ISOLATION AND PURE-CULTURE METHODS 37 



perfect, the species may be more or less readily differentiated, and 

 with a sterile needle transfers may be made of each of the one or 

 more promising sorts to such media as may have been prepared 

 for the purpose. This method suffices, of course, when the desire 

 is merely to get cultures of different fungi. When, however, one 

 wishes to get the product of a certain kind of spore, it is absolutely 

 essential to follow the germination of this spore in the Petri dish, 

 to locate germinating spores at a distance from any other organ- 

 isms, and then to mark the glass and observe these from day to day 

 or to directly remove these isolated spores with some of the sur- 

 rounding agar by means of a sterile needle, or scalpel point, to 

 tubes of prepared media. In the latter case a considerable number 

 of such cultures should be made, and the results may not be taken as 

 entirely conclusive unless there is agreement between the cultures 

 thus made. When the fungus is one possessing characters by means 

 of which it may be readily determined, the problem is not difficult. 



Frequently the spores which are to be located are so small that 

 it will be necessary to remove the cover of the Petri dish, and to 

 examine it fearlessly with the agar surface exposed. If carefully 

 done, the contaminations resulting are practically negligible. It 

 will be necessary to use an objective with a long working distance 

 and the |-inch or ^-inch is preferable. A rough examination, 

 where the spores are large, may be given by inverting the dish and 

 cover, making the examination from the bottom, and then the 

 location of spores may be indicated by ink marks. 



Establishing pure cultures: subcultures. The process of trans- 

 planting bacteria, spores, or mycelial masses from an isolation cul- 

 ture to sterile tubes of prepared media is properly that of establishing 

 pure cultures. Frequently it is desirable to make a large number 

 of such subcultural transplantings to be used as the stock cultures 

 from which, in future, any necessary series of experiments may 

 proceed. 



Under ordinary laboratory conditions, tube cultures may begin to 

 dry out in from six weeks to several months, and must therefore be 

 renewed or transferred. This consists merely in inoculating fresh 

 tubes from the old cultures. A record of such transfers is, for 

 physiological purposes at least, important, and may be indicated on 

 the label, or in the record book. 



