252 PROCEEDINGS OF THE AMERICAN ACADEMY. 



the others that zygospores might be expected to appear where some of 

 the mycelial colonies came into contact. Four crystallizing dishes 

 with paste, containing each six transfers, and two stenders with potato 

 agar containing respectively two and three transfers were placed in the 

 warm oven at 26° to 28° C, while three crystallizing dishes with 

 paste containing four transfers each and two stenders with potato agar 

 containing respectively two and three transfers were left at the room 

 temperature. When these cultures matured there was no evidence of 

 zygospores, even though a careful microscopic examination was made 

 of the low-lying hyphae at the areas of contact of all the colonies. On 

 the assumption that Rhizopus belonged to the heterothallic type it seemed 

 strange that out of the forty-six contrasts between pure transfers from 

 zygosporic cultures, in addition to the contrasts previously made, not a 

 single zygospore should have been found. Nevertheless, although the 

 frequent yellow appearance of the young sporangia was suggestive of a 

 varietal difference in the zygosporic strains, the action of the fungus in 

 the cultures above described, as well as the fact that the zygophoric 

 hyphae could never be traced to the same filament, convinced the writer 

 that the species could not be other than heterothallic. 



By running a large number of van Tieghera cell cultures into which a 

 small number of zygospores connected with the aerial mycelium had been 

 transferred, it was hoped that the hyphae connected with the two sides 

 of the zygospore would continue their growth and that by dissecting these 

 out under a microscope it might be possible to obtain cultures which one 

 could be sure had the same origin as the two respective gametes, and 

 further that by a contrast on the proper nutrients of the two cultures 

 thus obtained it might be possible to demonstrate the presence or absence 

 of a heterothallic condition. Unfortunately when the hyphae were suf- 

 ficiently few in number to make out their connections with the suspeu- 

 sors, they failed to grow, and though when larger masses of hyphae were 

 used in the inoculation zygospores were generally developed, it was not 

 possible to trace their mycelial connections. 



Finally, in one cell containing a single young zygospore, it was found 

 that the suspensors themselves had germinated, and by transferring the 

 mycelial growths which had thus arisen, two cultures derived from 

 mycelia of different origin, both of which were concerned in the forma- 

 tion of the original zj'gospore, were obtained. These strains, which for 

 reasons to be mentioned later have been called (+) and (— ) respectively, 

 were derived, as we have seen, from the germinations of opposite sus- 

 pensors. They were inoculated December 1, side by side in a van 



