Folia Parasitologica, vol. 61 (2014), issue 2

Folia Parasitologica 61[2] 113-119 (2014) | 10.14411/fp.2014.017

Spontaneous cystogenesis of Toxoplasma gondii in feline epithelial cells in vitro

Renata M. de Muno1, Marcos A. Moura1, Luciany C. de Carvalho1, Sergio H. Seabra2, Helene S. Barbosa1
1 Laboratório de Biologia Estrutural, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, RJ, Brazil;
2 Laboratório de Tecnologia em Cultura de Células, Centro Universitário Estadual da Zona Oeste, RJ, Brazil

Toxoplasma gondii Nicolle et Manceaux, 1908 is an obligate intracellular parasite with the ability to infect mammals and birds. The only definitive hosts for T. gondii are felids, as the parasites form immature oocysts that are shed in the faeces. Here we introduce cat cells as a model for the study of experimental toxoplasmosis. We selected epithelial cells derived from cat kidneys (CRFK) as a target to determine the intracellular fate of bradyzoites of the T. gondii ME49 strain. In parallel, we compared this infection using epithelial cells from the rat intestine (IEC-6), considering the enteroepithelial development that occurs in the cat. Different ratios of parasites to host cells were assayed over the course of a 14-day-infection. The intracellular development of T. gondii was dependent on the source of the epithelial cells and also on the parasite/host cell ratio. Cystogenesis was well established in the CRFK cell line at a ratio of 1 : 10 after 10-14 days of infection. This cellular model system opens a new field of investigation into the molecular aspects of the interactions between T. gondii and feline epithelial cells. The CRFK cell line appears to be a potential cellular model for large scale cyst production in vitro, which would allow a reduction in the number of animals used and/or replacement of animals by in vitro cultures.

Keywords: bradyzoites, felids cells, tachyzoites, tissue cysts in vitro, toxoplasmosis

Received: July 16, 2013; Accepted: November 11, 2013; Prepublished online: January 10, 2014; Published: April 14, 2014


