42 MANUAL OF MICROBIOLOGICAL METHODS 



in water and milk) ''standard media" have been designated by other 

 organizations (American Public Health Association, 1946, 1948). 



The peptone^ listed as an ingredient in some of the formulas is a product 

 derived by digestion of proteinaceous materials of either plant or animal 

 origin, by use of acid, alkali, or added or natural proteolytic enzymes 

 (Asheshov, 1941; Brewer, 1943; Gladstone and Fildes, 1940; Leifson, 

 1943; Mueller and Johnson, 1941). Since the composition of peptone 

 varies with the origin and the method of preparation, not all types may 

 be suitable in all instances ; any type of bacteriological grade found to give 

 best results for a particular purpose may be used. Data available at 

 present indicate that peptone prepared by pancreatic digestion of casein 

 (for example, B.B.L. "trypticase" and Difco "casitone") will often con- 

 tain growth-promoting substances, required by many organisms, which 

 are not found in some other peptones. In the formulas listed below in 

 which peptone is specified, a particular type is indicated in a few instances; 

 when this procedure is not followed, the worker may choose the type 

 giving more satisfactory results. 



Agar,^ a complex carbohydrate refined from marine algae, is the usual 

 agent for solidification of media. This material should be free from 

 starch and debris and capable of producing a clear solution when hot ; the 

 exact concentration to be used to give the desired degree of solidity may 

 vary with the degree of purification, although usually 1.5 per cent is 

 sufficient. Agar media should not be adjusted to a pH lower than 6.0 

 prior to sterilization, since the agar is hydrolyzed under these conditions. 

 When such agar media are required, the pH is adjusted after sterilization 

 by the aseptic addition of acid. 



All laboratory-prepared infusions, especially those from meat, should 

 be checked microscopically to assure freedom from bacterial growth, 

 especially if the infusion is held for even a few hours at temperatures 

 which. will permit microbial reproduction. 



Beef -extract peptone broth (often called nutrient hroth) ordinarily has 

 the following composition: beef extract, 3 g; peptone, 5.0 g; distilled 

 water, 1,000 ml. The Avater is heated to 60°C to promote solution of 

 ingredients; after cooling, the pH is adjusted to 7.0-7.2. After dispens- 

 ing in tubes or other containers, it is autoclaved at 121°C for 15 min. 

 This medium, still used quite widely for the general cultivation of 

 aerobic organisms and as a basal medium for a variety of physiological 

 tests, is now recognized to be nutritionally inadequate for many types of 

 fastidious organisms. In many instances addition of 5 g of yeast extract 

 will support growth of such types. For heef-extract agar (nutrient agar), 

 1.5 per cent of agar is added to the above medium before dispensing and 

 autoclaving. 



^ See appendix to this chapter for specifications of some peptones and agar. 



