COCCIDIOSIS IN BIRDS AND MAMMALS 283 



cent solution of potassium bichromate is very satisfactory for 

 this purpose. To provide a sufficient exposure to atmospheric oxy- 

 gen, the mixture of bichromate and feces should be contained in 

 flat-bottomed dishes such as moist chambers of Petri dishes. The 

 layer of the mixture should be not over one centimeter in depth. 

 Fluid lost by evaporation should be replaced daily as desiccation 

 will destroy the oocysts. The receptacle must 7iot be covered as 

 air is necessary for the sporulating oocysts. Kept at room tem- 

 perature, mature oocysts may appear any time after forty-eight 

 hours. The feces containing the oocysts may be washed to remove 

 dirt, straw, etc., either before or after sporulation. Washing is 

 easily accomplished by mixing the material with from five to ten 

 volumes of water and straining through four thicknesses of cheese- 

 cloth. The original volume of feces may be approximately re- 

 stored by centrifugation or sedimentation. 



A second method of sporulation is as follows : large Petri dishes 

 or moist chambers are prepared by lining the bottom with at 

 least six layers of filter paper which has been saturated with a 

 two per cent solution of formalin. Fresh, unwashed feces are 

 placed upon the wet filter paper. The formalin vapor prevents the 

 interference of bacterial putrefaction, and the fact that the mate- 

 rial is not submerged in fluid facilitates the ventilation of the mass 

 and thus promotes sporulation. The contents should be sprinkled 

 daily with water or with a one per cent formalin solution. The 

 container must not be covered. If the oocysts are kept at room 

 temperature, they will commence to segment within forty-eight 

 hours. After sporulation is complete, the feces may be washed ofif 

 the filter paper, strained, and concentrated. It is important to re- 

 member that oocysts prepared by this method for the infection of 

 animals must be thoroughly washed to remove all traces of 

 formalin which might injure the animal receiving them. 



Either of the above methods will work faster at incubator tem- 

 perature. The difficulty is that unless carefully watched, the mate- 

 rial will dry out due to the rapid evaporation occasioned by the 

 higher temperature. The following method has been designed to 

 circumvent this problem, and has frequently yielded mature 

 oocysts after incubation over night. Feces containing oocysts are 

 strained, sedimented, mixed with five per cent potassium bichro- 

 mate and placed in large Petri dishes in a layer not more than a 

 half centimeter in depth (Fig. 18, C). A 250 cubic centimeter 