References

  1. Balls M. 2007: Animal experimentation and the Three Rs: the need for hard heads and soft hearts and to confront the Three "I"s. Altern. Lab. Anim. 35: 547-548 Go to PubMed...
  2. Black M.W., Boothroyd J.C. 2000: Lytic cycle of Toxoplasma gondii. Microbiol. Mol. Biol. Rev. 64: 607-623 Go to original source... Go to PubMed...
  3. Cantin A., Woods D.E. 1930: Protection by antibiotics against myeloperoxidase-dependent cytotoxicity to lung epithelial cells in vitro. J. Clin. Invest. 91: 38-45 Go to original source...
  4. Crandell R.A., Fabricant C.G., Nelson-rees W.A. 1973: Development, characterization, and viral susceptibility of a feline (Felis catus) renal cell line (CRFK). In Vitro 9: 176-185 Go to original source... Go to PubMed...
  5. Darde M.L., Bouteille B., Leboutet M.J., Loubet A., Pestre-Alexandre M. 1989: Toxoplasma gondii: ultrastructural study of cystic formations observed in human fibroblast culture. Ann. Parasitol. Hum. Comp. 64: 403-411 Go to original source... Go to PubMed...
  6. Dubey J.P. 1973. Feline toxoplasmosis and coccidiosis: a survey of domiciled and stray cats. J. Am. Vet. Med. Assoc. 162: 873-877 Go to PubMed...
  7. Dubey J.P. 1998: Advances in the life cycle of Toxoplasma gondii. Int. J. Parasitol. 28: 1019-1024 Go to original source... Go to PubMed...
  8. Dubey J.P. 2006: Comparative infectivity of oocysts and bradyzoites of Toxoplasma gondii for intermediate (mice) and definitive (cats) hosts. Vet. Parasitol. 140: 69-75 Go to original source... Go to PubMed...
  9. Dubey J.P., Frenkel, J.K. 1972: Cyst-induced toxoplasmosis in cats. J. Protozool. 19: 155-177 Go to original source... Go to PubMed...
  10. Dubey J.P., Frenkel J.K. 1973: Experimental Toxoplasma infection in mice with strains producing oocysts. J. Parasitol. 59: 505-512 Go to original source... Go to PubMed...
  11. Dubey J.P., Lindsay D.S., Speer C.A. 1998: Structures of Toxoplasma gondii tachyzoites, bradyzoites, and sporozoites and biology and development of tissue cysts. Clin. Microbiol. Rev. 11: 267-299 Go to PubMed...
  12. Dubey J.P., Navarro I.T., Sreekumar C., Dahl E., Freire R.L., Kawabata HH, Vianna M.C., Kwok O.C., Shen S.K., Thulliez P., Lehmann T. 2004: Toxoplasma gondii infections in cats from Parana, Brazil: seroprevalence, tissue distribution, and biologic and genetic characterization of isolates. J. Parasitol. 90: 721-726 Go to original source... Go to PubMed...
  13. Dzierszinski F., Nishi, M., Ouko L., Roos D.S. 2004: Dynamics of Toxoplasma gondii differentiation. Eukaryot. Cell 3: 992-1003 Go to original source... Go to PubMed...
  14. Ferguson D.J. 2004: Use of molecular and ultrastructural markers to evaluate stage conversion of Toxoplasma gondii in both the intermediate and definitive host. Int. J. Parasitol. 34: 347-360 Go to original source... Go to PubMed...
  15. Ferguson D.J. 2009: Toxoplasma gondii: 1908-2008, homage to Nicolle, Manceaux and Splendore. Mem. Inst. Oswaldo Cruz 104: 133-148 Go to original source... Go to PubMed...
  16. Ferguson D.J., Hutchison W.M., Dunachie J.F., Siim J.C. 1974: Ultrastructural study of early stages of asexual multiplication and microgametogony of Toxoplasma gondii in the small intestine of the cat. Acta Pathol. Microbiol. Scand. B. Microbiol. Immunol. 82: 167-181 Go to PubMed...
  17. Ferguson D.J., Hutchison W.M., Siim J.C. 1975: The ultrastructural development of the macrogamete and formation of the oocyst wall of Toxoplasma gondii. Acta Pathol. Microbiol. Scand. B. 5: 491-505
  18. Ferreira da Silva M.F., Barbosa H.S., Gross U., Lueder C.G. 2008: Stress-related and spontaneous stage differentiation of Toxoplasma gondii. Mol. Biosyst. 4: 824-834 Go to original source... Go to PubMed...
  19. Ferreira da Silva M.F., Rodrigues R.M., Andrade, E.F., Carvalho, L., Gross U., Lueder C.G., Barbosa H.S. 2009a: Spontaneous stage differentiation of mouse-virulent Toxoplasma gondii RH parasites in skeletal muscle cells: an ultrastructural evaluation. Mem. Inst. Oswaldo Cruz 140: 196-200 Go to original source... Go to PubMed...
  20. Ferreira da Silva M.F., Takacs A.C., Barbosa H.S., Gross U., Lueder C.G. 2009b: Primary skeletal muscle cells trigger spontaneous Toxoplasma gondii tachyzoite-to-bradyzoite conversion at higher rates than fibroblasts. Int. J. Med. Microbiol. 299: 281-288 Go to original source... Go to PubMed...
  21. Fischer H.G., Nitzgen B., Reichmann G., Gross U., Hadding U. 1997: Host cells of Toxoplasma gondii encystation in infected primary culture from mouse brain. Parasitol. Res. 83: 637-641 Go to original source... Go to PubMed...
