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. 1995 Sep;63(9):3559–3566. doi: 10.1128/iai.63.9.3559-3566.1995

Leishmania pifanoi amastigote antigens protect mice against cutaneous leishmaniasis.

L Soong 1, S M Duboise 1, P Kima 1, D McMahon-Pratt 1
PMCID: PMC173494  PMID: 7642292

Abstract

In the search for a leishmaniasis vaccine, extensive studies have been carried out with promastigote (insect stage) molecules. Information in this regard on amastigote (mammalian host stage) molecules is limited. To investigate host immune responses to Leishmania amastigote antigens, we purified three stage-specific antigens (A2, P4, and P8) from in vitro-cultivated amastigotes of Leishmania pifanoi by using immunoaffinity chromatography. We found that with Corynebacterium parvum as an adjuvant, three intraperitoneal injections of 5 micrograms of P4 or P8 antigen provided partial to complete protection of BALB/c mice challenged with 10(5) to 10(7) L. pifanoi promastigotes. These immunized mice developed significantly smaller or no lesions and exhibited a 39- to 1.6 x 10(5)-fold reduction of lesion parasite burden after 15 to 20 weeks of infection. In addition, P8 immunization resulted in complete protection against L. amazonensis infection of CBA/J mice and partial protection of BALB/c mice, suggesting that this antigen provided cross-species protection of mice with different H-2 haplotypes. At different stages during infection, vaccinated mice exhibited profound proliferative responses to parasite antigens and increased levels of gamma interferon production, suggesting that a Th1 cell-mediated immune response is associated with the resistance in these mice. Taken together, the data in this report indicate the vaccine potential of amastigote-derived antigens.

