In Vitro Filaricidal Properties of Hydro-Methanolic Extracts of Powdery Fractions of Khaya senegalensis (Meliaceae) on Onchocerca ochengi.

Abstract

Onchocerciasis is a neglected tropical disease that remains endemic in sub-Saharan African countries. Unfortunately, only a few microfilaricidal agents have been approved so far. This study aimed to assess the in vitro macro and microfilaricidal potentialities of the hydro-methanolic extracts of the different powdery fractions of Khaya senegalensis against Onchocerca ochengi. Adult male worms and microfilariae (mf) of O. ochengi were isolated from cowhides in Ngaoundere II, Cameroon. Parasites were incubated for 4h (mf) or 48h (adult worms) in RPMI-1640 medium in the presence or absence of ivermectin, flubendazole, or hydro-methanolic extracts of different plant powdery fractions obtained by controlled differential sieving. The filaricidal effect was evaluated using motility (mfs) and mortality tests (worms) and oxidative stress parameters. Cytotoxicity and acute toxicity tests were performed on monkey-derived kidney cell lines (LLC-MK2) and Swiss albino mice, respectively, and selectivity indexes were determined. Phytochemical screening was also carried out using high-performance liquid chromatography/UV (HPLC/UV), molecular networking, and through quantification of phenolic contents. The hydro-methanolic extracts of 0-63µm fractions from leaves and barks exhibited the strongest macrofilaricidal activities with lethal concentrations 50 of 162.4 and 208.8µg/mL respectively versus 22.78µg/mL for flubendazole. These two fractions also showed the fastest microfilaricidal activities (T1/2 of 1h), although it was low when compared to ivermectin (T1/2 < 1h). Their macrofilaricidal effects were accompanied by a significant inhibition of nitric oxide secretion and a significant increase of glutathione and catalase activity compared to the untreated group. However, no effect was found on superoxide dismutase activity, the GABAergic and glutamatergic receptors. Although neither extract was toxic to Swiss mice until a dose of 2000mg/kg body weight, the 0-63µm leaf fraction hydro-methanolic extract was selectively more effective on worms than bark extract (SI = 1.28 versus 0.34). Both extracts were found to contain some flavonoids including procyanidin-, rutin-, myricetin-, and naringenin derivatives as well as new unknown compounds. However, the total polyphenol, flavonoid and tannin contents of the leaf extract were significantly greater (P < 0.05) than that of the bark extract. These results support the anti-filarial effect of K. senegalensis leaves and highlight stress oxidative markers as new therapeutic targets in O. ochengi. Further, in vivo experiments are required in understanding their anti-parasitic properties, and testing combinations of fine fractions.