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New advances in the in-vitro culture of Dientamoeba fragilis

Published online by Cambridge University Press:  16 February 2012

VARUNI S. MUNASINGHE ,
D. STARK  and
J. T. ELLIS
VARUNI S. MUNASINGHE
Affiliation:
University of Technology Sydney, School of Medical and Molecular Biosciences and the Ithree Institute, Broadway, NSW, Australia
D. STARK
Affiliation:
University of Technology Sydney, School of Medical and Molecular Biosciences and the Ithree Institute, Broadway, NSW, Australia Division of Microbiology, SydPath, St. Vincent's Hospital, Darlinghurst, NSW, Australia
J. T. ELLIS*
Affiliation:
University of Technology Sydney, School of Medical and Molecular Biosciences and the Ithree Institute, Broadway, NSW, Australia
*
*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
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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.

Keywords

Dientamoeba fragilisin vitro cultureEarle's balanced salt solutionferric ammonium citratecholesterol
Type
Research Article
Information
Parasitology , Volume 139 , Issue 7 , June 2012 , pp. 864 - 869
Copyright
Copyright © Cambridge University Press 2012

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