ordinary media. The cultural characteristics of each organism are given 

 in table No. 1. 



It will be seen from an examination of the table that acid is produced 

 readily by some of the cultures, while others produce very little or none 

 at all. If the acid producing cultures and the non-acid producing 

 organisms of this group belong to the same species, it was thought 

 possible to produce by selection from the descendants of the acid pro- 

 ducing variety an organism which would produce acid readily and one 

 which would not produce acid at all. Cultures 1 and 2 were selected. 

 These were plated out in gelatin and typical colonies were fished out 

 and transferred to slant agar tubes. These tubes were incubated for 

 24 hours and at the end of that time a sub-culture in broth was made, 

 and from each of the 24 hour broth cultures a single bacillus was ob- 

 tained by Barber's method (22). This was done in order to make 

 sure that all of the organisms would be descendants of a single cell, 

 so that the individual inheritance might be studied instead of averages. 



Goodman, in his experiments with diphtheria bacilli made transfers 

 direct from sugar broth to sugar broth. The direct influence of environ- 

 ment, as has been pointed out by Win slow and Walker (23), was 

 not entirely excluded. These authors attempted to exclude this factor 

 in their study of the variations in the paratyphoid bacillus by plating 

 out each culture on a series of gelatin plates and inoculating 100 agar 

 tubes from 100 separate colonies of each strain. Dextrose broth tubes 

 were then inoculated from the agar slants and the acidity in each of 

 the sugar broth tubes was determined by titration after 72 hours. 

 Instead of making further inoculations from the broth tubes, those agar 

 cultures were selected which in broth had shown the highest acidity for 

 their respective types. These were plated out and from the colonies on 

 the plates new agar streaks were made. 



Win slow and Walker (23), while excluding the factor of environ- 

 ment, carried their cultures through three generations only and thus did 

 not get the benefit of the accumulation of differences which might have 

 taken place if the number of transfers had been increased. 



In the present investigation, the cultures derived from a single cell 

 were plated out and three series of cultures were made. In the first 

 series, twenty separate colonies were isolated and inoculated into twenty 

 sugar-free broth tubes to which 1% of dextrose had been added. After 

 three days incubation at 37, the acidity was determined by titration 



with :r~:NaOH, using phenolphthalein as an indicator. The tube giving 



the maximum acidity and that showing the minimum acidity were plated 

 out and the dextrose broth tubes were inoculated from each of the 

 plates, a separate colony being fished out for each tube. 



In the second series, the cultures were plated out and 10 agar tubes 

 were inoculated from ten separate colonies. These tubes where incubated 

 for 24 hours at 37 C. and a tube of 1% dextrose broth was inoculated 

 from each. The dextrose tubes were incubated at 37 C. for three days 

 and titrated as in the first series. The tubes giving the maximum and 

 minimum of acid were selected. Wishing to avoid all chances of dealing 

 with organisms that might have acquired some tolerance to degrees of 

 acidity produced in the medium by their own growth, all such were 

 discarded; but the two agar streaks derived from the races that had 

 shown the highest and the lowest acidity for their respective types, were 



