312 ROBERT T. HANCE 



d. Conjugation 



A cycle of conjugation was awaited for over a year and then a 

 method of bringing it about at will was hit upon accidentally. 

 Two cultures which had been started with 200 cc. of hay infu- 

 sion were allowed to stand uncovered. In seven days the fluid 

 had evaporated to one half and a strong epidemic of conjugation 

 was observed. The experiment was repeated successfully eight 

 times, conjugants always appearing when the fluid had been 

 reduced to approximately one half its original volume. With 

 the containers used this usually required a week. 



From these results it seemed probable that conjugation had 

 been produced by the rapid concentration of metabolic products. 

 To. test this theory further a number of cultures were set up. 

 Some jars were covered and others were allowed to evaporate. 

 The jars used were much smaller than the original containers 

 and the liquid surface exposed was not more than one third as 

 great. Consequently the evaporation was much slower and three 

 weeks were required to reduce the liquid to the point that one 

 week produced in the larger jars. In three jars that contained 

 (1) pure hay infusion, (2) hay infusion plus 0.05 per cent sea 

 salt, and (3) hay infusion plus 0.1 per cent sea salt, although 

 evaporation did not take place conjugation began in (3) in thirty- 

 six days, in (2) in thirty-eight days and in (1) in thirty-nine 

 days. These cultures had been started in drinking glasses. 

 Three cultures similarly made at the same time were allowed to 

 evaporate. No conjugants appeared. When the medium be- 

 came very low in the container it was replenished wdth fresh 

 infusion and again allowed to evaporate. This was repeated 

 five times and no sign of conjugation was observed. 



During the year previous to these experiments cultures had 

 been made up using 500 or 600 cc. of hquid and in this time as 

 has been stated no conjugation took place. Conjugation has 

 been obtained in eighteen cultures when the exposed surface of 

 the fluid was sufficient to permit rapid evaporation at room 

 temperature or when the quantity of fluid was small and the 

 multiplication of the animals rapid. 



