Infectivity of amastigotes of Trypanosoma cruzi

Carvalho, Tecia Ulisses de; Souza, Wanderley de

doi:10.1590/S0036-46651986000400001

Abstracts

The infectivity amastigotes of Trypanosoma cruzi, isolated from the supernatant of the J774G8 macrophage-like cell line infected with trypomastigotes to normal macrophages in vitro was tested. After a period of 1 h of T. cruzi-macrophage interaction about 2% of the mouse peritoneal macrophages had ingested amastigotes. In contrast 12% of the macrophages had ingested epimastigotes. Treatment of the amastigotes with trypsin did not interfere with their ingestion by macrophages. Once inside the macrophages the amastigotes divided and after some days transformed into trypomastigotes. When i.p. inoculated into mice the amastigotes were highly infective, inducing high levels of parasitaemia and tissue parasitism. As previously described for trypomastigotes, amastigotes were not lysed when incubated in the presence of fresh guinea-pig serum. Contrasting with what has been described for trypomastigotes, the resistance of amastigotes to complement-mediated lysis persisted after treatment with trypsin.

Chagas'disease; Trypanosoma cruzi; Experimental infection in mice; Infectivity of amastigotes

A infecciosidade da forma amastigota do Trypanosoma cruzi, isolada do sobrenadante de culturas da linhagem tumoral de macrófago J774G8 previamente infectada com formas tripomastigotas, para macrófagos normais manti dos in vitro, foi analisada. Apenas 2% dos macrófagos ingeriram amastigotas quando o período de interação parásito-célula era de 1 hora enquanto que para este mesmo período 12% dos macrófagos ingeriram formas epimastigotas. Tratamento prévio do amastigotas com tripsina não interferiu na sua ingestão por macrófagos. Uma vez no interior dos macrófagos os amastigotas se dividiam e posteriormente transformavam-se em tripomastigotas. A forma amastigota mostrou-se altamente infectiva quando inoculada na cavidade peritoneal de camundongos, induzindo altos níveis parasitêmicos e parasitismo tecidual. Os amastigotas, não são lisados quando incubados na presença de soro fresco de cobaio à semelhança do que tem sido descrito para tripomastigotas. No entanto, ao contrário do que ocorre com tripomastigotas, os amastigotas não são lisados mesmo após prévia tripsinização.

Infectivity of amastigotes of Trypanosoma cruzi

Infecciosidade da forma amastigota do Trypanosoma cruzi

Tecia Ulisses de Carvalho; Wanderley de Souza(^* (* ) For all correspondence. )

Laboratório de Ultra-estrutura Celular. Instituto de Biofísica. Universidade Federal do Rio de Janeiro. Centro de Ciências da Saúde Bloco G. Cidade Universitária CEP 21941 Rio de Janeiro, RJ Brasil

SUMMARY

The infectivity amastigotes of Trypanosoma cruzi, isolated from the supernatant of the J774G8 macrophage-like cell line infected with trypomastigotes to normal macrophages in vitro was tested. After a period of 1 h of T. cruzi-macrophage interaction about 2% of the mouse peritoneal macrophages had ingested amastigotes. In contrast 12% of the macrophages had ingested epimastigotes. Treatment of the amastigotes with trypsin did not interfere with their ingestion by macrophages. Once inside the macrophages the amastigotes divided and after some days transformed into trypomastigotes. When i.p. inoculated into mice the amastigotes were highly infective, inducing high levels of parasitaemia and tissue parasitism. As previously described for trypomastigotes, amastigotes were not lysed when incubated in the presence of fresh guinea-pig serum. Contrasting with what has been described for trypomastigotes, the resistance of amastigotes to complement-mediated lysis persisted after treatment with trypsin.

Key words: Chagas'disease Trypanosoma cruzi Experimental infection in mice Infectivity of amastigotes

RESUMO

A infecciosidade da forma amastigota do Trypanosoma cruzi, isolada do sobrenadante de culturas da linhagem tumoral de macrófago J774G8 previamente infectada com formas tripomastigotas, para macrófagos normais manti dos in vitro, foi analisada. Apenas 2% dos macrófagos ingeriram amastigotas quando o período de interação parásito-célula era de 1 hora enquanto que para este mesmo período 12% dos macrófagos ingeriram formas epimastigotas. Tratamento prévio do amastigotas com tripsina não interferiu na sua ingestão por macrófagos. Uma vez no interior dos macrófagos os amastigotas se dividiam e posteriormente transformavam-se em tripomastigotas. A forma amastigota mostrou-se altamente infectiva quando inoculada na cavidade peritoneal de camundongos, induzindo altos níveis parasitêmicos e parasitismo tecidual. Os amastigotas, não são lisados quando incubados na presença de soro fresco de cobaio à semelhança do que tem sido descrito para tripomastigotas. No entanto, ao contrário do que ocorre com tripomastigotas, os amastigotas não são lisados mesmo após prévia tripsinização.

