56 CARBON NUTRITION 



must initiate and maintain growth at the expense of the test com- 

 pound. A second, more narrow, meaning of utilization is the ability 

 of viable active mycelium to make further growth at the expense of 

 a test compound. Here, the design usually involves providing a small 

 amount of a utilizable carbon source and measuring the additional 

 growth, if any, obtained from a second compound. It is often found 

 that materials which are not by themselves satisfactory carbon sources 

 are used in this kind of experiment; examples in particular fungi in- 

 clude lactose (135), sugar alcohols (150), and cellobiose (142). As 

 discussed later, a compound which is used only in association with 

 another carbon source is probably dependent for its metabolism on 

 the induced synthesis of an adaptive enzyme system or upon the oc- 

 currence of mutants possessing the enzyme system. 



So long as these two experimental designs are not confused, each has 

 its place. Confusion, however, does arise if an author proposes to 

 use the first design — one compound as the sole source of carbon — but 

 includes other carbon sources in the medium, either as complex ex- 

 tracts or as amino acids. Unless the additional carbon is negligible, 

 and this must be shown by control experiments without carbohydrate, 

 there is the possibility that the experiment measures utilization in its 

 second meaning rather than the one intended. Fortunately, most 

 fungi can be grown in a synthetic medium with an inorganic nitrogen 

 source and with pure growth factors at levels which do not contribute 

 significant amounts of carbon. 



There are several methodological errors which are common enough 

 to deserve comment. A cluster of such errors is associated with acidity. 

 The initial pH should be determined after sterilization, to guard 

 against the possibility that compounds in the medium are broken down 

 by autoclaving. Some compounds, particularly organic acids, may not 

 enter the cell at the pH chosen, in which case it is necessary to deter- 

 mine whether or not the organism can tolerate a pH at which pene- 

 tration is more likely. Finally, the organism itself may produce or- 

 ganic acids which in a poorly buffered medium lower the pH so far 

 as to restrict growth. Media may be buffered with phosphate for most 

 organisms; for those fungi which cannot tolerate the level of phosphate 

 required for effective buffer action (45), the pH may be controlled by 

 addition of sterile solid calcium carbonate after autoclaving or by in- 

 clusion of an indicator and periodic adjustment of the pH with sterile 

 alkali (58). The only way in which these problems can be detected in 

 a particular instance is to determine the culture pH at frequent inter- 

 vals and to exercise judgment in the interpretation of growth results 

 which may be affected by pH changes. 



