Bacteriological Studies oe the Fixation of Nitrogen. 25 
from this spot as a focal center that the malady had spread so as to 
involve eight acres in an entirely new part of the orchard. On April 
18, 1911, I looked over this proposition again. There was not a shadow 
of a doubt in my mind but that the whole fifteen acres were destined 
to go sooner or later. A conservative figure of the loss up to this 
date was thirteen acres and the balance were dying. The property 
had changed hands three times since my first acquaintance with it 
and the present owner, believing that drainage would relieve the diffi¬ 
culty, expended over $4000 in putting in some 15,000 feet of drain 
tile. Some little water flows from this drain but as far as ameliorat¬ 
ing the condition of the orchard, there seems to be practically no 
change. Sample No. 30 was taken from one of the black, wet spots 
mentioned. Since these were of very recent appearance I was inter¬ 
ested in knowing whether this intense black color was necessarily an in¬ 
dication of the concentration of the nitrates. On the one hand, if that 
was true, I should expect to get only a very slight fixation of nitrogen 
in the culture; on the other hand, if it bore no immediate relation to 
excessive quantities of the salt, then I would look for larger results. 
Sample No. 31 was collected by Dr. Headden, January 5, 1911, 
from the eight acre section where the nitrates had developed very 
rapidly since December, 1909. When he handed me the material, he 
remarked that he would not be at all surprised if I got no results from 
this soil for it was as brown and mealy as could be- In culture solution 
all three of these samples gave typical brown, wrinkled surface mem¬ 
branes which a microscopic examination showed to be rich in Azoto- 
bacter. With No. 29, taken from near the single affected tree, the in¬ 
crease in nitrogen in thirty days amounted to 10.2273 m. g.; with No. 
30, from the black spot, 10.15725 m. g. and with No. 31, 7.9857 m. g. 
The result from No. 30 would seem to indicate that the black color does 
not necessarily mean extremely concentrated nitrates. (Figs. 1, 4 
and 5.) 
Sample No. 32. 
The next sample was taken October 29, 1910, along the road side 
near the fence, where the soil looked mealy but where there 
was no discoloration, which could be accounted for probably by the 
dry condition of the soil. This spot was selected because the road 
mentioned ran alongside a young orchard where the trees had been 
dying from some unknown cause. There. was such a combination 
of factors at work in the orchard, namely, neglect, drought and 
possibly nitre that it was hardly safe to venture an opinion on the 
cause of the death of the trees. An alfalfa field adjoining this tract 
showed many barren spots which were suggestive at least. In culture, 
this soil developed a heavy, tough, brownish membrane, rich in Azoto- 
bacter and after thirty days gave an increase of 15.411 m. g. of nitro- 
