BACTERIA IN RELATION TO PLANT DISEASES. 



mountain regions must use autoclaves. Agar, potato, etc., in test-tubes, may be 

 steamed twenty minutes on each of three consecutive days. Gelatin, beef-bouillon, 

 and all other fluids likely to be injured by long heating should be steamed only ten or 

 fifteen minutes on each of three consecutive days, if in tubes. The writer frequently 

 steams such media fifteen minutes the first day, ten minutes the second, and five 

 minutes the third. Agar, gelatin, bouillon, etc., stored in flasks in large quantity 

 must be steamed a longer time usually thirty to forty-five minutes on each day. 



The first steaming, when softened gelatin is added 

 to bouillon, usually requires thirty minutes. To 

 melt flasked agar quickly, shake it into fragments 

 or break it with a sterile glass rod before putting 

 it into the steamer. 



Oversteaming should be carefully avoided. It 

 softens gelatins or altogether prevents their solidi- 

 fication, and is very apt to cause troublesome pre- 

 cipitates in a variety of media. Precipitates in 

 bouillon often occur if the tubes are not clean, or 

 if the bouillon was not well boiled at first before 

 filtering and placing in tubes. If the beef-broth 

 looks greenish in the beaker or flask, rather than 

 a clear yellow, it may be assumed that it needs 

 more boiling and that if tubed in this condition it 

 will throw down whitish particles on subsequent 

 steaming. The writer prefers to obtain his ordi- 

 nary + bouillons by incomplete neutralization 

 with sodium hydrate rather than by addition of 

 hydrochloric acid after full neutralization. The 

 adding of hydrochloric acid precipitates out certain 

 nutrient substances and also seems to interfere 

 with the growth of some organisms. Distilled 

 water and river water should be sterilized in 

 quantity in the autoclave. For details concern- 

 ing the making of particular media the student 

 should consult the standard text-books, a dozen or more of which should be kept 

 within easy reach in every laboratory. Some formulse are given in the middle 

 part of this volume. The autoclave may be used for the preparation of sterile 

 water and some media, but, in general, I prefer media which has not been heated 

 above 100 C., especially for use with sensitive organisms. Media should be 

 heated in the autoclave only for a brief time and at a minimum pressure, generally 

 not more than ten minutes and at not more than 110 C. Milk, gelatin, and 

 media containing sugars should never be sterilized in the autoclave. Sugars 



Fig. 83 * 



*Fic. 83. Apparatus for rapidly filling test-tubes with 10 cc. portions of agar, bouillon, etc. By 

 means of tins device an expert assistant can fill 500 tubes an hour. Made to order by Emil Greiner. 

 Height, 23 inches. The bulb above X is essential. 