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Selected References

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  1. Afonso L. C., Scharton T. M., Vieira L. Q., Wysocka M., Trinchieri G., Scott P. The adjuvant effect of interleukin-12 in a vaccine against Leishmania major. Science. 1994 Jan 14;263(5144):235–237. doi: 10.1126/science.7904381. [DOI] [PubMed] [Google Scholar]
  2. Antunes C. M., Mayrink W., Magalhaes P. A., Costa C. A., Melo M. N., Dias M., Michalick M. S., Williams P., Lima A. O., Vieira J. B. Controlled field trials of a vaccine against New World cutaneous leishmaniasis. Int J Epidemiol. 1986 Dec;15(4):572–580. doi: 10.1093/ije/15.4.572. [DOI] [PubMed] [Google Scholar]
  3. Barbiéri C. L., Giorgio S., Merjan A. J., Figueiredo E. N. Glycosphingolipid antigens of Leishmania (Leishmania) amazonensis amastigotes identified by use of a monoclonal antibody. Infect Immun. 1993 May;61(5):2131–2137. doi: 10.1128/iai.61.5.2131-2137.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Barral-Netto M., Reed S. G., Sadigursky M., Sonnenfeld G. Specific immunization of mice against Leishmania mexicana amazonensis using solubilized promastigotes. Clin Exp Immunol. 1987 Jan;67(1):11–19. [PMC free article] [PubMed] [Google Scholar]
  5. Bates P. A. Axenic culture of Leishmania amastigotes. Parasitol Today. 1993 Apr;9(4):143–146. doi: 10.1016/0169-4758(93)90181-e. [DOI] [PubMed] [Google Scholar]
  6. CONVIT J., KERDEL-VEGAS F. DISSEMINATED CUTANEOUS LEISHMANIASIS; INNOCULATION TO LABORATORY ANIMALS, ELECTRON MICROSCOPY AND FLUORESCENT ANTIBODIES STUDIES. Arch Dermatol. 1965 May;91:439–447. doi: 10.1001/archderm.1965.01600110025007. [DOI] [PubMed] [Google Scholar]
  7. Champsi J., McMahon-Pratt D. Membrane glycoprotein M-2 protects against Leishmania amazonensis infection. Infect Immun. 1988 Dec;56(12):3272–3279. doi: 10.1128/iai.56.12.3272-3279.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Convit J., Pinardi M. E., Rondón A. J. Diffuse cutaneous leishmaniasis: a disease due to an immunological defect of the host. Trans R Soc Trop Med Hyg. 1972;66(4):603–610. doi: 10.1016/0035-9203(72)90306-9. [DOI] [PubMed] [Google Scholar]
  9. Coombs G. H., Baxter J. Inhibition of Leishmania amastigote growth by antipain and leupeptin. Ann Trop Med Parasitol. 1984 Feb;78(1):21–24. doi: 10.1080/00034983.1984.11811768. [DOI] [PubMed] [Google Scholar]
  10. Doyle P. S., Engel J. C., Pimenta P. F., da Silva P. P., Dwyer D. M. Leishmania donovani: long-term culture of axenic amastigotes at 37 degrees C. Exp Parasitol. 1991 Oct;73(3):326–334. doi: 10.1016/0014-4894(91)90104-5. [DOI] [PubMed] [Google Scholar]
  11. Duboise S. M., Vannier-Santos M. A., Costa-Pinto D., Rivas L., Pan A. A., Traub-Cseko Y., De Souza W., McMahon-Pratt D. The biosynthesis, processing, and immunolocalization of Leishmania pifanoi amastigote cysteine proteinases. Mol Biochem Parasitol. 1994 Nov;68(1):119–132. doi: 10.1016/0166-6851(94)00157-x. [DOI] [PubMed] [Google Scholar]
  12. Eperon S., McMahon-Pratt D. Extracellular cultivation and morphological characterization of amastigote-like forms of Leishmania panamensis and L. braziliensis. J Protozool. 1989 Sep-Oct;36(5):502–510. doi: 10.1111/j.1550-7408.1989.tb01086.x. [DOI] [PubMed] [Google Scholar]
  13. Frommel D., Ogunkolade B. W., Vouldoukis I., Monjour L. Vaccine-induced immunity against cutaneous leishmaniasis in BALB/c mice. Infect Immun. 1988 Apr;56(4):843–848. doi: 10.1128/iai.56.4.843-848.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Handman E., Mitchell G. F. Immunization with Leishmania receptor for macrophages protects mice against cutaneous leishmaniasis. Proc Natl Acad Sci U S A. 1985 Sep;82(17):5910–5914. doi: 10.1073/pnas.82.17.5910. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Howard J. G., Nicklin S., Hale C., Liew F. Y. Prophylactic immunization against experimental leishmaniasis: I. Protection induced in mice genetically vulnerable to fatal Leishmania tropica infection. J Immunol. 1982 Nov;129(5):2206–2212. [PubMed] [Google Scholar]
  16. Ilg T., Fuchs M., Gnau V., Wolfram M., Harbecke D., Overath P. Distribution of parasite cysteine proteinases in lesions of mice infected with Leishmania mexicana amastigotes. Mol Biochem Parasitol. 1994 Oct;67(2):193–203. doi: 10.1016/0166-6851(94)00126-x. [DOI] [PubMed] [Google Scholar]
