A new medium containing antibiotics for the xenic cultivation of Entamoeba gingivalis

Diamond's trypticase-yeast extract-serum-gastric mucin (TYSGM-9) medium was studied for its suitability to support the xenic growth of the oral protozoan Entamoeba gingivalis. Amoebic growth was found to be best when the inoculum for transfer was 0.1 ml, the incubation temperature was 35 degrees C, and the interval between transfers was 48 h. These parameters were also useful for controlling the growth of accompanying bacteria. In addition, bacterial growth in xenic stock cultures, which had a direct effect on the activity of E. gingivalis trophozoites, was kept to a minimum by addition of the antibiotics piperacillin, erythromycin, neomycin, and penicillin. Varying substitutions and selected supplementations of TYSGM-9 medium led to the development of an improved medium for E. gingivalis. Supplements most beneficial for the growth of E. gingivalis trophozoites were ascorbic acid, ferric ammonium citrate, and special NCTC 107 vitamin mixture. As compared with TYSGM-9 medium (6-10 E. gingivalis trophozoites observed per field), the new E. gingivalis medium supported excellent growth (16-20 E. gingivalis trophozoites observed per field during optimal growth) of the oral protozoan. The medium is suitable for clinical isolation of E. gingivalis.