Detection of Fasciola-specific excretory/ secretory (E/S) protein fraction band (49.5 kDa) and its utilization in diagnosis of early fascioliasis using different diagnostic techniques

Abstract

The objective of the present work is to evaluate Fasciola E/S antigens for diagnosis of early fascioliasis utilizing different diagnostic techniques. Using enzyme-linked immunoelectro-transfer blot (EITB), Fasciola-specific E/ S protein fraction band (49.5 kDa) was determined and electroeluted. The mono-specific antibodies against this specific fraction band were prepared by immunizing pathogen-free rabbit. Assessment of the prepared mono-specific antibodies in diagnosis of human fascioliasis was performed through the detection of E/S copro-antigens by enzyme-linked immunosorbent assay (ELISA) in stool eluates obtained from patients with confirmed fascioliasis, other parasites as well as from other healthy individuals. Serum samples were collected and tested to detect serum antibodies against Fasciola E/S antigen using EITB and counter immunoelectrophoresis (CIEP). Analysis of Fasciola adult worm E/S products by SDS/PAGE revealed a number of bands, the molecular weight (MW) of which ranged from 14-200 kDa; with three major bands (27.5, 32.5 and 55 kDa). Fasciola EIS 49.5 kDa protein fraction proved to be specific to F. gigantica. Cross reaction with S. mansoni was observed at higher MW (110-120 kDa). The sensitivity, specificity and diagnostic accuracy of EITB were 45.2%, 100% and 70.7%, respectively, while those of CIEP were 38.7%, 100% and 67.2%, respectively. ELISA technique using mono-specific anti-49.5 kDa to detect copro-antigens proved to be practical and reliable. It showed higher sensitivity (91.4%) and higher diagnostic accuracy (91.8%), while the specificity was 92.3%. In addition, ELISA had higher negative predictive value (88.9%) and fair positive predictive value (94.1%).