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Abstract Fascioliasis is a food-borne parasitic disease caused by Fasciola spp. (species); Fasciola hepatica (F. hepatica) and Fasciola gigantica (F. gigantica). The disease is an important zoonosis that can infect humans as incidental hosts. The available treatment for fascioliasis is triclabendazole (TCBZ) which is a benzimidazole derivative with antihelminthic activity and it is known to be highly effective against F. hepatica and F. gigantica. However, over-reliance on this drug in the treatment of human fascioliasis (HF) and animal fascioliasis (AF) has led to the development of resistance. This emerging resistance necessitates identification of novel and alternative therapeutic measures. The aim of the present study was to evaluate the effect of Thioredoxin reductase inhibitor (auranofin) on adult Fasciola spp. in vitro through a series of parasitological, biochemical, molecular and ultrastructure studies. This study was designed using 75 adult Fasciola worms of nearly equal size to study the antihelminthic effects of auranofin against adult Fasciola worms. Five groups were included (each group included 15 F. gigantica adult worms). GI: adult worms were incubated in RPMI-1640 medium and served as a normal control group (NC), GII: worms were incubated in RPMI-1640 medium containing TCBZ at a concentration of 20 μL/mL and served as the TCBZ- exposed group, GIII: worms were incubated in RPMI-1640 medium containing auranofin at a concentration of 3 μg/mL were served as the auranofinexposed group a, GIV: worms were incubated in RPMI-1640 medium containing auranofin at a concentration of 5μg/mL and served as the auranofin- exposed group b and GV: worms were incubated in RPMI-1640 medium containing auranofin at a concentration of 10 μg/mL and served as the auranofin- exposed group c. The five groups were incubated for 3 hours at room temperature in an atmosphere of 5 % CO2 and then were subjected to motility assay, egg hatching, scanning & transmission electron microscope, histopathological, immunohistochemical studies for evaluation of the expression of caspase-3, assay of GST, analysis of SOD activity and real- time PCR for cathepsin-L gene expression. On assessment of worm motility changes among the studied groups, no major loss of activity in the control worms was observed. The decrease in the motility of worms exposed to auranofin was both time and concentration dependent. The highest concentration (10 μg/ml) of the auranofin produced inhibition of the motility of worms as compared to the normal control and TCBZ- exposed group. After three hours, all worms in auranofin- exposed three groups (a, b & c) were dead (score 0). In normal control group (GⅠ) and TCBZ- exposed group, only 3 and 5 worms respectively were dead (score 0) (Table 1). Also, decrease in motility of worms was significant in relation to time in auranofin- exposed groups (GⅢ, GⅣ and GⅤ) (P<0.001) (Table 8). Results of egg hatching in the studied groups showed that in both control (GⅠ) and TCBZ- exposed group (GⅡ), there was normal immature yellowish brown operculated eggs with thin shell and normal egg hatching. In auranofin- exposed groups (GⅢ, GⅣ and GⅤ), there was decrease in egg hatching with damage, disintegration, shrinking and collapse of egg. Also, there was significant decrease in hatching of F. gigantica eggs in normal control group (GⅠ) with increase in duration of incubation (P: 0.001), TCBZexposed (GⅡ) (P: 0.003), auranofin- exposed a (GⅢ) (P: 0.041), auranofin- exposed b (GⅣ) (P: 0.013) and auranofin- exposed c (GⅤ) with percentage of 40%, 13.3%, 13.3% |