KLUYVER S CONTRIBUTIONS TO MICROBIOLOGY AND BIOCHEMISTRY 



The opportunity to carry out such an investigation in Kluyver's 

 laboratory presented itself when Mayer [1938TI1] there encountered 

 a typical case of dissociation during his studies on the microbiology 

 of the 'Tibi grain'. This material, like the 'ginger beer plant' in Eng- 

 land, was used in some countries for the preparation of mildly alco- 

 holic, effervescent beverages from sugar solutions flavoured with var- 

 ious fruits. Mayer showed that these grains, which multiply during 

 the fermentation, are composed of a yeast, isolated in pure culture and 

 identified as Saccharomyces intermedins, and of a rod-shaped hetero- 

 fermentative lactic acid bacterium characterized by its ability to pro- 

 duce large capsules of dextran in sucrose media. This bacterium was 

 also isolated in pure culture and named Betabacterium vermiforme ; it 

 represented the first instance of a rod-shaped, dextran-forming lactic 

 acid bacterium. Mayer had moreover accomplished the synthesis of 

 typical 'Tibi grains' from the pure cultures of the two components. 



Starting with fresh 'Tibi grains' from fermenting solutions, Mayer 

 had frequently observed that the lactic acid bacteria colonies develop- 

 ing on sucrose-gelatin plates were all of one kind. But during the iso- 

 lation of Betab. vermiforme it had also been found that different kinds 

 of colonies appeared on glucose-agar plates streaked with suspensions 

 prepared from single, well-isolated colonies of uniform appearance 

 that had grown on solid media. In addition to the more numerous 

 'rough' colonies, representing the initial type, irregular flat ones were 

 encountered, and it was shown that every suspension of a 'rough' 

 colony invariably gave rise to a certain proportion of colonies of the 

 flat type. It is important to note that this behaviour was observed with 

 cultures on glucose-containing media; here capsule formation never 

 occurs, and this precludes the possibility that contaminating bacteria 

 would be regularly entrapped in the stiff mucus. By the side of the 

 flat, irregular colonies, smooth elevated ones were also found occa- 

 sionally. Associated with the changes in colonial structure was a loss 

 in ability to produce dextran from sucrose; this applied to both the 

 flat and the smooth types. 



A meritoriously critical evaluation of the sequence of events and of 

 many other aspects of the observations finally led to the conviction 

 that the observed 'dissociation' could best be explained as the result 

 of mutations, even though at the time this interpretation was prob- 

 ably the least favoured by microbiologists. The reasoning that led to 



!5! 