  17. Jardim A., Alexander J., Teh H. S., Ou D., Olafson R. W. Immunoprotective Leishmania major synthetic T cell epitopes. J Exp Med. 1990 Aug 1;172(2):645–648. doi: 10.1084/jem.172.2.645. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kahl L. P., Scott C. A., Lelchuk R., Gregoriadis G., Liew F. Y. Vaccination against murine cutaneous leishmaniasis by using Leishmania major antigen/liposomes. Optimization and assessment of the requirement for intravenous immunization. J Immunol. 1989 Jun 15;142(12):4441–4449. [PubMed] [Google Scholar]
  19. Kimsey P. B., Theodos C. M., Mitchen T. K., Turco S. J., Titus R. G. An avirulent lipophosphoglycan-deficient Leishmania major clone induces CD4+ T cells which protect susceptible BALB/c mice against infection with virulent L. major. Infect Immun. 1993 Dec;61(12):5205–5213. doi: 10.1128/iai.61.12.5205-5213.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Locksley R. M., Louis J. A. Immunology of leishmaniasis. Curr Opin Immunol. 1992 Aug;4(4):413–418. doi: 10.1016/s0952-7915(06)80032-4. [DOI] [PubMed] [Google Scholar]
  21. McMahon-Pratt D., Rodriguez D., Rodriguez J. R., Zhang Y., Manson K., Bergman C., Rivas L., Rodriguez J. F., Lohman K. L., Ruddle N. H. Recombinant vaccinia viruses expressing GP46/M-2 protect against Leishmania infection. Infect Immun. 1993 Aug;61(8):3351–3359. doi: 10.1128/iai.61.8.3351-3359.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Mitchell G. F., Curtis J. M., Handman E. Resistance to cutaneous leishmaniasis in genetically susceptible BALB/c mice. Aust J Exp Biol Med Sci. 1981 Oct;59(Pt 5):555–565. doi: 10.1038/icb.1981.48. [DOI] [PubMed] [Google Scholar]
  23. Mitchell G. F., Handman E., Spithill T. W. Examination of variables in the vaccination of mice against cutaneous leishmaniasis using living avirulent cloned lines and killed promastigotes of Leishmania major. Int J Parasitol. 1985 Dec;15(6):677–684. doi: 10.1016/0020-7519(85)90015-3. [DOI] [PubMed] [Google Scholar]
  24. Mosser D. M., Rosenthal L. A. Leishmania-macrophage interactions: multiple receptors, multiple ligands and diverse cellular responses. Semin Cell Biol. 1993 Oct;4(5):315–322. doi: 10.1006/scel.1993.1038. [DOI] [PubMed] [Google Scholar]
  25. Murray P. J., Spithill T. W., Handman E. Characterization of integral membrane proteins of Leishmania major by Triton X-114 fractionation and analysis of vaccination effects in mice. Infect Immun. 1989 Jul;57(7):2203–2209. doi: 10.1128/iai.57.7.2203-2209.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Nascimento E., Mayrink W., da Costa C. A., Michalick M. S., Melo M. N., Barros G. C., Dias M., Antunes C. M., Lima M. S., Taboada D. C. Vaccination of humans against cutaneous leishmaniasis: cellular and humoral immune responses. Infect Immun. 1990 Jul;58(7):2198–2203. doi: 10.1128/iai.58.7.2198-2203.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Pan A. A., Duboise S. M., Eperon S., Rivas L., Hodgkinson V., Traub-Cseko Y., McMahon-Pratt D. Developmental life cycle of Leishmania--cultivation and characterization of cultured extracellular amastigotes. J Eukaryot Microbiol. 1993 Mar-Apr;40(2):213–223. doi: 10.1111/j.1550-7408.1993.tb04906.x. [DOI] [PubMed] [Google Scholar]
  28. Pan A. A. Leishmania mexicana: serial cultivation of intracellular stages in a cell-free medium. Exp Parasitol. 1984 Aug;58(1):72–80. doi: 10.1016/0014-4894(84)90022-5. [DOI] [PubMed] [Google Scholar]
  29. Pan A. A., McMahon-Pratt D. Monoclonal antibodies specific for the amastigote stage of Leishmania pifanoi. I. Characterization of antigens associated with stage- and species-specific determinants. J Immunol. 1988 Apr 1;140(7):2406–2414. [PubMed] [Google Scholar]
  30. Pan A. A., Pan S. C. Leishmania mexicana: comparative fine structure of amastigotes and promastigotes in vitro and in vivo. Exp Parasitol. 1986 Oct;62(2):254–265. doi: 10.1016/0014-4894(86)90030-5. [DOI] [PubMed] [Google Scholar]
  31. Petersen E. A., Neva F. A., Barral A., Correa-Coronas R., Bogaert-Diaz H., Martinez D., Ward F. E. Monocyte suppression of antigen-specific lymphocyte responses in diffuse cutaneous leishmaniasis patients from the Dominican Republic. J Immunol. 1984 May;132(5):2603–2606. [PubMed] [Google Scholar]
