Anthelmintic action of plant cysteine proteinases against the rodent stomach nematode, Protospirura muricola, in vitro and in vivo

Cysteine proteinases from the fruit and latex of plants, including papaya, pineapple and fig, were previously shown to have a rapid detrimental effect, in vitro, against the rodent gastrointestinal nematodes, H eligmosomoides polygyrus (which is found in the anterior small intestine) and Trichuris miti,is (which resides in the caecum). Proteinases in the crude latex of papaya also showed anthelmintic efficacy against both nematodes in vivo. In this paper, we describe the in vitro and in vivo effects of these plant extracts against the rodent nematode, Protospirua muricola, which is found in the stomach. As in earlier work, all the plant cysteine proteinases examined, with the exception of actinidain from the juice of kiwi fruit, caused rapid loss of motility and digestion of the cuticle, leading to death of the nematode in vitro. In vivo, in contrast to the efficacy against H. polygyrus and T. muris, papaya latex only showed efficacy against P. muricola adult female worms when the stomach acidity had been neutralized prior to administration of papaya latex. Therefore, collectively, our studies have demonstrated that, with the appropriate formulation, plant cysteine proteinases have efficacy against nematodes residing throughout the rodent gastrointestinal tract

In vitro and in vivo anthelmintic efficacy of plant cysteine proteinases against the rodent gastrointestinal nematode, Trichuris muris

We examined the mechanism of action and compared the anthelmintic efficacy of cysteine proteinases from papaya, pineapple, fig, kiwi fruit and Egyptian milkweed in vitro using the rodent gastrointestinal nematode Heligmosomoides polygyrus. Within a 2 h incubation period, all the cysteine proteinases, with the exception of the kiwi fruit extract, caused marked damage to the cuticle of H. polygyrus adult male and female worms, reflected in the loss of surface cuticular layers. Efficacy was comparable for both sexes of worms, was dependent on the presence of cysteine and was completely inhibited by the cysteine proteinase inhibitor, E-64. LD50 values indicated that the purified proteinases were more efficacious than the proteinases in the crude latex, with purified ficin, papain, chymopapain, Egyptian milkweed latex extract and pineapple fruit extract, containing fruit bromelain, having the most potent effect. The mechanism of action of these plant enzymes (i.e. an attack on the protective cuticle of the worm) suggests that resistance would be slow to develop in the field. The efficacy and mode of action make plant cysteine proteinases potential candidates for a novel class of anthelmintics urgently required for the treatment of humans and domestic livestock

The anthelmintic efficacy of plant-derived cysteine proteinases against the rodent gastrointestinal nematode, Heligmosomoides polygyrus, in vivo

Gastrointestinal (GI) nematodes are important disease-causing organisms, controlled primarily through treatment with synthetic drugs, but the efficacy of these drugs has declined due to widespread resistance, and hence new drugs, with different modes of action, are required. Some medicinal plants, used traditionally for the treatment of worm infections, contain cysteine proteinases known to damage worms irreversibly in vitro. Here we (i) confirm that papaya latex has marked efficacy in vivo against the rodent gastrointestinal nematode, Heligmosomoides polygyrus, (ii) demonstrate the dosedependent nature of the activity (>90% reduction in egg output and 80% reduction in worm burden at the highest active enzyme concentration of 133 nmol), (iii) establish unequivocally that it is the cysteine proteinases that are the active principles in vivo (complete inhibition of enzyme activity when pre-incubated with the cysteine proteinase-specific inhibitor, E-64) and (iv) show that activity is confined to worms that are in the intestinal lumen. The mechanism of action was distinct from all current synthetic anthelmintics, and was the same as that in vitro, with the enzymes attacking and digesting the protective cuticle. Treatment had no detectable side-effects on immune cell numbers in the mucosa (there was no difference in the numbers of mast cells and goblet cells between the treated groups) and mucosal architecture (length of intestinal villi). Only the infected and untreated mice had much shorter villi than the other 3 groups, which was a consequence of infection and not treatment. Plant-derived cysteine proteinases are therefore prime candidates for development as novel drugs for the treatment of GI nematode infections

