170 



(My experience has been that less than y 8 voulme of the first 

 reagent ought not to be taken, but about 7 3 volume of the second 

 may be used without harm. By entirely leaving out the last, which 

 several authors do, the reactions do not always seem to be so 

 strong as when both reagents are used). When indol is present 

 a purple-red colour develops. The result is not read until 10—15 

 minutes have elapsed, as the colour often appears rather slowly. 



The series „I" was made in Jan. 1920, with the exception of 

 I 101—104, I 32-37, all the Ms's, and H 164—167, which were ex- 

 amined a couple of months later. They were inoculated in ordi- 

 nary broth containing lo/ Witte peptone and with the addition 

 of enough haemolysed horse blood corpuscles to produce a weak 

 pink colour (about l°/ 00 blood corpuscles), henceforward referred 

 to os ,,haemoglobin brolh". Before inoculating, the tubes were kept 

 in the incubator for 24 hours to be sure they were sterile. This 

 changed the weak red colour to a still weaker brownish tint which 

 in no way hampered the judging of the indol reaction. 



After inoculation the tubes were placed very obliquely in the 

 incubator for 5 days. (The same time was allowed for growth 

 in the following series when the contrary is not indicated. Both 

 in this and the following series cultivations were made, before 

 the indol test was carried out, from all the tubes on to haemoglo- 

 bin or Fildes agar. Only those tubes from which a growth of 

 other bacteria than Pfeiffer's bacillus occurred are omitted from 

 the table. 



For series II (March 1920), a sterilised watery solution of l<y 

 Liebig s meat extract, lo/ peptone „Chapotaut", and i/ 2 % salt 

 was the medium used. For series III, ordinary broth with lo/ 

 peptone, „Chapotaut", instead of the commonly used Witte's pep- 

 lone, was employed. For series IV a watery solution of 2<>/o pep- 

 tone „Chapota.ut" and i/ 2 % salt, was used. Haemoglobin was added 

 lo all three media, as described above. All the three series were 

 investigated at the same time. 



The medium in series V was the same as in series I, but the 

 cultures with which it was inoculated had previously been subcul- 

 tured 25 times on ascitic agar with a little haemoglobin, in sym- 

 biosis" with a large number of different species of bacteria. The 

 object was lo discover whether by cultivating Pfeiffer's bacillus 

 under variable nutritive conditions, its biological characters could 

 be altered (indol formation; for agglutination, see later). 



In series VI (May 1920) haemoglobin broth was used which was 

 inoculated with cultures that had been subcultured daily for 96 

 days on haemoglobin agar. (It is not known whether Witte pep- 

 tone, or „Peplonc Berna" which is said to be prepared „par le 

 proceed Witte", was used in this case). 



In series VII (Sept. 1920) and Vila (Oct. 1920) trypsin<ligested 

 casein prepared according to Cole & Onslow's instruction, was 

 used. One part of „stock broth" -f- 2 parts of water containing 



