.0003)).


The most probable explanation for disagreement was subjective endpoint reading. Using pictorials as training materials may be a useful tool to reduce disparity in results and promote more standardized reading of DAT, without compromising diagnostic sensitivity.

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