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Full text available only in PDF format.

ACKNOWLEDGEMENTS

This work has been supported by UNDP/ World Bank/WHO Special Programme for Research and Training in Tropical Diseases, Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Financiadora de Estudos e Projetos (FINEP) and Conselho de Ensino para Graduados da UFRJ.

Recebido para publicação em 22/7/1985.

1. ABRAHAMSON, I. A.; KATZIN, A. M. & MILDER, R. V. A method ior isolating Trypanosoma cruzi amastigotes from spleen and liver using two-step discontinuous gradient centrifugation. J. Parasit., 69: 437-439. 1983.
2. ANDRADE, P, P. de Clonagem de tripanosomatideos em meio nutriente sólido Rio de Janeiro, 1983. (PhD. Thesis Instituto de Biofísica da Universidade Federal do Rio de Janeiro).
3. ARAÚJO, F. G.; HANDMAN, E. & REMINGTON, J. S. Binding of lectins to the cell surface of Trypanosoma cruzi. J. Protozool., 27: 397-400, 1980.
4. ARAÚJO-JORGE, T. C. & SOUZA, W. de Effect of carbohydrates, periodate and enzymes in the process of endocytosis of Trypanosoma cruzi by macrophages. Acta trop. (Basel), 41: 17-28, 1984.
5. BEHEBEHANI, K. Developmental cycles of Trypanosoma (Schizotrypanum) cruzi (Chagas, 1909) in mouse peritoneal macrophages in vitro. Parasitology. 66: 343-353, 1973.
6. CARVALHO, R. M. G.; MEIRELLES, M. N. L.; SOUZA, W. de & LEON, W. Isolation of the intracellular stage of Trypanosoma cruzi and its interaction with mouse macrophages in vitro. Infect. Immun., 33: 546-554, 1981.
7. CARVALHO, T. U. de & SOUZA, W. de Separation of amastigotes and trypomastigotes of Trypanosoma cruzi from cultured cell. Z. Parasitenk., 69: 571-575, 1983.
8. CARVALHO, T. U. de; SOUTO-PADRÓN, T. & SOUZA, W. de Trypanosoma cruzi: surface charge and freezefracture of amastigotes. Exp. Parasit., 59: 12-23, 1985.
9. ENGEL, J. C.; DVORAK, J. A. & SEGURA, E. L. Simple cell-free culture protocol allows production and characterization of Trypanosoma cruzi amastigotes. Molec. Biochem. Parasit., (suppl.): 589, 1982, (Abstract) .
10. GUTTERIDGE, W. E.; COVER, B. & GABORAK, M. Isolation of blood and intracellular forms of Trypanosoma cruzi from rats and other rodents and preliminary studies of their metabolism. Parasitology, 76: 159-176, 1978.
11. HUDSON, L.; SNARY, D. & MORGAN, S. J. - Trypanosoma cruzi: continuous cultivation with murine cell lines. Parasitology, 88: 233-294, 1984.
12. KIPNIS, T, L.; DAVID, J. R.; ALPER, C. A.; SHER, A. & DIAS DA SILVA, W. Enzymatic treatment transforms trypomastigotes of Trypanosoma cruzi into activators of alternative complement pathway and potentiates their uptake by macrophages. Proc. nat. Acad. Sci. (Wash.), 78: 602-605, 1981.
13. LANAR. D. E. Growth and differentiation of Trypanosoma cruzi cultivated with a Triatoma infectans embryo cell line. J. Protozool., 26: 437-462, 1979.
14. McCABE, R. E.; REMINGTON, J. S. & ARAÚJO, P. G. Mechanisms of invasion and replication of the intracellular stage in Trypanosoma cruzi. Infect Immun., 46: 372-376, 1984.
15. MARIA, T. A. Estudo comparativo da ultra-estrutura de amastigotas do Trypanosoma cruzi obtidas do vertebrado e em meio líquido Belo Horizonte, 1975. (MSc Thesis Universidade Federal de Minas Gerais).
16. MUNIZ, J. & BORRIELO, A. Estudo sobre a ação lítica de diferentes soros sobre as formas de cultura e sanguícolas do Schizotrypanum cruzi. Rev. bras. Biol., 5: 563-576, 1945.
17. NOGUEIRA, N.; BIANCO, C. & COHN, Z. Studies on the selective lysis and purification of Trypanosoma cruzi. J. exp. Med., 142: 224-229, 1975.
18. NOGUEIRA, N.; CHAPLAN, S. & COHN, Z. Trypanosoma cruzi: factors modifying ingestion and fate of blood form trypomastigotes. J. exp. Med., 152: 447-451, 1980.
19. NOGUEIRA, N. & COHN, Z. Trypanosoma cruzi: mechanism of entry and intracellular fate in mammalian cells. J. exp. Med., 143: 224-229, 1976.
20. PAN, C. T. Cultivation of the leishmaniform stage of Trypanosoma cruzi in cell-free media at different, temperatures. Amer. J. trop. Med. Hyg., 17: 823-832, 1968.
21. PAN, C. T. Cultivation and morphogenesis of Trypanosoma cruzi in improved liquid media. J. Protozool., 18: 556-560, 1971.
22. PAN, C. T. Trypanosoma cruzi: intracellular stages grown in a cell-free medium at 37°C. Exp. Parasit.. 45: 215-224, 1978a.
23. PAN, S. C. Trypanosoma cruzi: In vitro interactions between cultured amastigotes and human skin-muscle cells. Exp. Parasit., 45: 274-286, 1978b.
24. RIBEIRO, R. D.; BELDA NETO, P. M. & FERRIOLI FILHO, P. Sub-amostras do T. cruzi obtidas por centrifugação diferencial de formas de cultura em meio líquido. I. Amostra Bolívia Comportamento no hospedeiro vertebrado. In: CONGRESSO INTERNACIONAL SOBRE DOENÇA DE CHAGAS, Rio de Janeiro, 1919. Anais. p. 31.
25. SEGURA, E. L.; SUBIAS, E,; ESTEVA, M.
26. UMEZAWA, E. S.; MILDER, R. V. & ABRAHAMSON, I. A. Trypanosoma cruzi amastigotes; development in vitro and infectivity in vivo of the forms isolated from spleen and liver. Acta trop. (Basel), 42: 25-32, 1985.
27. VILLALTA, F. & KIERSZENBAUM, F. Growth of isolated amastigotes of Trypanosoma cruzi in cell free medium. J. Protozool., 29: 570-576, 1982.
28. WARREN, L. Metabolism of Schlzotrypanum cruzi, Chagas. Effect of culture age and substrate concentration on respiratory rate. J. Parasit., 46: 529-539, 1960.

