NYCIII medium

This medium is used for growth of fastidious anaerobes like Gardnerella vaginalis and Lactobacillus iners. In my work I use it also for growth of other lactobacilli such as Lactobacillus crispatus although they will also grow on regular MRS medium. The reason for this is that in most cases I want to compare growth of these different bacteria, and want to keep as many other parameters in the experiment constant.

I adjusted this recipe from the ATCC protocol:

For 500 mL NYCIII medium:

• HEPES (CellGro) 1,2 gram
• Proteose Peptone No. 3 7.5 gram
• Yeast Extract 1.9 gram
• NaCl 2.5 gram
• Glucose 2.5 gram (I use monohydrated glucose which means that I need 10% more = 2.75 gram)
• water 450 mL

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Compared to the ATCC recipe I use less HEPES, and do not adjust the pH. The pH is always around 6.7 before autoclaving and addition of the Horse Serum. In case I want to poor plates, I add 5 grams of agar (1%). I autoclave this mixture at 121°C for 20 minutes. After autoclaving and cooling down I add 50 mL of heat inactivated horse serum.

NYC III medium without glucose, 1.1x

I use this medium in case I want to test carbon sources other than glucose. I use the same recipe as above except that I leave out glucose and only add 400 mL of water. In order to supplement with alternative carbon sources I add 10% of the final volume of a carbohydrate solution (such as glycogen or glucose dissolved in water), with water as the control. The concentration of the carbon source is 10x higher concentration than the final concentration.
So for a 1 mL culture in NYCIII medium with 0,5% of a carbon source I add:
900 uL NYCIII medium without glucose 1.1x
100 uL of a 5% glucose of glycogen solution in water or water as a control.