08 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 formule 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 thts 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. 
