Abstract The aim of this study was to assess the in vitro ovicidal activity of an ethyl acetate extract from Arceuthobium vaginatum (EtOAc-E) and their subfractions (AvR5-AvR14) against Haemonchus contortus using the egg hatching inhibition (EHI) test. The EtOAc-E and subfractions were tested at 0.12-2.00 and at 0.015-2.0 mg/mL, respectively. Distilled water and methanol (2%) were used as negative controls and Thiabendazole (0.10 mg/mL) as a positive control. Treatments with a dependent effect on concentration were subjected to regression analysis to determine the effective concentrations (EC50 and EC90). The major secondary compounds present in the extract and subfractions were identified by high performance liquid chromatography (HPLC). The EtOAc-E and AvR9 exhibited the best ovicidal effect recording 97.5 and 100% of EHI at 0.25 mg/mL, respectively. The EtOAc-E and AvR9 displayed an EC50= 0.12 and 0.08 mg/mL, respectively. The HPLC analysis in the EtOAc-E and bioactive fractions indicated the presence of a polyphenol, glycosylated flavanones, quercetin glucoside, cinnamates, coumarin, cinnamic acid derivative, ferulic acid, coumarate, naringenin, protocatechuic acid and naringin. Results demonstrated that A. vaginatum extract and fraction is able to inhibit the egg hatch process of H. contortus and could be a viable option for the control of small ruminant haemonchosis.

Resumo O objetivo deste estudo foi avaliar a atividade ovicida in vitro de um extrato de acetato de etila de Arceuthobium vaginatum (EtOAc-E) e suas subfrações (AvR5-AvR14) contra Haemonchus contortus, utilizando-se o teste de inibição da eclosão de ovos (EHI). O extrato foi testado em 0.12-2.00 mg/mL, e as subfrações foram testadas em 0.15-2.0 mg/mL. Água destilada e metanol (2%) foram utilizados como controles negativos e Tiabendazol (0.10 mg/mL) como controle positivo. Os tratamentos com efeito concentração dependente foram submetidos à análise de regressão para determinação das concentrações efetivas (CE50 e CE90). Os principais compostos secundários, presentes no extrato e nas subfrações, foram identificados por cromatografia líquida de alta eficiência (HPLC). EtOAc-E e subfração AvR9 exibiram o melhor efeito ovicida, registrando-se 97.5 e 100% do EHI a 0.25 mg/mL, respectivamente. O EtOAc-E e AvR9 apresentaram CE50= 0,12 e 0,08 mg/mL, respectivamente. A análise por HPLC nas frações EtOAc-E e bioativas indicou a presença de um polifenol, flavanonas glicosiladas, glicosídeo de quercetina, cinamatos, cumarina, derivado de ácido cinâmico, ácido ferúlico, cumarato, naringenina, ácido protocatecuico e naringina. Os resultados demonstraram que extrato e frações de A. vaginatum são capazes de inibir o processo de eclosão dos ovos de H. contortus, podendo ser uma opção viável para o controle da hemoncose em pequenos ruminantes.

Abstract The anthelmintic resistance problem is widely recognized in sheep production. Therefore, new methods of control against gastrointestinal nematodes (GIN) need to be integrated. The aim of this work was to assess the toxicity of A. cina 30 CH as a homeopathic product against Haemonchus contortus in in vitro and in vivo assays. A. cina 30 CH was obtained from a commercial laboratory, and confirmation of artemisinin as a key ingredient was performed with mass spectrophotometry. The A. cina 30 CH and the artemisinin pure reagent were used for the inhibition of egg hatching (IEH) and for the inhibition of larval migration of H. contortus L3 (ILM). In addition, three groups of 10 naturally infected lambs with GIN were treated with A. cina 30 CH and albendazole, and 10 were used as control. The parasitic infection was monitored at 0, 7, 14 and 28 d postreatment (PT) to determine the number of eggs per gram (epg) and FAMACHA index. The in vitro data showed 100 % IEH and 64.7 % ILM by A. cina 30 CH, and nonlethal activity was observed with the artemisinin pure reagent. The toxicity of A. cina 30 CH against H. contortus in infected lambs was observed after 7 d of infection. Administration of the A. cina 30 CH yielded a 69 % reduction in the epg at 28 d PT, similar to the albendazole (P<0.05). In conclusion, A. cina 30 CH had the ability to IEH and ILM of H. contortus in in vitro assays and reduced the number of eggs of H. contortus, which is the primary parasitic nematode in grazing lambs, thereby reducing infection.

