CHAPTER XXXVIII— LABORATORY AND TEACHING 

 METHODS (CONTINUED) 



LESSON 30 



Inoculation Experiments. — The experiments recorded below need not be rigidly 

 followed by the m^cologic teacher. Other organisms and other hosts can be used 

 just as satisfactorily. The types used must be determined by locality and by other 

 considerations of cultural methods and laboratory facilities. The directions below 

 may be taken as samples. 



Potato Rot {Fusarium trichothecoides) . — Take Green Mountain potato tubers and 

 sterilize surface by soaking in 2 per cent, formalin for two hours. The tubers are 

 then held with towels that have been boiled in water, and are wrapped in these steri- 

 lized wet towels after having been inoculated with Fusarium trichothecoides by 

 pricking the surface of the tubers and dipping them in distilled water which holds 

 the spores of the fungus in suspension. The potato tubers wrapped with wet towels 

 are then surrounded with oiled paper and kept at a temperature not lower than 

 10° to 1 2°C. Tubers of several varieties can be used, such as Up-to-date, Early Rose, 

 Irish Cobbler. If the inoculation has been successful, results will be noted in ten 

 to fifteen days. A transfer to potato slant test-tubes will result in a fungus which 

 has powdery-rosy appearance. Consult Jamison, C. O., and Wollenwebee, 

 H. W.: An External Dry-rot of Potato Tubers caused by Fusarium trichothecoides. 

 Journal of the Washington Academy of Science, II, No. 6, March 19, 1912. 



After the normal lesions have been obtained and the fungus studied mor- 

 phologically under the microscope, take small slices of potato tuber showing healthy 

 and diseased tissues in proximity and fix in chromacetic acid. Wash off the fixa- 

 tive in running water, and carry through the alcohol, etc., to paraffin. After im- 

 bedding in paraffin, section and mount as usual (see Lesson 42). 



Crown-gall {J'seudomonas tumefaciens) (Fig. 227). — Inoculate the stem of a 

 geranium {Pelargonium zonale) with the organism in pure culture by first 

 washing the stem at the intended point of infection with i per cent, formalin and 

 then with distilled water. Place some of the pure culture on the stem by means of 

 a platinum needle and prick the organism into the stem with a sterile needle mounted 

 in a wooden handle. The part of the geranium stem selected should be a young 

 actively growing leader (consult the Biilletin of Erwin F. Smith, and the book 

 of Dtjggar, Diseases of Plants, pp. 114-118). 



This organism can be successfully grown on beef agar which is made as follows. 

 To 1000 c.c. of peptonized beef bouillon add i per cent, of agar flour, steam three- 

 quarters of an hour and cool down below 6o°C. Then add neutralized white of two 

 eggs to clarify. Made to -f- 15 Fuller's scale by adding 4NaOH. The test-tubes 

 are autoclaved fifteen minutes at iio°C. 



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