[HARRISON & BARLOW] NODULE ORGANISM OF THE LEGUMINOSAE 179 



The final examination of this plant was made on j\Iay 7th, 1906. 

 There were two mould colonies present in the flask, and an abundant 

 growth of Ps. radicicola over the surface of the agar, this film was 

 mucilaginous, drawing out in slimy threads when touched with the 

 needle. A stain made with saturated alcoholic gentian violet showed 

 that the cells were of characteristic appearance. Ko bacterial con- 

 tamination was detected by staining, but spores and mycelium of the 

 mould were seen. 



The medium was still alkaline except near the roots, where the 

 colour was largely reduced. The roots showed both general invasion 

 and nodule formation, the nodules appearing al>ove and below the swol- 

 len and infected parts of the roots. All the main roots were swollen 

 and contained Ps. radicicola in great numbers. A selected nodule 

 showed numerous short and long rods, some swollen and clavate with 

 banded protoplasm, and others with branching forms. Cultures were 

 from the agar, from a nodule and from an infected root. The plate 

 containing medium E82 inoculated from the surface growth in the 

 flask developed only numerous and typical colonies of Ps. radicicola. 

 Stained preparations from these mucilaginous colonies showed the usual 

 single polar flagellum. 



Cultures from a nodule handled in a manner already described, 

 in medium E102 produced numerous typical colonies of Ps. radicicola 

 between the fourth and tenth days. The colonies were mucilaginous 

 and drew out in slimy thread and the cells were uniflagellate. In 

 plates containing medium E82 numerous colonies developed in two days, 

 and sub-cultures in other media showed in all respects the. character- 

 istic gro\\i:h of Ps. radicicola. 



The same process was carried out in detail with material from 

 an infected root, and cultures and stain confirmed the presence of Ps. 

 radicicola in this root. 



This method of flask culture, inoculation and isolation was carried 

 out in detail with all our flasks, thus working out thoroughly the four 

 cardinal requirements as set forth by Koch in his postulates. 



Flash IX. — A control was planted September 12th, 1905, with one 

 germinating pea seed. The plant grew until May 7th, 1906, when 

 an examination was made. The medium was acid in the vicinity of 

 the roots. The growth was moderate. The flask had not been inocu- 

 lated and no nodules formed on the roots. There was no evidence at 

 any time of any contamination in the flask. On May 7th, 1906, stains 

 were made from agar in the flask, but these were negative. A piece 

 of the living root was crushed on a slide and stained but no bacteria 

 were seen. 



