STUDIES ON GROWTH CYCLE OF AZOTOBACTER 329 



SYMPLASTIC STAGE 



With regard to the theory of the symplastic stage in the life 

 cycle of azotobacter described by Lohnis and Smith, our obser- 

 vations of the four varieties of azotobacter studied lead us to 

 conclude that there are good grounds for accepting the theory. 

 Such clusters of cells referred to by Lohnis and Smith as "sym- 

 plasm" have been observed by us in various stages from clusters 

 in which the individual cells were well defined to clusters in 

 which it was difficult to distinguish individual cells but in which 

 minute granules were present in considerable numbers, evidently 

 corresponding to the reproductive granules of Lohnis and Smith. 

 These granules were readily responsive to some stains, particu- 

 larly Heidenhain's iron haematoxylin, which stains them black, 

 and Neisser's blue, which stains them dark blue, and makes 

 them quite distinct from the surrounding substance. In other 

 masses of "symplasm" these granules were larger in size and 

 were assuming the appearance of individual cells, and in still 

 others they had become small individual azotobacter cells, 

 multiplying by fission. 



Previous to the publication of Lohnis' and Smith's symplastic 

 theory, we had considered these clusters to be mechanical ag- 

 glomerations of cells, their association being not vital but acci- 

 dental, and the formation of the reproductive granules with 

 their subsequent liberation being identical with that which we 

 described in 1914 as occurring in individual azotobacter cells. 



However, from the observations that we have made this year 

 in connection with our cultures of azotobacter, we have come to 

 the conclusion that there are good grounds for accepting Lohnis' 

 and Smith's theory regarding the fusion or mixing together of 

 the protoplasm of those cells which constitute these symplastic 

 clusters. We have observed these symplastic masses in stained 

 preparations from cultures ranging in age from a few days to 

 several months, on beef peptone agar, Ashby's agar and in 

 Ashby's solution, and, notwithstanding the fact that at first 

 we were antagonistic to the theory, we were finally led to accept 

 it by our repeated observations. 



