148 Journal of Agricultural Research voi. xvi.no. s 



FIXING AND STAINING METHODS 



Material was obtained from the different parts of infected plants at 

 various stages of the development of the disease and killed, preferably in 

 Flemming's medium fluid. The material was put through the regular 

 process of washing until it had been passed through 70 per cent alcohol. 

 It was then placed in a 10 per cent solution of hydrofluoric acid and 

 allowed to remain for three or four days to remove any silicates which 

 might be present. Sections not treated in this manner injured the knife 

 and could not be sectioned satisfactorily. The material was then returned 

 to 70 per cent alcohol and the regular process followed until it was 

 embedded in paraffm. 



Three different stains were used for differential staining of host and 

 fungus tissue — namely, Delafield's hematoxylin, and Flemming's, triple 

 and Pianeze stains. Both the triple and Pianeze stains proved to be 

 desirable for this purpose, while the hematoxylin was not satisfactory. 

 The orange G in the triple stain was taken up very readily by the fungus 

 and was very desirable fer staining the small hyphae. The Pianeze stain 

 gave the fungus tissue a pinkish color and served to bring out more of 

 the details of structure, especially in the reproductive bodies. Sections 

 from both corn and teosinte were stained, and the more successful sections 

 were obtained from the thicker parts of the sheath tissue of teosinte. 



ARTIFICIAL INOCULATIONS 



Artificial inoculations were first tried in the greenhouse at Madison, 

 Wis., in the winter of 1916-17. A special section of the house was ob- 

 tained for this work, one in which a fine spra)^ of water was kept going 

 to keep the room damp. The temperature was kept at as near 30° C. 

 as possible, which was perhaps a little too high for the germination of the 

 sporangia. The plants were inoculated by spraying a suspension of 

 sporangia behind the sheaths and in the bud. None of these pla,nts were 

 infected, for some reason. Inoculations were made in a similar manner 

 on plants in a small isolated plot at West Raleigh, N. C, on July 23, 191 7. 

 At the same time these inoculations were made there was a daily occur- 

 rence of summer showers which continued three days after the date of 

 inoculation. These plants were kept under observation by Dr. F. A. 

 Wolf, Pathologist of the Station, who observed the first signs of the dis- 

 ease 10 days after the inoculation had been made. Two weeks after the 

 inoculations had been made, he reported the presence of the dark-brown 

 spots caused by the presence of abundant sporangia. The writer received 

 specimens from these plants for his collections. In May of this year 

 (191 8) inoculations were made on corn growing in the greenhouse at the 

 Arlington Experimental Farm at Arlington, Va. The weather was very 

 warm, and it was necessary to spray these plants twice daily in order to 

 keep the bud and sheath water from evaporating. On the writer's return 



