Newly defined conditions for thein vitrocultivation and cryopreservation ofDientamoeba fragilis: new techniques set to fast track molecular studies on this organism

Abstract

SUMMARYDientamoeba fragilisis a pathogen of the human gastrointestinal tract that is a common cause of diarrhoea. A paucity of knowledge on thein vitrocultivation and cryopreservation ofDientamoebahas meant that few studies have been conducted to investigate its biology. The objective of this study was to define, for the first time,in vitroculture conditions able to support the long-termin vitrogrowth ofDientamoeba. Also, we aimed to define a suitable method for cryopreserving viableDientamoebatrophozoites. A modified BD medium, TYGM-9, Loeffler's slope medium, Robinson's medium, Medium 199, Trichosel and aTritrichomonas fetusmedium were compared, using cell counts, for their ability to support the growth ofD. fragilisat various temperatures and atmospheric conditions. Loeffler's slope medium supported significantly better growth compared to other media. A temperature of 42°C and a microaerophilic atmosphere were also optimum forDientamoebagrowth. To our knowledge, this is the first study to describe and compare different culture media and conditions for the growth of clinical isolates ofD. fragilis.This new technology will aid the development of diagnostics for dientamoebiasis as well as facilitate large-scale sequencing projects that will fast track molecular studies onD. fragilis.