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Parasitología al día

versão impressa ISSN 0716-0720

Parasitol. día v.23 n.3-4 Santiago jul. 1999 


Modification and adaptation in semi-defined media for
cultivation of flagellate Tetratrichomonas didelphidis
(Trichomonadidae trichomonadinae) from the Didelphis

and IVELI ROSSET******


Tetratrichomonas didelphidis (Hegner and Ratcliffe, 1927) Andersen and Reilly, 1965 is a flagellate protozoan from the intestine, cecum, and colon of Didelphis marsupialis Linnaeus, 1758. The parasite was found and isolated in the rectal glands in Pavlova starch-containing media in Florianópolis, SC, Brasil from D. marsupialis.
Key words: Tetratrichomonas didelphidis, Didelphis marsupialis, in vitro cultivation.

* This research was supported in part by Faculdade de Farmácia, PUCRS and FAPERGS (grant Nº 96/50021.9), Porto Alegre, RS, Brasil.
** Faculdade de Farmácia e Programa de Pós-Graduação em Biociências, PUCRS e Bolsista do CNPq, Porto Alegre, RS, Brasil.
*** Faculdade de Farmácia, PUCRS, Av. Ipiranga 6681, 90619-900 Porto Alegre RS, Brasil. + Corresponding author. E-mail:
**** Departamento de Microbiologia e Parasitologia, UFSC, Florianópolis, SC, Brasil.
***** Bolsista de Inciação Científica, PUCRS-FAPERGS, Porto Alegre, RS, Brasil.
****** Bolsista BPA-PUCRS, Porto Alegre, RS, Brasil.


Tetratrichomonas didelphidis (Hegner and Ratcliffe, 1927) Andersen and Reilly, 1965 is a flagellate protozoan from the intestine, cecum, and colon of Didelphis marsupialis Linnaeus, 1758. Up to the present, only one paper on the occurrence and anatomy of trichomonads fromopossum exist1. The protozoa T. didelphidis used in this study was isolated in the rectal glands in Pavlova starch-containing media by Prof. Dr. Mario Steindel, in Florianópolis, SC, Brasil from Didelphis marsupialis. The aim of this communication was to describe modifications and adaptation of trichomonads isolated from the opossum in axenic culture.


Trichomonads from opossum used in this study was provided in monoxenic culture with a large number of bacteria. Meropenen (100 µg/ml) was employed for the elimination of bacteria in order to make a comparison with the mensural data given by Andersen and Reilly1. In the present study, complementary techniques were used to analyze the nutrition and physiology of trichomonads in Diamond's trypticase-yeast extract-maltose (TYM) medium2 without agar and phosphate buffer; TYM modified,3 and Stuart`s medium4.The bacteria mask the trichomonads in the microscopic observations of stained specimens in Pavlova starch-containing media. Living cells were observed in a light microscope. Material for the stained smears was obtained from parasites harvested (250 x g for 20 min) from a 72 hr culture in Diamond's medium without maltose (TYS). All photomicrographs were made with a Olympus AX70 photomicroscope equipped with a 100 X apochromatic oil immersion objective with Kodak Proimage 135 mm ASA 100.


Microscopic observations of trichomonads in Pavlova starch-containing media presents many difficulties because it has a great quantity of starch granules. TYS, pH 7,5, support a very good growth of T. didelphidis when was added starch solution (5 mg/ml), and supplemented with 10% (v/v) heat inactivated bovine serum, in air, at a temperature of 28°C (± 0.5). Isolates were subcultured every 72 hr in TYS media. Of the various media and temperature ranged tested, the organisms grew best in TYS media at 28°C. Andersen and Reilly1 with the TYM medium without agar had only a moderate growth. The second most successful medium tested was the TYM modified3 in which growth occurred best at 28°C. The organisms grew best in Pavlova starch-containing media but only in monoxenic culture. Direct microscopic examination of smears from culture media revealed a vast numbers as actively motile flagellate protozoa. These were classified as trichomonads because of their elongate ellipsoid shape which could be accurately counted only when the trichomonads had slowed down or stopped moving. The body is very plastic, but not particularly ameboid. Most of these morphological features could be recognized in air-dried smears of culture media, fixed in methanol and stained with Giemsa, and iron hematoxylin. As shown by our study, this trichomonad grew best in Diamond's medium without maltose and with starch solution (TYS), pH 7,5 at 28°C. There are few reports on temperature employed in cultivating parasites from marsupials1 . In a descriptive account it is frequently difficult to make a valid comparison of the specimen with other similar organisms. Not only are the current staining techniques difficult to equate because of the modifications introduced by various investigators, but many of the early descriptions were completed before some of the present day techniques had been developed. Our preliminary results suggest that the protozoa studied is T. didelphidis. Further investigations is needed to determine the significance of our results and its comparations with the data of Andersen and Reilly1


1.- ANDERSEN F L, J R REILLY J R. The anatomy of Tetratrichomonas didelphidis (Hegner and Ratcliffe, 1927) comb. nov. from the opossum. J Parasitol 14: 27-35, 1965.         [ Links ]

2.- DIAMOND L S. The stablishment of various trichomonads of animals and man in axenic cultures. J Parasitol 43: 488-490, 1957.         [ Links ]

3.- KULDA J, HONIGBERG B M, FROST J K, HOLLANDER D H. Pathogenicity of Trichomonas vaginalis. Am J Obstet Gynecol 108: 908-918, 1970.         [ Links ]

4.- STUART R D. The preparation and use of a single culture medium for leptospirosis. J Path and Bact 58: 343-349, 1946.         [ Links ]




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