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

Published online by Cambridge University Press:  03 May 2007

School of Biology, University Park, University of Nottingham, Nottingham NG7 2RD, UK
School of Biology, University Park, University of Nottingham, Nottingham NG7 2RD, UK
Section of Molecular Medicine, University of Sheffield Medical School, Sheffield S10 2RX, UK
School of Biology, University Park, University of Nottingham, Nottingham NG7 2RD, UK
School of Biology, University Park, University of Nottingham, Nottingham NG7 2RD, UK
*Corresponding author: School of Biology, University of Nottingham, University Park, Nottingham NG7 2RD, UK. Tel: 44 115 951 3208. Fax: 44 115 951 3251. E-mail:


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 dose-dependent 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.

Research Article
Copyright © Cambridge University Press 2007

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Albonico, M., Crompton, D. W. T. and Savioli, L. (1999). Control strategies for human intestinal nematode infections. Advances in Parasitology 42, 277341.Google Scholar
Albonico, M., Bickle, Q., Ramsan, M., Montresor, A., Savioli, L. and Taylor, M. (2003). Efficacy of mebendazole and levamisole alone or in combination against intestinal nematode infections after repeated targeted mebendazole treatment in Zanzibar. Bulletin of the World Health Organization 81, 343352.Google Scholar
Athanasiadou, S., Kyriazakis, I., Jackson, F. and Coop, R. L. (2001). Direct anthelmintic effects of condensed tannins towards different gastrointestinal nematodes of sheep: in vitro and in vivo studies. Veterinary Parasitology 99, 205219.Google Scholar
Bansemir, A. D. and Sukhdeo, M. V. K. (1994). The food resource of adult Heligmosomoides polygyrus in the small intestine. Journal of Parasitology 80, 2428.Google Scholar
Barrett, A. J., Kembhavi, A. A., Brown, M. A., Kirschke, H., Knight, C. G., Tamai, M. and Hanada, K. (1982). L-trans-epoxysuccinyl-leucylamido(4-guanidino)butane (E-64) and its analogues as inhibitors of cysteine proteinases including cathepsins B, H and L. The Biochemical Journal 201, 189198.Google Scholar
Behnke, J. M. and Parish, H. A. (1979). Nematospiroides dubius: arrested development of larvae in immune mice. Experimental Parasitology 47, 116127.Google Scholar
Berger, J. and Asenjo, C. F. (1939). Anthelmintic activity of fresh pineapple juice. Science 90, 299300.Google Scholar
Berger, J. and Asenjo, C. F. (1940). Anthelmintic activity of crystalline papain. Science 91, 387388.Google Scholar
Bryant, V. (1973). The life cycle of Nematospiroides dubius, Baylis, 1926 (Nematoda: Heligmosomoidae). Journal of Helminthology 47, 263268.Google Scholar
Chan, M.-S. (1997). The global burden of intestinal nematode infections – fifty years on. Parasitology Today 13, 438443.Google Scholar
de Clercq, D., Sacko, M., Behnke, J., Gilbert, F., Dorny, P. and Vercruysse, J. (1997). Failure of mebendazole in treatment of human hookworm infections in the southern region of Mali. American Journal of Tropical Medicine and Hygiene 57, 2530.Google Scholar
Coles, G. C. (1995). Chemotherapy of human nematodes: learning from the problems in sheep. Journal of the Royal Society of Medicine 88, 649P651P.Google Scholar
Coles, G. C. (2005). Anthelmintic resistance–looking to the future: a UK perspective. Research in Veterinary Science 78, 99108.Google Scholar
Coles, G. C., Warner, A. K. and Best, J. R. (1996). Triple resistant Ostertagia from angora goats. Veterinary Record 139, 299300.Google Scholar
