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Research Project: Genetic Improvement of Crop Plants for Use with Low Quality Irrigation Waters: Physiological, Biochemical and Molecular Approaches

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Title: Terminalia catappa: chemical composition, in vitro and in vivo effects on Haemonchus contortus

Author
item KATIKI, L - Instituto De Zootecnia
item GOMES, C - Instituto De Zootecnia
item BARBIERI, A - Instituto De Zootecnia
item PACHECO, P - Instituto De Zootecnia
item RODRIGUES, L - Instituto De Zootecnia
item VERISSIMO, C - Instituto De Zootecnia
item GUTMANIS, G - Instituto De Zootecnia
item PIZA, A - Instituto De Zootecnia
item LOUVANDINI, H - Instituto De Zootecnia
item Ferreira, Jorge

Submitted to: Veterinary Parasitology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/8/2017
Publication Date: 9/28/2017
Citation: Katiki, L.M., Gomes, C.P., Barbieri, A.M., Pacheco, P.A., Rodrigues, L., Verissimo, C.J., Gutmanis, G., Piza, A.M., Louvandini, H., Ferreira, J.F. 2017. Terminalia catappa: chemical composition, in vitro and in vivo effects on Haemonchus contortus. Veterinary Parasitology. 246:118-123. doi: 10.1016/j.vetpar.2017.09.006.

Interpretive Summary: Gastrointestinal parasites (GIP) of sheep and goats have developed resistance to most commercial drugs in several countries. There is no new drug being developed to control these GIP and animals with excess parasite burden (mainly goats) can die within a week. Terminalia catappa (leaf, fruit pulp, and seed extracts) were evaluated in vitro against a multi-drug resistant strain of the ‘barber pole’ worm (Haemonchus contortus) through the egg hatch assay (ovicidal effect), and in vivo (pulp and seed powder) using lambs experimentally infected with the barber pole worm. Crude extracts from leaves, fruit pulp, and seeds were obtained with 70% acetone, concentrated, and lyophilized. Seed extracts killed 50% of the parasite eggs (LC50)at the concentration of 2.48 µg/mL, while the pulp extract had LC50 = 4.62 µg/mL. When compared in vitro to the effect of the commercial parasiticide drug Thiabendazole (LC50 = 1.31 µg/mL), seed and pulp extracts had LC50 values of 2.48 and 4.62 µg/mL, while the leaf extract had LC50 = 20 µg/mL. Condensed tannins were present in greater quantities in pulp extract (183,92g of leucocyanidin/kg dry matter) than in leaf (4,6 g) and seed (35,13 g) extracts, while leaves were highly concentrated in total tannins, indicating high concentration of hydrolysable tannins and flavonoids. Phytochemical qualitative tests established that all extracts contained alkaloids, flavonoids, saponins, phenols, and terpenoids. In vivo tests were performed to evaluate the anthelmintic activity of T. catappa pulp + seed. Male Santa Ines lambs were artificially infected with multidrug-resistant barber pole worms. Control animals were infected and treated with Thiabendazole, while animals treated with crude plant extracts were infected with worms and received with T. catappa pulp + seed (whole fruit) powder at 2g/kg of live body weight (LBW). Whole fruit powder was mixed with feed concentrate and provided for five days. At the end of the experiment, animals underwent parasitological analysis, kidney profile (urea and creatinine), liver profile (aspartate aminotransferase), and had their weight recorded. In vitro, seed extracts had anti-parasitic effect similar to Thiabendazole (LC50 ranged from 2.48 vs. 1.31 µg/mL), but in vivo studies with whole fruit powder did not corroborate in vitro results obtained with extracts. Our results agreed with those of several others in that in vivo results did not replicate in vitro results, indicating that tannins and other phenolic compounds may suffer bacterial and ruminal degradation after ingestion by ruminant animals. However, at the used dose, there was no toxicity of T. catappa crude extracts to sheep. Our results will provide sheep farmers with important information on the safety and efficacy (or lack of) of plant-based extracts that may help livestock cope with important parasites such as the barber pole worm.

Technical Abstract: Haemonchus contortus is the most important nematode in small ruminant systems, and has developed tolerance to all commercial anthelmintics in several countries. In vitro (egg hatch assay) and in vivo tests were performed with a multidrug strain of Haemonchus contortus using Terminalia catappa leaf, fruit pulp, and seed extracts (in vitro), or pulp and seed powder in lambs experimentally infected with H. contortus. Crude extracts from leaves, fruit pulp and seeds obtained with 70% acetone were lyophilized until used. In vitro, the extracts had LC50= 2.48 µg/mL (seeds), LC50=4.62 µg/mL (pulp), and LC50=20 µg/mL (leaves). In vitro, seed and pulp extracts had LC50 similar to Thiabendazole (LC50= 1.31 µg/mL). Condensed tannins were more concentrated in pulp extract (183.92 g of leucocyanidin/kg dry matter) than in either leaf (4.6 g) or seed (35.13 g) extracts. Phytochemical tests established that all extracts contained alkaloids, flavonoids, saponins, phenols, and terpenoids. Based on these results, in vivo tests were performed to evaluate the anthelmintic activity of T. catappa whole fruit (pulp + seed) powder. Male Santa Ines lambs were artificially infected with multidrug-resistant H. contortus and divided, according to similar fecal egg count (FEC) and weight, into two groups: Control (infected/untreated) and treated (infected/treated with whole fruit powder). Whole fruit powder was mixed with concentrate and provided at 2 g/kg of body weight (BW) for five days. After treatment, parasitological analysis (FEC and egg hatch assay), renal profile (urea and creatinine), liver profile (aspartate aminotransferase) and BW were determined. In vitro (based on LC50), seed/pulp extracts had ovicidal effect similar to Thiabendazole but whole fruit powder had no anthelmintic effect on adult nematodes in the abomasum. We discuss the plausible causes of the lack of in vivo activity.