Identification of plumbagin and sanguinarine as effective chemotherapeutic agents for treatment of schistosomiasis

Abstract

Schistosomiasis, a snail-borne parasitic disease, affects more than 200 million people worldwide. Currently the treatment of schistosomiasis relies on a single therapy of praziquantel, a drug developed over 30 years ago. Thus, there is an urgent need to develop alternative antischistosomal drugs. In the pursuit of novel antischistosomal drugs, we examined the antischistosomal activities of 45 compounds that had been reported to exhibit antimicrobial and/or antiparasitic activities. Two plant-derived compounds, plumbagin and sanguinarine, were found to possess potent antischistosomal activities in vitro. For both the compounds, a concentration of 10 μM (equivalent to 1.88 μg/ml for plumbagin and 3.68 μg/ml for sanguinarine) resulted in 100% mortality at 48 h, which meets the World Health Organization's (WHO) criterion of "hit" compounds for the control of schistosomiasis. Morphological changes and tegumental alterations of the dead worms treated by the two compounds were quite different. The significant morphological changes of worms after treatment by the two compounds suggest the two compounds target different biological pathways, both of which result in parasite's death. This study provides evidence to suggest plumbagin and sanguinarine have real potential as effective alternative chemotherapeutic agents for the treatment of schistosomiasis.

Keywords: Chemotherapy; Plumbagin; Sanguinarine; Schistosomiasis.

Graphical abstract

Fig. 1

In vitro antischistosomal activities. Effect of compounds at various concentrations on the mortality of schistosome worms under culture conditions. The data was generated from three independent studies and the time (day) of death was determined based on two or three consistent experiments. No obvious mortality in control groups (DMSO) was observed within 4 days of worm culture or 4 h of cercariae culture. The final concentration of DMSO is 1% (v/v) for all experiments. Note that the intervals on the X-axis are not equal.

Fig. 2

Effect of various concentrations of plumbagin and sanguinarine on the survival of worms. The average percent of live worms is shown. The average percentages were generated from three independent experiments and the bars show the range of live worm percentages in the experiments. Note that the intervals on the X-axis are not equal.

Fig. 3

Effect of compounds on the survival of cercariae. (A) The effect of various concentrations of compounds on cercariae mortality. The data was generated from three independent studies and the time (hour) of death was determined based on two or three consistent experiments. Note that the intervals on the X-axis are not equal. (B) Upper (a) and lower (b) images showing cercariae after treatment with plumbagin and sanguinarine, respectively.

Fig. 4

Comparison of morphological differences of the dead worms after treatment with plumbagin (PBG) and sanguinarine (SGR). (A) Morphological appearance of dead schistosome worms after treatment with sanguinarine (left) and plumbagin (right) (under a microscopic field). (B) Actual average size of dead worms (male and female) after treatment with either sanguinarine, plumbagin or DMSO (control). No significant difference was found between sanguinarine and DMSO control in either sex.

Fig. 5

SEM images showing teguments of male (left panel) and female worms (right panel) after culture with DMSO (A, B), plumbagin (C, D) and sanguinarine (E, F). (A) and (B) show SEM image of the normal tegument of male and female worms. Male teguments shows hill-shaped tubercles covered with pointed spines and a convoluted surface of the tegumental membrane between tubercles. The female tegument does not have tubercles and spines and appears like a field plowed surface. (C) Shows a SEM image of a male worm exposed in vitro to plumbagin at a concentration of 10 μM for 24 h. Tegumental surfaces are damaged usually with disintegration of tubercles accompanied by a decrease in the number of spines. In addition, there were varying degrees of disintegration of the tegumental surfaces including peeling, wrinking and blebbing. (D) Among the female worms treated with plumbagin at a concentration of 10 μM for 24 h, the worms displayed moderate damage when compared to male damage of the tegumental surface. Holes and divots are prevalent throughout the tegument. (E) SEM image shows the tegument of an adult worm exposed in vitro to sanguinarine at a concentration of 10 μM for 24 h. Distortions of the tegumental surfaces can be seen usually with severe erosion and disintegration of the tegumental surface between tubercles. The holes and broken surface are visible. (F) Female worms treated with 30 μM sanguinarine for 8 h displayed varying degrees of swelling and cracking ranging from minimal to moderate throughout the anterior and posterior tegumental ridges/fissures.