STUDIES ON GROWTH CYCLE OF AZOTOBACTER 331 



temperature or changes in the culture media in which they are 

 growing, and that cells of this type are weakened or degenerated 

 and sometimes devitalized. This being the case, the writer 

 finds that with azotobacter, in addition to there being a "regular 

 occurrence of many different forms and stages of growth con- 

 nected with each other by constant relations," there is also an 

 irregular occurrence of more or less abnormal forms which merit 

 the term "involution forms." .These forms, in great variety 

 of size and shape, are fairly common in old cultures (one to two 

 months) of azotobacter grown in Ashby's solution or on Ashby's 

 agar at room temperature, but when cultures are incubated at 

 37°C, they become numerous in a few days. Numerous Ashby's 

 agar hanging block cultures in moist chambers were made by 

 the writer in which various involution forms were included in the 

 inoculum. The development of these cultures was observed 

 under the oil immersion lens and in only a very small percentage 

 of cases did the involution forms show any tendency to develop 

 or reproduce. In most cases the involution forms remained 

 dormant either until they were overgrown by colonies of azoto- 

 bacter developing near them from normal cells included in the 

 inoculum, or until the cultures gradually dried out. Some of 

 these moist chamber cultures were held under observation for 

 fourteen days, sterile water being added to the chamber from 

 time to time to prevent, as long as possible, the drying out of 

 the culture. 



In those cases where the abnormal forms reproduced, the 

 process of reproduction at first more closely resembled budding 

 than ordinary simple fission. Irregular rounded projections 

 would develop, apparently from any part of the cell, and in 

 course of a few hours abstriction of these projections would 

 take place, the abstricted portions then proceeding to develop 

 and reproduce by fission. 



SUMMARY AND CONCLUSIONS 



1. The four varieties of Azotobacter studied by us, which were 

 isolated from garden soil at Guelph, Ontario, in 1910, and were 

 reported on in 1913 and 1914, have a complex life cycle. 



