Methods in Rust Investigations 245 



being made this method has the objection that considerable time and 

 work is necessary to prepare the cells and remove the vaseline from the 

 cover glasses in preparation for another test. 



A less laborious method is to use Syracuse watch glasses (fig. 1, B) 

 with ground glass margins. The bottom of the watch glass is covered 

 with water on which the aeciospores or urediniospores are floated. The 

 accession number of the collection or other identification mark is writ- 

 ten on the ground glass margin. Another dish is placed on top of this 

 in which another collection of spores is tested. A number of watch 

 glasses can be stacked thus and a number of such stacks placed in shal- 

 low dishes for convenience in carrying from place to place. 



Hanging drops also can be made with these watch glasses by placing 

 the spores in one or more drops of water on the bottom of a watch 

 glass, inverting it and placing it on another inverted watch glass which 

 has been covered with water. The beveled edge of one glass fits into 

 a groove in the bottom of the other so that there is little danger of 

 slipping. The hanging drop is by far the best method with teliospores 

 of Puccinia triticina, for they otherwise are likely to sink to the bottom 

 of the drop and either not germinate or give abnormal germination. 

 In this method, however, the drops can be examined to advantage only 

 under the lower powers of the microscope. This method has the ad- 

 vantage of requiring but little time in the preparation and labeling of 

 the tests. Hursh (1922) also has used and described this method. 



Still another method is to make hanging drops on the lid of a petri 

 dish, the water in the bottom providing the humidity. Spores have been 

 tested also by placing the drops on glass slides in petri dishes raised 

 above the wet substratum (moistened filter paper or paper toweling) 

 by glass slips (fig. 1, A) as described by Melhus (1915) and Melhus 

 and Durrell (1919). The petri dishes take up somewhat more room 

 than watch glasses, but the latter method has the advantage that a 

 cover glass can be placed on the drop, enabling examination under the 

 higher powers of the microscope. Each method has features of advant- 

 age in different types of investigations. 



4. Environmental Conditions Governing Spore Germination. As 

 inoculation with the rust fungi depends upon the germination of their 

 spores, the conditions influencing spore germination must be taken into 

 consideration in devising methods for the culture of these fungi. Per- 

 haps the most essential condition is a humidity of 100 per cent. In 

 many rusts, the spores must also be in contact with water. While 

 spores may germinate through a fairly wide range of temperatures, 

 for many rusts a temperature of 16-20° C, is most favorable for germi- 

 nation, few spores germinating below 5° C, or above 30° C. Different 

 species, however, vaiy somewhat as to the cardinal temperatures for 

 spore germination. Fromme (1913),Doran (1922) and Maneval (1922) 

 give summaries of the results obtained by themselves and other investi- 

 gators in the studies of the effect of various conditions on the spore 

 germination of a number of rusts. 



While these are the principal factors governing germination, still 

 other factors are of some importance in the greenhouse investigations. 



