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New advances in the in-vitro culture of Dientamoeba fragilis

  • VARUNI S. MUNASINGHE (a1), D. STARK (a1) (a2) and J. T. ELLIS (a1)

Summary

Dientamoeba fragilis is an intestinal protozoan in humans that is commonly associated with diarrhoea and other gastrointestinal complaints. Studies conducted to investigate the biology of this parasite are limited by methods for in vitro cultivation. The objective of this study was to improve a biphasic culture medium, based on the Loeffler's slope, by further supplementation in order to increase the yield of trophozoites in culture. The current in vitro culture of D. fragilis is a xenic culture with a mix of bacteria. Three different liquid overlays were evaluated including Earle's balanced salt solution (EBSS), PBS and Dulbecco's modified PBS (DPBS), for their ability to support the in vitro growth of D. fragilis trophozoites. Out of these 3 overlays EBSS gave the highest increase in the trophozoite numbers. The effect of supplementation was analysed by supplementing EBSS with ascorbic acid, ferric ammonium citrate, L-cysteine, cholesterol and alpha-lipoic acid and quantification of in vitro growth by cell counts. A new liquid overlay is here described based upon EBSS supplemented with cholesterol and ferric ammonium citrate that, in conjunction with the Loeffler's slope, supports the growth of D. fragilis trophozoites in vitro. This modified overlay supported a 2-fold increase in the numbers of trophozoite in culture from all 4 D. fragilis isolates tested, when compared to a PBS overlay. These advances enable the harvest of a larger number of trophozoites needed for further studies on this parasite.

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Corresponding author

*Corresponding author: School of Medical and Molecular Biosciences, University of Technology Sydney, P.O. Box 123, Broadway, NSW 2007, Australia. Tel: +61 2 9514 4161. Fax: +61 2 9514 8206. E-mail: john.ellis@uts.edu.au

References

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Amin, A., Neubauer, C., Liebhart, D., Grabensteiner, E. and Hess, M. (2010). Axenization and optimization of in vitro growth of clonal cultures of Tetratrichomonas gallinarum and Trichomonas gallinae. Experimental Parasitology 124, 202208.
Banik, G., Barratt, J., Marriott, D., Harkness, J., Ellis, J. and Stark, D. (2011). A case-controlled study of Dientamoeba fragilis infections in children. Parasitology 1, 15.
Barratt, J. L. N., Banik, G. R., Harkness, J., Marriott, D., Ellis, J. T. and Stark, D. (2010). Newly defined conditions for the in vitro cultivation and cryopreservation of Dientamoeba fragilis: new techniques set to fast track molecular studies on this organism. Parasitology 137, 18671878.
Barratt, J. L. N., Harkness, J., Marriott, D., Ellis, J. T. and Stark, D. (2011 a). The ambiguous life of Dientamoeba fragilis: the need to investigate current hypotheses on transmission. Parasitology 138, 557572.
Barratt, J. L. N., Harkness, J., Marriott, D., Ellis, J. T. and Stark, D. (2011 b). A review of Dientamoeba fragilis carriage in humans: several reasons why this organism should be considered in the diagnosis of gastrointestinal illness. Gut Microbes 2, 312.
Bryant, J. C. (1975). Earle's balanced salt solution: Preparation of the saline. Methods in Cell Science 1, 185187.
Farthing, M. J., Keusch, G. T. and Carey, M. C. (1985). Effects of bile and bile salts on growth and membrane lipid uptake by Giardia lamblia. Possible implications for pathogenesis of intestinal disease. Journal of Clinical Investigation 76, 17271732.
Gillin, F. D., Gault, M. J., Hofmann, A. F., Gurantz, D. and Sauch, J. F. (1986). Biliary lipids support serum-free growth of Giardia lamblia. Infection and Immunity 53, 641645.
Lehker, M. W. and Alderete, J. F. (1992). Iron regulates growth of Trichomonas vaginalis and the expression of immunogenic trichomonad proteins. Molecular Microbiology 6, 123132.
Lujan, H. D., Mowatt, M. R. and Nash, T. E. (1996). Lipid requirements and lipid uptake by Giardia lamblia trophozoites in culture. Journal of Eukaryotic Microbiology 43, 237242.
Sawangjaroen, N., Luke, R. and Prociv, P. (1993). Diagnosis by faecal culture of Dientamoeba fragilis infections in Australian patients with diarrhoea. Transactions of the Royal Society of Tropical Medicine and Hygiene 87, 163165.
Stark, D., Barratt, J., Roberts, T., Marriott, D., Harkness, J. and Ellis, J. (2010 a). Comparison of microscopy, two xenic culture techniques, conventional and real-time PCR for the detection of Dientamoeba fragilis in clinical stool samples. European Journal of Clinical Microbiology and Infectious Diseases 29, 411416.
Stark, D., Barratt, J., Roberts, T., Marriott, D., Harkness, J. and Ellis, J. (2010 b). A review of the clinical presentation of dientamoebiasis. American Journal of Tropical Medicine and Hygiene, 82, 614619.
Stark, D., Beebe, N., Marriott, D., Ellis, J. and Harkness, J. (2007). Dientamoeba fragilis as a cause of travelers’ diarrhea: report of seven cases. Journal of Travel Medicine 14, 7273.

Keywords

New advances in the in-vitro culture of Dientamoeba fragilis

  • VARUNI S. MUNASINGHE (a1), D. STARK (a1) (a2) and J. T. ELLIS (a1)

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