CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



203 



subcultures add fresh normal rabbit 

 blood.) 

 (6) Cover with a layer of parafHn 1.5 cm. 

 high. 

 Sterilization: Not specified. 

 Use: Cultivation of Spironema duttoni. 

 Incubate for 24 hours at 37°C., and then 

 at room temperature. 

 Reference: Kligler (1922 p. 215). 



727. Hiss' Basal Serum Peptone Solution 

 (Buerger et al.) 



Constituents : 



1. Distilled water 1000.0 cc. 



2. Serum (Beef) 500.0 cc. 



3. Peptone 30.0 g. 



4. Litmus (Kahlbaum's) 

 Preparation: 



(1) Mix a liter of distilled water and 

 500.0 cc. of beef serum. 



(2) Flask and steam for 10 minutes. 



(3) Dissolve 30.0 g. of peptone in a little 

 water (100 to 150.0 cc.) by heating 

 over a small flame. 



(4) Filter (3) and cool. 



(5) Cool (2) and add to (3). 



(6) Add one of the added nutrients and 

 sufficient Kahlbaum's litmus to give 

 the desired color. 



Sterilization: Sterilize for 15 minutes on 

 each of 3 or 4 successive days in streaming 

 steam. 



Use: To study fermentation ability of 

 pneumococci, gonococci, streptococci, etc. 



Added nutrients and variants: 



(a) Buerger added 1.0% inulin. 



(b) Ruediger added 5.0 g. NaCl, used 

 20.0 g. Witte's peptone and prepared 

 the medium as follows: 



(1) Dissolve 5.0 g. NaCl, 20.0 g. Witte's 

 peptone and 2.0% inulin or other 

 carbohydrate in a liter of water. 



(2) Add 20.0 cc. litmus. 



(3) Tube in 2.0 cc. lots. 



(4) Sterilize in the autoclave if using 

 inulin; using other carbohydrates 

 sterilize intermittently. 



(5) Collect beef serum without special 

 precautions and dilute with an 

 equal volume of water. 



(6) Pass (5) thru a Berkefeld filter^ 

 flask in 50.0 to 100.0 cc. quantities 

 and heat at 65°C. for 30 minutes on 

 each of 2 successive days. 



(7) Add 2.0 cc. of (6) to each tube of 

 (4) under aseptic conditions, 

 (c) Watabiki added 5.0 g. NaCl, used 

 10.0 g. Witte's peptone and prepared 

 the medium as follows: 



(1) Dissolve 10.0 g. Witte's peptone, 

 5.0 g. NaCl in a liter of distilled 

 water. 



(2) Boil for 30 minutes and filter. 



(3) Add 15.0 g. of one of the following 

 materials dissolved in a little water 

 and 20.0 cc. of a 5.0% litmus solu- 

 tion to (2). 



mannitol dulcitol 



maltose inulin 



glucose lactose 



dextrin levulose 



sucrose galactose 



(4) Distribute in 5.0 cc. quantities in 

 test tubes. 



(5) Sterilize for 30 minutes on each of 

 3 successive days. 



(6) Add 5.0 cc. of fresh horse serum 

 heated to 55 to 60° for 40 minutes to 

 each tube. 



(7) Incubate for 24 hours to test 

 sterility. 



(d) Park, Williams and Krumwiede pre- 

 pared the medium as follows: 



(1) Dilute serum with 2 or 3 times its 

 volume of distilled water. 



(2) Sterilize (1) in the Arnold. 



(3) Prepare a 10.0% solution of pep- 

 tone. 



(4) Sterilize (3), method not given. 



(5) Prepare a 10.0 or 20.0% solution 

 of any desired carbon source or 

 fermentable material. 



(6) Heat (5) in small containers in the 

 Arnold sterilizer for 30 minutes on 

 each of 3 successive days. When 

 using inulin, sterilize in the auto- 

 clave. 



(7) Add, under aseptic conditions, suffi- 

 cient (4) to (2) to give a 1 .0% peptone 

 concentration. 



(8) Add, under aseptic conditions, suffi- 

 cient (6) to (7) to give a 1.0% con- 

 centration of carbon source. (Gen- 

 erally 5.0% glycerol is employed 

 instead of 1.0%.) In routine work 

 with glucose, lactose, sucrose, man- 

 nitol and dulcitol it is generally 

 sufficient to add the sugar to the 



