BACTERIA WITH MYXOMYCETES. 5 By (ht 
The presence of aérobic bacteria seems to be necessary to the life of the myxomycete outside 
of the host cells. Thus among the great number of tubes of agar sowed with the spores, the greater 
part of those containing bacteria gave at once a development of the fungus (formation of the amceboid 
individuals which soon perished) while in two containing no bacteria, the spores, though perfectly 
preserved did not germinate. 
Pinoy thinks, therefore, that the bacteria introduced with the parasite contribute to the decay 
of the tumor of the cabbage when the conditions are favorable to their multiplication. 
Pinoy continued his interesting studies, publishing a monograph in 1907 wherein he 
considered at length the relations of various bacteria to several species of Myxomycetes 
in the Group Acrasieae; then to several species of the endospore-bearing forms, and finally 
to Plasmodiophora brassicae. I summarize as follows: 
Pinoy, like Nadson, found Bacterium fluorescens liquefaciens associated with Dictyostelium 
mucoroides. 
On agar poured plates the spores of this slime mold germinate only in the presence of bacterial 
colonies. Elsewhere the spores do not germinate. But those spores which do not germinate can 
not for that reason be assumed to be bacteria-free. They also bear bacteria which can be resusci- 
tated by putting them into bouillon, which clouds after 6 to 8 days. Nadson, Potts, and Vuillemin 
did not take into account these tardily developing bacteria. 
B. fluorescens is killed by exposure to 50° C. for 1 hour. On the contrary, 80 per cent of the 
spores of Dictyostelium mucoroides are still able to germinate after such an exposure. It is possible 
also to purify the spores by heating them for 2 minutes at 56° C. Spores about 8 days old are most 
resistant. Spores thus purified will never germinate on any culture medium whatsoever, unless 
suitable bacteria are added. 
Potts’ statement that he was able to grow Dictyostelium mucoroides on the dead bodies of his 
Bacterium fimbriatum is regarded as illusory, 7. e., the purity of his spores is questioned. Pinoy 
showed the need of living bacteria quite clearly as follows: Into a flask of culture medium he plunged 
a collodion sack attached to a projecting tube. After proper sterilization, the medium inside the 
collodion sack was inoculated with purified spores of D. mucoroides, that outside with Bact. fluorescens. 
The spores of the Myxomycete germinated but the amoebae soon rounded off and died. ‘The dead 
bodies of Bact. fluorescens (killed by heat, ether and chloroform) were also placed in the collodion 
sack without result. 
“En résumé, le D. mucoroides ne peut vivre qu’en association avec une Bactérie vivante. ‘Toutes 
les Bactéries ne conviennent pas également.” 
The culture medium exerts a marked influence, e. g., on potato sowed with mixtures of Dicty. 
mucoroides and Bact. fluorescens the slime mold does not develop. The same is true on this medium 
whatever bacterium isused. With most bacteria this is also true on peptone agar, or meat broth agar. 
The best culture medium found was flax-seed agar. On this the bacteria grow abundantly and the 
harvest of Dictyostelium mucoroides reaches its maximum. 
Bacteria that stain by Gram are not suitable for such cultures. In general, bacteria which do 
not stain by Gram allow the Dictyostelium mucoroides to grow. ‘The Dicty. mucoroides will not grow 
in the presence of pure cultures of Bacillus megaterium, but if this organism is added to Dictyostelium 
mucoroides with Bact. fluorescens growth may be had on beef agar. 
The growth of the Dictyostelium mucoroides may be regarded as a parasitism on the bacteria. 
They are absorbed into the vacuoles and digested. Pinoy confirms Metchnikoff’s observation that 
the liquid in the vacuoles is acid. 
Neutral red is recommended as a stain for the bacteria in process of digestion. It does not 
stain the living bacteria nor kill the Myxomycete unless too strong. Vesuvin may also be used. 
For details see paper. Potts did not find bacteria in the interior of the vacuoles of Dicty. mucoroides 
because his technic of fixation and staining was insufficient, he therefore formed the erroneous 
hypothesis of an extra-cellular diastase. 
Grown with Bacillus coli, the enzyme isolated at the end of 40 hours liquefied gelatin. It acts 
in neutral or feebly alkaline liquids. The acidity of methyl orange inhibits. It is therefore related 
to trypsin rather than to pepsin. It is destroyed at 55°C. Its maximum of activity is about 38° C. 
It has scarcely any action on fibrin or on albumen coagulated by heat. 
“‘ Acradiastase’’ does not act on bacteria killed by heat, but readily dissolves those killed by 
ether or chloroform. 
“The best bacterial test is also B. coli which is not self-autolytic, and a chloroformed emulsion 
of which remains cloudy. 
