414 HANDBOOK OF PROTOZOOLOGY 



Peranema, Chilomonas, and other holozoic or saprozoic Masti- 

 gophora. — Although ripe hay infusion is frequently adequate for 

 these flagellates, it is often worth while to prepare the following 

 medium. Mix 10 gm. of peptone and 5 gm. of sodium chloride 

 and dissolve in 1000 c.c. of distilled water. Add 3 gm. of Liebig's 

 beef-extract and 1.5 gm. of shredded agar, to the solution and 

 boil the whole slowly, stirring frequently until the agar is dis- 

 solved. Add distilled water to make up for the loss by evapora- 

 tion. The reaction should be slightly alkaline (pH 7.4), Filter 

 while hot through cotton in a steam sterilizer. Then divide 

 the filtrate into a number of test tubes, insert cotton plugs, 

 and autoclave for 15 minutes under 15 pounds pressure. The 

 tubes thus sterilized can be kept indefinitely. Before using, 

 each test tube is heated and the melted contents poured into 

 a Petri dish and allowed to cool. 



Amoeba proteus and allied forms.- — These amoebae feed on 

 diatoms, bacteria, protozoans, and not infrequently small meta- 

 zoans. Hay infusion makes a satisfactory medium if a few 

 wheat grains are added. Knop's and Benecke's solutions men- 

 tioned above are also suitable for culturing these free-living 

 holozoic amoebae. Some workers recommend cutting and boil- 

 ing aquatic plants, such as Elodea, for this purpose. Upon 

 cooling, pour the plants and extract into a deep, wide-mouthed, 

 glass jar and add aerated rain water. A weaker solution is, 

 on the whole, much better than a stronger one, although the 

 development of amoebae may be slow. After leaving the jar 

 standing exposed for several days, introduce into it the amoebae 

 that have been previously collected. The inoculated jar should 

 be kept in a cool, well-lighted place. 



Vahlkampfia, Naegleria, etc.- — These small mono- or di-phasic 

 amoebae which feed on bacteria can easily be cultivated on the 

 agar medium mentioned above for Peranema and others. A sim- 

 ilar medium, devised by Musgrave and Clegg, and modified by 

 Walker, consists of the following: 



Agar 2.5 gm. 



Sodium chloride 0.05 gm. 



Liebig's beef-extract 0.05 gm. 



Normal NaOH solution 2.0 gm. 



Distilled water 100 c.c. 



