L-Canavanine Glycine Bromothymol Blue (CGB) Agar

For differentiation of C. neoformans and C. gattii (Kwon-Chung et al. 1982).

Solution A

• Glycine Univar 10 g
• KH2PO4 1 g
• MgSO4 1 g
• Thiamine HCl 1 mg
• L-canavanine sulphate 30 mg
• Distilled water 100 mL

1. Dissolve ingredients in small beaker and adjust pH to 5.6
2. Filter sterilise solution using 0.22 μm filter.
3. Store in refrigerator.

Solution B (Aqueous Bromothymol Blue)

• Bromothymol Blue 0.4 g
• 0.01N NaOH 64 mL
• Distilled water 36 mL

1. Dissolve the Bromothymol Blue in the NaOH
2. Add to the water.

To prepare medium (1 litre for plates)

• Distilled water 880 mL
• Solution B 20 mL
• Bacto agar (BD 214010) 20 g

1. Autoclave to 121C for 15 minutes, cool to 48C.
2. For plates add 100 mL of the filtered solution A and mix. Dispense as plates.