1919] 



ALLEN — AZOTOLACTER CHROOCOCCUM 



27 



of this suspension was then used as the inoculum. This pro- 

 cedure was designated as method A. 



In the other method of inoculation a spiral of the growth 

 on an agar slant was introduced into a regular 10-cc. water 

 blank, well shaken, and a spiral of this used at once as the 

 inoculum. This procedure was designated as method B. 



Twenty 250-cc. flasks wer -a prepared, each containing 20 cc. 

 of Ashby's solution (plus CaCOs). Ten of these were inocu- 

 lated by method A and 10 by method B. For preparing the 

 suspension in the former, the growth from a 72-hour Ashby 

 soil extract agar slant of F4 generation was used, while for 

 method B, material from the same slant 76 hours old was 

 used ; that is, all flasks were seeded on the same date. After in- 

 oculation the cultures were incubated on the shaking machine 1 

 in the warm room which remained at 28-30° C. except one 

 night when it dropped to 22° C. for several hours. Cultures 

 inoculated according to method A showed a distinct and uni- 

 form turbidity on the third day, while those inoculated accord- 

 ing to method B showed no visible growths until the fourth 

 day, and these were less distinct and less uniform than those 

 observed in the other set at 3 days. An incubation period of 

 only 5 days was used, as it was believed that a short period 



TABLE VI 



COMPARISON OF METHODS OF INOCULATION 







Culture 



Method 



no. 



1st dist. 



(mgs. N) 





1 



2.26 





2 



1.59 





3 



2.01 





4 



1.64 





5 



6.08 





6 



4.05 





7 



2.31 





8 



3.34 





9 



3.07 





10 



1.74 





Ave. 







1 & V V-* « 







Inoculated according to: 



A 



Method B 



2nd dist. 



1st dist. 



2nd dist. 



(mgs. N) 



(mgs. N) 



(mgs. N) 



1.31 



2.41 



0.97 



1.26 



2.44 



0.93 



1.14 



1.18 



1.10 



1.10 



1.97 



0.92 



1.14 



Lost* 



0.81 



1.51 



1.10 



0.96 



1.03 



1.71 



0.78 



1.12 



1.11 



0.97 



1.21 



1.81 



0.83 



1.03 



1.67 



1.04 



1.185 





0.93 



* Visible amount of alkali carriec over mechanically. 



1 At the time of setting up this experiment, the mechanical difficulties attend- 

 ing the construction of a satisfactory and reliable rotary shaker had not been 

 overcome, so an ordinary laboratory shaker was geared down so as to tilt the 

 flasks back and forth at a rate of 3 complete excursions each 2 minutes. 



