APPENDIX 415 



By subcultures, almost a pure culture with bacteria can be 

 obtained. Hay infusion is also excellent for these small free- 

 living amoebae. 



Arcella and other Testacea. — These forms commonly multi- 

 ply in a mixed culture in hay infusion. Hegner cultivated 

 Arcella by the following method: Pond water and weeds are 

 collected. In the laboratory they are shaken up violently and 

 filtered through eight thicknesses of cheese cloth, which ap- 

 parently prevents passage of particles larger than Arcella. The 

 filtrate is distributed among Petri dishes, and when the sus- 

 pended particles settle down to the bottom of the dish, Arcella 

 are introduced. They multiply rapidly. 



Actinophrys, Actinosphaerium, etc. — Belaf cultivated these 

 heliozoans successfully in Knop's solution. 



Paramecium, Euplotes, Stylonychia, and other ciliates. — Ail 

 these ciliates are easily cultivated in a weak solution of hay 

 infusion, with or without wheat grains. The crystallization 

 jars containing the medium should be left standing uncovered 

 for several days to allow a rich bacterial growth in them. Seed 

 them with material such as submerged leaves or scum contain- 

 ing these ciliates. When once started, separate cultures can be 

 easily made and maintained by the isolation method. If 

 desired, a culture can be started with a single individual. Sub- 

 cultures should be made from time to time. Mixtures of beef- 

 extract and hay infusion have also been used for cultivating 

 free-living holozoic ciliates. 



Cultivation of Parasitic Forms 



Embadomonas , Trichomonas, Chilomastix, etc — There are 

 numerous media which have been used for successful cultiva- 

 tion of these flagellates in vitro. For Trichomonas hominis and 

 T. vaginalis. Lynch used nutrient broth with 0.05 per cent 

 acetic acid, incubated at 30° C, making subcultures every 

 two to three days. Hogue's ovo-mucoid medium is made as 

 follows: Whites of six eggs are broken in a sterile flask with 

 beads. Add 600 cc. of 0.7 per cent salt solution and cook the 

 whole for 30 minutes over a boiling water bath, shaking the 

 mixture constantly. Filter through a coarse cheese cloth and 

 through cotton-wool with the aid of a suction pump. Put 5 cc. 



