SPORE GERMINATION WITHOUT BACTERIA 87 



to be tested was then placed on the under surface of each of the 

 ten cover-glasses with the help of a sterilised platinum loop ; and 

 each drop was inoculated with spores removed from a pure spore- 

 deposit with a sterilised needle. After the ten rings had all been 

 provided with hanging drops containing spores, a little specially 

 distilled water was placed in the bottom of the Petri dish and the 

 dish cover again applied. The whole apparatus was then placed 

 in the dark. The spore-deposits used for inoculating the drops 

 were collected by placing sterilised glass slides just beneath pilei 

 which were shedding spores in a small damp-chamber ; and they 

 were kept dry in a sterilised box for from 6 to 38 days before being 

 used. Duggar's arrangement for the glass cells has various good 

 features of which ease of sterilisation and the avoidance of either 

 wax or vaseline are the chief. In a few experiments ordinary 

 glass-ring cells were employed. A single ring was attached to a 

 glass shde with paraffin wax, distilled water was placed in the 

 bottom of the cell, and the cover-glass with its hanging drop 

 containing spores was sealed with vaseline. The ring and cover- 

 glass were sterilised with a flame before being used. The drops 

 into which the spores were introduced remained free from bacteria 

 for as long as they were observed, i.e. for several days. Had 

 bacteria developed, they would have shown themselves under the 

 microscope in the nutrient gelatine as colonies, and in the liquid 

 media as a cloud of individual particles ; but no traces of these 

 organisms ever appeared in any of the critical experiments. 



In one series of experiments, the drop cultures, made in five 

 series with the five different kinds of culture media, were kept in 

 a chamber in which the temperature was maintained at about 30° C. 

 Under these conditions the first signs of germination usually 

 occurred from 23 to 37 hours after the spores had been sown ; and 

 from 80 to 90 per cent, of the spores were found to have germinated 

 at the end of about 48 hours. No bacteria appeared in any of the 

 drops. This series of experiments seems to prove conclusively that 

 the presence of bacteria is not required for the germination of the 

 spores. 



Further evidence that the spores will germinate in the absence 

 of bacteria was obtained in the following manner. A series of test- 