(*

) For all correspondence.

Publication Dates

Publication in this collection
16 Oct 2012
Date of issue
Aug 1986

History

Received
22 July 1985

This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.

[1] 1. ABRAHAMSON, I. A.; KATZIN, A. M. & MILDER, R. V. A method ior isolating Trypanosoma cruzi amastigotes from spleen and liver using two-step discontinuous gradient centrifugation. J. Parasit., 69: 437-439. 1983.

[2] 2. ANDRADE, P, P. de Clonagem de tripanosomatideos em meio nutriente sólido Rio de Janeiro, 1983. (PhD. Thesis Instituto de Biofísica da Universidade Federal do Rio de Janeiro).

[3] 3. ARAÚJO, F. G.; HANDMAN, E. & REMINGTON, J. S. Binding of lectins to the cell surface of Trypanosoma cruzi. J. Protozool., 27: 397-400, 1980.

[4] 4. ARAÚJO-JORGE, T. C. & SOUZA, W. de Effect of carbohydrates, periodate and enzymes in the process of endocytosis of Trypanosoma cruzi by macrophages. Acta trop. (Basel), 41: 17-28, 1984.

[5] 5. BEHEBEHANI, K. Developmental cycles of Trypanosoma (Schizotrypanum) cruzi (Chagas, 1909) in mouse peritoneal macrophages in vitro. Parasitology. 66: 343-353, 1973.

[6] 6. CARVALHO, R. M. G.; MEIRELLES, M. N. L.; SOUZA, W. de & LEON, W. Isolation of the intracellular stage of Trypanosoma cruzi and its interaction with mouse macrophages in vitro. Infect. Immun., 33: 546-554, 1981.

[7] 7. CARVALHO, T. U. de & SOUZA, W. de Separation of amastigotes and trypomastigotes of Trypanosoma cruzi from cultured cell. Z. Parasitenk., 69: 571-575, 1983.

[8] 8. CARVALHO, T. U. de; SOUTO-PADRÓN, T. & SOUZA, W. de Trypanosoma cruzi: surface charge and freezefracture of amastigotes. Exp. Parasit., 59: 12-23, 1985.

