BY R. GREIG SMITH. 155 



cholerae, kc. In fact, as I pointed out in the paper already- 

 quoted, "Extract of lupins or other leguminous plant does not 

 seem a necessity for the culture media. Grass will do quite as 

 well, and for that matter the plant extract might be left out 

 entirely.'' Fairly luxuriant cultures were obtained upon a 

 gelatine medium containing glucose and inorganic salts, and 

 since that time I have cultivated the organism upon a medium 

 containing faintly acid agar (2 %), glucose (2%) and inorganic 

 salts (CaCL. and KH^ PO^) nearly neutralised with KOH."^ 

 On the latter medium there is no nitrogen except that which may 

 be present as impurity in the washed agar, the glucose or the tap 

 water. I have also grown the organism in an agar-free fluid 

 medium prepared exactly as the agar medium. Such a fluid 

 after inoculation becomes turbid and forms a slight sediment of 

 organisms together with a bulky zoogloea cloud or sedimentary 

 film. 



The apparent growth in this very poor medium led to the belief 

 that fixation of nitrogen might have occurred, but this was dis- 

 pelled when the experimental flasks holding 250 c.c. of culture 

 fluid were found to contain exactly the same amount of nitrogen 

 (0 6 milligrm.) as was contained in control flasks. 



* The method of preparing the faintly acid medium is described in these 

 Proceedings, 1899, Part 4, p. 663. 



