Anthelmintic efficacy of agro-industrial by-products against gastrointestinal nematodes of sheep: In vitro tests

Abstract

Gastrointestinal nematodes (GINs) are ubiquitous in grazing small ruminants and cause significant costs due to production losses. Moreover, anthelmintic resistance (AR) is now widespread throughout Europe and poses a major threat to the sustainability of modern small ruminant livestock farming. Trichostrongylus colubriformis, an intestinal parasite of small ruminants, is the second nematode to Haemonchus contortus in terms of distribution and magnitude in several regions. The control of this helminth is traditionally achieved with the use of anthelmintic drugs, however due to regulations in organic farming and the rise in anthelmintic resistance (AR), alternatives are sought after. The aim of the present study was to evaluate the in vitro anthelmintic effects of olive (Olea uropaea), carob (Ceratonia siliqua), pomegranate juice ( unica granatum), wine (Vitis vibifera), citrus (Citrus Senensis), hazelnut (Corylys avellana) and tomato (Solanum lycopersicum) by-products extracts on T. colubriformis of sheep. Extracts of seven by-products were tested in vitro on two development stages of T. colubriformis (eggs and infective larvae) using the Egg Hatch Assay (EHA) and the Larval Exsheathment Inhibition Assay (LEIA). The egg hatching rate was measured after incubation with each by-product extract (concentrations: 150, 300, 600, 1200 mg/mL) for 48 h at 26 °C. nsheathed infective larvae were incubated for 3h at 20 °C with each by-product extract (concentrations: 150, 300, 600, 1200 mg/mL). Artificial exsheathment was induced in vitro by adding sodium hypocloride solution (2%w/v) diluted in 1 to 300 in PBS to the larval suspension. The progress of exsheathment over time was measured by repeated observations at 20-min intervals for 60 min (Moreno-Gonzalo et al., 2013. Vet Parasitol, 197:235-43). Among the 7 extracts tested using the two in vitro tests (EHA and LEIA), those with the greatest anthelmintic potential against T. colubriformis were the by-product extracts of wine, pomegranate and hazelnut. In particular, the inhibition in the development of eggs using EHA was 2.3%-48.5% for wine, 6.1%-48.1% for hazelnut and 18.6%-42.4% for pomegranate by-product extracts. The inhibition of exsheathment using LEIA was 87.5%-100% for wine, 54.5%-98.7% for hazelnut and 62.3%-100% for pomegranate by-product extracts. Results showed that by-product extracts of wine, pomegranate and hazelnut exhibits in vitro anthelmintic activity, suggesting that, these by-products can also be an ally for T. colubriformis control in sheep.