240 
Journal o f Agricultural Research 
Vol. XXIX, No. 5 
chamber. The seeds were treated 
with formaldehyde by the presoak 
method 9 described by Braun (1) and 
put into the damp chamber November 
24, 1920. Two days later, having 
germinated, they were inoculated by 
spraying with a water suspension of a 
2-day old culture of Bact. phaseoli EFS. 
The following day they were sprayed 
again. No leaves were yet visible. 
On the third day after inoculation 
some of the leaves were beginning to 
appear and the seedlings were trans¬ 
ferred to large moist chambers. 
On November 30 there were no signs 
of infection and five of the seedlings 
(Nos. 16 to 20) were transferred to 
Sachs' nutrient solution and placed 
under a bell jar in a south window in 
the laboratory. On December 2 they 
were reinoculated by spraying. The 
leaves were just beginning to be visible 
between the cotyledons in some cases; 
in others they were beginning to push 
out. 
Eight days after the first spraying 
in the Petri dish the cotyledons of 
many of the seedlings remaining in the 
damp chamber had water-soaked spots 
and bacterial ooze therefrom (PI. 7, 
N, O). In some cases the seed coat 
was still firmly attached and the spots 
were visible through it. Seven of 
these seedlings with infected cotyle¬ 
dons (Nos. 45-51) were transferred to 
Sachs' solution and placed under a 
bell jar in a north window in the labora¬ 
tory. They were sprayed heavily 
with sterile H 2 0 in an attempt to 
spread the infection by scattering the 
bacteria oozing from the spots. 
On December 9 seedlings Nos. 16 to 
20, reinoculated December 2 (two days 
after their transfer to Sachs' solution), 
showed a pronounced browning and 
curling of the leaf tips or margins (like 
PI. 7, P). Five days later Bad. 
phaseoli EFS. was isolated from the 
browned curled margin of a leaf of 
No. 20 in which microscopic examina¬ 
tion showed the bacteria present in 
abundance. 
Also on December 9 seedlings Nos. 
45 to 51, transferred to Sachs' solution 
on December 4 but not reinoculated, 
were all very much stunted. The 
main shoot was shriveling in Nos. 48 to 
51 and there were irregular brown 
spots on the leaves of the other three 
(Nos. 45 to 47). Bad. phaseoli. EFS. 
was reisolated December 11 from dark 
brown spots on the cotyledons of plant 
No. 46. On December 31, all the 
plants transferred to Sachs' solution 
were dead, doubtless partly because of 
the abnormal conditions under which 
they had been grown. 
Similar results were obtained on 
Ito San soybean seed treated with 
sulphuric acid 10 and inoculated by 
spraying in a Petri dish damp chamber 
immediately after germination therein. 
In a third experiment presoaked soy¬ 
bean seeds of the Ito San variety, 
treated with formaldehyde, were ger¬ 
minated in a damp chamber and trans¬ 
ferred as soon as the seedlings had made 
sufficient growth to Sachs' solution 
and inoculated with Bad. phaseoli 
EFS. 8 days after the sowing of the 
seed in the damp chamber. In some 
cases the leaves had not begun to 
push out, but »the cotyledons were 
spreading and the culture was sprayed 
between them upon the folded leaves 
at 2.15 p. m. They were placed under 
a low bell jar in a south window of the 
laboratory; at 4.15 they were sprayed 
again and twice the next day, at 10.20 
a. m. and 4.30 p. m. The moisture held 
well for 48 hours. The low bell jar was 
then replaced by a taller one and this 
was removed 2 days later. A week after 
the inoculation there was a curling 
and yellowing or browning of the 
margin of many of the first leaves 
(like PI. 7, P), and Bad. phaseoli 
EFS. was isolated from two of them. 
No infection of cotyledons was ob¬ 
served. 
Ito San soybean seedlings grown 
like the preceding but taken to the 
greenhouse, after their transfer to 
Sachs' solution, and there inoculated 
eight days after the seed was sown in 
the damp chamber, became similarly 
infected. One of these photographed 
eight days after inoculation, is shown 
in Plate 7, P. 
These experiments were repeated 
several times and always resulted in 
infection. Bad. phaseoli EFS. was 
repeatedly isolated both from the 
cotyledons and the brown curled leaf 
margins. No pustules were ever pro¬ 
duced. 
Another attempt was made on De¬ 
cember 10, 1920, to produce infection 
with Bacterium phaseoli EFS. on young 
seedlings of Ito San and Mammoth 
Yellow soybeans growing in pots in the 
greenhouse, but the only positive results 
were obtained on 11 Ito San seedlings, 
the leaves of which had been rubbed 
9 H 2 O for 10 minutes in bags of double thickness surgeon’s gauze; moist chamber (still in bags), 
1:300 formaldehyde for 15 minutes; moist chamber (wet with 1:300 formaldehyde), 45 minutes; spi 
hours; 
spread out 
^lo^SO^Slmcentrated) for 1 minute; H 2 S0 4 poured off and sterile water added; sterile water added and 
poured off several times. 
