Chapter 4 

 SPORE FORMATION IN THE COLONY 



Knowledge of life cycles and variations in microorganisms has 

 developed almost exclusively from the study of isolated individual 

 cells, without consideration of the structural organization of the 

 colony. Legroux and Magrou (1920) studied the structure of colo- 

 nies of Vibrio cholerae and of a number of other bacteria. They 

 discovered that rod-shaped variants of the Vibrio appear in per- 

 pendicularly arranged packets on the exterior of the colonies, held 

 together by a transparent substance apparently exuded from the 

 cells. They assumed that this substance gave the rough colonies 

 their rigidity. A special stain was used: 



Eosinate of methylene blue . 7.0 gm. 



Eosinate of toluidine blue . . 1.5 



Toluidine blue 5.0 



Methyl alcohol (99.5%) .... 490 cc. 



The rod -shaped organisms in the outer layer of the colony were 

 made up of a violet central region and an azure exterior. The cen- 

 tral region underwent divisions into two or four rounded violet par- 

 ticles, often of unequal size. This phenomenon was found in Vibrio, 

 in the typhoid bacillus, in the diphtheria bacillus, and in the tuber- 

 cle bacillus. Kahn and Nonidez (1933), apparently unacquainted with 

 the work of Legroux and Magrou, confirmed their findings in a study 

 of the tubercle bacillus but concluded that the outer layer was the 

 younger part of the colony. The work of Legroux and Magrou proves 

 this view untenable. 



Pisova (1930) found that when a yeast colony is grown on agar, 

 a pseudo-mycelial growth of long fibrous cells penetrates the agar, 

 especially at a high sugar concentration. After a few days or weeks, 

 the surface cells begin to autolyze, continuing until an outer layer 

 of autolyzed cells is formed. 



Lindegren and Hamilton (1944) grew yeast colonies on malt- 

 extract yeast agar, and cut out portions of the agar containing colo- 

 nies which were then dropped into Flemming's solution. After 24 

 hours the killing fluid was washed out in tap water, and the material 

 was imbedded in paraffin. Sections were made and stained with a 

 variety of dyes. Direct smears were also made by cutting the fresh 

 colony in half vertically with a razor and pressing the exposed sec- 

 tion gently against the slide. The cells were fixed on the slide by 

 heat and stained. 



4-1 



