In vitro effect of heather extracts on Trichostrongylus colubriformis eggs, larvae and adults

Abstract

This study was carried out to evaluate the in vitro effects of different heather species on Trichostrongylus colubriformis eggs, larvae and adult worms, and obtain scientific evidence to attribute these effects to the action of their phenolic compounds and/or tannins. Total phenolic extracts of three heather species (Calluna vulgaris, Erica cinerea, and Erica umbellata) and an equal mixture of these three extracts were tested in vitro in the three development stages of T. colubriformis using an egg hatching assay (EHA), larval exsheathment inhibition assay (LEIA), and adult motility inhibition assay (AMIA). The egg hatching rate was measured after incubation with heather extracts for 48h at 25°C. Infective third-stage larvae (L3) were incubated for 3h at 25°C with heather extracts. The evolution of artificial exsheathment over time was measured with repeated observations at 20-min intervals for 60min. Adult worms were obtained from one donor goat and incubated with the extracts at 37°C for 5 days in 48-multiwell plates. Worm motility was measured at 0, 19, 24, 43, 48, 67, 72, 96 and 115h after the beginning of the experiment. The extracts were tested at concentrations of 75, 150, 300, 600 and 1200μg/ml. All extracts significantly (P<0.001) inhibited egg hatching and the effect was dose dependent. All extracts inhibited or delayed the exsheathment of T. colubriformis L3, and the effect was dose dependent for C. vulgaris. Incubation with heather extracts induced a reduction in adult worm motility compared to control, although significant (P<0.05) differences were only found at the highest concentrations. Additional studies showed that purified tannins of the same heather species disturbed T. colubriformis larval exsheathment. All these results confirm the anthelmintic properties of heather against T. colubriformis, and suggest that not only tannins but also some other phenolic compounds might be involved.