186 THE MICHIGAN' ACADEMY OF SCIENCE. 



METHODS USED IN CULTIVATING PURE STRAINS OF 



PARAMOECIUM. 



FRANCES J, DUNBAR, 



In some experiments which are being carried on in this laboratory, 

 by the writer, it was necessary to obtain a pure strain of Paramoecium 

 from a single ancestor, with the individnals in numbers such that from 

 600 to 800 could be had for each of a considerable number of experi- 

 ments. It was, therefore, necessary to place^ isolated individuals in 

 the environment most conducive to rapitl and continued division. 



At the outstart of the work individual Paramoecia were isolated in 

 a hay infusion made according to the following formula : Thirty grams 

 of hay were heated to 90° C, in one liter of water. The mixture was 

 allowed to cool slowly, and then was strained through cheese-cloth. 

 In making the hav infusion distilled water was used. 



The cultures thus started promised well for a time, but just as the 

 numbers of individuals became suflBcient so that one could begin to think 

 of setting up an experiment the cultures invariably began to show 

 signs of deterioration, and in a day or so every individual would be 

 dead. It seemed probable from the appearance of the cultures that 

 the Paramoecia died of starvation ; the medium appearing to be in some 

 way unsuited to the continued growth of bacteria. Gelatin was laid 

 upon the surface of the hay infusion with no markedly beneficial effect. 



Upon setting up cultures in some of which the hay infusion had 

 been made slightly alkaline, and in some of which it had been made 

 slightly acid, it was found that the individuals in the alkaline solution 

 died within a few hours, while those in the acid solution flourished well 

 for a time, but still did not persist long enough to attain to numbers 

 suflflcient for our purpose. At last it was thought that the lack of 

 inorganic salts in the distilled water might inhibit the continued growth 

 of bacteria; and cultures were set up in hay infusion which had been 

 diluted about one-half with tap-water, which had passed through a clay 

 filter, supposed to be bacteria proof. At the same time, following a 

 suggestion of Prof. A, W. Greely, a small pellet of fresh wheat bread 

 was added to each culture. In this medium our Paramoecia flourished 

 as the green bay tree, so that soon in 5 c.c. of the liquid 293 individuals 

 were counted, as compared with fifty-six in 5 c.c. of the media tried 

 before. 



The tap-water (filtered to prevent a possible contamination of the 

 pure culture with individuals contained in it) provided the organic salts 

 necessary to the continued growth of bacteria; while, according to the 

 theory of Prof. Greeley, the yeast-spores in the bread, grown active in 

 the stimulating medium, provided a very minute but favorable amount 

 of acid in the shape of COo. 



In the carrying on of the experiments for which these Paramoecia 

 were desired it was found that the rate of division was markedly in- 

 creased in those experiments set up in from an N/100 to an N/300 sugar 



