Mycoplasma orale Taylor-Robinson et al.

1. Follow instructions as suggested for the culturing of Mollicutes:

                PROCEDURES FOR PROPAGATING MOLLICUTES:

1. Open thawed vial.
2. Make serial dilutions by transferring 0.5 mL from the original tube to a tube containing 5.0 mL. Repeat process by transferring 0.5 mL from the second to a third tube, etc. Dilutions are important, not only for titration purposes, but also to keep culture in varying stages of growth. Many strains will die out rapidly once acid or alkaline conditions are reached. It is recommended to prepare several dilutions from the initial tube as the cryoprotectant used in the freeze-drying process often inhibits growth.
3. Plates may be inoculated to check colony morphology. You can also spot each dilution on the surface of plate (4 or more/plate) to determine the number of colony forming units. However, not all strains do well on solid medium.
4. Incubate all tubes and plates under the recommended conditions and appropriate temperature. The time necessary for growth will vary from strain to strain. Growth on plates generally requires additional incubation. 
5. Depending on the medium used, growth will be indicated by increased turbidity, a color change, or both. 

2. Tubes and plates are incubated under anaerobic conditions using an anaerobe jar or other appropriate method. The incubation temperature is 37°C. 
3. Medium to light turbidity will appear in the first few dilution tubes within 72 hour to 7 days. Colonies generally appear on solid medium within 7 days. 
4. Subsequent, fresh transfer will grow more rapidly than the original rehydrated culture; but usually still requires 48 hours between transfers.

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