18 DIFCOMANUAL 



or prevent them from proceeding with the normal functions of their metabolic 

 processes. 



The usual range of temperature suitable for the growth of microorganisms lies 

 between 15° and 43 °C. Microorganisms have, however, been known to develop 

 at 0°G., and others, such as the soil organisms, may grow at 80°C. The patho- 

 genic organisms in general are limited by a comparatively narrow range of tem- 

 perature around 37°G. while the saprophytes usually have a much broader 

 latitude. 



All organisms exhibit three cardinal points in their thermic relations : 



(a) A minimum below which development ceases. 



(b) An optimum at which growth is luxuriant. 



(c) A maximum above which growth ceases and death occurs. 



In addition to a suitable temperature for growth of microorganisms it is 

 necessary to provide sufficient moisture in the atmosphere. Some organisms re- 

 quire a moist surface for growth. For example, media in plates inoculated with 

 the gonococcus, may fail to show growth of the organism if placed in an 

 ordinary incubater at 35-37 °G. A duplicate inoculation, in contrast, in a sealed 

 container in which is placed moist cotton or a wet towel to provide moisture, 

 will show profuse growth. Incubators should have open containers filled with 

 water at all times to provide sufficient moisture for growth and prevent drying of 

 media. Growth of most microorganisms is obtained in the absence of light. Sun- 

 light is to be avoided. 



The media upon which microorganisms are grown must be sterile or free from 

 all other forms whose development might influence or prevent the normal growth 

 of the inoculated type. The usual method for immediate sterilization of culture 

 media is by means of the autoclave in which steam under pressure is the steriliz- 

 ing agent. The proper operation of the autoclave to insure sterilization of media 

 requires careful manipulation. 



Autoclave sterilization for 15 minutes at 15 pounds pressure (121°G.) is recom- 

 mended for quantities of liquid media up to one liter. If larger volumes are to 

 be sterilized in one container, and especially if the medium is not hot when 

 placed in the autoclave, a longer period should be employed. The medium is 

 prepared according to formula, distributed in tubes or flasks which are then 

 plugged with nonabsorbent cotton or loosely capped and placed in the autoclave. 

 Tubes should be placed in racks or packed loosely in baskets. Flasks should never 

 be more than two-thirds full. 



In the operation of the autoclave, all the enclosed air must be allowed to 

 escape and must be completely replaced by steam. Pressure-temperature relations 

 of a properly operated autoclave are shown in the table below. 



Pressure-Temperature Relations in Autoclave 

 (Figures based upon complete replacement of air by steam.) 



Pressure in Pounds Temperature 



o G op 



5 108 226 



10 116 240 



15 121 250 



20 127 260 



25 131 267 



30 134 274 



