EXPERIMENT STATION BULLETINS. 475 



meability very much. Naturally, the air surrouuding the soil particles 

 will uever contain more oxygen than the air over a peptone solution. 

 If the bacteria in a sand culture find more available oxygen than in a 

 I)lain solution, this is the result of a faster gas exchange. This ex- 

 change depends entirely upon the surface exposed and upon the oxygen 

 concentration in the liquid and the gas phase. The amount of oxygen 

 in the soil air dejjends directly upon the ventilation. But the surface 

 between liquid and gas, or the depth of the liquid, is also very essential. 

 The culture in peptone solution in a layer about 2 cm. deep produced 

 in two days 2.7 mg NH., (Table VII) ; the same culture spread over 

 sand in a layer about 20 p. deep, produced in the same time 79.5 mg 

 NH3 ; in the latter culture, the surface between gas and liquid was ap- 

 jn'oximately a thousand times larger than in the former, while all 

 other conditions were practically identical. The oxygen concentration 

 in a sterile solution is the same wiiether it is spread over soil or sand 

 particles or in a flask by itself. After inoculation, the bacteria begin 

 to use up the dissolved oxygen and the further growth of obligate 

 aerobic bacteria depends upon the rate of oxygen replacement. If the 

 oxygen-content of the air remains constant, the oxygen replacement is 

 in direct ratio to the surface, or in invert ratio to the depth of the 

 liquid. 



Even the decrease in the depth of liquid from 30 fi to 20 /x (medium 

 sand with 15% and 10% moisture) yields an increase of ammonia 

 (see Table VII). A further decrease of the depth to 10 fx (5% water) 

 results in a decrease of ammonia. This has been explained in the 

 former paragraph by the too slow diffusion of the food in the very 

 thin films. The most abundant oxygen supply is of no benefit to the 

 cells if they have no food to be oxidized. 



It must be remembered that the oxygen supply does not depend solely 

 upon the contact surface between liquid and gas, but also upon the 

 oxygen concentration in this gas. The oxygen of the soil air will be 

 used up very fast, and the further development depends entirely upon 

 the ventilation. If there is no ventilation, the growth of strictly aerobic 

 bacteria T\all cease as soon as the last trace of oxygen is used up, re- 

 gardless of the film thickness. 



It should be pointed out briefly how little oxygen is actually available 

 to aerobic microorganisms in our ordinary liquid media in flasks or 

 test tubes. For experiments concerning the actual oxygen require- 

 ments of microorganisms, it seems necessary to use sand cultures rather 

 than the customary liquid cultures, because only in this way an even 

 oxygen concentration can be secured. 



The best conditions of existence for B mycoides in sand of 1 mm. 

 grain diameter were found to be at about 10% moisture, i. e. at a 

 moisture film of about 20 /x thickness. A soil or sand with 0.5 mm. 

 grainslze would require 20% moisture to give the optimal moisture 

 film. If the grainsize is 0.1 mm., the moisture necessary to produce 

 the optimum film thickness would be 100 parts of water to 100 parts of 

 dry soil. Such soil is, of course, water-logged and has no ventilation 

 whatever. Most agricultural soils have a grainsize nearer 0.1 mm. 

 than 1.0 mm. The consequence is that in these soils, the strictly 

 aerobic bacteria can never have optimal conditions of existence. If 

 the oxygen supply is sufficient, the food-supply is inadequate on account 



