18 
Resi arch Bulletin No. 1 
adequate method if one has to determine positively whether one 
is dealing Avith single spores or not. The writers found that 
the following method is quite easy, absolutely safe, and rigidly 
scientific: A small amount of culture material is put into about 
1 c.c. of sterilized water and then well shaken and stirred. The 
suspension is then poured into a petri dish, and a few drops 
about the size of a pin point, which are removed with a platinum 
wire with a minute loop, are examined and the spore content 
determined. If so dilute that one gets one or no spores, single 
drops are then put upon sterile cover-glasses and examined on a 
sterile hanging drop slide with the microscope. If a spore is 
present, the cover-glass is transferred to an agar plate by shoving 
it into the agar medium, drop side up, until the drop touches the 
surface of the medium. A little sterilized water is then poured 
on the agar, and the plate covered and rolled so as to distribute 
the suspension. If this work is done in an inoculation cage* and 
if one works cleanly and carefully, it can be done without danger 
of contamination. If contaminations do set in, transfers are 
made as soon as the colonies appear, and in this manner ab- 
solute single spore cultures are gotten. Plantings made in this 
manner when more than one spore was counted in the drop show 
the required number of colonies. The contaminations which 
ordinarily appear are Mucor, Penicillium, and Aspergillus. 
If one allows the spores to germinate in the petri dish before 
the actual mounts are made, detection of the spores under the 
lens is facilitated, and in this manner the smallest of spores 
rnn be found. Sooner or later all who expect to have pure cultures 
which are really pure cultures will have to follow the advice of 
Brefeld (14 : 87) .f 
The color determinations were made according to the Re- 
pertoire de Conleurs published by the Societe Francaise des 
rhrysrmtheniists 1005. 
MACROSCOPIC CHARACTERS. 
Studies for the determination of the gross aspect of the fungus 
were made with cultures on glucose agar and on beef bouillon. 
* Wilcox, E. M , and Link, G. K. K. 1912. A new form of pure culture 
chamber. Phytopathology 2:120. Pig. 1. 
t "Fur die Durchfiihrung zuverlassiger mycologischer Untersuehungen 
1st die Gewinnung des Sporenmaterials von den verschiedenen Pilzen in 
reiner Form das erste und unerlasslichste Erfordernis. Von dem reiuen 
Sporenmaterial kann nur durch die Aussaat der einzelnen Spore und ihre 
continuierliche Verfolenng in alien Stadien der Entwicklung in den 
geeigneten, durchsichtisren pilzfreien Nahrmedien ein sicheres Rpsultat 
gewonnen werden, welches in den einzelnen Fallen der weiteren Erganz- 
ung bedarf durch die Aussaat rein gewonnenen Sporenmaterials in sicher- 
^terilisiprten und znsagenden. nach aussen geschiitzten Massensubstraten." 
