178 CONTRIBUTIONS TO OUR KNOWLEDGE Of SOIL-FERTILITY, xvi., 



exhibit any pronounced degree of toxicity as judged by the effect 

 of boiling, or dilution, or by the relation of the raw extract to 

 the water-control. They are of the indefinite type which leads 

 one to further experimenting in the hope of obtaining something 

 more definite. 



In the soil-tests, we have a direct evidence of toxicity in the 

 twelve days' culture. The filtered extract was directly toxic, 

 reducing the number of bacteria added at the start from 188 to 

 100. The effect of boiling the extract for an hour under a con- 

 denser, and of diluting the raw extract is also well shown in the 

 increased nutritive effect. The toxic nature of the twelve days' 

 culture is confirmed by that of the fifteen days', for, although the 

 added bacteria are not decreased, they did not increase. 



A trial was made to see if humus, such as vegetable-mould, 

 would be useful for maintaining an approximate neutrality. 

 Four grams of lime-treated mould were tried against 50 grams 

 of sterile soil. In fourteen days, the humus test showed a reac- 

 tion of - 0-4, and the soil - 003. Humus was therefore useless 

 for the purpose. There was no evidence of toxicity in the 

 extracts. 



While dilute hay-infusion has been found to be very good for 

 growing the amoebae, and for obtaining evidence of toxin-forma- 

 tion, it seemed advisable to see if a simple nutrient would be as 

 good. The preliminary tests in the cultivation of the amoeba- 

 had shown that they grew well in urea and ammonium chloride 

 and, with these, calcium nitrate was included as a nutrient, 

 which would probably not produce an alteration in the reaction 

 of the culture-medium. Solutions of urea, 0*02%, ammonium 

 chloride, 0036%, and calcium nitrate, 0-056% in tap-water, were 

 prepared. These contained equivalent quantities of nitrogen. 

 To 500 c.c. of these solutions, 50 grams of soil and 25 c.c. of an 

 amoeba-culture containing 75,000 mobile forms were added. 

 Unfortunately the amoebae did not increase in numbers and were 

 rarely more than 330 per c.c. Extracts were prepared on the 

 seventeenth da}^ when all hope of their increase had been given 

 up. The extracts had the following reactions, urea, - 1-36, 

 ammonium chloride, +0*24, and calcium nitrate, +004. There 



