COLLECTION, CULTIVATION, OBSERVATION 889 



To inhibit bacterial growth in cultures of Entamoeba, various 

 antibiotics have been tried. For example, Spingarn and Edelman 

 (1947) found that when streptomycin was added in the amount of 

 1000-3000 units per cc. to culture of E. histolytica, the survival of the 

 amoebae in culture was prolonged from an average of 8 days to 33.7 

 days, which effect was apparently due to the inhibition of bacteria. 



Encystment of Entamoeba histolytica is usually brought about by 

 first cultivating the organisms in starch-free media and then by 

 transferring them into media with starch. Balamuth (1951) recom- 

 mends a diphasic medium of the following composition: 2 gm. of 

 Wilson liver concentrate powder is brought to boiling in 80 ml. dis- 

 tilled water and filtered. Then 6.4 ml. of 0.25 molar Na 3 P0 4 . 12 H 2 

 and 7.6 ml. of 1.0 molar potassium phosphate buffer (in the ratio of 

 4.7 parts K 2 HP0 4 to 0.3 part KH 2 P0 4 ) are added. By adding dis- 

 tilled water in a volumetric flask bring the mixture to 100 ml. Trans- 

 fer it to a beaker and add 3 gm. Bacto-agar. Heat gently until agar 

 dissolves; then autoclave for 20 minutes at 15 lbs pressure. The pH 

 should be about 7.2. The overlay is prepared by mixing double- 

 strength eggyolk and normal horse serum (10:1) and rice starch is 

 added last. 



Plasmodium. — Bass and John's (1912) culture is as follows: 10 cc. 

 of defibrinated human blood containing Plasmodium and 0.1 cc. of 

 50% sterile dextrose solution are mixed in test tubes and incubated 

 at 37-39°C. In the culture, the organisms develop in the upper layer 

 of erythrocytes. Since that time a number of investigators have 

 undertaken cultivation of different species of Plasmodium. For in- 

 formation the reader is referred to Geiman, Anfinsen et al. (1946) and 

 Trager (1950). 



Balantidium coli. — Barret and Yarbrough (1921) first cultivated 

 this ciliate in a medium consisting of 16 parts of 0.5% NaCl and 1 

 part of inactivated human blood serum. The medium is tubed. 

 Inoculation of a small amount of the faecal matter containing the 

 trophozoites is made into the bottom of the tubes. Incubation at 

 37°C. Maximum development is reached in 48-72 hours. Subcul- 

 tures are made every second day. Rees used a mixture of 16 parts 

 of Ringer's solution and 1 part of Loeffler's derrydrated blood serum. 



Atchley (1935) employed a medium composed of 4 parts of Ringer's 

 solution and 1 part of faeces, which is filtered after 24 hours, centri- 

 fuged and sterilized by passage through a Seitz filter. Nelson (1940) 

 also used 1 part of caecal contents of pig in 9 parts of Ringer's solu- 

 tion, which mixture is passed through a sieve and then filtered 

 through a thick absorbent cotton. Balantidium which shows posi- 



