July 2i, 1923 
Crowngall Organism and Its Host Tissue 
129 
absolute alcohol the sections were cleaned and stained by a saturated 
solution of light green in clove oil, rinsed in xylol, and mounted in 
balsam. 
Slides made in this manner showed the walls acted on by the bacteria 
and the xylem walls to be stained red, while the rest of the tissue appeared 
green. The bacteria for the most part appeared red along with the 
bordering tissue (Pi. 3, C, D; 4, C, D; and 5, A-D). The crystals of 
calcium oxalate remained uncolored. The previously mentioned gran¬ 
ules of pectic substance were never observed in stained preparations. 
The bacteria found in these sections are present in larger numbers 
than was previously supposed. Repeated statements have appeared 
in the literature that the organisms were scarce ( 12 , p. 193; u, p • 18). 
These appeared to be confirmed by the writer's earlier isolations. In 
view of the comparisons of microscopic and plate counts made from soil 
by Conn (1, p. jo), it seemed logical to expect a poured plate to show the 
presence of no more than one-tenth of the number of bacteria that might 
be distinguished under the microscope. But even with this discount 
the results from earlier platings showed a comparatively small number 
of colonies. 
The chance examination of a io-day-old isolation plate revealed an 
interesting phenomenon. This plate had been poured from gall tissue 
which had been treated with mercuric chlorid and washed and crushed 
before mixing with the agar in a manner similar to that described by 
Smith ( i2, p. 22). A bacterium-free zone surrounded the portions of 
tissue for a radius of more than a centimeter, while outside of this region 
a very large number of colonies had appeared. These were proved by 
successful inoculation into tomato to be the gall organisms. It seemed 
quite clear that the treated tissue had an inhibitory influence on the 
bacteria, due either to the diffusion of the disinfectant used or to some 
detrimental product of its own. 
To determine whether or not a short treatment of the tissue with the 
mercuric chlorid was inhibiting to the growth of the bacteria, three 
plates were poured from a suspension of a pure culture of the crowngall 
organism at a dilution which would produce about 1,000 colonies in 
each plate. Small pieces of normal tomato tissue which had been 
dissected out under aseptic conditions and treated with mercuric 
chlorid were placed in the first and second plates. In the first 
the tissue was placed 45 seconds in mercuric chlorid 1 to 1,000, as 
Smith ( 12, p. 24) describes and washed a minute and a half in sterile 
water, while in the second the treatment with the disinfectant lasted 3 
seconds and the washing 10 seconds, as recommended later by Smith 
(jo, p. 434). The tissue in the third plate was not treated with the 
disinfectant, but was placed in sterile distilled water for 30 seconds. 
After five days the portions of tissue which had been treated with mer¬ 
curic chlorid for either period had bacterium-free zones around them 
for a radius of about 1 pi cm. Outside of these the bacteria appeared in 
great abundance. The other fragment, which had been treated only 
with water, appeared to have exerted no inhibitory effect on the 
organisms. This experiment was repeated twice with the variation that 
gall tissue was used as well as normal tissue. The results were con¬ 
firmatory in every respect. 
A few contaminating organisms were secured in the plates with the 
gall tissue. Some of these were able to tolerate the inhibiting influence. 