  22. Flood P.P. 1975: Scanning electron microscope observations on the muscle innervation of Oikopleura dioica Fol (Appendicularia, Tunicata) with notes on the arrangement of connective tissue fibres. Cell Tissue Res. 164: 357-369 Go to original source... Go to PubMed...
  23. Freyre A. 1995: Separation of Toxoplasma cysts from brain tissue and liberation of viable bradyzoites. J. Parasitol. 81: 1008-1010 Go to original source... Go to PubMed...
  24. Guimaraes E.V., Acquarone M., de Carvalho L., Barbosa H.S. 2007: Anionic sites on Toxoplasma gondii tissue cyst wall: expression, uptake and characterization. Micron 38: 651-658 Go to original source... Go to PubMed...
  25. Guimaraes E.V., de Carvalho L., Barbosa H.S. 2008: Primary culture of skeletal muscle cells as a model for studies of Toxoplasma gondii cystogenesis. J. Parasitol. 94: 72-83 Go to original source... Go to PubMed...
  26. Hoff R.L., Dubey J.P., Behbehani A.M., Frenkel J.K. 1977: Toxoplasma gondii cysts in cell culture: new biologic evidence. J. Parasitol. 63: 1121-1124 Go to original source... Go to PubMed...
  27. Hogan M.J., Yoneda C., Feeney L., Zweigart P., Lewis A. 1960: Morphology and culture of Toxoplasma. Arch. Ophthalmol. 64: 655-667 Go to original source... Go to PubMed...
  28. Jones T.C., Bienz K.A., Erb P. 1986: In vitro cultivation of Toxoplasma gondii cysts in astrocytes in the presence of gamma interferon. Infect. Immun. 51: 147-156 Go to PubMed...
  29. Lehmann T., Blackston C.R., Parmley S.F., Remington J.S., Dubey J.P. 2000: Strain typing of Toxoplasma gondii: comparison of antigen-coding and housekeeping genes. J. Parasitol. 86: 960-971 Go to original source... Go to PubMed...
  30. Lindsay D.S., Blagburn B.L., Dubey J.P., Mason W.H. 1991: Prevalence and isolation of Toxoplasma gondii from white-tailed deer in Alabama. J. Parasitol. 77: 62-64 Go to original source... Go to PubMed...
  31. Matsubayashi H., Akao S. 1963: Morphological studies on the development of the Toxoplasma cyst. Am. J. Trop. Med. Hyg. 12: 321-333 Go to original source... Go to PubMed...
  32. McHugh T.D., Gbewonyo A., Johnson J.D., Holliman R.E., Butcher P.D. 1993: Development of an in vitro model of Toxoplasma gondii cyst formation. FEMS Microbiol. Lett. 114: 325-332 Go to original source... Go to PubMed...
  33. Moura M.A., Amendoeira M.R., Barbosa H.S. 2009: Primary culture of intestinal epithelial cells as a potential model for Toxoplasma gondii enteric cycle studies. Mem. Inst. Oswaldo Cruz 104: 862-864 Go to original source... Go to PubMed...
  34. Popiel I., Gold M.C., Booth K.S. 1996. Quantification of Toxoplasma gondii bradyzoites. J. Parasitol. 82: 330-332 Go to original source... Go to PubMed...
  35. Shimada K., O'connor G.R., Yoneda C. 1974: Cyst formation by Toxoplasma gondii (RH strain) in vitro. The role of immunologic mechanisms. Arch. Ophthalmol. 92: 496-500 Go to original source... Go to PubMed...
  36. Soete M., Camus D., Dubremetz J.F. 1994: Experimental induction of bradyzoite-specific antigen expression and cyst formation by the RH strain of Toxoplasma gondii in vitro. Exp. Parasitol. 78: 361-370 Go to original source... Go to PubMed...
  37. Speer C.A., Dubey J.P. 2005: Ultrastructural differentiation of Toxoplasma gondii schizonts (types B to E) and gamonts in the intestines of cats fed bradyzoites. Int. J. Parasitol. 35: 193-206 Go to original source... Go to PubMed...
  38. Weiss L.M., Kim K. 2000: The development and biology of bradyzoites of Toxoplasma gondii. Front. Biosci. 5: 391-405 Go to original source... Go to PubMed...
  39. Weiss L.M., Laplace D., Takvorian P.M., Tanowitz H.B., Cali A., Wittner M.A. 1995: A cell culture system for study of the development of Toxoplasma gondii bradyzoites. J. Eukaryot. Microbiol. 42: 150-157 Go to original source... Go to PubMed...
  40. Zhang Y.W., Halonen S.K., Ma Y.F., Tanowtiz H.B., Weiss L.M. 2010: A purification method for enrichment of the Toxoplasma gondii cyst wall. J. Neuroparasitol. 1: 1-6 Go to original source...
  41. Zhang Y.W., Halonen S.K., Ma Y.F., Wittner M., Weiss L.M. 2001: Initial characterization of CST1, a Toxoplasma gondii cyst wall glycoprotein. Infect. Immun. 69: 501-507 Go to original source... Go to PubMed...
  42. Zhang Y.W., Halonen S.K., Ma Y.F., Wittner M., Weiss L.M. 2001: Initial characterization of CST1, a Toxoplasma gondii cyst wall glycoprotein. Infect. Immun. 69: 501-507 Go to original source... Go to PubMed...