EXPERIMENTAL PRODUCTION OF PARASITES. 45 



The pulp of an infected potato was mixed with water and filtered through the Chamberland 

 bougie. The reaction was distinctly alkaline. After neutralization with HC1, it was divided into 

 12 parts, 2 of which were used as checks. To the others was added 0.5 per cent of one of the follow- 

 ing acids : formic, acetic, tartaric, or lactic, or an equal quantity of soda, and a drop of essence of 

 mustard to prevent the growth of microbes. Pieces of potato were immersed in these. After 12 

 hours the cells of the checks were disassociated to a depth of 2 to 3 mm. and their protoplasm con- 

 tracted. The use of o. 5 per cent tartaric acid and 0.5 per cent and 1 per cent soda gave the same 

 result. In the case of 1 per cent tartaric acid, 0.5 per cent lactic acid, 0.5 per cent and 1 per cent 

 acetic acid, the disintegration was feeble, and in other cases the tissues were not attacked. Turnips 

 were affected more rapidly and completely. The reaction given by the pulp and the liquid when 

 diluted in water varies with the nature of the plant attacked radish and turnip pulp gave an acid 

 reaction, and rotted onion was still more acid. 



Active soluble substances are secreted by the microbe in cultures in organic solutions as well 

 as in the tubers. 



Exposure to a temperature of 62 for 5 minutes or to sunlight for 8 hours destroyed the solvent 

 activity of the diluted pulp of infected potatoes. 



The existence of a variety of cytase was established. Flocculent alcoholic precipitates, dis- 

 solved in distilled water, and in the presence of essence of mustard caused the characteristic soften- 

 ing of the tissues of potatoes which were kept in it 12 hours. It acts on the potato in an alkaline 

 but not in an acid medium. 



The substances causing the death of the protoplasm were not determined by Laurent. The 

 alkaline secretions which kill the protoplasm are more resistant to heat than the cytase, but are 

 destroyed at ioo. Even after the destruction of these substances the juice is able to diminish the 

 resistance of the most rebellious varieties of potatoes. This indicates that toxic substances are still 

 present. 



Laurent states that the second bacillus used in his experiments is a form of Bacillus colt. It 

 does not liquefy gelatin, and emits gas bubbles when inoculated by pricking into must of beer gelatin. 

 It develops better in the presence of oxygen than in vacuo, but is anaerobic to a certain extent. 

 Deprived of air, it reduces the nitrates with great rapidity. Several races of this organism were 

 isolated by the author. The essential characteristics of the bacillus of all these races are not modified 

 in any lasting manner by cultivation in bouillon, on cooked slices of potato, on must of beer or 

 bouillon gelatin or agar. All developed in mineral solutions containing saccharose, lactose, glucose, 

 mannite, glycerin, potassium succinate, potassium lactate, potassium citrate, ammonium bimalate, 

 sodium butyrate, sodium hippurate, asparagin and peptone, but not in those containing potassium 

 tartrate, ammonium tartrate, potassium acetate, and sodium formate. 



Inoculations made with authentic cultures of B. coli (obtained from Calmette, Malvoz, and 

 Van Ermengem) and with related forms, gave infections on potato tubers treated with 1 per cent 

 soda solution. All afterwards became parasites it is said on untreated tubers. Experiments were 

 also made with the following bacilli: Typhoid bacillus from Gand, Liege, and Lille, Gaertner's 

 Bacillus cuteritidis, Moorzeele and van Ermengem's bacillus from the meat of calf, Friedlander's 

 bacillus, Lambert and van Ermengem's bacillus from liver. 



All these strongly resembled B. coli when cultivated in gelatin and on cooked potato, but 

 showed different chemical capacities. Bacillus fluoresceins putidus, B. fluorescein liquefaciens and 

 a bacillus forming yellow colonies, isolated from rotten tomatoes were also used. 



All of these when inoculated into normal potatoes gave negative results. When tubers treated 

 with soda solution were used, however, all attacked the tissues, and from that time were able to 

 live as real parasites on several varieties of tubers. The typhoid bacillus was more virulent than the 

 strains of B. coli. A second passage of this organism through potato produced a pulpy layer 10 to 

 12 mm. thick after 24 hours in the thermostat at 35 . When again cultivated in gelatin, the various 

 organisms had the same characteristics as in the original cultures. 



In the course of these experiments Laurent observed a gummy disease of the tubers of 

 Cattleya mossiae (orchid). He isolated from these in cultures on must of beer gelatin a short bacillus 

 which became parasitic on potatoes, and which he identifies as a form of B. coli. The gummy dis- 

 ease he attributes to excess of nitrogenous fertilizers. The signs in the orchid are the softening of 

 the tissues of the tubercles which become deep brown and then black. 



Laurent also isolated a bacillus supposed to be the cause of black-rot in tomato fruits. This 

 differed much from B. coli. It did not grow in mineral solutions. On potato it formed golden yellow 

 colonies, and it did not liquefy bouillon gelatin. 



In another case, B.fluoresccns liquefaciens was isolated from badly rotted tomato plants, which, 

 according to the grower, had been cultivated on ground dosed with enormous quantities of manure 

 and liquid fertilizers. 



