Summary: Progressive stages in the germination at 60° of the highly refractile spores of were observed by means of phase-contrast microscopy. The development of individual spores was followed in slide cultures. Mass cultures were also prepared in liquid and solid media from spore suspensions in phosphate buffer which had been heated at 100° for periods of 1-20 min. and 30 min.-4 hr. The liquid cultures derived from heated and unheated spores were sampled at frequent intervals and examined in wet preparations containing dilute toluidine blue. Throughout, loss of refractility preceded germination, which occurred in 1-3 hr. Germinating spores appeared as dense opaque bodies by phase contrast and were then capable of absorbing the dyestuff to which non-germinating spores were impermeable.

Heat activation at 100° for 1 min. enhanced the initial germination rate within the first 3 hr. period. No differences were observed between heated and unheated spores during the processes of germination. Indications of an eccentric ‘nucleus’ were occasionally found in phase-contrast examination of very lightly stained fixed spores from material heated for short periods, or in electron micrographs of unheated growths. Heating at 100° for the shorter periods (1-10 min.) resulted in a higher proportion of the spores becoming darkened than was the case with the longer exposures. Artificial darkening of the spores in the manner successfully used for eubacterial spores was not accomplished. Sectored colonies—the sectors showing loss of aerial mycelium and impaired viability—frequently developed from spores which had been subjected for more than 1 hr. to a temperature of 100°.


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