V49 22 MANUAL OF METHODS FOR PURE CULTURE STUDY 



TABLE 2 



DEGREES OF ACIDITY EASILY RECOGNIZED IN MILK 



'Neutral" 



'Weak" 



'Moderate"... . 



'Strong" 



'Very strong" . 



INDICATOR. REACTION, ETC. 



Same color with brom cresol purple as sterile inilk- 



i. e. blue to gray-green 

 Color with brom cresol purple lighter than in steril 



milk — i. e. gray-green to greenish yellow 

 Yellow with brom cresol purple. Not curdled 

 Curdled. Blue or green to brom phenol blue 

 Yellow to brom phenol blue 



approximate- 

 pH-value 



6.2-6 8 



5.2-6.0 



4.7-60 



3.4-4 6 



Under 3.4 



During the second World War, stimulated by the unavailability of 

 litmus, Ulricli (1944) proposed using instead of litmus a mixture 

 of methylene blue and chlor phenol red. This combination added 

 to milk shows, for many species, all that litmus does and in addition 

 shows a distinction between acid reduction and alkaline reduction; 

 but in using it one must accustom himself to alkalinity being indicated 

 by red, acid by yellow or green. When using litmus or the Ulrich 

 combination, one must distinguish between reduction before and 

 after coagulation, as the latter is often of little significance. 



It is possible to recognize the five degrees of acidity listed in Table 

 'i by the use of brom cresol purple (either in the milk before inocu- 

 lation or added after incubation), the subsequent addition of brom 

 phenol blue, and observation as to the presence of curdling. This is 

 only a rough method of measurement; but in the routine study of 

 milk cultures it will often be found valuable. 



H. C. Brown (1922) proposed condensed milk diluted with 4 parts water containing 

 phenol red. The reaction is adjusted by addition of alkali until first appearance of a 

 brick red. Subsequent changes of reaction in either direction can be observed. 



Rennet Production 



The production of the enzyme, rennet (lab), can sometimes be recog- 

 nized in litmus milk by noticing the occurrence of coagulation with- 

 out the appearance of acid. It is often obscured by simultaneous di- 

 gestion, however, and two other methods have been proposed which 

 often show rennet production with cultures that fail to show it when 

 inoculated directly into milk. 



Conn (1922) grows bacteria in milk sterilized in the usual manner; after the appear- 

 ance of whey or peptonized milk, 0.5 ml. is transferred to 10 ml. of unsterilized milk 

 and placed in a 37° incubator. Examinations are made every 5 minutes for the first half 

 hour, and at less frequent periods thereafter for a few hours longer. First appearance of 

 coagulation is noted. 



Gorini (1932) obtains vigorous growth on an agar slant, then covers the growth with 



