174 CONTRIBUTIONS '10 OUR KNOWLEDGE OF SOIL-FERTILITY, xvi., 



hay-infusion, in which the bacteria had grown for some time, had 

 changed from having a faint original acidity of -I-0-25 to an 

 alkalinity of —2*4. It was naturally assumed that the alka- 

 linity developed overnight had been the cause of the disappear- 

 ance of the ama3b8e, but, when either saccharose, lactose, lactic 

 acid, humic acid, or sodium phosphate was added, the trouble 

 still remained, although it was not so pronounced in the presence 

 of humic acid. The addition of potassium nitrate, mono- or 

 di-hydrogen phosphate, or ammonium phosphate to make a 005% 

 solution with the hay-infusion, also resulted in the disappear- 

 ance of the amcebse. It seemed at one time as if the smallest 

 droplets gave the most satisfactory growths, that one amoeba in 

 a small droplet had a better chance of growing than one cell in 

 a large droplet, and it was concluded that the hay-infusion was 

 too strong. It is a recognised rule, in growing amoebae, that a 

 poor medium should be used in order to prevent the protozoon 

 being overwhelmed with bacteria, but 1% hay-infusion is by no 

 means considered to be a rich medium, especially when made 

 from a rather poor sample of couch-grass. However, experiments 

 j showed that 0-2% hay-infusion was well adapted for growing 

 amoebae in mass-culture, although it produced rather delicate 

 forms in single-cell work. One of my most successful starter- 

 cultures was obtained by gradually adding 1% hay-infusion so 

 that the bacteria were kept under as much as possible. An 

 equally good starter was obtained by growing the cells in a 2% 

 infusion of exhausted leaf-mould, the same as was used in some 

 experiments about to be recorded. 



During this part of the investigation, the effect of adding 

 sodium chloride to the hay-infusion was tested. In one instance, 

 the addition of 0-2% was beneficial, while a larger quantity de- 

 stroyed the amoebae. In another instance, the addition was 

 injurious. A good culture-fluid was found in tap-water contain- 

 ing 0-05% asparagin with 0-11% K^HPO^. Although two of 

 the original five cells died overnight, the remaining three cells 

 increased to 39 in another day. In view of this, an experiment 

 was made in which the asparagin was replaced by chloride, 

 sulphate or nitrate of ammonia, nitrate of potash and urea. A 