  32. Rainey P. M., Spithill T. W., McMahon-Pratt D., Pan A. A. Biochemical and molecular characterization of Leishmania pifanoi amastigotes in continuous axenic culture. Mol Biochem Parasitol. 1991 Nov;49(1):111–118. doi: 10.1016/0166-6851(91)90134-r. [DOI] [PubMed] [Google Scholar]
  33. Russell D. G., Alexander J. Effective immunization against cutaneous leishmaniasis with defined membrane antigens reconstituted into liposomes. J Immunol. 1988 Feb 15;140(4):1274–1279. [PubMed] [Google Scholar]
  34. Russell D. G. Immunity to leishmaniasis: what properties delineate a protective antigen? Ann Inst Pasteur Immunol. 1987 Sep-Oct;138(5):774–781. doi: 10.1016/s0769-2625(87)80037-5. [DOI] [PubMed] [Google Scholar]
  35. Russell D. G., Talamas-Rohana P. Leishmania and the macrophage: a marriage of inconvenience. Immunol Today. 1989 Oct;10(10):328–333. doi: 10.1016/0167-5699(89)90188-6. [DOI] [PubMed] [Google Scholar]
  36. Sadick M. D., Raff H. V. Differences in expression and exposure of promastigote and amastigote membrane molecules in Leishmania tropica. Infect Immun. 1985 Feb;47(2):395–400. doi: 10.1128/iai.47.2.395-400.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Scharton T. M., Scott P. Natural killer cells are a source of interferon gamma that drives differentiation of CD4+ T cell subsets and induces early resistance to Leishmania major in mice. J Exp Med. 1993 Aug 1;178(2):567–577. doi: 10.1084/jem.178.2.567. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Scott P. IFN-gamma modulates the early development of Th1 and Th2 responses in a murine model of cutaneous leishmaniasis. J Immunol. 1991 Nov 1;147(9):3149–3155. [PubMed] [Google Scholar]
  39. Scott P., Pearce E., Natovitz P., Sher A. Vaccination against cutaneous leishmaniasis in a murine model. I. Induction of protective immunity with a soluble extract of promastigotes. J Immunol. 1987 Jul 1;139(1):221–227. [PubMed] [Google Scholar]
  40. Scott P., Pearce E., Natovitz P., Sher A. Vaccination against cutaneous leishmaniasis in a murine model. II. Immunologic properties of protective and nonprotective subfractions of soluble promastigote extract. J Immunol. 1987 Nov 1;139(9):3118–3125. [PubMed] [Google Scholar]
  41. Straus A. H., Levery S. B., Jasiulionis M. G., Salyan M. E., Steele S. J., Travassos L. R., Hakomori S., Takahashi H. K. Stage-specific glycosphingolipids from amastigote forms of Leishmania (L.) amazonensis. Immunogenicity and role in parasite binding and invasion of macrophages. J Biol Chem. 1993 Jun 25;268(18):13723–13730. [PubMed] [Google Scholar]
  42. Taswell C. Limiting dilution assays for the determination of immunocompetent cell frequencies. I. Data analysis. J Immunol. 1981 Apr;126(4):1614–1619. [PubMed] [Google Scholar]
  43. Titus R. G., Marchand M., Boon T., Louis J. A. A limiting dilution assay for quantifying Leishmania major in tissues of infected mice. Parasite Immunol. 1985 Sep;7(5):545–555. doi: 10.1111/j.1365-3024.1985.tb00098.x. [DOI] [PubMed] [Google Scholar]
  44. Traub-Cseko Y. M., Duboise M., Boukai L. K., McMahon-Pratt D. Identification of two distinct cysteine proteinase genes of Leishmania pifanoi axenic amastigotes using the polymerase chain reaction. Mol Biochem Parasitol. 1993 Jan;57(1):101–115. doi: 10.1016/0166-6851(93)90248-v. [DOI] [PubMed] [Google Scholar]
  45. Turco S. J., Sacks D. L. Expression of a stage-specific lipophosphoglycan in Leishmania major amastigotes. Mol Biochem Parasitol. 1991 Mar;45(1):91–99. doi: 10.1016/0166-6851(91)90030-a. [DOI] [PubMed] [Google Scholar]
  46. Yang D. M., Fairweather N., Button L. L., McMaster W. R., Kahl L. P., Liew F. Y. Oral Salmonella typhimurium (AroA-) vaccine expressing a major leishmanial surface protein (gp63) preferentially induces T helper 1 cells and protective immunity against leishmaniasis. J Immunol. 1990 Oct 1;145(7):2281–2285. [PubMed] [Google Scholar]
  47. Yang D. M., Rogers M. V., Liew F. Y. Identification and characterization of host-protective T-cell epitopes of a major surface glycoprotein (gp63) from Leishmania major. Immunology. 1991 Jan;72(1):3–9. [PMC free article] [PubMed] [Google Scholar]

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