BY K. (jREirx SMITH. 661 



half a minute in ether. Th(> nodules were picked out of the 

 ether, held with the forceps till the ether had evaporated, and cut 

 open with a sterile knife. The cut surface was rubbed over the 

 solidified agar in the Petri dish. After several davs' incubation 

 at 22° C many growths appeared on the plates, but in none oi 

 them could the typical orgaiiisras be obsei'ved. This is not extra- 

 ordinary, for Marshal] Ward complained that it was not so easy 

 to obtain a culture from the nodules as the description of Beijer- 

 inck would lead one to believe. 



There is a considerable difference of opinion with regard to 

 the medium best suited to the organism. Beijerinck in his later 

 papers recommended a very poor medium, and ascribed the want 

 of success that experimenters had experienced in their endeavours 

 to obtain the organism, to the employment of media rich in 

 albuminoids. Atkinson found that it grew well in ordinary meat 

 agar. Gonnermann used a plant infusion with 3 per cent, peptone. 

 Maze recommends a plant extract with 3 per cent, saccharose. 

 Beijerinck did not neutralise the natural acidity of the extract, 

 while Laurent and also Maze advised a neutral ov slightly alkaline 

 medium. 



In the plates containing the simple unneutral ised medium, no 

 colonies of the organism could be obtained, but after about a week 

 a dark coloured smudge vvas noticed on one of the plates. An 

 examination of this slight stain showed a few iri'egular forms of 

 the organism, and several tubes of dift'erent media were inoculated. 

 The only medium in which growth took place was one recom- 

 mended by Hansen for cultivating yeast. As advised by him, 

 however, it is too acid, and consequently it was neutralised.* 

 The culture in the faintly acid medium was purified by inoculating 

 a series of three liquefied ordinary nutrient gelatine tubes, and 



* The peptone -glucose medium eventually used contained : — Peptone, 

 10 grams ; glucose, 50 grams; calcium chloride (cryst. ), 5 grams; mono- 

 potassium phosphate, 2'5 grams ; tap water, 1000 c.c. Neutralise with 

 caustic potash until 10 c.c. contain an acidity equal to 0"7c.c. tenth normal 

 acid. Boil, filter and sterilise. 



