FORMATE AND FUMARATE
Prepare a solution containing sodium formate and fumaric acid at a concentration of 6% each, adjust the pH to 7.0 and filter-sterilize. Prior to inoculation, add 0.05 ml to a tube containing approximately 6.5 ml of ATCC Medium 593 (see below).
ATCC Medium 593:
Ground beef (free of fat)........500.0 g
Distilled water....................1.0 L
N NaOH............................25.0 ml
Use lean beef or horse meat. Remove fat and connective tissue before grinding. Mix meat, water, and NaOH and bring to a boil while stirring. Cool to room temperature, skim fat off surface, and filter, retaining both meat particles and filtrate. Add sufficient distilled water to filtrate to restore volume to one liter.
To the filtrate add:
Peptone...........................30.0 g
Yeast extract......................5.0 g
K2HPO4 .............................5.0 g
0.025% Resazurin solution..........4.0 ml
Boil, cool and add 0.5 g L-cysteine . HCl; adjust pH to 7.0. Under 97% nitrogen, 3% hydrogen, dispense 7 ml over 1 part meat particles to 4-5 parts fluid per test tube. Cap with butyl rubber stoppers under N2, H2 and autoclave in press for 15 minutes under fast exhaust.