38 BULLETIN 934, U. S. DEPARTMENT OE AGRICULTURE. 
readily at certain times of the year was done before the development of pure- 
culture methods. Water cultures kept in the dark and in the light, at constant 
and at varying temperatures, with nutrient substrata consisting of steamed 
or outoclaved fragments of potato, carrot, sweet potato, turnip, sugar beet, corn 
meal or rice, nutrient agar, sugar-beet seedlings, and insects have all produced 
only sexual fruiting bodies and chlamydospores (the so-called conidia). 
(3) The successful cross-inoculations, those which Edson (38) used on sugar- 
beet strains and the writer had found parasitic on pine and had used on pine. 
strains which Hawkins had found parasitic on potato tubers and Edson on 
sugar beets, confirm the work of Hofmann (77) in indicating that the Pythium 
which causes the damping-off pine is a parasite on entirely unrelated species 
of host plants, a commonly recognized characteristic of Pythium debaryanutn. 
The organism is easily isolated from recently damped-off conifer- 
ous seedlings or from soil direct by placing the seedlings or a lump 
of soil at the edge of a Petri dish of solidified prune agar and transfer- 
ring to tubes mycelium from the advancing edge of the resulting 
growth. It has been found in or obtained from damped-off conifers in 
California, Kansas, Nebraska, Minnesota, and the District of Columbia, 
as well as in cultures made by Mr. Glenn G. Hahn in Michigan. Picea 
engelmanni, P. sitchensis, Tsuga mertensiana, Pinus nigra austriaca, 
and Pseudotsuga taxifolia are the coniferous hosts from which cul- 
tures of Pythium debaryanum have been obtained. It has been iso- 
lated directly from soil not only in coniferous seed beds but from 
open grassland in California not adjacent to any seed bed or culti- 
vated crop. Unless Mucor is abundant, Pythium is commonly ob- 
tained in apparently pure condition on the first transfer from the 
plate, as prune agar appears unfavorable foT most bacterial growth 
while allowing rapid spread of the Pythium. On media made from 
prunes which taste sweet and with a total gross weight of not more 
than 40 or 50 grams per liter of medium, the Pythium will make a 
rapid growth, often extending radially 1 mm. per hour at tempera- 
tures in the neighborhood of 22° C. and produce both chlamydospores 
and oospores. A less valuable medium for isolation work, but more 
convenient for subcultures than any other which has been tested, is 
autoclaved corn-meal agar. The growth is not luxuriant, but spores 
are always formed and the cultures seem to be as long lived as those 
on any other medium, retransfer being rarely necessary more often 
than twice a year. Much stronger growth and more abundant fruit- 
ing is obtained on such media as sugar-beet or rice-stem agar, but 
the leathery surface of the culture on such media makes transferring 
difficult. On rice grains, corn-meal mush, beef agar, and on corn- 
meal agar plus 2 per cent dextose or sucrose no spores are formed 
and the cultures are short lived, though, growth is heavy and on 
the last-named medium extremely rapid. On agar containing the 
juice from sour prunes or on corn-meal agar prepared without sub- 
jecting it to the high temperature of the autoclave, both growth and 
fruiting have been very poor or even lacking. 
