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Isolation, fractionation and partial characterization of the tegumental surface from protoscoleces of the hydatid organism, Echinococcus granulosus

Published online by Cambridge University Press:  06 April 2009

D. P. McManus  and
N. J. Barrett
D. P. McManus
Affiliation:
Department of Pure and Applied Biology, Imperial College of Science and Technology, London SW7 2BB
N. J. Barrett
Affiliation:
Department of Pure and Applied Biology, Imperial College of Science and Technology, London SW7 2BB
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Extract

Several approaches were adopted for the disruption and removal of the tegumental surface from protoscoleces of the horse strain of the hydatid organism, Echinococcus granulosus. The effectiveness of each method and the purity of subsequent microthrix-enriched fractions obtained by differential centrifugation were evaluated by electron microscopy, by the amount of protein released and by the degree of enrichment of surface plasma membrane marker enzymes. Incubation in saponin for 10 min produced the purest microtriche preparation, but in low yield; freeze/thawing, incubation in Triton X-100 for 10 mm or in saponin for 20 min produced fractions containing significant amounts of relatively pure microtriches, but mild homogenization was a poor method for surface disruption and subsequent isolation of microtriches. Phosphodiesterase, adenosine triphosphatase (total and ouabain-inhibited), leucine aminopeptidase and glutamyltransferase were active in the protoscoleces but none were enriched in any of the microthrix fractions. In contrast, alkaline phosphatase, acid phosphatase, 5′ nucleotidase and maltase were enriched significantly in all of the isolated microtriche preparations, which suggests that these enzymes are predominantly surface membrane bound. The protein profiles of the microthrix-enriched fractions, following SDS—PAGE, were basically similar, although there were some qualitative and quantitative differences in the proteins released by each isolation procedure. Three major PAS-staining components were present in all the preparations and these probably originated from the glycocalyx. One of these PAS-positive components, with an approximate molecular weight of 110 kDa, may be a glycoprotein specific to the horse strain of E. granulosus.

Type
Research Article
Information
Parasitology , Volume 90 , Issue 1 , February 1985 , pp. 111 - 129
Copyright
Copyright © Cambridge University Press 1985

