Protozoa hi Relation to Soil Bacteria. 



301 



these were made into a suspension in sterile distilled water, a measured 

 •volume of which was used for moistening one of the soil samples. This 

 particular lot of soil, together with one to act as a control, had been 

 ■submitted to a further treatment by heating to 75-80° C. for 5 or 

 6 hours with a view to getting rid of the amoebae and flagellates which had 

 survived the previous heating to 65° C. 



In order to estimate the numbers of protozoa in soil, cultures were made 

 in 1 per cent, hay-infusion and on agar plates, as described in my previous 

 communication (p. 442).* It has been found impracticable to weigh smaller 

 ■quantities of soil than 0-0002 grm., and for estimating protozoa further than 

 this point a dilution method was employed in which O'OOl grm. of soil is 

 -shaken up with 10 c.c. of sterile normal salt solution, and then measured 

 quantities of the suspension are transferred to the surface of nutrient* agar 

 plates, three plates being used for each dilution ; 1 c.c. of such a suspension 

 is equivalent to - 0001 gram of soil, and so on. I may add that this method 

 has yielded satisfactory results. The agar-plate method has been found the 

 more serviceable, being especially good for amoebae, whilst the hay-infusion 

 is better for ciliated protozoa, as illustrated by the fact that whilst from the 

 Toluened plus ciliates (T + C) soil I could obtain no Colpoda cucullus by the 

 -agar-plate method, by the use of hay-infusion this organism appeared in 

 •culture regularly down to a dilution of 1 in 1000. Bacterial counts were 

 .made periodically by the gelatine-plate method. 



The complete set of soils as finally put up so as to include various 

 controls consisted of the following :— Untreated (17), Toluened plus sterile 

 distilled water (T + W), Toluened plus 5 per cent, untreated soil (T-f-5 per 

 cent. IT), Toluened plus ciliates in filtered hay-infusion (T + C), Toluened plus 

 mixed culture of protozoa and bacteria in hay-infusion (T + M), Toluened 

 plus filtered culture, hay-infusion without protozoa and bacteria (T + F), 

 Toluened and twice heated plus amoebae (T + A), Toluened and twice heated 

 4TH 2 ). 



Results. 



The results of the periodical bacterial counts are given in the Tables on 

 p. 302, and are represented graphically in figs. 2, 4 and 5. 



* ' Roy. Soc. Proc.,' B, vol. 83 (1915). 



