444 BULLETIN 387 



Smith (1911) and Eycleshymer (1894), both careful workers, state that 

 they saw these bacteria. This is also in accordance with what Maire and 

 Tison (1911) claim to be true for certain parasitic slime molds that are 

 able to ingest unicellular algas present in their aquatic host; and with 

 what Kunkel (1915) has demonstrated in the case of Spongospora sub- 

 terranea grown on agar in which pure cultures* of plasmodia become 

 abnormal and die, while those with which bacteria are present live and 

 thrive. 



All of Pinoy's (1905) experiments appear to corroborate his idea that 

 a coccus form enters the root with the swarm-spore and lives in constant 

 association with the parasite thruout its entire life cycle. He stained 

 sections of the root and observed bacterial forms within the cells. They 

 appeared so much like parts of the cell contents that the microscopical 

 analysis had to be accompanied by cultural study. For this he procured 

 diseased roots of Brassica sinensis measuring from eight to ten centi- 

 meters in diameter, seared the outside, and cut plugs from the interior 

 by means of a flamed pipette. When these plugs were planted in agar 

 media, numerous colonies of bacteria soon appeared. To prove that 

 these organisms were necessary for the development of the myxomycete, 

 Pinoy placed spores of Plasmodiophora Brassicae in a large number of test 

 tubes containing sterilized extract of roots. In two tubes the spores were 

 accidentally not associated with bacteria and they failed to germinate, 

 while in all the other tubes the spores did germinate. 



Pinoy's results are interesting, altho the work does not appear to be 

 extensive enough to warrant the conclusion he has drawn. The following 

 experiments were undertaken by the writer in further quest for facts bearing 

 on this problem : 



Thruout three years of study almost five hundred petri-dish and test- 

 tube cultures have been made from diseased roots of all sizes and ages, 

 grown under various conditions and in widely separated localities. The 

 ordinary method of procedure was to place the roots for ten minutes in 

 mercuric chloride i-iooo; then, after rinsing them several times in sterilized 

 distilled water, to break the roots open and remove bits of the diseased 

 tissue from the broken surface by means of a flamed scalpel. The bits 

 of roots were placed in a sterilized petri dish, where they were teased apart 

 in a few drops of sterilized distilled water. Two successive dilutions 

 were made, and these, together with the original drop in which the tissue 

 had been crushed, were poured into nutrient agar media. 



All the results were uniform in that no bacterial colonies were obtained 

 from the roots with young swellings. From the medium-sized swellings 

 occasional colonies developed; and from the larger galls, especially when 

 the epidermis had been broken, numerous colonies always appeared. 



