418 Agricultural Experiment Station, Ithaca, N. Y. 



brotli,lias been used, except in one or two cases where tlie organism 

 refused to grow and produce spores on tliis substance. In 

 these cases potato agar, made from potato and agar only, was 

 employed. 



To separate a fungus a small portion containing spores is 

 removed usually to a slide and teased apart in a little water. 

 Meanwhile three tubes of agar have been placed in the water-bath 

 and heated until the agar is melted. They are then cooled in water 

 to 43° Centigrade, and the spores and pieces of fungus on the slide 

 are washed into the first tube and well shaken. A. drop or two is 

 now poured into tube number two and shaken, and the process is 

 repeated with tubes numbers two and three. It is now probable 

 that by this repeated dilution only a few organisms have been trans- 

 ferred to tube number three. In separating spores of fungi this 

 method of poring a drop from one tube to the next, is preferable to 

 dipping in a sterilized needle as is usually done in transferring 

 bacteria, for the spores of fungi are so much larger than bacteria that 

 the number of fungus spores, which would cling to the needle, would 

 be small in proportion to the number of bacteria which would also be 

 transferred with them. By pouring out a drop, the proportion of 

 fungus spores to the bacteria present remains unchanged. The con- 

 tents of each tube is now poured into a sterilized Petrie-dish. 

 The melted agar spreads evenly over the bottom of the dish and 

 soon hardens into a iirm, jelly-like layer, thus fixing in position any 

 organisms present. Bacteria and spores of fungi grow freely in this 

 layer, and the plate may be placed at any time under the microscope 

 and the progress examined. Moreover the organism, being fixed, 

 is obliged to confine its growth to a relatively small area. Isolated, 

 growths or colonies starting from a single organism or spore may 

 usually be found in the second plate and often in the first. It is 

 well to make studies from such isolated growths, for if two run 

 together it is possible that they will contain more than one species. 

 A small portion may now be removed on the point of a sterilized 

 platinum needle and transferred to a stick of sterilized potato or 

 other suitable medium. A pure culture is thus obtained. It is 

 also sometimes possible to obtain the pure culture direct from the 

 insect by touching a sterilized needle to the" spore-bearing stroma 

 and then to a tube of potato. 



