REVIEW OF FUSIFORM RUST INOCULATION TECHNIQUES 345 



with tent chambers constructed over nursery beds or in greenhouses. 

 Although it is time consuming for large studies and difficult to control 

 inoculum density, the method can be used to good advantage to provide 

 abundant inoculum for each seedling and, thereby, increase the chance for 

 successful inoculation. 



Inoculation chamber . --In this proceedings, a chamber for inocu- 

 lation of individual seedlings is described (see Snow and Kais) . This 

 two -compartment plexiglass box is designed to control inoculum density, 

 temperature, and relative humidity for epidemiological studies. The 

 disadvantages of this chamber arise from its single plant capacity. 

 However, for many studies, the advantages which accrue from the control 

 of important variables far outweigh the disadvantages. 



8. Water suspensions of sporidia .--Sporidia cast from germinated 

 telial columns can be suspended in water and sprayed onto susceptible pine 

 foliage. This technique, which is not wholly unnatural, usually did not 

 result in the successful establishment of the fungus in pine tissues. In 

 addition to separating telial germination from sporidial germination, this 

 technique provides a means of quantifying inoculum density and, therefore, 

 is a candidate for further study. 



9. Direct placement of precast sporidia onto pine tissue . --Place- 

 ment of precast sporidia onto pine has resulted in successful inoculations. 

 Sporidia are cast into water, collected on "Millipore" filters and trans- 

 ferred to pine tissues. Small numbers of sporidia (5 to 10) can be trans- 

 ferred from filters having an even distribution of spores via finely drawn 

 glass rods. This technique has two important advantages; it provides for 

 good inoculum density control and allows critical placement of inoculum. 



In addition, telial germination is separated from the inoculation technique. 

 Although this technique is not suited to large resistance screening trials, 

 it has promise for studies of penetration, tissue susceptibility, or 

 inoculum density. 



10. Stem insertion . --C. fusi forme , in the form of telia-bearing oak 

 leaves, telial columns, and diseased gall tissue, is placed in wounds made 

 in pine stems. Although these methods are only fairly successful, some 

 advantage is gained via a relatively short incubation period for the 

 formation of stem galls. This is a time consuming, unnatural method which 

 bypasses possible needle resistance. Perhaps, stem insertion techniques 

 could identify modes of disease resistance in the stems of trees immune 

 due to foliar resistance factors. These "bark resistant factors" which, 

 otherwise, would be masked by resistance to initial establishment of the 

 fungus could provide for the establishment of useful differentially 

 resistant lines. 



11 and 12. Injection of sporidia into pine stems and needles . -- 

 Sporidia collected on water beneath germinating telial columns, centrifuged, 

 and drawn into a syringe are injected into pine tissues. Both stem and 

 needle tissues can be inoculated in this manner. Because inoculum of 

 varying densities can be placed in any desired location, this is a useful 

 technique to study infection courts, symptom development, and modes of 

 resistance. Insofar as disease resistance screening is concerned, 

 obvious objections arise because injection techniques are time consuming 

 and unnatural. Needle injections bypass mechanical plant barriers which 

 might be involved in resistance. Stem inoculations bridge all types of 

 needle resistance including mechanical and physiological barriers to both 

 germinatioiand penetration of sporidia and subsequent colonization prior 



