Novel anthelmintic compounds and molluscicides from medicinal plants.

This review assesses the role that can be played by allelochemicals (bioactive secondary compounds) from medicinal and other plants in the control of human helminthic diseases. In the search for new anthelmintics among plant allelochemicals, 3 practical issues have considerable significance. They are the range and capacity of anthelmintic bioassays utilised in preclinical studies in vitro on plant extracts, the phenomenon of coexistent allelochemicals with overlapping activity spectra within single plants, and the problem of non-specific cytotoxins among plant allelochemicals. These topics are discussed in the context of the present absence of any clinically useful plant anthelmintics. In the search for new plant molluscicides for schistosomiasis control, the characteristics of a range of molluscicidal plants are measured against those of the synthetic molluscicide of choice, niclosamide, and against the postulated attributes of practically useful plant molluscicides.

Molluscicidal activity in the seeds of Millettia thonningii (Leguminosae: Papilionoideae).

Seeds of the leguminous plant Millettia thonningii were shown to possess promising molluscicidal activity against Bulinus trunctatus. The size of the snails was an important determinant of their susceptibility to the molluscicide; specimens with shell lengths of 2 to 3 mm being more susceptible than snails 5 to 6 mm long.

A comparison of topical formulations for the prevention of human schistosomiasis.

Recently, a dimeticone formulation has been shown to be effective at preventing Schistosoma cercariae infecting skin, while DEET (N,N-diethyl-m-toluamide), a highly effective insecticide, has been shown to have activity against cercariae. Seven formulations, 3 containing DEET, were prepared and applied to excised human skin in Franz cells for 1 h. Schistosoma cercariae were applied for 30 min at 1 and 24 h, and the number that penetrated the skin calculated (n = 9). DEET could not be incorporated into the dimeticone formulation, yet it remained the most effective at preventing cercarial penetration, both 1 and 24 h after application. The ointments that contained DEET did prevent penetration but their mode of action was due to the toxicity of DEET against the cercariae. The persistence of the protection afforded by the dimeticone formulation after washing suggests that the formulation may be interacting with the stratum corneum to prevent cercarial recognition of skin.

Activity of the molluscicidal plant Millettia thonningii (Leguminosae) toward Biomphalaria glabrata eggs.

A dichloromethane extract of the seeds of the molluscicidal west African legume Millettia thonningii was tested for ovicidal activity toward Biomphalaria glabrata egg masses. The extract was found to be highly ovicidal at concentrations as low as 5 mg/L. Embryonic development of the snails in egg masses was monitored using photomicrographs from which embryonic diameters were estimated. Such measurements revealed that ovicidal effects were developmentally stage specific and normally induced a curtailment of development during the gastrula to trochophore transition.

Antischistosomal bioactivity of isoflavonoids from Millettia thonningii (Leguminosae).

Extracts of the seeds of the leguminous tree Millettia thonningii are molluscicidal and schistosomicidal and contain a mixture of isoflavonoids and 3-phenylcoumarins. Robustic acid and a pure mixture of alpinumisoflavone and dimethylalpinumisoflavone, as characterized by nuclear magnetic resonance, were isolated from a dichloromethane extract. The bioactivity of these compounds against Schistosoma mansoni miracidia, cercariae, and adult worms was tested in vitro. Both robustic acid and the alpinumisoflavone mixture killed cercariae and adult schistosomes in vitro. At sublethal concentrations of the isoflavonoids, reductions in miracidial motility were measured using a video-imaging technique. The inhibition of mitochondrial electron transport at site I by the isolated isoflavonoids was also investigated. The results suggest that robustic acid and at least 1 alpinumisoflavone compound from the dichloromethane extract of the seeds of M. thonniogii are responsible for some of the observed bioactivity of this extract against schistosomes.

The plant molluscicide Millettia thonningii (Leguminosae) as a topical antischistosomal agent.

The West African legume Millettia thonningii is used in Ghana as an anthelmintic and as a purgative agent. A chloroform extract of the seeds of Millettia thonningii which is known to be molluscicidal and cercaricidal was topically applied to mouse skin 2 and 24 h prior to exposure to Schistosoma mansoni cercariae. The presence of Millettia thonningii extract components on the surface of the skin appeared to be effective in preventing subsequent establishment of infection. The compound responsible for the activity is thought to be the isoflavonoid alpinumisoflavone.

Control of water-borne parasitic diseases with natural products: the potential of Dialium guineense as a molluscicide.

