148 BACTERIA IN RELATION TO PLANT DISEASES. 



This view is favored by the fact that a very considerable time is required to bring about this 

 change (56 to 82 hours in case of fibrin) and also by the fact that gelatin, the most easily digested 

 substance is known to be readily liquefied by a great number of micro-organisms. 



In all, Tischutkin made over 150 experiments. 



In 1892 Tischutkin published a second paper on carnivorous plants in which he 

 attempted to show that micro-organisms are a constant feature in the fluid excreted by 

 such plants, and that these organisms are able to dissolve albumen. 



For his experiments, Pinguecula vulgaris, Drosera longifolia, D. rotundijolia, Dionaea 

 muscipula, and Nepenthes mastersiana were used. The following is a synopsis of his paper. 



Microscopic examination of the excreted sap 24 and 48 hours after stimulation, and also 3 to 5 

 days later, revealed the presence of bacteria and moulds. Myriads of bacteria were found in the 

 extruded fluid of Drosera and Dionaea in 20 to 24 hours. A drop of the sap, 24 hours after stimula- 

 tion of the glands, was placed by means of a flamed platinum loop, in sterile flesh-peptone-gelatin and 

 after 3 dilutions in such gelatin Petri-dish poured plates were made from each of the 4 tubes. When 

 colonies had developed, small portions of them were removed with a sterile platinum wire, placed in 

 similar gelatin and kept at room temperature. After cultivation for 9 or 10 generations in this 

 medium, test tubes of weakly acid, pepton-free, flesh bouillon and of weakly acid sterile water were 

 inoculated from the cultures and a piece of albumen was added. The solution of albumen was rapid. 

 Several species of such bacteria were found, one or more kinds on each insectivorous plant, 4 sorts on 

 Pinguicula. 



The sap from unopened but fully developed pitchers of Nepenthes was unable to dissolve albumen. 

 A piece of albumen was inserted into two such pitchers and the opening carefully closed with animal 

 plaster. One of these was N. distillatoria, the other N. hirsuta. In 4 days the pitchers opened. At 

 this time the sap contained no pepton and extremely few bacteria. The albumen was unmodified. 

 In Rothert's review of Tischutkin's Russian paper it is also stated that the sap had a strongly acid 

 reaction. This same sap when poured into test tubes and kept at 20 to 21 , dissolved the albumen 

 in 4 to 5 days but then the bacteria had multiplied enormously. 



From these results he draws the conclusion that the solution of albumen is due, not to the 

 secretion of the plant, but to the action of peptonizing bacteria living in it, and beginning only when 

 these organisms have become very numerous. They are not present in the unopened pitchers. 



The dry residue left by evaporation of pitcher liquid, according to Volker's analysis, varies from 

 0.92 per cent to 0.85 per cent, and is composed of: malic and a little citric acids, 38.61; potassium 

 chloride, 50.42; carbonate of soda, 6.36; lime, 2.59; magnesia, 2.59; organic material, a trace. 



The action of the extruded fluids has nothing in common with animal digestive fluids. We may 

 regard the relation of the bacteria to these plants as a symbiosis in which the higher plant supplies 

 the food for the bacteria and takes in return soluble animal products digested by the latter. 



In 1897 Vines published a paper on the proteolytic enzym of Nepenthes, in which he endeavored 

 to confirm his earlier statements and to refute the views of Dubois and Tischutkin. 



In his experiments he used, chiefly, as material for digestion, well-washed blood-fibrin which had 

 been preserved in a mixture of one part pure glycerin and two parts distilled water. He says that 

 such fibrin may be regarded as free from bacteria in view of the antiseptic action of glycerin (Copeman 

 and Blaxall, Br. Asso. Rep., 1896). He never failed to obtain digestion of fibrin by the pitcher liquid 

 when an adequate amount of acid (0.25 per cent solution of hydrochloric) was present. In control 

 tubes of dilute hydrochloric acid and fibrin, the latter was softened in the time required for digestion 

 with pitcher liquid, but never digested. 



Experiments were made to determine whether this digestion would go on without the presence 

 of bacteria. 



"Three test tubes were prepared as follows: Each tube contained some pitcher liquid acidified 

 with 0.25 per cent hydrochloric acid, and a shred of fibrin. To ( 1 ) was added some potassium cyanide ; 

 to (2) some thymol; to (3) a few drops of chloroform.* At 1 p. M. the tubes were placed in the incu- 

 bator at 35 C. The fibrin in tube (1) was completely dissolved by 5 p. m. ; that in the other two tubes 

 was broken up by that time and was found to have entirely dissolved by the following morning at 



9 A. M." 



Similar experiments using egg albumen and others using fibrin and corrosive sublimate gave 

 satisfactory results (10 cc. pitcher liquid, 5 cc. 0.25 per cent hydrochloric acid, 5 cc. 1 per cent mercuric 



"Thymol and chloroform, it should be remembered do not restrain all bacteria, nor is potassium cyanide generally 

 reckoned as a strong germicide. 



