Endogenous respiration of spores of the fungus Myrothecium verrucaria can be stimulated up to over-10 fold by diverse chemicals or by physical treatments. Greatest effects were caused by azide (12-fold at 250 ,Mm) and by 2,4-dinitrophenol (7-fold at 300 ,uM). Marked stimulation was also caused by 10 AM silver (5-fold), 30 gM pentachlorophenol (6-fold), 10 ,uM carbonyl cyanide m-chlorophenyl hydrazone (4.5-fold) and 10 ,uM merthiolate (4-fold). Physical treatments such as heat (50 C), freezing, and sonication at sublethal levels were also The studies reported here were initiated following the discovery that azide had a remarkably high stimulatory effect on the endogenous respiration of spores of the fungus Myrothecium verrucaria (11) and that trehalose constituted about 20% of the spore dry weight, thus providing sufficient reserves to support germination (14). It was hoped that an investigation of the action of azide and comparison with effects of other chemicals and physical treatments on endogenous metabolism would contribute to an understanding of the mechanisms of dormancy and germination in these spores.Instances of metabolic stimulation by compounds usually having inhibitory action are not uncommon. Webb (22) has discussed this in general terms and has proposed a number of hypothetical mechanisms to account for the phenomenon. Slater (19) gives additional discussion relating to agents which stimulate metabolism through uncoupling oxidative phosphorylation, the mechanism proposed by most workers.In essentially all published reports of stimulation we have encountered (2,15,16,18), the 2-to 3-fold increases reported have been much less dramatic than those observed here. The only exceptions are the studies of Desser and Broda (4, 5) who used labeled Bacillus cereus spores to permit detection of extremely low rates of respiration. They reported 80-fold increases in the presence of azide. It should be noted, however, that the absolute increase was extremely low, i.e., Q.2 = 0. 110 compared with 0.0013 for control spores.MATERIALS AND METHODS Cultural and procedural methods were similar to those reported previously (10,14). Briefly, spores of M. verrucaria (Alb. and Schw.) Ditm. ex Fr. QM460 were harvested from surface inoculated agar cultures grown at 29 C with filter paper as carbon source, and 0.005% yeast extract was added to improve sporulation. Sporulation was complete in about 5 days. Figures given as spore age are actually culture age, true spore age being about 5 days less. Spore suspensions were prepared freshly before use by washing with water and suspending in buffer (50 mM KH,PO-KMHPO4, pH 5.5) at concentrations on the order of 5 mg/ml. Dry weights were determined on aliquots filtered through fine sintered glass crucibles. Cell volume changes were determined by centrifuging in protein tubes (12).Respirometry was by standard Warburg techniques at 30 C with 1.0 ml of spores in 50 mm potassium phosphate buffer at pH 5.5 in the vessel, 0.2 ml of 10% KOH in the center well, and 0.5 ml of additive ...