[9] 9. ENGEL, J. C.; DVORAK, J. A. & SEGURA, E. L. Simple cell-free culture protocol allows production and characterization of Trypanosoma cruzi amastigotes. Molec. Biochem. Parasit., (suppl.): 589, 1982, (Abstract) .

[10] 10. GUTTERIDGE, W. E.; COVER, B. & GABORAK, M. Isolation of blood and intracellular forms of Trypanosoma cruzi from rats and other rodents and preliminary studies of their metabolism. Parasitology, 76: 159-176, 1978.

[11] 11. HUDSON, L.; SNARY, D. & MORGAN, S. J. - Trypanosoma cruzi: continuous cultivation with murine cell lines. Parasitology, 88: 233-294, 1984.

[12] 12. KIPNIS, T, L.; DAVID, J. R.; ALPER, C. A.; SHER, A. & DIAS DA SILVA, W. Enzymatic treatment transforms trypomastigotes of Trypanosoma cruzi into activators of alternative complement pathway and potentiates their uptake by macrophages. Proc. nat. Acad. Sci. (Wash.), 78: 602-605, 1981.

[13] 13. LANAR. D. E. Growth and differentiation of Trypanosoma cruzi cultivated with a Triatoma infectans embryo cell line. J. Protozool., 26: 437-462, 1979.

[14] 14. McCABE, R. E.; REMINGTON, J. S. & ARAÚJO, P. G. Mechanisms of invasion and replication of the intracellular stage in Trypanosoma cruzi. Infect Immun., 46: 372-376, 1984.

[15] 15. MARIA, T. A. Estudo comparativo da ultra-estrutura de amastigotas do Trypanosoma cruzi obtidas do vertebrado e em meio líquido Belo Horizonte, 1975. (MSc Thesis Universidade Federal de Minas Gerais).

[16] 16. MUNIZ, J. & BORRIELO, A. Estudo sobre a ação lítica de diferentes soros sobre as formas de cultura e sanguícolas do Schizotrypanum cruzi. Rev. bras. Biol., 5: 563-576, 1945.

[17] 17. NOGUEIRA, N.; BIANCO, C. & COHN, Z. Studies on the selective lysis and purification of Trypanosoma cruzi. J. exp. Med., 142: 224-229, 1975.

[18] 18. NOGUEIRA, N.; CHAPLAN, S. & COHN, Z. Trypanosoma cruzi: factors modifying ingestion and fate of blood form trypomastigotes. J. exp. Med., 152: 447-451, 1980.

[19] 19. NOGUEIRA, N. & COHN, Z. Trypanosoma cruzi: mechanism of entry and intracellular fate in mammalian cells. J. exp. Med., 143: 224-229, 1976.

[20] 20. PAN, C. T. Cultivation of the leishmaniform stage of Trypanosoma cruzi in cell-free media at different, temperatures. Amer. J. trop. Med. Hyg., 17: 823-832, 1968.

[21] 21. PAN, C. T. Cultivation and morphogenesis of Trypanosoma cruzi in improved liquid media. J. Protozool., 18: 556-560, 1971.

[22] 22. PAN, C. T. Trypanosoma cruzi: intracellular stages grown in a cell-free medium at 37°C. Exp. Parasit.. 45: 215-224, 1978a.

[23] 23. PAN, S. C. Trypanosoma cruzi: In vitro interactions between cultured amastigotes and human skin-muscle cells. Exp. Parasit., 45: 274-286, 1978b.

[24] 24. RIBEIRO, R. D.; BELDA NETO, P. M. & FERRIOLI FILHO, P. Sub-amostras do T. cruzi obtidas por centrifugação diferencial de formas de cultura em meio líquido. I. Amostra Bolívia Comportamento no hospedeiro vertebrado. In: CONGRESSO INTERNACIONAL SOBRE DOENÇA DE CHAGAS, Rio de Janeiro, 1919. Anais. p. 31.

[25] 25. SEGURA, E. L.; SUBIAS, E,; ESTEVA, M.

[26] 26. UMEZAWA, E. S.; MILDER, R. V. & ABRAHAMSON, I. A. Trypanosoma cruzi amastigotes; development in vitro and infectivity in vivo of the forms isolated from spleen and liver. Acta trop. (Basel), 42: 25-32, 1985.

[27] 27. VILLALTA, F. & KIERSZENBAUM, F. Growth of isolated amastigotes of Trypanosoma cruzi in cell free medium. J. Protozool., 29: 570-576, 1982.

[28] 28. WARREN, L. Metabolism of Schlzotrypanum cruzi, Chagas. Effect of culture age and substrate concentration on respiratory rate. J. Parasit., 46: 529-539, 1960.