CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



507 



3. Peptone (Defresne) 10.0 g. 



4. Agar 5.0 g. 



5. Neutral red 0.1 cc. 



Preparation : 



(1) Macerate 500.0 g. of finely divided 

 beef with one liter distilled water for 

 12 hours. 



(2) Boil slowly for 15 minutes. 



(3) Allow to cool, skim and filter thru 

 filter paper. 



(4) Adjust the reaction to 1.5. 



(5) Add 5.0 g. of agar and heat for 15 

 minutes at 118°C. 



(6) Filter. 



(7) Prepare a solution of 0.1 cc. neutral 

 red in 20.0 cc. distilled water. 



(8) Add 5.0 cc. of (7) to one liter of (6). 



(9) Tube. 



Sterilization: Sterilize at 112°C for 15 

 minutes. 



Use: Cultivation of Bacillus leprae. 

 Author reported that a change of color 

 or fluorescence may be present or absent. 



Variants: Harvey added 5.0 cc. of a 1.0% 

 solution of neutral red and 5.0 g. of any 

 desired carbohydrate, alcohol, etc., to 

 medium 1661, variant (v). 



References: Emile-Weil (1917 p. 379), Har- 

 vey (1921-22 p. 89). 



1679. Lentz and Tietz's Malachite Greea 

 Infusion Agar 



Constituents : 



1. Loeffler's 2.0% meat infusion peptone 

 agar. 



2. Malachite Green. 

 Preparation: 



(1) Add 1:1000, 1:2000, 1:4000, 1:6000, 

 1:8000 or 1:10,000 malachite green to 

 Loeffler's 2.0% meat infusion peptone 

 agar (method of preparation not 

 given) . 



(2) Pour into petri dishes. 

 Sterilization: Not specified. 



Use: Isolation colon-typhoid bacteria from 

 feces. Author reported that coli grew 

 weakly in 1:10,000 malachite green, 

 typhoid well in 1:4000 and para B 1:2000. 



Variants: Loeffler gave the following 

 method of preparation: 



(1) Prepare beef infusion 1 pound in two 

 liters water. 



(2) Add 30.0 g. agar. 



(3) Add 7.5 cc. N/1 HCl. 



(4) Boil 30 minutes. 



(5) Add 7.0 cc. N/1 NaOH. 



(6) Neutralize to litmus with sodium 

 carbonate. 



(7) Add 5.0 cc. N/1 sodium carbonate. 



(8) Add nutrose to make 1% solution. 

 (0.5% peptone may be used instead 

 of nutrose). 



(9) Boil. 



(10) Distribute in 500.0 cc. flasks. 



(11) Sterilize several hours on each of two 



successive days in streaming steam , 



(12) Let cool slowly in sterilizer. 



(13) Decant clear supernatant liquid. 



(14) To each 100.0 cc. of agar add 2-2.5 cc. 

 of a sterile 2.0% aqueous malachite 

 green solution. 



(15) Pour 15.0 to 20.0 cc. in each petri 

 dish. 



(16) Leave dishes open until agar is cool 

 and hard. 



References: Lentz and Tietz (1903 p. 2139), 

 Loefiler (1906p. 289). 



1680. Zielleczky's Phenolphthalein Infusion 



Agar 



Same as medium 783 but solidified by the 

 addition of agar. 



1681. Werbitzki's China Green Infusion 



Agar 



Constituents : 



1. Distilled water 1000.0 cc. 



2. Beef 500.0 g. 



3. Peptone, Witte (1.0%) 10.0 g. 



4. NaCl (0.5%) 5.0 g. 



5. Agar (3.0%) 30.0 g. 



6. China green (Boyer's) 

 Preparation : 



(1) Add 500.0 g. of finely chopped bone 

 and sinew free lean beef to one liter 

 of distilled water. Mix well. 



(2) Weigh (1). 



(3) Boil for 45 minutes stirring con- 

 stantly. 



(4) Weigh, and make up the loss in 

 weight by the addition of distilled 

 water. 



(5) Filter thru a towel and measure the 

 filtrate. 



(6) Add 1.0% Witte's peptone and 0.5% 

 NaCl to (5). 



(7) Boil in the steamer. 



(8) Cool in a cold water bath. 



