BY R. GREI« SMITH. 



66a 



The earlier cultures in agar media made from the unneutralised 

 infusion were not at all successful, a circumstance due partly to 

 the acidity and partly to the agar surface which was very soft 

 owing to the action of the acid wliich, us sterilisation proceeded, 

 made the medium less and less gelatinous. This was obviated by 

 neutralising the medium immediately after the agar or gelatine 

 was dissolved. Potassium hydrate suggests itself as the best 

 alkali to use in neutralising a plant extract, especially when one 

 remembers how much the Leguminosae are benefited Ijy potash 

 salts. In some of the cultures, as for example lupin-agar, with 1 

 per cent, potassium chloride, it seemed as if the salt had stimu- 

 lated the grow*-h of the organism. According to Maze, sodium 

 chloride acts as a poison towards the nodule bactex'iimi paralysing 

 its development. A plate seeded with the organism and dotted 

 with solutions of varit)us salts showed the greatest amount of 

 growth between a potassium phosphate and a calcium chloride 

 manuring. This suggested a means of clarifying the various 

 plant-extract media which are alwa^'s more or less turbid from 

 the gradual precipitation of organic matter. When the agar or 

 gelatine is dissolved in the plant extract 5 c.c. each of a 10 per 

 cent, solution of monopotassium phosphate and of a 20 per cent, 

 solution of crystallised calcium chloride are added to every 100 c.c. 

 of the hot gelatine oi' agar medium, which is then neutralised 

 with 10 per cent, potassium hydrate to faint acidity. Ten c.c. of 

 the solution are pipetted out and neutralised with tenth normal 

 potash, using phenolphthalein as an indicator, and normal potash 

 is added to the bulk of the medium in projiortion to make every 

 100 c.c. possess an acidity equal to 0'7 c.c. of normal acifl. This 

 acidity is equal to 0'05 per cent, tartaric acid. 



The organism is a strong aerobe, and grows most freely when 

 started upon the surface of a medium. It does not grow under 

 anaerobic conditions in peptone-glucose fluid, a medium which 

 seems best suited to its needs. Laurent maintained that it could 

 grow anaerobically, while Maze, denying this, assumed that 

 oxygen had not been thoroughly eliminated from Laurent's culture 

 media. 