Dekeyser, P. M., Buttle, D. J., Devreese, B., van Beeumen, J., Demeester, J. and Lauwers, A. (1995). Kinetic constants for the hydrolysis of aggrecan by the papaya proteinases and their relevance for chemonucleolysis. Archives of Biochemistry and Biophysics 320, 375379.Google Scholar
Geerts, S. and Gryseels, B. (2000). Drug resistance in human helminths: current situation and lessons from livestock. Clinical and Microbiological Reviews 13, 207222.Google Scholar
Gill, J. H. and Lacey, E. (1998). Avermectin/milbemycin resistance in trichostrongyloid nematodes. International Journal for Parasitology 28, 863877.Google Scholar
Githiori, J. B., Höglund, J., Waller, P. J. and Baker, R. L. (2004). Evaluation of anthelmintic properties of some plants used as livestock dewormers against Haemonchus contortus infections in sheep. Parasitology 129, 245253.Google Scholar
Hale, L. P. (2004). Proteolytic activity and immunogenicity of oral bromelain within the gastrointestinal tract of mice. International Immunopharmacology 4, 255264.Google Scholar
Hansson, A., Veliz, G., Naquira, C., Amren, M., Arroyo, M. and Arevalo, G. (1986). Preclinical and clinical studies with latex from Ficus glabrata HBK, a traditional intestinal anthelmintic in the Amazonian area. Journal of Ethnopharmacology 17, 105138.Google Scholar
Hong, C., Hunt, K. R. and Coles, G. C. (1996). Occurrence of anthelmintic resistant nematodes on sheep farms in England and goat farms in England and Wales. Veterinary Record 139, 8386.Google Scholar
Martin, R. J. and Robertson, A. P. (2000). Electrophysiological investigation of anthelmintic resistance. Parasitology 120, (Suppl.) S87S94.Google Scholar
Molyneux, D. H., Hotez, P. J. and Fenwick, A. (2005). “Rapid-impact interventions”: How a policy of integrated control for Africa's neglected tropical diseases could benefit the poor. PLoS Medicine 2, 101107.Google Scholar
Mwamachi, D. M., Audho, J. O., Thorpe, W. and Baker, R. L. (1995). Evidence for multiple anthelmintic resistance in sheep and goats reared under the same management in coastal Kenya. Veterinary Parasitology 60, 303313.Google Scholar
Nelde, A., Teufel, M., Hahn, C., Duschl, A., Sebald, W., Bröcker, E. B. and Grunewald, S. M. (2001). The impact of the route and frequency of antigen exposure on the IgE response in allergy. International Archives of Allergy and Immunology 124, 461469.Google Scholar
Nieuwhof, G. J. and Bishop, S. C. (2005). Costs of the major endemic diseases of sheep in Great Britain and the potential benefits of reduction in disease impact. Animal Science 81, 2329.Google Scholar
Paolini, V., Frayssines, A., de la Farge, F., Dorchies, P. and Hoste, H. (2003). Effects of condensed tannins on established populations and on incoming larvae of Trichostrongylus colubriformis and Teladorsagia circumcincta in goats. Veterinary Research 34, 331339.Google Scholar
Reynoldson, J. A., Behnke, J. M., Pallant, L. J., MacNish, M. G., Gilbert, F., Giles, S., Spargo, R. J. and Thompson, R. C. A. (1997). Failure of pyrantel in treatment of human hookworm infections (Ancylostoma duodenale) in the Kimberley region of North West Australia. Acta Tropica 68, 301312.Google Scholar
Robbins, B. H. (1930). A proteolytic enzyme in ficin, the anthelmintic principle of Leche de Higueron. Journal of Biological Chemistry 87, 251257.Google Scholar
Rowan, A. D., Buttle, D. J. and Barrett, A. J. (1990). The cysteine proteinases of the pineapple plant. The Biochemical Journal 266, 869875.Google Scholar
