# 149: MEGADESULFOVIBRIO MEDIUM


## Main sol. 149

970	ml	**Solution A**	
20	ml	**Solution B**	
1	ml	**Solution C**	
10	ml	**Solution D**	

1. Sparge solution A with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic, then dispense under the same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. Solution B is autoclaved under 80% N2 and 20% CO2 gas atmosphere. Solution C is prepared under 100% N2 gas atmosphere and sterilized by filtration. Solution D is autoclaved separately under 100% N2 gas. To complete the medium appropriate amounts of solutions B to D are added to the sterile solution A in the sequence as indicated. Final pH of the medium should be 7.0 - 7.2.
2. Note: Addition of 10 - 20 mg sodium dithionite per liter (e.g. from 5% (w/v) solution, freshly prepared under N2 and filter-sterilized) may stimulate growth of some strains at the beginning. For transfers use 5 - 10% inoculum.


## Solution A

1	g	KH2PO4	
0.5	g	NH4Cl	
0.4	g	MgSO4 x 7 H2O	
2	g	Na2SO4	
0.1	g	CaCl2 x 2 H2O	
1	ml	**Trace element solution SL-6**	
1	ml	H2SO4 (2 N)	
2	g	Na-L-lactate	
0.5	ml	Sodium resazurin (0.1% w/v)	
970	ml	Distilled water	


## Solution B

1	g	Na2CO3	
20	ml	Distilled water	


## Solution C

1	ml	**Seven vitamins solution**	


## Solution D

0.3	g	Na2S x 9 H2O	
10	ml	Distilled water	


## Trace element solution SL-6

0.1	g	ZnSO4 x 7 H2O	
0.03	g	MnCl2 x 4 H2O	
0.3	g	H3BO3	
0.2	g	CoCl2 x 6 H2O	
0.01	g	CuCl2 x 2 H2O	
0.02	g	NiCl2 x 6 H2O	
0.03	g	Na2MoO4 x 2 H2O	
1000	ml	Distilled water	


## Seven vitamins solution

100	mg	Vitamin B12	
80	mg	p-Aminobenzoic acid	
20	mg	D-(+)-biotin	
200	mg	Nicotinic acid	
100	mg	Calcium pantothenate	
300	mg	Pyridoxine hydrochloride	
200	mg	Thiamine-HCl x 2 H2O	
1000	ml	Distilled water