256 APPLIED BIOLOGY 



Sterilization. It is probable that both the bread inside the tubes 

 and the cotton have spores of molds on their surfaces, because the 

 spores are so abundant in ordinary buildings that practically every- 

 thing exposed to air will have them. It is therefore necessary to kill 

 all spores inside the plugged tubes, that is, to sterilize or to make the 

 tubes sterile. This is best done by boiling or steaming (100 C.) 

 in a sterilizer. There are many sterilizers for laboratory and home 

 use on the market, but one can be easily made from any tin 

 bucket with a cover. A sheet of perforated tin (e.g., an inverted 

 pan two inches deep), or a piece of wire-netting, should be supported 

 about two inches from the bottom, and on this place the tubes to be 

 sterilized. Put in water two inches deep. For convenience in 

 keeping mouths of tubes upward, they may be tied into bunches, 

 or placed in small tin cans perforated with numerous holes. A layer 

 of cloth or cotton between the glass tubes and the metal will prevent 

 breakage. 



Keep the water boiling (100 C.) for a half -hour. From time to 

 time add more water (hot) so that the sterilizer will not " boil dry." 



Label some of the tubes "sterilized once, 30 minutes." Set 

 aside for observation from day to day. Do any molds appear on the 

 bread in these ? Conclusions ? 



Sterilize the other tubes again two days after the first sterilizing, 

 and label "sterilized twice, total 60 minutes." Set some of these 

 aside and observe from day to day. Sometimes a third sterilizing 

 is necessary to kill all the spores of molds. 



Inoculation. Take a tube which two or three days after the 

 second or third sterilizing shows no sign of molds on the bread, and 

 hence is probably sterile, and inoculate . it as follows : Sterilize an 

 inoculating needle (or a hat-pin, or piece of wire) by passing it 

 quickly several times through the flame of a gas- or alcohol-lamp. 

 Allow the needle to cool for a moment, and then touch the sterilized 

 end carefully to a spot on moldy bread where there is only one kind 

 of mold. Now, holding one of the sterile tubes in a horizontal 

 position, quickly pull out the cotton plug, insert the needle, wipe it 

 along one side of the piece of sterile bread, and then quickly replace 

 the plug. Inoculate other tubes from other spots of molds, of 

 different kinds if available, but take care to heat the wire of the 

 inoculating needle before and after each tube is inoculated, otherwise 

 you may mix several kinds of spores. Label tubes "black mold," 

 "green mold," etc. 



Do not be surprised if with all these precautions two kinds of 

 molds appear in some tubes ; for spores may have been left alive 



