Evaluation and comparison of indirect immunofluorescent antibody test (IFAT) and Direct agglutination test (DAT) using standardized antigen in diagnosis of visceral leishmaniasis

Background and Human visceral leishmaniasis (HVL) is endemic in several foci in IRAN, such as Ardebil and Fars provinces (in North western and southern parts of IRAN) and in some regions as sporadic. Visceral leishmaniasis in Iran is Mediterranean type and the causative agent is Leishmania infantum and its main reservoir is dog. In this study direct agglutination test (DAT) was compared with indirect fluorescent antibody test (IFAT) for the diagnosis of visceral leishmaniasis in patients suspected of kala-azar. A total of 70 serum samples were collected from suspected kala-azar patients mainly in the kala-azar endemic areas. The Leishmania infantum antigens (MHO/TN/80/IPTi) were prepared in Department of parasitology, School of Medicine, Isfahan University of Medical Sciences. The principal phases of the procedure from making DAT antigen were mass production of promastigotes of leishmania in the RPMI1640 + fetal bovine serum, Trypsiniznation of parasites, staining with Comassie Blue and fixing with formaldehyde. The human serum samples were tested by DAT, as well as, by IFAT, with the L.infantum antigen prepared in our laboratory. The sero positive rate (SPR) with DAT in titers of ≥ 1:3200 was 91.4% and with IFAT in titers of ≥ 1:80 was 94.3%. Geometric means of reciprocal titers (GMRT) were 6309 for DAT and 692 for IFAT. Therefore, as the titers of ≥1:3200 are usually considered positive in DAT, the titers of ≥1:80 were regarded as position in IFAT. The coincidence of the two tests was 92%. These results showed that a simple local laboratory with one or two trained technicians is quite sufficient for DAT, sero-diagnosis and serological surveys of kala-azar in an endemic area. According to the results of this study, it seems that in Kala azar endemic areas, the clinical symptoms of Visceral leishmaniasis, particularly among the children with DAT antibody titers ≥1:3200 is a good indication for specific treatment of Kala-azar.