108 



requirements of Pfeiffers bacillus are not greatly altered by 

 prolonged cultivation. In order to investigate this more closely 

 I repeatedly inoculated all the strains which were still living 

 at the time, on haemoglobin-free media. 



On 26—28. XII. 1919 the following strains were subcullured in 

 pure culture on ordinary agar (weakly alkaline to azolitmin paper: 

 I 1, 5, 6, 20, 21, 22, 23, 27, 29, 33, 38, 43, 51, 57, 59, 71, 72, 91, 

 100; I 5, 21; P 14, 15, 16, 22, 23, 24, 25, 23; H 14, 19, 34, 35a, 

 37a, 37c, 49b, 122, 127, 133, 151, 152, 154, 155, 156, 157, 158, 159, 160, 

 161; Pa 4, 5. In no case was there any growth after 1 days 

 in the incubator. The same cultures were inoculated on ordinary 

 agar together with various other bacteria (while air-coccus, Strepto- 

 coccus, B. faecalis alcaligenes) in the same way as was done in 

 the symbiosis reaction described later. Some of the strains were 

 inoculated directly from haemoglobin agar, and showed a quantity 

 of growth but only in the immediate neighbourhood of the al- 

 caligenes culture. The other strains were inoculated from sym- 

 biosis" cultures on plates poor in haemoglobin. None or only 

 very slight growth took place. We thus see what a marked effect 

 the minute quantity of haemoglobin which is transferred with the 

 culture, can exert when it is combined with the action of the other 

 bacteria. 



On 2. III. 1920 the same strains were inoculated on agar and 

 ascitic agar in company with a haemolytic Streptococcus which 

 with suitable experimental technique, has a good growthpromoting 

 action on Pfeiffer's bacillus. Both Pfeiffers bacillus and the Strepto- 

 coccus were inoculated in parallel streaks at about % cm. distance, 

 alternately. The agar was alkaline to azolitmin paper, but did not 

 react to a-naphlhol-phlhalein. For ascitic fluid the weakly haemo- 

 globin-containing ,;1,017" (see p. 106) was used. The ascitic agar 

 mixture was slightly alkaline to azolitmin paper. In order to avoid 

 transferring a greater amount of haemoglobin than necessary, cul- 

 tures of all the strains were first produced on agar with dissolved 

 blood corpuscles in a concentration of Viooooj m „symbiosis" with 

 the same Streptococcus. After IV2 days in the incubator there was 

 a very rich growth on this medium. Subcultures were then di- 

 rectly made for the actual test. As a control that the agar and 

 ascitic agar were efficient, some haemoglobin was added to a number 

 of them, and they were inoculated with some of the cultures of 

 Pfeiffer's bacillus, and they showed particularly good growth. On 

 agar plates with the streaked „symbiosis" cultures a distinct though 

 slight growth appeared after an interval of 4 days. From these, 

 subcultures were made on to new plates, using the same technique. 

 (The agar plates used in this second culture were made on the 

 day before, while the ascitic agar plates had been kept 5 days in 

 the icesafe). No growth of Pfeiffer's bacilli occurred this time 