The molluscicidal activity of the fruit and leaves of Dialium guineense was found to be due to glycosides of the triterpenoid oleanolic acid. Three glycosides were isolated from the fruit and a fourth from the leaves and are known compounds. The amount of total saponins present in D. guineense makes it a good candidate for a readily available molluscicide in Nigerian villages.

Interactions of the cysticercoids of Hymenolepis diminuta and raillietina cesticillus in their intermediate host, Tribolium confusum.

This study was based on the experimental exposure of beetles to homologous and heterologous infections of Hymenolepis diminuta and Raillietina cesticillus. The results demonstrated that, for both species over the range of parasite densities employed, the presence of a primary infection had no effect on the establishment success of an homologous challenge infection. The establishment success of R. cesticillus cysticercoids was not affected by the presence of an H. diminuta infection. The existence of an R. cesticillus infection, however, severely reduced the establishment success of an H. diminuta infection. The reduction in H. diminuta establishment was greatest in R. cesticillus infections less than 10 days old. The plausibility of some of the mechanisms that are potentially responsible for the reduction in H. diminuta establishment success is discussed, as is the possible ecological significance of these results.

Pallisentis rexus from the Chiang Mai Basin, Thailand: ultrastructural studies on egg envelope development and the mechanism of egg expansion.

Pallisentis rexus Wongkham & Whitfield, 1999 (Eoacanthocephala: Quadrigyridae) infects the freshwater snakehead fish, Channa striata, in the Chiang Mai Basin, Thailand. All stages of egg development within the body cavity of the female parasite were observed, using transmission electron microscopy. Changes in mature eggs after contact with water were also investigated. The mature egg has five egg envelopes separated from each other by four gaps. The fertilization membrane, which is formed first, is pushed centrifugally by other, subsequently formed, envelopes and gaps, which produces a final total shell thickness of 8-36 microm around the acanthor. The disappearance of the outermost layer and the unpleating of an adjacent inner layer causes the expansion of eggs on contact with water. The volume of an expanded egg is approximately 27 times that of an unexpanded one, but the density of eggs is reduced from a value greater than water to one almost equal to water. This is believed to aid the dispersion of eggs.

Vasoactive intestinal polypeptide-like immunoreactive tegumental cells in the digenean helminth Echinostoma liei: possible role in host-parasite interactions.

Immunocytochemical investigations were carried out on the tegument of the digenean platyhelminth Echinostoma liei. Tests with well-characterised antisera to mammalian vasoactive intestinal polypeptide (VIP) reveal a subpopulation of tegumental cells which contain material immunologically similar to VIP. Control experiments with antisera for a related peptide as well as absorption of primary antisera with VIP or secretin confirm the specificity of the reaction. Immunoreactive processes may be traced from the tegumental cells towards the distal cytoplasm of the tegument. Immunoreactivity is also concentrated in the outer layers of that cytoplasm. The finding of VIP-like immunoreactive material in peripheral nonneuronal, nonendocrine cells in this gastrointestinal parasite of mammals is discussed in terms of its implication for both parasite biology and host pathophysiology.

A comparison of freshwater and marine/estuarine strains of Pomphorhynchus laevis occurring sympatrically in flounder, Platichthys flesus, in the tidal Thames.

Collections of flounder, Platichthys flesus, at two sites on the tidal River Thames in 1994 and 1995 have, for the first time, revealed the sympatric occurrence of the freshwater and marine/estuarine strains of the acanthocephalan parasite Pomphorhynchus laevis. This natural co-occurrence of the strains has been employed to compare infection levels and a range of parasite attributes of the two strains under conditions of sympatry. At both Lots Road (Chelsea) and Tilbury the marine/estuarine strain was present at far higher infection levels than the freshwater form. In a detailed comparison of worms from Tilbury flounder, a range of differences was revealed between the two strains. In single strain infections in individual fish, freshwater and marine/estuarine worms had distinct but overlapping gut microhabitat use patterns, with the former having a central intestinal bias and the latter a bias for the posterior region of the gut. In mixed strain infections, niche contraction occurred so that no overlap occurred. Freshwater worms were larger and had more eggs, more ovarian balls, and a higher percentage of fully developed eggs than the marine/estuarine worms. These differences are thought to reflect intrinsic, presumably genetically determined, differences between the two strains as they occurred in the same fish host species collected at the same place and time. Apparent differences in strain reproductive potential in flounder in the tidal Thames are discussed in the context of previous studies and the intermediate host segment of the parasite life cycle.

Inhibition of Site I mitochondrial electron transport by an extract of the seeds of Millettia thonningii: a potential mechanism for the plant's molluscicidal and schistosome larvicidal activity.

