Chlamydospore-bearing colonies of Candida albicans were placed under nutritive conditions favorable for germination at various stages of colony development. Only young chlamydospores germinated; yet those from 14-20hr cultures could not be distinguished by light microscopy from large round cells which later budded again on the sporulation medium and became suspensor cells. Mature chlamydospores, i.e., those in the center of colonies 60hr old, could not be induced to germinate under a wide variety of physical or cultural conditions.
The synthesis of ribosomal RNAs in Bacillus subtilis 111 and an asporogenous mutant strain 61, derived from 111, was examined. The RNA synthesis decreased at the early stationary phase in both strains. The addition of either chloramphenicol or casein acid hydrolyzate stimulated the RNA synthesis in the parent strain. In the mutant strain, like the parent, the addition of casein acid hydrolyzate stimulated the RNA synthesis, but chloramphenicol was always without effect. The degradation of pre-formed proteins occurred in the parent strain at the stationary phase. However, no significant degradation was observed in the mutant strain. Although serine and metal proteases were found both in the culture medium and in the cell extract, these proteases were likely not to be functioning in the mutant in viva during the stationary phase. Because of the deficiency in the amino acid pool due to the lack of protease activity, the asporogenous mutant presumably failed to synthesize spore proteins.
The effects of glucose and sodium chloride supplemented in culture media, initial pH of the media and cultural temperature upon bacterial growth were examined in connection with inoculum size using a Biophotometer (Jouan, Paris) which monitors turbidity of the cultures continuously. The bacterial growth was estimated on the basis of lag time and growth rate. The growth rate was derived from L10, which is the time of reduction of lag time when inoculum size is increased ten times. In higher concentrations, glucose and sodium chloride greatly influenced the lag time and growth rate of bacteria, but the concentrations affecting the growth varied with strains. The effect similar to those of glucose and sodium chloride was observed in lower initial pHs. Even when inoculum size was changed, growth rate of bacteria was not affected by these environmental factors. Temperature also influenced both lag time and growth rate. In the range of temperatures tested, lag time became longer and growth rate smaller at lower temperatures than at higher temperatures. Moreover, the growth rate in a large inoculum size was greater than that in a small inoculum size. The effect of inoculum size was discussed from the viewpoint of bacterial growth.
The mechanism of death of yeast cells due to biotin deficiency in the presence of aspartic acid (Asp medium) was further studied. Total fatty acids of yeast cells per culture or per dry cell weight grown in biotin-sufficient Asp medium increased with time, but they did not increase in biotin-deficient condition, although the growth was observed. Each of fatty acids such as palmitic, palmitoleic, stearic, and oleic acids, which are main component of yeast cells, scarcely increased for 8hr in biotin-deficient Asp medium, although they increased gradually in biotin-sufficient condition. The effect of various fatty acids on cells grown under biotin-free Asp medium was examined and a supplementation of palmitoleic, oleic, or linolenic acid promoted the cell growth, same as that of biotin, and saved the yeast from death completely. It was confirmed by using oleic acid-requiring mutant of Saccharomycescerevisiae that cell death also occurred by oleic acid deficiency even in the biotin-sufficient condition. From these results, a primary cause of death due to biotin deficiency was assumed to be the insufficient synthesis of the main component of cell fatty acids such as palmitic, palmitoleic, stearic, and oleic acids during the growth of cells induced by the addition of aspartate.