68 METHODS OF CULTIVATION OF BACTERIA 



is advisable to practise a massive inoculation by pouring into 

 medium part of an actively growing bouillon culture.) The flask 

 is then connected with the hydrogen apparatus by means of a short 

 piece of sterile indiarubber tubing, and hydrogen is passed 

 through for half an hour. In the case of flask (1), the lateral 

 nozzle is plugged with melted paraffin and covered with alternate 

 layers of cotton wool and paraffin, the whole being tightly 

 bound on with string. The entrance tube is now completely 

 drawn off in the flame before being disconnected from the 

 hydrogen apparatus. In the case of flask (2), first the exit 

 tube and then the entrance tube are sealed off in the flame 

 before the flask is disconnected from the hydrogen apparatus. 



It is well in the case of both 



to run some melted 

 paraffin all over the rubber 

 stopper. Sometimes much gas 

 is evolved by anaerobes, and 

 in dealing with an organism 

 where this will occur, provi- 

 sion must be made for its 

 escape. This is conveniently 

 done by leading down the exit 

 tube, and letting the end just 

 dip into a trough of mercury 

 (Fig. 23), or into mercury in 

 a little bottle tied on to the 

 Fig. 23.— Flask arranged for culture of en( i f the exit tube. The 

 anaerobes which develop gas. pressure of gas ^thin causes 



6isatrou Si?tSb e e r S intOWhi0h an escape at the mercury 



contact, which at the same 

 time acts as an efficient valve. The method of culture in fluid 

 media is used to obtain the soluble products of such anaerobes 

 as the tetanus bacillus. 



The Method of Tarozzi. — This observer has found that if 

 small pieces of fresh sterile organs are added to ordinary bouillon, 

 growth of anaerobes takes place under ordinary atmospheric con- 

 ditions. For this purpose, portions of liver, spleen, or kidney 

 are most suitable. The method has been used in the cultivation 

 of spirochsetes, organism of poliomyelitis, etc. It has been 

 shown, by Douglas, Fleming, and Colebrook that the addition 

 of pieces of vegetable (even after being boiled), bran, and even 

 asbestos wool, is effective in making a fluid medium suitable for 

 the growth of anaerobes, the presence of fine interstices in the 

 material being an important factor in aiding the growth. ' It 