Nematicidal effects of cysteine proteinases against sedentary plant parasitic nematodes

Cysteine proteinases from the fruit and latex of plants, such as papaya, pineapple and fig, have previously been shown to have substantial anthelmintic efficacy, in vitro and in vivo, against a range of animal parasitic nematodes. In this paper, we describe the in vitro effects of these plant extracts against 2 sedentary plant parasitic nematodes of the genera Meloidogyne and Globodera. All the plant extracts examined caused digestion of the cuticle and decreased the activity of the tested nematodes. The specific inhibitor of cysteine proteinases, E-64, blocked this activity completely, indicating that it was essentially mediated by cysteine proteinases. In vitro, plant cysteine proteinases are active against second-stage juveniles of M. incognita and M. javanica, and some cysteine proteinases also affect the second-stage juveniles of Globodera rostochiensis. It is not known yet whether these plant extracts will interfere with, or prevent invasion of, host plants

In Vitro and In Vivo Efficacy of Monepantel (AAD 1566) against Laboratory Models of Human Intestinal Nematode Infections

Soil-transmitted helminthiases affect more than one billion people among the most vulnerable populations in developing countries. Currently, control of these infections primarily relies on chemotherapy. Only five drugs are available, all of which have been in use for decades. None of the drugs are efficacious using single doses against all soil-transmitted helminths (STH) species and show low efficacy observed against Trichuris trichiura. In addition, the limited availability of current drug treatments poses a precarious situation should drug resistance occur. Therefore, there is great interest to develop novel drugs against infections with STH. Monepantel, which belongs to a new class of veterinary anthelmintics, the amino-acetonitrile derivatives, might be a potential drug candidate in humans. It has been extensively tested against livestock nematodes, and was found highly efficacious and safe for animals. Here we describe the in vitro and in vivo effect of monepantel, on Ancylostoma ceylanicum, Necator americanus, Trichuris muris, Strongyloides ratti, and Ascaris suum, five parasite-rodent models of relevance to human STH. Since we observed that monepantel showed only high activity on one of the hookworm species and lacked activity on the other parasites tested we cannot recommend the drug as a development candidate for human soil-transmitted helminthiases

Anthelmintic activity of trans-cinnamaldehyde and A- and B-type proanthocyanidins derived from cinnamon (Cinnamomum verum)

Cinnamon (Cinnamomum verum) has been shown to have anti-inflammatory and antimicrobial properties, but effects on parasitic worms of the intestine have not been investigated. Here, extracts of cinnamon bark were shown to have potent in vitro anthelmintic properties against the swine nematode Ascaris suum. Analysis of the extract revealed high concentrations of proanthocyanidins (PAC) and trans-cinnamaldehyde (CA). The PAC were subjected to thiolysis and HPLC-MS analysis which demonstrated that they were exclusively procyanidins, had a mean degree of polymerization of 5.2 and 21% of their inter-flavan-3-ol links were A-type linkages. Purification of the PAC revealed that whilst they had activity against A. suum, most of the potency of the extract derived from CA. Trichuris suis and Oesophagostomum dentatum larvae were similarly susceptible to CA. To test whether CA could reduce A. suum infection in pigs in vivo, CA was administered daily in the diet or as a targeted, encapsulated dose. However, infection was not significantly reduced. It is proposed that the rapid absorption or metabolism of CA in vivo may prevent it from being present in sufficient concentrations in situ to exert efficacy. Therefore, further work should focus on whether formulation of CA can enhance its activity against internal parasites

Bacillus thuringiensis Cry5B Protein Is Highly Efficacious as a Single-Dose Therapy against an Intestinal Roundworm Infection in Mice

