—"s ae 
80 BACTERIA IN RELATION TO PLANT DISEASES. 
Since the relation of the middle lamella to the dissolving enzym is of the greatest importance 
to bacterial infection, tissues from different sources and from plants of different ages were tested. 
Thin sections were immersed in an enzym solution from decayed potatoes and carrots. The plants 
tested were potatoes, carrots, onions, roots of yellow turnips, red beets, apples, seeds of Phaseolus 
vulgaris green and ripe, the pods of Phaseolus vulgaris both green and dried, date seeds, stem of 
Phaseolus vulgaris, Vicia faba, Cucumis sativus, Zea mays, Avenea sativa, Brassica acephala, young and 
old plants, stem and leaf stalks of Solanum tuberosum, and Lycopersicum esculentum, leaf stalks of 
Daucus carota, Petroselinum sativum and A pium graveolens. 
The results were as follows: At the end of several days’ immersion in the solution the stems of 
maize and oats, and the endosperm of date were entirely unchanged. The breaking up of the tissue 
of red beet was extraordinarily slow. At the end of 60 hours the first evidence of softening was 
observed in the thin sections and it must for the present be considered a question whethera complete 
breaking up of the tissue can occur. Of all the other plants tested, the parenchyma tissue was 
completely broken up into single cells with unchanged cell-membrane, while on the other hand 
all the suberized and woody tissues were unaffected by an immersion of long duration in the enzym 
solution. The youngest tissue was broken up most rapidly, thus sections of the green seeds of Phase- 
olus vulgaris were disintegrated in about 20 minutes; those of the ripe ones in about 2 hours. Green 
pods of the same plant were broken up much more quickly than those completely dried ones gathered 
in the late fall. Likewise the parenchyma of stems of Brassica acephala was disintegrated more 
easily in June than at the end of November. 
Experiments were then carried on to test the action of acidified juice from decayed plants 
sterilized with formalin, on the tissues of a single plant, 7. e., the carrot root. Acidification was 
made with malic, tartaric, and hydrochloric acid. The juice was mixed with the acid, left standing 
4 hours at room temperature, 18 to 20°, and then tested as to its dissolving power. Potato juice 
was always used. This titrated + 5 to phenolphthalein. 
The action of the enzym is gradually diminished by the acidity of a solution titrating+ 10, 
and it entirely ceases when the solution titrates over +20. Citric acid and hydrochloric acid have the 
same effect while malic acidis seemingly not as effective. 
The enzym, even in a very weak dilution remains active. The rapidity of a solution of the 
tissue decreases proportionally to the amount of dilution of the enzym solution. 
The power of the enzym to destroy the middle lamella is destroyed by boiling. A much lower 
temperature is also sufficient for this. Experiments show that the action of the enzym is destroyed 
if kept at 60° for any length of time. 
According to the investigations of various experimenters the middle lamellais composed of the cal- 
cium salt of pectic acid. Experiments were carried on to show whether this could serve as a nutrient 
substance for the bacteria. Calcium pectate from carrots and yellow beets to which was added vari- 
ous nitrogenous substances was inoculated with the bacteria. A nutrient solution containing 2 grams 
of KH,PO, and 1 gram of MgSO, to the liter was used. As sources of nitrogen 1 per cent asparagin, 
peptone, potassium nitrate and ammonium phosphate were added. If the medium was not neutral 
it was neutralized with soda,—temperature 28°. The flask containing the saltpetre showed no 
growth. On the contrary, in all the others there was very evident growth in 48 hours. ‘The gela- 
tinous content of the flask cleared in the upper portion to a thin liquid, which was sharply differen- 
tiated from the ever diminishing pectate layer. In 14 days the flask containing ammonium phosphate 
was slightly cloudy throughout and the contents were changed into a thin fluid with a small amount 
of precipitate. The fluid had a slightly aromatic odor, was weakly alkaline and very quickly dissolved 
into their elements sections of carrot treated with 0.2 per centformalin. The solution of the calcium 
pectates was somewhat slower in the flask containing the asparagin, and still slower in that. containing 
the peptone. The contents of the control flasks remained gelatinous. The fluid in the flask con- 
taining the ammonium salt gave the following reactions: 
ADsOlUte alcohol ociiaineaveavatec ns attncve orartsipiee Maisie ees hide arbalravele TS arene No precipitation. 
Iie UVATOCHIOLIC ACIG ners vested ee tc eee Ok arene elodeee an Lana bate dae No precipitation. 
Alcohol:-Pihydrockloriciacidy. 5 enc sis ss sic aic ogo Od ois sibs ad bea ties been No precipitation. 
Boiled with Fehling’s solution.................0 ahuiw lasabne Ne o abun DRE ee EE No reduction. 
Boiled with concentrated hydrochloric acid. ........ ...Furfurol reaction. 
Boiled with concentrated hydrochloric acid and with Fehling’s solution. . ...Very strong reduction. 
Slightly heated with dilute caustic potash lye......... 0. ccc cece cece eee e eens Yellow color. 
On evaporating, a brown horny mass remains. ‘The solution then contains a body, one of the 
pentoses—perhaps one of the hexose group—which is distinguished from calcium pectate only by 
its solubility. It is thus probable that the calcium pectate has been converted into metapectate 
by the enzym of the bacterium. A further splitting up of the metapectate into sugar does not appear 
to take place outside of the bacterial cell. Alcohol is also present, apparently. At least the solution 