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References

REFERENCES

Atkinson, B. G. & Podesta, R. B. (1982). Two dimensional electrophoretic separation and fluorographic analysis of the gene products synthesised by Hymenolepis diminuta with particular reference to the parasite-specific polypeptides in the brush border membrane. Molecular and Biochemical Parasitology 6, 33–42.Google Scholar
Ames, B. N. (1966). Assay of inorganic phosphate, total phosphate and phosphatases. In Methods in Enzynmlogy, vol. 8 (ed. Colowick, S. P. and Kaplan, N. O.), pp. 115–18. New York and Amsterdam: Academic Press.Google Scholar
Befus, A. D. & Podesta, R. B. (1976). Intestine. In Ecological Aspects of Parasitology (ed. Kennedy, C. R.), pp. 303–25. Amsterdam: North Holland.Google Scholar
Boutetlle, M., Dubuy-Coin, A. M. & Moyne, G. (1975). Techniques of localisation of proteins and nucleoproteins in the cell nucleus by high resolution autoradiography and cytochemistry. In Methods in Enzymology, Vol. 40 (ed. Colowick, S. P. and Kaplan, N. O.), pp. 3–41. New York and London: Academic Press.Google Scholar
Bråten, T. (1968). The fine structure of the tegurnent of Diphyllobothrium latum. A comparison of the plerocercoid and adult stages. Zeitschrifl für Parasitenkunde 30, 104–12.Google Scholar
Coltorti, E. A. & Varela-Diaz, V. M. (1974). Echinococcus granulosus: penetration of macromolecules and their localisation on the parasite membranes of cysts. Experimental Parasitology 35, 225–31.CrossRefGoogle Scholar
Coltorti, E. A. & Varela-Diaz, V. M. (1975). Penetration of host IgG molecules into hydatid cysts. Zeitschrift für Parasitenkunde 48, 47–51.Google Scholar
Evans, W. H. (1982). Subcellular membranes and isolated organelles: preparative techniques and criteria for purity. In Techniques in Lipid and Membrane Biochemistry, Part 1, B40/a, pp. 1–46. County Clare, Ireland: Elsevier/North Holland.Google Scholar
Friedman, P. A., Weinstein, P. P., Davidson, L. A. & Mueller, J. F. (1982). Spirometra mansonoides: Lectin analysis of tegumental glycopeptides. Experimental Parasitology 54, 93–103.Google Scholar
Gamble, H. R. & Pappas, P. W. (1981). Type 1 phosphodiesterase in the isolated brush border membrane of Hymenolepis diminuta. Journal of Parasitology 67, 617–22.Google Scholar
Hopkins, C. A., Law, L. M. & Threadgold, L. T. (1978). Schistocephalus solidus: pinocytosis by the plerocercoid tegument. Experimental Parasitology 44, 161–72.Google Scholar
Hustead, S. T. & Williams, J. F. (1977). Permeability studies on taeniid metacestodes: 1. Uptake of proteins by larval stages of Taenia taeniaeformis, T. crassiceps and Echinococcus granulosus. Journal of Parasitologoy 63, 314–21.Google Scholar
Khorsanni, H. O. & Tabibi, V. (1978). Similarities of human hydatid cyst fluid components and the host serum. Acta Medica Iranica 21, 161–72.Google Scholar
Kilejian, A., Schinazi, L. A. & Schwabe, C. W. (1961). Host–parasite relationships in Echinococcosis. V. Histochemical observations on Echinococcus granulosus. Journal of Parasitology 47, 181–5.Google Scholar
Knowles, W. J. & Oaks, J. A. (1979). Isolation and partial biochemical characterisation of the brush border plasma membrane of the cestode Hymenolepis diminuta. Journal of Parasitology 65, 7157–31.CrossRefGoogle ScholarPubMed
Kusel, J. R. (1972). Protein composition and protein synthesis in the surface membranes of Shistosoma mansoni. Parasitology 65, 55–69.CrossRefGoogle Scholar
Laemmli, U. K. (1970). Cleavage of structural proteins during the assembly of the head of the bacteriophage T4. Nature, London 227, 680–5.Google Scholar
Lumsden, R. D. (1975). Surface ultrastructure and cytochemistry of parasitic helminths. Experimental Parasitology 37, 267–339.CrossRefGoogle ScholarPubMed
Lumsden, R. D. & Murphy, V. A. (1980). Morphological and functional aspects of the cestode surface. In Cellular Interactions in Symbiosis and Parasitism (ed. Cook, C. B., Pappas, P. W. and Rudolph, E. D.), pp. 95–130. Columbus: Ohio State University Press.Google Scholar
Lumsden, R. D., Voge, M. & Sogandares-Bernal, F. (1982). The metacestode tegument: fine structure, topochemistry, and interactions with the host. In Cysticercosis: Present State of Knowledge and Perspectives (ed. Fusser, A., Wilims, K., Laclette, J. P., Larralde, C., Ridaura, C. and Beltran, F.), pp. 307–61. New York and London: Academic Press.Google Scholar
Markwell, M. A. K., Haas, S. M., Bieber, L. L. & Tolbert, N. E. (1978). A modification of the Lowry procedure to simplify protein determination in membrane and lipoprotein samples. Analytical Biochemistry 87, 206–10.CrossRefGoogle ScholarPubMed