A dichloromethane extract of the seeds of Millettia thonningii (Leguminosae) which contains a mixture of isoflavonoids (predominantely robustic acid, alpinumisoflavone and dimethylalpinumisoflavone) is known to have larvicidal activity towards the miracidia and cercariae of schistosomes and to possess significant molluscicidal activity. The present investigation has assayed the effects of this extract on the electron transport systems of isolated rat liver mitochondria. The extract was found to inhibit mitochondrial electron transport at Site I (NADH dehydrogenase) at concentrations of 30-159 mg x l(-1). Although the extract is not as potent an inhibitor at Site I as rotenone, a known inhibitor of NADH dehydrogenase, such observations could explain the molluscicidal and schistosomicidal activity of dichloromethane extracts of the seeds of M. thonningii.

Intestinal helminth parasites in flounder Platichthys flesus from the River Thames: an infracommunity analysis.

An analysis was undertaken of intestinal helminth communities in flounder Platichthys flesus from two sites on the River Thames. A comparison was made between helminth community richness and diversity from these sites at the component and infracommunity levels. At the component community level, a richer and more diverse parasite community was found in flounder from the Tilbury location (marine influence) than that from the Lots Road location (freshwater influence). At the infracommunity level, more parasite species and parasite individuals per host were found at Lots Road and the percentage of similarity values were low at both locations. Helminth species with high prevalence values in the parasite communities of the flounder are the dominant species in any individual fish, harbouring multi-specific infections. The presence of more invertebrate species, which are intermediate hosts in the helminth life cycle in the Thames, fish vagility and the high prevalence and abundance values of Pomphorhynchus laevis in the flounder, may explain the differences between the two locations.

Currently available molluscicides.

Views about the importance of the role of molluscicides in the integrated control of human schistosomiasis have passed through cyclical changes over the past 15 years. For a time, it was hoped that chemotherapy alone would achieve significant morbidity control; it has since become clear that molluscicides cannot be easily excluded from the anti-schistosome armoury. In this review, Sheena Perrett and Phil Whitfield summarize the evidence for this conclusion and provide an overview of currently available synthetic molluscicides and those natural product molluscicides currently under active investigation.

Atanine (3-dimethylallyl-4-methoxy-2-quinolone), an alkaloid with anthelmintic activity from the Chinese medicinal plant, Evodia rutaecarpa.

Phytochemical studies using a range of chromatographic and spectroscopic techniques coupled with in vitro bioassays against larval Schistosoma mansoni, L4 larvae of Ostertagia circumcincta, and adults and larvae of Caenorhabditis elegans have led to the isolation of an active anthelmintic compound in the Chinese medicinal plant Evodia rutaecarpa (Rutaceae) and its identification as atanine (3-dimethylallyl-4-methoxy-2-quinolone). Atanine has not previously been found to possess antiparasitic activity.

Parthenogenesis and asexual multiplication among parasitic platyhelminths.

Among flatworms with parasitic and commensal modes of existence, parthenogenesis and asexual multiplication appear to be largely confined to the Digenea and Cestoda, the only parasitic platyhelminths that routinely utilize indirect life-cycles. Parthenogenesis is apparently restricted to a minority of adult digeneans and cestodes inhabiting their final hosts, and a survey is made of the particular modes of parthenogenesis (i.e. apomictic, automictic and generative) which are employed by such adults. Asexual (amictic) multiplication, in the form of fissioning, is demonstrated by young adults of the cyclophyllidean cestode, Mesocestoides corti, but is otherwise not exhibited by adult cestodes or digeneans, other than in the perplexing phenomenon of proglottid formation in polyzoic tapeworms. Secondary multiplication is of ubiquitous occurrence in digenean life-cycles in the form of the proliferation which takes place within sporocysts and rediae (germinal sacs) located in the first intermediate host. The controversy concerning the nature of this multiplication is reconsidered in the context of recent findings which have centred on cellular aspects. On the basis of present evidence germinal sac multiplication should be regarded as an asexual rather than a parthenogenetic process. The cestode asexual multiplication which occurs in intermediate hosts is a function of the metacestode stage of development. Metacestode proliferation is only known from about 20 species and 6 families of polyzoic cestodes with approximately half the described instances occurring in the family Taeniidae. The organization of these proliferative metacestodes, findings concerning their totipotent stem cells and the ontogeny of buds and new scolices are all reviewed. Finally, the capacity for population expansion of multiplicative larval digeneans and metacestodes are compared, while the ecological roles and the genetical consequences of both parthenogenesis and amictic multiplication in the two taxa are also examined.

