Description of an improved method for Blastocystis hominis culture and axenization

Abstract

An improved method for Blastocystis hominis culture and axenization was developed in the present study. Stool samples were cultured in prereduced Boeck-Drbohlav NHI modified medium (with several modifications) supplemented with antibiotics (0.4% ampicillin, 0.1% streptomycin, 0.0006% amphotericin B). Axenization was performed by the combination of partial purification of B. hominis by Ficoll-metrizoic acid gradient and inoculation in fresh medium containing active antibiotics against remaining bacteria. A total of 25 strains were obtained by this procedure. The time required for axenization ranged between 3 and 5 weeks. The generation time of axenic strains ranged from 6.6 to 12.1 h (mean +/- SD 110.0 +/- 1.8 h) and the mean number of generations was 2.5 +/- 0.6 h per 24 h. The size of vacuolar and ameboid forms found in stools and in culture was similar. The special formulation of the medium used reduced the generation time and did not modify the cellular size as compared with fecal forms.