Resumen El problema de la resistencia a los antihelmínticos es ampliamente reconocido en la producción ovina. Por lo tanto, es necesario integrar nuevos métodos de control contra los nematodos gastrointestinales (NGI). El objetivo de este estudio fue evaluar la toxicidad de Artemisia cina 30 CH como un producto homeopático contra Haemonchus contortus en ensayos in vitro e in vivo; A. cina 30 CH se obtuvo de un laboratorio comercial, y la confirmación de la artemisinina como ingrediente clave se realizó con espectrometría de masas. A. cina 30 CH y la molécula registrada como antihelmíntica de la artemisinina se usaron para inhibir la eclosión del huevo (IEH) y la migración larval (L3) de H. contortus (IML). Además, tres grupos de 10 corderos infectados naturalmente con NGI se trataron con A. cina 30 CH y albendazol, y 10 fueron utilizados como control. La infección parasitaria se controló a los 0, 7, 14 y 28 días post-tratamiento (PT) para determinar el número de huevos por gramo (HPG) y el índice FAMACHA. Los datos in vitro mostraron 100 % de IEH y 64.7 % de IML por A. cina 30 CH, y se observó actividad no letal con la artemisinina. La toxicidad de A. cina 30 CH contra H. contortus en corderos infectados se observó después de 7 días de infección. La administración de A. cina 30 CH produjo una reducción de 69 % de EPG a los 28 días PT, similar al albendazol (P<0.05). En conclusión, A. cina 30 CH tuvo la capacidad de IEH e IML de H. contortus en ensayos in vitro y redujo el número de huevos de H. contortus, que es el nematodo parásito primario en corderos en pastoreo, con lo que se redujo la infección.

The aim of this study was to evaluate the predatory behavior in vitro of the mite Lasioseius penicilliger on 3 nematode species: Teladorsagia circumcincta (L3) (a sheep-parasitic nematode), Meloidogyne sp. (J2) (a plant-parasitic nematode), and on various developmental stages of Caenorhabditis elegans (a free-living nematode). The coincubation of mites and nematodes was individually assessed in 2% water agar placed in plastic Petri dishes (2 cm x 1 cm diameter). One thousand nematodes of a species and 5 mites were placed into each plate (10 replicates) and incubated for 5 days at room temperature (18-25ºC). L. penicilliger showed predatory behavior against the 3 assessed nematode species. The percentages of predatory activity recorded were 95.1, 80.5 and 79.3 against Meloidogyne sp., C. elegans, and T. circumcincta, respectively (P ≤ 0.05). These results suggest that L. penicilliger has important potential as a biological control agent of parasitic nematodes.

El objetivo de este estudio fue evaluar el comportamiento depredador in vitro del ácaro Lasioseius penicilliger sobre tres especies de nemátodos: Teladorsagia circumcincta (L3, nemátodo parásito de ovejas), Meloidogyne sp. (J2, nemátodo parásito de plantas), y sobre varios estadios de desarrollo de Caenorhabditis elegans (nemátodo de vida libre). La coincubación de los ácaros y los nemátodos se evaluó en 2% agar-agua colocados en cajas de Petri de plástico (2 cm x 1 cm diámetro). Se colocaron 1 000 nemátodos de cada especie y cinco ácaros en cada caja (10 réplicas) y se incubaron por cinco días a temperatura ambiente (18-25ºC). L. penicilliger mostró comportamiento depredador contra las tres especies de nemátodos evaluadas. Los porcentajes de actividad depredadora registrados fueron 95.1, 80.5 y 79.3, contra Meloidogyne sp., C. elegans y T. circumcincta, respectivamente (P ≤ 0.05). Estos resultados sugieren que L. penicilliger tiene un importante potencial como agente de control biológico contra nemátodos parásitos.