For xenic (i.e. non-sterile) in-vitro cultivation of Blastocystis, Jones’ Medium is required:

Modified Jones’ Medium for in-vitro culture of Blastocystis

Stock solution

Na2HPO4              9.46 g in 1 L distilled water

KH2PO4                 9.08 g in 1 L distilled water

NaCl                     9.00 g in 1 L distilled water

To prepare Jones' Medium

1. Mix 93.8 mL Na2HPO4 with 31.3 mL KH2HPO4 and 562.5 mL NaCl.

2. To the buffered solution, add yeast extract (Oxoid) to 0.1%.

3. Autoclave at 15 lb inch-2 for 15 min.

4. Before use, add 10% horse serum (heat inactivated at 56ºC for 30 min) to Jones’ Medium.

For each faecal specimen transfer with a steril swab 50 mg faeces (or so) to a centrifuge tube with screw cap containing Jones’ Medium (3 ml will do). The culture is incubated at 37 °C for 48-72 h. (Note: The presence of bacteria in the sample will be enough for creating the anaerobic environment needed for Blastocystis to grow.) When subculturing, transfer about 50-100 uL of the sediment from each used culture into 3 ml fresh Jones’ medium containing 10% (heat inactivated) horse serum. Subculture every 3—4 days.

Examination

Cultures are examined with or without iodine at x 200 and x400 LM magnification and evaluated for the presence of especially vacuolar (the main stage seen) but also other stages of Blastocystis.

References

Zaman et al., 1994.  A comparison of direct microscopy with culture for the diagnosis of Blastocystis hominis.  Southeast Asian Journal of Tropical Medicine and Hygiene, 25: 792-793.

Leelayoova, S., et al. 2002. In-vitro cultivation: a sensitive method for detecting Blastocystis hominis. Annals of Tropical Medicine and Parasitology, 96: 803-807.

Personal communication Dr. Huw Smith, Glasgow (Suresh, K and Smith, H, 2004. Comparison of methods for detecting Blastocystis hominis. European Journal of Clinical Microbiology and Infectious Diseases 23: 509-511)

The original "Jones’ solution" (Jones, W. R. 1946):

Sterile horse serum                                        0,5 mL

1 % marmite solution**                                 1,0 mL

Buffer saline solution (pH 7,2)                        8,5 mL

Rice starch                                                     30 mg

** Marmite is autolysed yeast extract made by Marmite Limited, London

Reference:

Jones, W. R. 1946. The experimental infection of rats with Entamoeba histolytica; with a method for evaluating the anti-amoebic properties of new compounds. Annals of Tropical Medicine and Parasitology, 40: 130-140.