Salih, E., Malthouse, J. P. G., Kowlessur, D., Jarvis, M., O'Driscoll, M. and Brocklehurst, K. (1987). Differences in the chemical and catalytic characteristics of two crystallographically “identical” enzyme catalytic sites. Characterisation of actinidin and papain by a combination of pH-dependent substrate catalysis kinetics and reactivity probe studies targeted on the catalytic-site thiol group and its immediate environment. The Biochemical Journal 247, 181193.Google Scholar
Sangster, N. C. and Gill, J. (1999). Pharmacology of anthelmintic resistance. Parasitology Today 15, 141146.Google Scholar
Satrija, F., Nansen, P., Bjorn, H., Murtini, S. and He, S. (1994). Effect of papaya latex against Ascaris suum in naturally infected pigs. Journal of Helminthology 68, 343346.Google Scholar
Satrija, F., Nansen, P., Murtini, S. and He, S. (1995). Anthelmintic activity of papaya latex against patent Heligmosomoides polygyrus infections in mice. Journal of Ethnopharmacology 48, 161164.Google Scholar
Soto-Mera, M. T., López-Rico, M. R., Filgueira, J. F., Villamil, E. and Cidrás, R. (2000). Occupational allergy to papain. Allergy 55, 983984.Google Scholar
Stepek, G., Behnke, J. M., Buttle, D. J. and Duce, I. R. (2004). Natural plant cysteine proteinases as novel anthelmintics? Trends in Parasitology 20, 322327.Google Scholar
Stepek, G., Buttle, D. J., Duce, I. R., Lowe, A. and Behnke, J. M. (2005). Assessment of the anthelmintic effect of natural plant cysteine proteinases against the gastrointestinal nematode, Heligmosomoides polygyrus, in vitro. Parasitology 130, 203211.Google Scholar
Stepek, G., Lowe, A. E., Buttle, D. J., Duce, I. R. and Behnke, J. M. (2006). In vitro and in vivo anthelmintic efficacy of plant cysteine proteinases against the rodent gastrointestinal nematode, Trichuris muris. Parasitology 132, 681689.Google Scholar
Stepek, G., Lowe, A. E., Buttle, D. J., Duce, I. R. and Behnke, J. M. (2007). Anthelmintic action of plant cysteine proteinases against the rodent stomach nematode, Protospirura muricola, in vitro and in vivo. Parasitology 134, 103112.Google Scholar
Tagboto, S. and Townson, S. (2001). Antiparasitic properties of medicinal plants and other naturally occurring products. Advances in Parasitology 50, 199295.Google Scholar
Vandamme, T. F. and Ellis, K. J. (2004). Issues and challenges in developing ruminal drug delivery systems. Advanced Drug Delivery Reviews 56, 14151436.Google Scholar
van Wyk, J. A., Malan, F. S. and Randles, J. L. (1997). How long before resistance makes it impossible to control some field strains of Haemonchus contortus in South Africa with any of the modern anthelmintics? Veterinary Parasitology 70, 111122.Google Scholar
Varady, M., Praslicka, J., Corba, J. and Vesely, L. (1993). Multiple anthelmintic resistance of nematodes in imported goats. Veterinary Record 132, 387388.Google Scholar
Waller, P. J. (1986). Anthelmintic resistance in nematode parasites of sheep. Agricultural Zoology Reviews 1, 333373.Google Scholar
Waller, P. J. (1997). Nematode parasite control of livestock in the tropics/subtropics: the need for novel approaches. International Journal for Parasitology 27, 11931201.Google Scholar
Waller, P. J., Bernes, G., Thamsborg, S. M., Sukura, A., Richter, S. H., Ingebrigtsen, K. and Höglund, J. (2001). Plants as de-worming agents of livestock in the Nordic countries: historical perspective, popular beliefs and prospects for the future. Acta Veterinaria Scandinavica 42, 3144.Google Scholar
Zucker, S., Buttle, D. J., Nicklin, M. J. H. and Barrett, A. J. (1985). The proteolytic activities of chymopapain, papain, and papaya proteinase III. Biochimica et Biophysica Acta 828, 196204.Google Scholar