314 NINETEENTH REPORT. 



4. Any enzyme which can be separated from the culture liquid by 



precipitation with alcohol, etc., is not the toxic principle. 



5. An extract from the mycelium which produces the toxic sub- 



stance in tlie culture liquid, has no toxic effect upon the 

 leaves. 



6. The original isolation began suddenly to lose its virulence. New 



isolations were virulent. 



7. At the same time the filtrate from the attenuated culture lost 



its toxicity. 



We have to deal in the study of this organism with a pathogene in 

 wliicli the toxic principle is more evident than any lytic agent. This 

 toxic principle is either an excretion from the mycelium rather than an 

 integral part of it, or else it is the result of the cleavage of the culture 

 solution. At all events the toxic substance is able to produce a black- 

 ening quite like that produced in the normal course of the disease. It 

 is jirobably not an enzyme. It is somewhat thermolabile. Although 

 occurring in a medium strongly alkaline witli ammonia, the failure of 

 ammonia solutions of similar alkalinity to produce like results, seems to 

 indicate that the ammonia is not responsible for the toxicity. Associated 

 with the success or failure of tlie toxic substance, we find respectively 

 virulence or attenuation. 



METHODS OF DISSEMINATION. 



The possible methods of dissemination of the fungus were deduced 

 from extensive field observations under control conditions. There are 

 two ])hases to the process by which a jjlant pathogene may be dissemi- 

 nated: First, the translocation of the causal agent; second, the presence 

 of those conditions which are conducive toward the entrance of the host 

 by the organism. In the experiments wliich follow, these two factors 

 were considered wherever possible. 



Soil : 



In the dissemination of any disease, the soil merely plays the part of 

 a harboring agent. To determine whether the plants can be infected 

 directly from the soil whicli has grown a diseased crop, the following 

 experiment was performed: 



The upper two or three inches of soil from a field in which badly dis- 

 eased plants were growing, was selected. This soil, upon examination, 

 was found to contain bits of dried, diseased leaves. A little of the soil 

 was also shaken in water, which was then removed and centrifuged. 

 Spores of M. sarcinae forme were found in the sediment, thus making 

 positive the presence of the fungus. In this soil, red clover seed which 



