ROOT-NODULES OF LEGUMINOSAE. 127, 
Frank in his first paper on the dimorphism of the pea nodules stated that the albuminous nodules 
contain 6.936 per cent nitrogen while those containing the ‘‘amylodextrin’’ only 4.828 per cent 
nitrogen in the dry substance. 
Hiltner denies the existence of a dimorphism of pea nodules in the sense used by Frank, but says 
we might speak of a dimorphism of the bacteroids. He also takes issue with Moller that the end of 
the bacteroids in all nodules is fatty degeneration. Specimens examined by him showed plainly all 
degrees of transition between the two sorts of nodules described by Frank. The contents of normal 
pea nodules flow out readily when the nodule is cut, and are easily mixed with water. On the contrary 
the nodules in question which are distinguished outwardly by a somewhat darker color show in 
section a chalky consistency; even from cut cells the contents do not flow out into the water, but 
adhere so firmly together that it is difficult to separate the single elements from each other. Normal 
nodules, even when in decay, show no sign of this chalky consistency. The cells of the bacteroid 
tissue in these chalky nodules are filled with large starch-like grains which, however, stain red-brown 
with iodine instead of blue. An examination of these chalky nodules showed that almost without 
exception the septate mycelium of a fungus was present in the nodule or on that part of the root from 
which the nodule grew. The subsequent investigations at Dahlem verified the assumption that the 
aggregation of a fatty substance was to be referred to this fungus. Hiltner verified Frank’s statement 
that these abnormal nodules are likely to occur on land where the pea has been cultivated several 
years. This may, he says, be due to early exposure of the roots to soil fungi, or to the lack of a 
physiologically important nutrient substance. 
At Dahlem, in soil used for the first time for pea culture, often after 6 to 8 weeks’ growth, in a 
portion of the plants, the upper leaves of pea vines formerly green and healthy took on a pale color 
and a cessation of growth occurred. In another such field sickly plants appeared among the healthy 
ones. Investigation showed that healthy plants possessed normal nodules while great numbers of 
abnormal nodules were found on the roots of the sickly plants, so that one could tell by the appear- 
ance of the plants before uprooting them, whether the roots bore normal or abnormal nodules. From 
these facts it is easy to explain why abnormal nodules form on the pea, for the pea is known to 
experience Bodenmiidigkeit very rapidly and even to fail on some soils in its first year of cultivation 
after passing successfully the earlier stages of growth. Hiltner considers this a valuable addition to 
our knowledge of the causes of soil sickness. 
A close study of the waxy substances of bacteroids from artificial cultures, which substance is 
similar to that in the abnormal nodules, led Hiltner to agree with Moller in regard to its solubility 
but to disagree with Frank in that he found it unchanged by concentrated sulphuric acid. It can be 
shaken out of nodules or old cultures by means of chloroform and has rather the consistency of gutta- 
percha than of wax. ‘The red-brown color caused by iodine is not a reaction of the waxy substance 
but of a soluble substance present with it. When sections of waxy nodules were left in water for 2 
days the granules stained pure yellow instead of red-brown, without having otherwise changed in 
appearance. From his experiments Hiltner found no reason why the red-brown color should not be 
considered as a glykogen reaction. 
A qualitative analysis of this waxy substance from abnormal nodules showed that it was abso- 
lutely nitrogen-free. This was at first a disappointing discovery since Hiltner had considered that the 
waxy substance, so constant in its appearance in artificial cultures and in abnormal nodules repre- 
sented an accumulation of the products of nitrogen assimilation which the plants had lost the power 
to absorb. To test this point further, cultures of nodule bacteria from pea, soy-bean and Robinia 
were allowed to grow for 4 months in seven different nutrient solutions and then tested for increase 
in nitrogen. Though in suitable solutions the large outgrowths containing waxy inclusions had been 
formed, in no case was there a trace of nitrogen increase. These solutions contained 1 per cent grape- 
sugar, 0.02 per cent monopotassium phosphate and variable amounts of nitrogen in the form of 
peptone, asparagin and saltpeter—in all 50 experiments. 
It is further interesting to note that while, ordinarily, nodule bacteria stain very little or not at 
all by Gram’s method, the waxy outgrowths or germ-plasma of the bacteroids, arising in nutrient 
solutions containing carbon compounds, stain intensively by Gram soon after their differentiation. 
As this stage passes, the ability to retain the stain is gradually lost. The substance which holds this 
stain evidently passes largely into the bacterial slime for this stains readily while the almost colorless 
bacteroids show then only a few deeply colored granules which somtimes stick to their exterior. 
The following observations were made on the soy-bean, grown in pots for several years. 
In this plant the hunger stage lasts uncommonly long. ‘The plants suffer from it even after 
numerous large nodules have formed on the roots. On plants showing nitrogen hunger for about 
8 days, the bacteriods of these inactive nodules were small and only beginning to show plasma 
differentiation. In the nodules of other plants which the authors knew would in a few days show 
greening from the beginning of nitrogen assimilation, all the bacteroids possessed large, roundish 
