186 



the bacteria I had isolated in Marcli diffeieiit from tliose grown in 

 October, whilst the tubercles came from plants growing side by 

 side and being in ttie same state of development. With the yellow 

 lupine and serradella I iiad similar results. In most other cases, 

 however, for example with Pisum, Latkyrus, Vicia, and Irifolium^ 

 the similarity of the various mutually independently isolated stocks 

 is so complete and the image of B. radicicola can so distinctlj^ be 

 recognised, that the above observation requires nearer confirmation. 

 But we cannot now enter upon this point. 



When trying to isolate B. radicicola from materials other than 

 the nodules, for example from the soil and from the dying surface 

 cell-layers of the root, it proves very difficult to recognise this 

 species amid the numerous other saprophytes, especially when the 

 number of the germs of the different species is to be determined 

 quantitatively. B. jluorescens liquefaciens causes muc'h trouble by 

 the liquefying of the gelatin plates, and yet it is necessary to use 

 these plates as on them the colonies of ail the species lie free from 

 one anotiier, while on agar they are overgrown and rendered 

 unrecognisable by B. jiuorescens, which extends strongly sideways. 



Concerning the question if only one or more species of tubercle 

 bacteria exist the following. 



Already in 1892 experiments thereabout were made by the late 

 Hellriegel ^) in the experimental station at Bernburg with pure cul- 

 tures of the bacteria made b}' myself at Delft. Of his results Hkll- 

 RiKGET. sent me two reports. In the first, dated 24 July 1892, he 

 gives as "Augenblicl<.licl)es Hauptresultat : "Es gelingt mit den 

 Reinkulturen von B. radicicola var. Pisi oder von Vicia /aba, die 

 Erbsen und Bohnen, und mit deneji des Bac. radio, var. Lupin, oder 

 Ornithopodis Lupinen und Serradella erfolgreich zu infiziren und zum 

 Wachstum resp. der Assimilation des freien Stickstoffs zu bringen, 

 und das ist was unsere anfangliche Behauptung bestiitigt". Already 

 earlier Hetj-riegel held arrived at the conclusion that the bacteria 

 of Lupiiius and Ornithopus belong to a species different from tliat 

 of Pisum. and Vicia, which was also my own o [union. 



In later years many interesting experiments were made in this 

 direction, especially by Hiltiser. Yet the evidence is unsatisfac- 

 tory as it proved hitherto impossible in the sand cultures ') 

 to bi'ing Leguminosae to complete development by infection with 



1) He died 24 September 1895 of a stomach disease and was already suffering 

 when I visited him at Bernburg in 1892. 



2) It is a well-known fact that the Papilionaceae, when cultivated in liquids, do 

 not fix the atmospheric nitrogen indifferently whether they produce tubercles or not. 



