126 MORPHOLOGIC VARIATION 



servations of the degree of variation in form of the cells during the 

 death phase; to devise a measure of this variation which would con- 

 sider all of the changes in form that occur; to include in the in- 

 vestigation the decrease in number of living cells by plate counts, 

 as well as the decrease in total number of cells by microscopic 

 counts; and to carry on the study in a number of different media 

 with the idea of thus varying the death rate and so determining more 

 conclusively whether or not the variations in cell form were directly 

 correlated with the rate of death. Very little work has been pub- 

 lished upon factors influencing the natural death rate in cultures, 

 and since it could not be predicted that this would vary with the 

 size of seeding and the concentration of nutrients, as does the growth 

 rate, it was thought that more would be accomplished by using 

 media of different composition, such as has been done by most of 

 the authors who have described unusual cell forms, in an attempt to 

 obtain the same types of morphologic variation as they have ob- 

 served. 



With these aims in mind the following experiment was performed. 

 Five lots of agar were prepared, each having as nutrient 2 per 

 cent of peptone. These proved to be neutral (pH 7.4). One lot 

 was kept as such, another was made acid (pH 4.5) by the addition 

 of HCl, a third alkaline (pH 9.2) by the addition of NaOH. The 

 fourth had added to it 5 per cent of NaCl; the fifth, 3 per cent of 

 CaCP. These were inoculated with a stock culture of the colon 

 bacillus, the same strain as was used for the observations recorded 

 in Chapter V. It was not possible to make all of these observations 

 simultaneously, but conditions were kept as much alike as possible; 

 the media were all made from the same lot of peptone, the cultures 

 were inoculated from twenty-four-hour agar slants suspended in 

 water to about the same density. The tubes were incubated at 

 37° in a moist chamber. Samples were removed after one, two, 

 three, five, seven, ten, fourteen, nineteen, and twenty-five days (the 

 final sample of the NaCl series was removed on the twentieth day, 

 listed in Table XXVII as the nineteenth day). Five tubes were 

 removed for sampling, the growth of each suspended in 5 c.c. of 

 sterile water, and the contents of the five tubes were then pooled. 

 From this suspension a 1 c.c. sample was removed and serially 



