120 CROWN-GALL OF PLANTS. 



VITALITY ON CULTURE MEDIA. 



The life of tJiis organism on culture media is brief to moderate. 

 Often in hot summer weather agar streak cultures were found dead 

 after 15 to 25 days when exposed to room temperatures. In cooler 

 weather agar stab and streak cultures have lived for 4 or 5 weeks, 

 but many observations made in the course of the prolonged inocula- 

 tion experiments indicate that the organism is rather short lived on 

 agar. It lives somewhat longer on agar kept in the ice box. Cultures 

 freslily made from the galls have to be transferred as often as every 

 three weeks if one would be certain of keeping them alive. The 

 length of life of this organism is considerably prolonged by growing 

 it in liquid media, notably in milk, in which it will retain its vitality 

 for more than twice the length of time that it will on agar, whether 

 kept in the ice box or at room temperature. (See Milk, p. 112.) 



Flask cultures made in February, 1910, in river water containing 1 

 per cent dextrose, 1 per cent Witte's peptone, and some grams of 

 calcium carbonate, were alive at the end of 7 months. 



TEMPERATURE RELATIONS. 



THERMAL DEATH POINT. 



TJie death temperature is about 51° C, exposing for 10 minutes in 

 + 15 peptonized beef-bouillon. After several preliminary tests, e.g., 

 at 43° to 53° C.,it was concluded that the thermal death point must 

 lie between these two temperatures. The following tests were then 

 carried through in order to determine the point more accurately. 

 Three sets of 4 tubes each of peptonized beef broth were inoculated 

 with the daisy organism from 3-day-old slant agar cultures, and these 

 tubes were placed in water at constant temperatures of 50°, 51°, and 

 52° C. At the end of 10 miinutes the tubes were removed and kept for 

 several days at about 29° C. At the end of 4 days growth appeared 

 in all the tubes that had been exposed to 50° C. for 10 minutes. 

 SHght growth appeared in some of the tubes that had been kept at 

 51 C, but no growth appeared even after 10 days in the tubes that 

 had been kept at 52° C. for 10 minutes. This experiment was repeated 

 several times with the same result, indicating that under the condi- 

 tions named 51° C, is near the thermal death point of this organism. 



OPTIMUM TEMPEUATURE. 



This appears to lie between 25° and 28° C. Growth at 25° C. on 

 standard agar and in peptonized beef bouillon was three times better 

 than at 30° C, and decidedly better than at 12° C. At the latter 

 temperature growth was better in the bouillon. 



213 



