BIOLOGY OF CLOSTRIDIUM WELCHII 381 



remaining spores fail to germinate and are observed microscopic- 

 ally. On the other hand, it has never been possible to cause 

 sporulation of a vegetative strain in the presence of a fermentable 

 carbohydrate. 



4. Relation of spores to acidity. Spores of Clostridium Welchii 

 withstand a wide range of reaction, surviving + 12 to - 2 per 

 cent. Ordinarily spores are kept in neutral or slightly acid 

 media but in some cases where all the spores failed to germinate 

 in sugar media they could be recovered for a long time even if 

 the acidity had reached a maximum of + 10 or + 12 per cent. 

 Spores will develop in slightly acid or alkaline media but are 

 formed best in neutral media. An environment of 5 per cent 

 acid or 3 per cent alkali will inhibit the formation of spores 

 but if already formed they will survive such an environment for 

 a long time and can be preserved in such a condition. Media 

 of — 2 reaction did not inhibit the formation of spores from the 

 vegetative forms. 



VI. CULTURAL REQUIREMENTS 



1. Aerobiosis. Clostridium Welchii is an obligate anaerobe 

 which requires strict anaerobiosis for its growth. All anaerobes, 

 especially obligate anaerobes, have an optimum oxygen tension 

 above which growth ceases. Although this tension for Clostrid- 

 ium Welchii is greater than for some others of this class, anaero- 

 biosis must be fully maintained to obtain consistent and satis- 

 factory results. Milk tubes which are not rendered strictly 

 anaerobic before inoculation with this organism show either no 

 growth or simple coagulation without a trace of gas formation, 

 presenting that atypical reaction so often reported by investi- 

 gators. This same phenomenon is observed with stock media 

 which has been sterilized under a film of oil (albolin) . 



At the end of three months, the oil does not admit oxygen 

 in a sufficient quantity to inhibit the growth of Clostridium Welchii 

 when inoculated. At the end of five months, however, oxygen 

 has been absorbed by the medium to such an extent that the 

 optimum aerobic supply is exceeded and consequently growth 



