45 



indicator in the Rebipelagar is neutral red. Acid producing colonies are 

 distinctly red. This medium is very extensively used in England but has 

 not found favor in the United States for, as will be shown later in the 

 discussion of the presumptive test for Bact. coli, there is considerable evi- 

 dence that the bile salts may inhibit some strains of Bact. coli. Reference 

 to the foregoing table shows that in one of the fecal suspensions a count 

 of 450,000 on agar was reduced to 60,000 by bile. 



The isolation of the colon group by direct plating is recommended for 

 highly polluted waters and sewage where the number of coli strains are 

 quite large, being present in one c. c. or less. If, however, it is necessary 

 to employ large samples (5 c. c. or 10 c. c.) as is essential in routine 

 water work, it becomes extremely inconvenient to plate out. Furthermore, 

 if the incidence of the colon group is small compared to other forms, they 

 may be missed. Although theoretically the colon colonies are clearly dif- 

 ferentiated from others, it is the practical experience of all investigators 

 that when growing on a plate with a large number of alkali formers, the 

 acid of Bad. coli and related forms may be neutralized, and the characteris- 

 tic reaction thereby masked, so that a colon colony may thus escape de- 

 tection. On the other hand, in the process of sterilization of medium the 

 lactose may be decomposed in part and it will then be found that some 

 soil forms will produce an acid reaction on solid media. Acid production 

 on any of the solid media described above can therefore not be relied upon 

 to always indicate the colon group, and so further tests must be resorted 

 to as has been mentioned in discussing litmus lactose agar. 



For the isolation from relatively large quantities of water, or for an 

 admixture with numerous other forms, the best results are obtained by the 

 indirect or so-called preliminary enrichment method. The procedure con- 

 sists of three steps: 



1. Growth in a favorable liquid medium containing some constituent 

 which will indicate the probable presence of the colon group. This is 

 spoken of as a preliminary enrichment tube and is often employed as a pre- 

 sumptive test. 



2. Isolation from this preliminary enrichment tube on some one of 

 the solid media described above. If typical colonies are formed the pres- 

 ence of members of the colon group is considered partially confirmed. 



3. Suspicious colonies on (2) are examined, by fishing to agar and 

 then testing for coagulation of litmus milk, gas in lactose broth, V. P., etc. 

 This would constitute a completely confirmed test. 



The media most commonly employed for preliminary enrichment are: 

 Glucose broth 

 Phenol broth 

 Liver broth 

 Lactose broth 

 Lactose bile salts broth 



Eijkman test (glucose peptone water at 46 C.) 

 Lactose peptone bile 



