60 METHODS OF CULTIVATION OF BACTERIA 



erosion of the glass. The whole apparatus may be placed in the 

 incubator till growth occurs. 



It is often advisable in dealing with material suspected to 

 contain anaerobes to inoculate an ordinary deep glucose agar 

 tube with it, and incubating for 24 or 48 hours, to then apply an 

 anaerobic separation method to the resultant growth. Sometimes 

 the high powers of resistance of spores to heat may be taken 

 advantage of in aiding the separation (vide Tetanus). 



Cultures of Anaerobes. When by one or other of the above 

 methods separate colonies have been obtained, growth may be 

 maintained on media in contact with ordinary air. The media 

 generally used are those which contain reducing agents, and the 

 test-tubes containing the medium must be filled to a depth of 4 

 inches. They are sterilised as usual and are called " deep " 

 tubes. The long straight platinum wire is used for inoculating, 

 and it is plunged well down into the " deep " tube. A little air 

 gets into the upper part of the needle track, and no growth takes 

 place there, but in the lower part of the needle track growth 

 occurs. From such " deep " cultures growths may be maintained 

 indefinitely by successive sub-cultures in similar tubes. Even 

 ordinary gelatin and agar can be used in the same way if the 

 medium is heated to boiling-point before use to expel any 

 absorbed oxygen. 



Carroll's Method for Anaerobic Cultures. This may be used 

 with culture tubes containing any of the media suitable for 

 anaerobes, with Esmarch's roll-tubes, or with fermentation tubes. 

 There is required a dry tube of the same diameter as the culture 

 tube, a short U-shaped glass tube, and two pieces of rubber tub- 

 ing all of like diameter. The culture tube having been inoculated, 

 the plug is pushed home below the lip of the tube. The ends 

 of the U-tube are smeared with vaseline and a rubber tube 

 slipped over each ; the end of the culture tube being similarly 

 treated, the free end of one of the rubber tubes is pushed over it 

 till the glass of the U-tube is in contact with the glass of the 

 culture tube. In the dry tube one or two grammes of pyrogallic 

 acid are placed and the powder is packed down with a layer of 

 filter paper. Ten or twenty cubic centimetres of a ten per cent 

 solution of sodium hydrate are then poured in and the tube is 

 quickly connected up by the rubber tubing with the other end 

 of the U-tube. In this apparatus the oxygen is absorbed by the 

 sodium pyrogallate and the conditions for anaerobic growth 

 are fulfilled. 



Cultures of Anaerobes in Liquid Media. It is necessary to 

 employ such in order to obtain the toxic products of the growth 