McManus, D. P. & Macpherson, C. N. L. (1984). Strain characterisation in the hydatid organism, Echinococcus granulosus: current status and new perspectives. Annals of Tropical Medicine and Parasitology 78, 193–8.CrossRefGoogle ScholarPubMed
McManus, D. P. & Smyth, J. D. (1978). Differences in the chemical composition and carbohydrate metabolism of Echinococcus granulosus (horse and sheep strains) and E. multilocularis. Parasitology 77, 103–9.Google Scholar
McManus, D. P. & Smyth, J. D. (1979). Isoelectric focusing of some enzymes from Echinococcus granulosus (horse and sheep strains) and E. multilocularis. Transactions of the Royal Society of Tropical Medicine and Hygiene 73, 259–65.CrossRefGoogle ScholarPubMed
Morseth, D. (1967). Fine structure of the hydatid cyst and protoscolex of Echinococcus granulosus. Journal of Parasitology 53, 312–25.CrossRefGoogle ScholarPubMed
Oaks, J. A., Knowles, W. J. & Cain, G. D. (1977). A simple method of obtaining an enriched fraction of tegumental brush border from Hymenolepis diminuta. Journal of Parasitology 63, 479–85.CrossRefGoogle ScholarPubMed
Pappas, P. W. (1980 a). Structure, function and biochemistry of the cestode tegurnentary membrane and associated glycocalyx. In Endocytobiology (ed. Schwemmler, W. and Schenk, H. E. A.), pp. 587–603. Berlin and New York: Walter de Gruyter & Co.Google Scholar
Pappas, P. W. (1980 b). Phosphohydrolase activity of the isolated brush-border membrane of Hymenolepis diminuta (Cestoda) following sodium dodeeyl sulfate (SDS)-polyacrylamide gel electrophoresis. Journal of Parasitology 66, 914–19.Google Scholar
Pappas, P. W. (1981). Hymenolepis diminuta: partial characterisation of membrane bound nucleotidase activities (ATPase and 5′ nucleotidase) in the isolated brush border membrane. Experimental Parasitology 51, 209–19.Google Scholar
Pappas, P. W. & Narcisi, E. M. (1982). A comparison of membrane-bound enzymes of the isolated brush border plasma membranes of the cestodes Hymenolepis diminuta and H. microstoma. Parasitology 84, 39 1–6.Google Scholar
Rahman, M. S., Mettricx, P. F. & Podesta, R. B. (1981 a). Use of saponin in the preparation of brush border from a parasitic flatworm. Canadian Journal of Zoology 59, 911–17.Google Scholar
Rahman, M. S., Mettrick, P. F. & Podesta, R. B. (1981 b). Properties of a Mg2+-dependent, Ca2+-inhibited ATPase localised in the brush border of the surface epithelium of a parasitic flatworm. Canadian Journal of Zoology 59, 918–23.Google Scholar
Read, C. P. & Rothman, A. (1958). The role of carbohydrates in the biology of cestodes. VI. The carbohydrates metabolized in vitro by some cyclophyllidean species. Experimental Parasitology 7, 217–23.CrossRefGoogle ScholarPubMed
Roberts, S. M., MacGregor, A. N., Vojvodic, M., Wells, E., Crabtree, J. E. & Wilson, R. A.. (1983). Tegument surface membranes of adult Schistosoma mansoni: development of a method for their isolation. Molecular and Biochemical Parasitology 9, 105–27.CrossRefGoogle ScholarPubMed
Simpson, A. J. G., Schryer, M. P., Cesari, I. M., Evans, W. H. & Smithers, S. R. (1981). Isolation and partial characterisation of the tegumental outer membrane of adult Schistosoma mansoni. Parasitology 83, 163–77.CrossRefGoogle ScholarPubMed
Smyth, J. D. (1969). The Physiology of Cestodes. San Francisco: W. H. Freeman & Co.; Edinburgh and London: Oliver and Boyd.Google Scholar
Smyth, J. D. (1977). Strain differences in Echinococcus granulosus, with special reference to the status of equine hydatidosis in the United Kingdom. Transactions of the Royal Society of Tropical Medicine and Hygiene 71, 93–100.Google Scholar
Smyth, J. D. & Davies, Z. (1974). In vitro culture of the strobilar stage of Echinococcus granulosus (sheep strain): a review of basic problems and results. International Journal for Parasitology 4, 631–44.Google Scholar
Thompson, R. C. A., Houghton, A. & Zaman, V. (1982). A study of the microtriches of adult Echinococcus granulosus by scanning electron microscopy. International Journal for Parasitology 12, 579–83.Google Scholar
Threadgold, L. T. & Dunn, J. (1983). Taenia crassiceps: regional variations in ultrastructure and evidence of endocytosis in the cysticercus tegument. Experimental Parasitology 55, 121–31.CrossRefGoogle ScholarPubMed
Threadgold, L. T. & Hopkins, C. A. (1981). Schistocephalus solidus and Ligula intestimalis: pinocytosis by the tegument. Experimental Parasitology 51, 444–56.Google Scholar