Fasciola hepatica and Rumen Flukes - In Vitro Evaluation of Main Commercial Anthelmintics

Background: Trematode infections are of great importance as they affect the health of many species of mammals as cattle, sheep and goat. Fasciola hepatica represents the main trematode zoonosis and risks to human and cattle and param-phistomosis is one emerging parasitic diseases of ruminants widely distributed in the world. The economic expenses are incurred by the use of ineffective anthelmintics for trematode control. Besides to faecal egg count reduction test (FECRT) to determine the anthelmintic efficacy, can be used in vitro assays, by this the aim of the study was to determine the lethal doses (LD) with hatching egg test (EHT) of the main commercial anthelmintics used for the control of trematodes in cattle. Materials, Methods & Results: Liver and rumen were examined from cattle slaughtered in Tabasco, Chiapas and Campeche states from Mexico. F. hepatica eggs were recovered from gallbladder and rumen fluke eggs collected from adult parasites in saline solution. Subsequently, the hatching egg assays were performed placing 100 trematode eggs in distilled water in each one of 96 wells of polystyrene plates. After making the appropriate dilutions, several concentrations of commercial anthelmintics were evaluated, ranging from 0.04 to 80.63 mM for triclabendazole + 0.046 to 96.87 mM febendazole (TC+FBZ), from 0.04 to 91 mM for rafoxanide (RAFOX), from 0.02 to 43.74 mM for closantel (CLOS), from 0.036 to 76.18 mM for clorsulon + 0.002 to 3.31 mM ivermectin (CLORS+ IVM) and from 0.163 to 334.47 mM for nitroxynil (NITROX). A control group (water) was included in each plate. Lethal doses were obtained using the Probit procedure and analysis of the means with a one-way statistical design. Most drugs used against rumen fluke eggs presented a high LD 50 and therefore were ineffective to cause egg mortality, such was the case of RAFOX that presented LD 50 from 4,580 to 10,790 mu g/mL (7 to 17 mM). CLOS presented the lowest LD 50 (80 mu g/mL or 0.12 mM) on rumen fluke eggs. TC+FBZ was found to be effective drug against the development of F. hepatica eggs in many samples. In the same way NITROX showed a low LD 50 (37 to 63 mu g/mL or 0.13 to 0.22 mM), but RAFOX presented a highest LD 50 (1,450 mu g/mL or 2.32 mM). Discussion: The present study focused on screen the ovicidal activity and determining in vitro lethal doses 50 of main commercial anthelmintics used to control F. hepatica and rumen fluke as rapid tests in a tropical region from Mexico. The FECRT is the main method to detect effectiveness of anthelmintic and other method is the coproantigen reduction test (CRT) by ELISA. Both tests require many infected animals depending the number of treatments and by this the egg hatch assay (EHA) represent a complementary diagnosis of effectiveness of anthelmintic products to compare between regions and even between farms, because few animals are required from the farm to collect trematode eggs, and it is possible to know the effectiveness against various anthelmintics at the same time. Efficacy studies on trematodes using egg hatching tests are scarce, although they have the advantage that they can be applied to both F. hepatica and rumen fluke. TC+FBZ was one of the most effective products in inhibiting the development of F. hepatica eggs. However, RAFOX showed low effectiveness against trematode eggs, with very high lethal doses. These results agree with a study that show low efficacy against the development of Paramphistomum cervi eggs and with the FECRT test reductions of 75% and 80.58% were obtained, in times from 7 to 84 days after treatment with RAFOX. NITROX and CLORS were drugs that had good efficacy on the development of F. hepatica eggs. A differential response between liver and rumen fluke was observed. The anthelmintics used against rumen fluke eggs show low ovicidal activity and in Fasciola hepatica TC+FBZ show the best activity.