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      Trypanosoma cruzi: Maintenance in Culture Modify Gene and Antigenic Expression of Metacyclic Trypomastigotes

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          Abstract

          In this study we examined whether the maintenance of Trypanosoma cruzi by long-time in axenic culture produces changes in gene expression and antigenic profiles. The studies were made with a Dm30L-clone from a low-virulent strain and a non-cloned virulent EP-strain of T. cruzi. Both parasites were maintained, for at least seven years, by successive alternate passage triatomine/mouse (triatomine condition), or by serial passage in axenic medium (culture condition). The comparison of the [35S]methionine metabolic labeling products of virulent and non-virulent parasites by 2D-SDS-PAGE, clearly indicates that the expression of metacyclic trypomastigotes (but not of epimastigotes) proteins have been altered by laboratory maintenance conditions. Western blot analysis of EP and Dm30L-epimastigotes using a serum anti-epimastigotes revealed that although most of antigens are conserved, four antigens are characteristics of triatomine condition parasites and three other are characteristics of culture condition parasites. Anti-metacyclics serum revealed significative differences in EP- and Dm30L-metacyclic trypomastigotes from triatomine condition. However, avirulent metacyclic forms were antigenically very similar. These results suggest that besides a possible selection of avirulent subpopulation from T. cruzi strains genetically heterogeneous when maintained by long time in axenic culture, changes in virulence might be due to post-translational modifications of the antigens induced by the absence of the natural alternability (vertebrate-invertebrate) in the life-cycle of T. cruzi

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          Most cited references32

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          Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4

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            High resolution two-dimensional electrophoresis of proteins.

            A technique has been developed for the separation of proteins by two-dimensional polyacrylamide gel electrophoresis. Due to its resolution and sensitivity, this technique is a powerful tool for the analysis and detection of proteins from complex biological sources. Proteins are separated according to isoelectric point by isoelectric focusing in the first dimension, and according to molecular weight by sodium dodecyl sulfate electrophoresis in the second dimension. Since these two parameters are unrelated, it is possible to obtain an almost uniform distribution of protein spots across a two-diminsional gel. This technique has resolved 1100 different components from Escherichia coli and should be capable of resolving a maximum of 5000 proteins. A protein containing as little as one disintegration per min of either 14C or 35S can be detected by autoradiography. A protein which constitutes 10 minus 4 to 10 minus 5% of the total protein can be detected and quantified by autoradiography. The reproducibility of the separation is sufficient to permit each spot on one separation to be matched with a spot on a different separation. This technique provides a method for estimation (at the described sensitivities) of the number of proteins made by any biological system. This system can resolve proteins differing in a single charge and consequently can be used in the analysis of in vivo modifications resulting in a change in charge. Proteins whose charge is changed by missense mutations can be identified. A detailed description of the methods as well as the characteristics of this system are presented.
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              In vitro differentiation of Trypanosoma cruzi under chemically defined conditions.

              Metacyclic trypomastigotes of Trypanosoma cruzi have been obtained in chemically defined axenic culture. The differentiating medium, composed of artificial triatomine urine supplemented with proline, allows high yields of metacyclic trypomastigotes after 72-h incubation of T. cruzi cells at 27 degrees C. Morphological differentiation of the parasites is gradual under these chemically defined conditions and is preceded by the expression of stage-specific polypeptides. The yield of in vitro-induced metacyclic trypomastigotes depends upon the age of the epimastigote culture, the size of the inoculum and the depth of the medium. Metacyclic trypomastigotes differentiated in vitro from the Dm 28c clone of T. cruzi are both resistant to complement lysis and to macrophage digestion. They are able to infect mice with an efficiency similar to that obtained for natural metacyclic trypomastigotes obtained from triatomine excreta.
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                Author and article information

                Journal
                mioc
                Memórias do Instituto Oswaldo Cruz
                Mem. Inst. Oswaldo Cruz
                Instituto Oswaldo Cruz, Ministério da Saúde (Rio de Janeiro, RJ, Brazil )
                0074-0276
                1678-8060
                November 1998
                : 93
                : 6
                : 753-760
                Affiliations
                [01] orgnameUniversidad de Carabobo orgdiv1 Centro de Biología Molecular de Parásitos, Facultad de Ciencias de la Salud
                Article
                S0074-02761998000600011 S0074-0276(98)09300611
                10.1590/S0074-02761998000600011
                9921298
                608a9e61-e36f-4c35-8b28-576187ebb441

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

                History
                : 13 July 1998
                : 12 February 1998
                Page count
                Figures: 0, Tables: 0, Equations: 0, References: 33, Pages: 8
                Product

                SciELO Brazil

                Categories
                Systematics

                gene expression,Trypanosoma cruzi,metacyclic,metabolic labeling

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