98 METHODS OF PURE CULTURE 



the aid of an improved form of the dilution method, further mention of which 

 will be made in the second volume. 



The so-called fractional method of culture, as it was afterwards styled by 

 Klebs, was employed in particular in Pasteur's experiments on fermentation. 

 It consists in taking from a sample of fermenting liquid that has attained its 

 maximum of development a small aliquot portion and transferring this to a new, 

 sterile medium. By recalling the remarks made in the paragraphs of Section 

 II. dealing with symbiosis, it will be understood that, at the period of highest 

 fermentation in a natural liquid and therefore one rich in different species 

 that species which is the cause of the fermentation in question will preponderate. 

 Therefore, if merely a single droplet thereof be placed in a medium analogous in 

 composition to the original habitat, this species will be favourably situated from 

 the outset, and will increase at a relatively quicker rate than its associates. By 

 repeating this transference (" re-inoculation ") several times over, cultures will 

 finally be obtained wherein impurities, i.e. extraneous species, can only be 

 detected by more searching methods of separation, such as are described in the 

 next paragraph. 



These subjugated species will, however, come to the front again if the 

 (apparently pure) bacterial culture be inoculated in a different medium forming 

 a favourable environment for their development. Mention of this has already 

 been made in a previous section, when referring to the older evolutionary 

 labours of Lister, Lankester, Hallier, Billroth, and others. We are now in 

 possession of another more convenient method for the purposes of pure cultiva- 

 tion, which will be described in the succeeding paragraph, and consequently a 

 criticism of the dilution method can be omitted. 



At present, only a couple of words will be devoted, as supplementary to the 

 remarks already made, to the examination of brewery water for the presence of 

 dangerous organisms. For the brewer those water bacteria alone are important 

 that develop in wort and beer and are capable of producing injurious changes 

 therein. Consequently, sterilised samples of both these liquids are employed 

 in the biological analysis of brewing water. The method employed was first 

 proposed by E. CH. HANSEN (IV.), according to whom fifty small Freudenreich 

 flasks are used, twenty-five of them being charged with 15 c.c. of sterilised wort 

 apiece and the remainder with a similar quantity of sterilised beer. In each of 

 the first fifteen flasks in both series is placed one drop (0.04-0.05 c.c.), and in 

 each of the remaining flasks 0.25 c.c., of the water to be tested. These fifty 

 flasks are then kept at a temperature of 24-2^ C. for fourteen days, and are 

 examined to ascertain how many become turbid or throw up a skin, i.e. exhibit 

 signs of the development of organisms. The ratio of the number of flasks with 

 turbid contents to the total number is referred to i c.c. of water, and a standard 

 for determining the destructive capacity of the sample in question is thus 

 obtained. Assuming that, for instance, three out of the fifteen wort-flasks 

 (inoculated with o 04 c.c.) exhibit turbidity, then three growths have proceeded 

 from 15 x 0.04 c.c., or five from i c.c. of the water. Or, on the other hand, 

 suppose that of the beer-flasks only one has become turbid, and that this is one 

 inoculated with 0.25 c.c. In this case, then, there is but one growth per 

 10 x 0.25 = 2 5 c.c. 



H. WICHMANN (II.) attempted to add, as a co-factor influencing the con- 

 clusion arrived at, the length of time required for the turbidity to develop ; of 

 this fuller particulars will be found in the reference just given. Hansen showed 

 that a largo number of species of water-bacteria are incapable of developing 

 in the two solutions last named, and this is particularly the case with beer, the 

 flasks charged therewith seldom becoming turbid after inoculation with water. 

 J. CH. HOLM (I.) has, for several consecutive years, regularly examined the well- 



