Evaluation of antileishmanial activity of pyrethrin extracts of Chrysanthemum cinerariaefoliumTrevir (Asterales: Asteraceae) plant

Abstract

Treatments of leishmaniases have relied on antimonials drugs which are toxic, expensive and resistance has been reported. The pyrethrum plant, Chrysanthemum cinerariaefolium has been shown to have antiplasmodial, antitrypanosomal and insecticidal activity but little has been studied about its potential on anti-Leishmania effects. The objective of this study was to investigate the antileishmanial activity of water and methanol extracts of Chrysanthemum cinerariaefolium both in vitro and in vivo. In this study concentration between 31.25 to 1000µg/ml of methanolic and aqueous extracts of C. cinerariaefolium were tested for anti- parasitic effects against Leishmania major in in vitro cell culture and in BALB/c mice. The inhibitory concetrations (IC50) and Minimum inhibition concentration (MIC) on L. major promastigotes, percentage rates of macrophages infected by amastigotes and cytotoxicity on vero cells were determined. In vivo bioassays BALB/c mice were infected subcutaneously with 1×106 promastiogetes and kept for 4 weeks to allow disease manifestation. They were then treated with 250µg/ml of water extracts and 125µg/ml of methanol extracts, 20mg/kg/body weigth of pentostam and amphotericin B and Phosphate buffered saline (PBS) were subjected orally and intraperitoneally for four weeks. The lesions were measured weekly and at the 4th week mice were sacrificed and impression smears from spleen were made on slide and examined microscopically. Cytokine analysis was done from serum and nitric oxide production by infected macrophages treated with extracts in vitro was determined. The MIC of water extracts was was not significantly high (250µg/ml) than that of methanol extracts (125µg/ml). IC50 for both methanol (8.3µg/ml) and water extracts (13.6µg/ml) were lower than those of pentostam (14.1µg/ml) and Amphotericin B (8.623µg/ml) but not significant (P> 0.05). Water (238µg/ml) and methanol (121µg/ml) extracts had non significant toxicity (P˃0.05) against vero cells compared to Pentostam and Amphotericin B (106.8 and 68.6µg/ml respectively) and NO produced by macrophages stimulation by extracts was minimal. Treatment with both extracts significantly reduced the number of infected macrophages compared to untreated (P=0.042). The mean difference in multiplication index (number of amastigotes) in macrophages treated with methanolic extract was significantly reduced compared to water extract treatment (t =7.9994, P=0.0076). The infected macrophages treated with pentostam and amphotericin B had significantly reduced number of amastigotes than those treated with both the extracts (P<0.05). Reduction in lesion sizes, low parasite burden in the spleen and low toxicity exhibited by water and methanol of C. cinerariaefolium extracts makes them good candidate compounds for tests in the treatment of various Leishmania species. The study observed that IFN gamma cytokine produced by methanolic and water extracts (at concentration of 125µg/ml and 250µg/ml respectively) treated BALB/c mice were not significantly different from those treated with pentostam and Amphotericin B at 20mg/kg/body weigth. The anti-leishmanial activity and immunological modulation by water and methanol extracts of C. cinerariaefolium suggests that they can be a good candidate for management of leishmaniasis, therefore more studies is required before a conclusive policy on the use of these extracts of treatment of leishmaniasis.