The rate of commitment of bacterial spores to germinate after short exposure to L-alanine increases exponentially from the time of addition of L-alanine. This absence of a lag facilitates kinetic analysis and allows the dependence of commitment on temperature and pH to be determined. The pH profile of commitment has been compared with that obtained from measurements of absorbance decreases during germination, and the two profiles exhibit differing pK values. It is suggested that because the decrease in A600 of spore suspensions is a late event in germination, it is an unsuitable parameter for studying germination-triggering reactions. Commitment has been shown to be temperature-dependent, with an optimum at approx. 37 degrees C and an activation energy (mu) of 1.08 X 10(5) J/mol. The data obtained from the present studies have been used to develop a model for the triggering of germination.

SummaryIn spite of displaying the most extreme dormancy and resistance properties known among living systems, bacterial endospores retain an alert environmentsensing mechanism that can respond within seconds to the presence of specific germinants. This germination response is triggered in the absence of both germinant and germinant-stimulated metabolism. Genes coding for components of the sensing mechanism in spores of Bacillus subtilis have been cloned and sequenced. However, the molecular mechanism whereby these receptors interact with germinants to initiate the germination response is unknown. Recent evidence has suggested that in spores of Baciiius megaterium KM, proteoiytic activation of an autolytic enzyme constitutes part of the germination trigger reaction. Environmental sensing and spore dormancyMicrobial cells possess environment-sensing mechanisms that respond to a variety of stimuli, including chemotactic gradients, heat shock, changes in nutrient status and the presence of deletenous chemicals. Many of these mechanisms have been recognized to include a two-component protein system with conserved functional domains (Ronson et ai., 1987;Kofoid and Parkinson, 1988). Such responses are dependent on metabolic signalling events within the cell and some (e.g. sporulation and the heatshock response) require the transcription of new sets of genes mediated, at least in part, by alternative sigma factors. Bacterial endospores, produced as a result of differentiation of Bacillus and Clostridium species, possess a different and possibly unique class of environment-sensing mechanism. This mechanism, essential for the germination response, is the subject of this review.

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The germination response of spores of Bacillus megaterium KM, as measured by loss of A600, is more than 95% inhibited by 1 mM-HgCl2. Two Hg2+-sensitive sites (referred to as 'sites I and II') have been identified during germination. Site I represents a pre-commitment event and can be protected from HgCl2 by 50 mM-D-alanine, whereas site II represents a post-commitment event and is not D-alanine-protectable. At 1 mM-HgCl2, 25% of the spore population becomes committed to germinate, but an A600 loss of less than 5% occurs. In this system, loss of heat resistance was associated with commitment, whereas selective cortex hydrolysis, release of pyridine-2,6-dicarboxylic acid, Zn2+ and soluble peptidoglycan, as well as loss of refractility, were identified as post-commitment events. The commitment event was reversibly inhibited by several proteinase inhibitors and a membrane bulking agent. A model of spore germination based on these results is presented.