ISOLATION AND PURE-CULTURE METHODS 17 



in the chambered bottom is rapidly brought to the boiling point, 

 and then the gradual entrance through the holes of water from the 

 reservoir will supply the boiler for several hours, if it is necessary 

 to employ it so long. 



The Koch sterilizer is now less used. Aside from being a well- 

 made piece of apparatus, it has only the advantage that the regulation 

 of the water supply is automatic. 

 It is expensive and is only espe- 

 cially desirable in case of steriliza- 

 tion or digestion for many hours. 



Countless experiments have 

 shown that while "the vegetative 

 cells of most bacteria are usually 

 killed by a single sterilization of 

 from fifteen minutes to one hour 

 at 100° C, yet the spores of many 

 forms are not killed by one ex- 

 posure at this temperature. As a 

 matter of fact, an exposure for 

 a few minutes at ioo° C. in the steam sterilizer is usually suffi- 

 cient to kill the growing parts of most fungi. It is not, however, 

 such delicate parts which are to be reckoned with in the sterilization 

 of nutrient media, but rather the resistant spores of fungi and bac- 

 teria, and thick-walled mycelial parts. Forms which are strongly 

 heat-resistant may often be encountered in the preparation of such 

 media as potatoes and manure decoction. 



To Cohn is due the notable discovery of heat-resistance in the 

 spores of bacteria, and logically following this Tyndall demonstrated 

 the necessity of discontinuous or successive sterilization, after in- 

 tervals sufficiently long to permit such organisms, or parts of organ- 

 isms, as have remained as spores to germinate, and therefore to be 

 more readily killed at the next heating. In general, it is necessary 

 to sterilize on three successive days. As is well known, when one 

 is careful in making the medium a single sterilization of half an 

 hour is usually sufficient for tubes of agar, if the apparatus has 

 not become infested with particularly resistant spores. In no case, 

 however, should one depend upon a single sterilization unless the 

 material is to be kept several days previous to inoculation. Stock 



Fig. 4. Arnold Steam Sterilizer, 

 Square Type, showing Construc- 

 tion 



