DEHYDRATED CULTURE MEDIA 31 



3. The identification of Gram negative, non-sporulating, aerobic organisms 

 capable of producing gas when reinoculated into Lactose Broth (completed 

 test). 



Since 1917 "Standard Methods of Water Analysis," in the interests of greater 

 uniformity, has recommended the use of a Beef Extract Lactose Broth in place 

 of the infusion medium formerly employed. Bacto-Lactose Broth is prepared from 

 the ingredients recommended in "Standard Methods" and requires only solution 

 in distilled water and sterilization. No adjustment of reaction is required. 



Bacto-Lactose Broth prepared in the normal concentration contains 0.5 per 

 cent each of Bacto-Lactose and Bacto-Peptone together with 0.3 per cent Bacto- 

 Beef Extract. This concentration of sugar and of peptone yields an optimum 

 growth of the organisms of the coli-aerogenes group.^ "Standard Methods"^ 

 specify that when using inocula greater than 1 ml., multiple strength Lactose 

 Broth shall be prepared to maintain the concentration of nutriments as given in 

 the formula (1 ml. or less of sample may be added to 10 ml. of single strength 

 medium). 



"Standard Methods" procedures apply to the U.S.P.H.S. Drinking Water 

 Standards* in all details of technique in the bacteriological examination, includ- 

 ing apparatus and media. These specifications provide for the use of 10 ml. and 

 100 ml. as standard portions of water for the bacteriological test. They recom- 

 mend standard dilution water bottles of 160 to 180 ml. capacity as containers for 

 the presumptive test where 100 ml. water samples are to be tested; and also 

 authorize the use of either the ordinary inverted vial or a Cowles^ tube for col- 

 lection of gas produced in the medium. The inverted vial or Cowles tube must 

 be of sufficient length so that it will be in at least a 45° angle to the base of the 

 bottle. For convenience it is recommended that the bottles used for the test be 

 marked at the volume of medium to be used and also at the total volume of the 

 medium plus the water sample. For instance, if it is desired to use regularly 35 

 ml. of concentrated medium for each 100 ml. of water sample the bottle can be 

 marked at the 35 ml. and the 135 ml. points. 



In order to maintain a constant final concentration of nutriments in all tests, 

 it is suggested that the medium be prepared in the concentrations given in the 

 table below. 



To rehydrate the medium, dissolve 13 grams, or other specified quantity, of 

 Bacto-Lactose Broth in 1000 ml. of distilled water, distribute into test tubes or 

 bottles with fermentation vials and sterilize in an autoclave for 15 minutes at 

 15 pounds pressure (121° C). The final reaction of the medium will be pH 6.7. 



One pound of Bacto-Lactose Broth will make 34.9 liters of medium. 



Concentrations of Dehydrated Medium Required 

 to Maintain the Proper Concentration of Ingredients 



1 Standard Methods for the Examination 3 Public Health Reports, 44:2865:1929. 

 of Water and Sewage, 9th Edition: 186:1946. * Public Health Reports, 61:376:1946 



2 Standard Methods for the Examination (Reprint No. 2697). 

 of Dairy Products, gth Edition: i33:!048. ^^ J. Bact., 38:677:1939. 



