136 The Philippine Journal of Science 1919 
were always present, inoculation experiments were conducted 
with them in order to test their virulence. . 
The inoculations were made chiefly with seedling coconuts 
from 60 to 180 centimeters in height. They were prepared by 
stripping off the outermost leaves. Then the outside of the 
portion to be inoculated was washed with mercuric chloride, 1 
to 1,000, and after ten minutes was rinsed with sterilized water. 
With a sterile scalpel a stab was made into the growing point, 
and a pure culture of the bacterium introduced with a platinum 
needle. The injured portion was then covered with melted 
paraffin. Controls were prepared in the same manner except that 
‘sterile water was used in place of the bacteria. About three 
hundred inoculations have been made in this manner, and in a 
number of instances cases of bud-rot were produced (Plate II, 
figs. 5 and 6). Employing this method, saprophytic bacteria 
were eliminated, because of their failure to produce disease. 
After the elimination of these saprophytic forms, there remained 
one distinct kind of bacterium that would produce disease under 
certain favorable conditions, such as severe injury of the grow- 
ing point, and excessive dampness. Inoculations were not re- 
peatedly successful, but when the exact requirements were met 
a number of positive infections could be obtained. Bacteriolo- 
gical tests have shown that the organism thus isolated and used 
for inoculations was similar to Bacillus coli (Escherich) Mig., 
and apparently is identical with that considered by Johnston(7) 
to be the cause of coconut bud rot. 
Authenticated cultures of Bacillus coli (Escherich) Mig. were 
obtained from Dr. L. R. Jones, of the University of Wisconsin, 
and also from Dr. O. Schébl, of the Philippine Bureau of Science. 
The cultures from the United States were isolated from man, 
and those secured in the Philippines, from man and _ horse. 
Under extremely favorable conditions a bud rot could be pro- 
duced with each of these cultures. The infections produced 
from the first inoculations were very slight; but the organism, 
after it was reisolated and then used for reinoculation, appeared 
to produce a more rapid and severe case of rot. This may 
indicate that the bacteria increase somewhat in virulence after 
passing through a weakened tissue. 
Cytological studies also have shown the presence of bacteria 
in the tissues of the coconut. Sections of a typical case were 
made from parts of the young leaves near the growing point, 
from portions of the growing point, and from the woody tissue. 
These sections show that the organism is present not only in 
the parenchymatous tissue, but also in the vascular system. 