Intestinal parasitic nematode diseases infect over one billion people and cause significant disease burden in children (growth and cognitive stunting, malnutrition), in pregnant women, and via their dampening of the immune system in infected individuals. In over thirty years, no new classes of anti-roundworm drugs (anthelmintics) for treating humans have been developed. Because of limitations of the current drugs and the threat of parasite resistance, new anthelmintics are needed. The soil bacterium Bacillus thuringiensis (Bt) produces crystal (Cry) proteins that specifically target and kill insects and nematodes and is used around the world as a safe insecticide. Here we test the effects of the Bt Cry protein Cry5B on a chronic, natural intestinal roundworm infection in mice, namely the helminth parasite Heligmosomoides bakeri. We find that a single dose of Cry5B can eliminate 70% of the parasites and can almost completely block the ability of the parasites to produce progeny. Comparisons of Cry5B's efficacy with known anthelmintics suggest its activity is as good as or perhaps even better than those currently used. Furthermore, this protein is rapidly digested by simulated stomach juices, suggesting that protecting it from these juices would reveal a superior anthelmintic

Biological activities for 'Ficus carica' latex for potential therapeutics in Human Papillomavirus (HPV) related cervical cancers

Abstract Infection caused by high-risk human papillomaviruses (HPVs) are implicated in the aetiology of cervical cancer. Although current methods of treatment for cervical cancer can ablate lesions, preventing metastatic disseminations and excessive tissue injuries still remains a major concern. Hence, development of a safer and more efficient treatment modality is of vital importance. Natural products from plants are one of the principal sources of precursors to lead compounds with direct pharmaceutical application across all disease classes. One of these plants is Ficus carica, whose fruit latex, when applied on HPV-induced skin warts, has shown potential as a possible cure for this virus related lesions. This study explores the in vitro biological activities of fig latex and elucidates its possible mechanisms of action on cervical cancer cell lines CaSki and HeLa positive for HPV type 16 and 18, respectively. Our data shows that fig latex inhibits properties that are associated with HPV-positive cervical cancer transformed cells such as rapid growth and invasion and substantially downregulated the expression of p16 and HPV onco-proteins E6, E7. These findings suggest Ficus carica latex has the potential to be used in the development of therapeutic modalities for the possible treatment, cure and prevention of HPV related cervical cancer

Assessment of the anthelmintic activity of medicinal plant extracts and purified condensed tannins against free-living and parasitic stages of Oesophagostomum dentatum

Background: Plant-derived condensed tannins (CT) show promise as a complementary option to treat gastrointestinal helminth infections, thus reducing reliance on synthetic anthelmintic drugs. Most studies on the anthelmintic effects of CT have been conducted on parasites of ruminant livestock. Oesophagostomum dentatum is an economically important parasite of pigs, as well as serving as a useful laboratory model of helminth parasites due to the ability to culture it in vitro for long periods through several life-cycle stages. Here, we investigated the anthelmintic effects of CT on multiple life-cycles stages of O. dentatum. Methods: Extracts and purified fractions were prepared from five plants containing CT and analysed by HPLC-MS. Anthelmintic activity was assessed at five different stages of the O. dentatum life cycle; the development of eggs to infective third-stage larvae (L3), the parasitic L3 stage, the moult from L3 to fourth-stage larvae (L4), the L4 stage and the adult stage. Results: Free-living larvae of O. dentatum were highly susceptible to all five plant extracts. In contrast, only two of the five extracts had activity against L3, as evidenced by migration inhibition assays, whilst three of the five extracts inhibited the moulting of L3 to L4. All five extracts reduced the motility of L4, and the motility of adult worms exposed to a CT-rich extract derived from hazelnut skins was strongly inhibited, with electron microscopy demonstrating direct damage to the worm cuticle and hypodermis. Purified CT fractions retained anthelmintic activity, and depletion of CT from extracts by pre-incubation in polyvinylpolypyrrolidone removed anthelmintic effects, strongly suggesting CT as the active molecules. Conclusions: These results suggest that CT may have promise as an alternative parasite control option for O. dentatum in pigs, particularly against adult stages. Moreover, our results demonstrate a varied susceptibility of different life-cycle stages of the same parasite to CT, which may offer an insight into the anthelmintic mechanisms of these commonly found plant compounds