Presumptive ciliated receptors associated with the fibrillar glands of the spines of the echinoderm Amphipholis squamata.

The ultrastructure of the cuticle, fibrillar glands and stäbchen (see Reichensperger 1908) of the arm spines of the brittle star Amphipholis squamata are described. Stäbchen are shown to be cuticular projections into which pass one or more uniciliate cytoplasmic processes. The proximal tips of the cilia protrude into the external environment to a variable extent. Evidence is presented for considering these ciliated endings to be the terminations of sensory neurites. Stäbchen are, in consequence, presumed to have a receptor function. Groups of stäbchen are spatially associated with fibrillar gland openings on the sides of spines while the bases of fibrillar gland cells are in contact with axons from the spine nerve which are laden with 45-270 nm diameter, electron-dense, membrane-bounded granules. The possible functional significance of these relationships is discussed in the context of a hypothesis in which fibrillar gland secretions are considered to act as food-catching mucus.

Resistance to Millettia molluscicide in Biomphalaria glabrata: a quantitative genetical approach.

This paper demonstrates the use of a quantitative genetical technique to assess genetic variance in molluscicide resistance in a Millettia thonningii-Biomphalaria glabrata system. Results indicated that there were significant genetic variances in both snail size and survival time in 10 p.p.m. molluscicide suspension. The possibility of wider applications of this technique in examining the genetic basis of molluscicide or pesticide resistance in target species is discussed.

The epidemiology of Taenia pisiformis infections in domestic dogs in Cairo.

The epidemiology of Taenia pisiformis infections in domestic dogs in Cairo, Egypt. Over 5000 street dogs humanely killed in a culling programme organized by the Egyptian Ministry of the Interior in the winter of 1986 and the spring of 1987 have been utilized to examine the final host epidemiology of the cestode Taenia pisiformis in Cairo. In the winter months the prevalence of infection was 63.4% and the mean intensity 2.82 per dog: by the spring of 1987 these values had risen to 70.8% and 3.78 respectively. These parasite burdens probably represent the highest infection levels with T. pisiformis yet described from urban dog populations anywhere in the world. The dispersion of T. pisiformis within the street dog population was assessed and found to be significantly aggregated and well described by a negative binomial model of overdispersion (variance/mean ratio: 5.16, coefficient k = 0.795). A separate analysis was carried out of infection in each of 18 circumscribed regional areas of Cairo which revealed considerable area-to-area differences with mean intensities varying between 1.62 and 6.54 and prevalences between 43.3% and 91.2%. The size structure of the winter and spring worm samples were very similar with about 45% of worms less than 35 cm in length, about 35% between 35 and 59 cm, about 20% between 60 and 100 cm and less than 5% greater than 100 cm in length. The epidemiological factors which lead to high T. pisiformis infection levels in Cairo dogs have been considered and the consumption of discarded rabbit offal containing Cysticercus pisiformis by street dogs is regarded as a crucial mode of transmission.

Attenuation of Schistosoma mansoni cercariae with a molluscicide derived from Millettia thonningii.

A chloroform extract of molluscicidal plant secondary compounds from the seeds of a West African legume Millettia thonningii was used to attenuate cercariae of Schistosoma mansoni prior to infection of NMRI female mice. Exposure of cercariae to 0.3, 0.6, 1.3, 2.5, 10 or 20 mg/l concentrations of extract for 30 min, immediately before standardized infection, was associated with a concentration-dependent decline in worm establishment at 55 days post-infection. The mean numbers of adult worms established declined from about 17 worms/mouse with control cercariae and those exposed to 0.3 mg/l Millettia to 0.1 worms/mouse after 10 mg/l pre-treatment. Mice exposed to cercariae after 20 mg/l pre-treatment had no adult worms at 55 days post-infection. The activities of cercariae 30 min after exposure to Millettia extract at concentrations up to 2.5 mg/l were similar to those of control larvae: none was immobile. Exposure to higher concentrations of Millettia progressively reduced swimming activity and increased the proportion of immobile cercariae. After pretreatment with 20 mg/l Millettia the majority of larvae were immobile. Levels of anti-parasite antibodies (estimated by ELISA assay using an adult worm-based antigen preparation) rose between 21 and 55 days p.i. with control cercariae and those pre-treated with 2.5 mg/l Millettia. After 5 mg/l pre-treatment, cercariae induced a reduced antibody rise, while mice exposed to cercariae after 10 mg/l pre-treatment showed no rise in anti-parasite antibody levels. These results are discussed in the context of protocols which could possibly use Millettia-attenuated cercariae to induce useful levels of protection in mice towards further cercarial challenge.