INOCULATION OF EASTERN WHITE PINE WITH BLISTER RUST 381 



These results have been confirmed by other studies reported in 

 another paper in these proceedings; differentiation of germ tubes into 

 infection structures--i .e . , the vesicle and infection hypha similar to 

 those in the substomatal chamber of a needle at the point of initial 

 infection—occurred to any significant extent only when basidiospores 

 that germinated on collodion membranes were subjected to similar tempera- 

 ture fluctuations. Under such conditions vesicles were found on up to 

 10% of germinated spores and were typical in shape and size of those 

 formed in needles. Thus, although spore germination occurred readily at 

 a constant 16 or 20°C temperature, the formation of vesicles, which is 

 apparently an essential part of the infection process, was favored by a 

 diurnal temperature fluctuation of about 4.5 to 24°C. 



It appears that stimulation of vesicle formation and infection might 

 be the result of a high temperature shock, but further details on tempera- 

 ture limits or timing of fluctuation cycles have not yet been determined. 

 Trials so far have indicated that light has no effect on the differentiation 

 of infection structures, as opposed to the experience of Bromfield (1967) 

 and Emge (1958) with wheat rust. Their results indicated that light plus 

 elevated temperatures had a distinct effect on the later stages of the 

 infection process. Bromfield (1967) made the point, important to consider 

 in relation to white pine blister rust, that cardinal temperatures for 

 infection based on averages or continuous temperatures during dew periods 

 (or in this case, during the inoculation period) fail to permit adequate 

 estimates of the occurrence of infection. Thus the infection process is 

 much more complicated than germination, which is merely the initial step 

 in the sequence. 



Some interesting support for this concept is provided by Hirt's 

 (1942) experience with field inoculations of white pine. He found that 

 although pine infection might take place when fluctuating temperatures 

 ranged between about 18°C or below to 21°C or slightly above, temperatures 

 constantly near 20°C or above would inhibit pine infection even though 

 other factors were particularly favorable. Infection occurred usually 

 within a temperature range of 10 to 20°C, but only rarely when tempera- 

 tures averaged above 21°C. Thus, although a temperature of 20°C or below 

 was necessary for spore germination, some change or fluctuation from this 

 temperature was necessary for initiation of pine infection. 



SIM1ARY AND CONCLUSIONS 



The aim of inoculation techniques is to obtain infection, and several 

 methods have been used with C. ribiaola on eastern white pine that can 

 fulfill this objective. The major problem, however, is one of standardi- 

 zation; applying a known quantity of inoculum to a known amount of needle 

 surface under conditions favorable to spore germination and initiation of 

 infection so that the end result, either infection or resistance reactions, 

 can be evaluated in a quantitative manner. Most effort so far has made 

 sure that trees will become infected by application of massive doses of 

 inoculum. There has been little or no control of inoculum concentration 

 among trees within a test, or among different tests, and there is little 

 or no information at present on the relation of quantity of inoculum to 

 host response. 



Assurance of an adequate supply of inoculum is a prerequisite for 

 progeny testing. For large scale tests, it is almost imperative that a 

 "rust garden" be planted for production of inoculum to be used in artificial 



