ROOT-NODULES OF LEGUMINOSAE. I3I 
experiments on high moorland at Bremen. In this case the strikingly large nodules formed on the 
yellow lupin from spontaneous infection were completely inactive, because of the poor degree of 
virulence [word here used with wider meaning] of the invading bacteria. 
Experiments with soy-beans in pots confirmed this result and also explains how this increase in 
virulence can be brought about. Pots of Dahlem earth and sand, half of which were inoculated with 
Japanese soy-bean earth, were planted with soy-beans. Only late in the season did the difference 
appear between the inoculated and uninoculated plants. he greatest difference, however, was 
observable at maturity. The leaves of the uninoculated plants were then almost white, having lost 
almost completely their xanthophyll and their roots were free from nodules while the leaves of the 
inoculated plants were deep yellow and their lateral roots full of nodules as large as peas. The amount 
of dry substance from the inoculated plants was 33 per cent greater, and of seed about 74 per cent 
greater than that from the uninoculated plants. The next year plants in these same pots of soil with- 
out further inoculation showed an even greater difference. Those in the uninoculated soil remained 
completely nodule-free while the others showed extraordinarily strong nodule formation, so that every 
root on the upper third was closely covered with nodules the size of peas. Moreover the activity of 
the nodules the second year was remarkably increased. Even if it is very probable that the increased 
number of bacteria in the soil plays a réle here, and that because of the great lack of nitrogen in 
the soil the nodules became active earlier and hence had a longer season in which to work, nevertheless 
the most important factor must be sought in the increased virulence, for while the total amount of 
nodule formation was considerably greater the second year it was not so much so as to go parallel 
with the increase in activity. Increased virulence here, as in organisms pathogenic to animals, is 
due to repeated passage through the organism in question. The dry substance of the uninoculated 
_ the first year being taken as 100, we have the following yields: 
Year. | Uninoculated. Inoculated. 
1900 | 100 | 131 
1901 66.4 | 1458 
Hiltner took this into consideration in obtaining virulent cultures for inoculation for all import- 
ant species of legumes, and with few exceptions his cultures were passed several times through the 
appropriate host-plant. The better results of 1901 and 1902 in the open field he thinks is largely 
referable to this fact. 
The chain of evidence for the greater activity of virulent bacteria was completed by making 
simultaneous inoculation experiments with bacteria of differing virulence, mostly on peas. The 
results entirely confirmed the preceding observations. 
The effect of difference in virulence is also shown in the case where soil from Zehringer in Anhalt 
in which peas were known to form active nodules only on the deeper roots was used for inoculation 
in comparison with a third generation of pure cultures. The result was that the bacteria from the 
pure cultures penetrated the roots at once and formed numerous nodules on all the upper parts of 
the roots while those from the Zehringer earth produced nodules only on the deeper side roots and 
sparingly at that, 7. e., as in the fields. Only virulence can explain why the bacteria in the Zehringer 
soil did not penetrate the roots at once. Hiltner thinks that a moderate degree of virulence on the 
part of the bacteria is sufficient to cause leguminous plants to attain their maximum size. He thinks 
it also unquestionable that virulence may be increased in other ways than by repeated passage 
through the plant, 7. e., as Remy showed by feeding the inoculated plant with saltpeter. 
It is an important question for agriculture whether one can not obtain too great a degree of 
virulence. It has been shown that at first the bacteria act toward the plant as real parasites. If 
this is correct, as appears on more grounds than one, we have to reckon with the possibility that the 
fight will be the more severe for the plant as the bacteria become more virulent. Indeed Nobbe and 
Hiltner record an experiment in which highly virulent pea bacteria caused direct injury to the plants 
growing in pots of nitrogen-free sand. In this case the bacteria penetrated the roots in such numbers 
that almost every root-hair was infected, and everywhere numerous nodules appeared in which, 
however, bacteroid formation was suppressed owing to the excessive growth of the bacteria. Asa 
result no nitrogen assimilation occurred and the aerial parts of the plants were plainly weakened. 
Another experiment in 1902 at Dahlem on pot plants with especially virulent bacteria resulted in a 
decrease in the crop: Inoculated plants gave a dry substance amounting to 34.1 g. per pot.* while 
uninoculated plants gave 47.1 g. 
*Possibly in some of these cases Bact. tumefaciens may have been the cause of the nodules. See Crown-gall: Its 
cause andremedy. Bull. 213, B. P. Ind., U.S. Dept. Agric., 1911, plate xvI, 2a. 
