102 METHODS OF PURE CULTURE 



upon the medium, where they rapidly develop into such masses of branched 

 threads that the bacterial colonies are smothered, thus rendering all the care 

 bestowed upon the preparation of no avail. In order to prevent this, the flat 

 plates are replaced by double shallow glass dishes, in which the cultures can be 

 examined under low powers without being exposed to the air. These dishes 

 were first introduced into bacteriology by Salomonsen, but are generally known in 

 Germany as Petri dishes, this latter worker having been the first to test them. 

 Their use for this purpose can be recommended. Instead of pouring out the in- 

 oculated gelatin, the closed tube can be held almost horizontally under the stream 

 from the water-tap and slowly turned round on its axis, whereby the contents 

 are distributed uniformly over the walls, and will set as a thin stratum wherein 

 the germs then develop into colonies. These cultures are generally called Esmarch 

 tubes or roll cultures, and were first proposed by W. Hesse. 



Pkte cultivation affords useful assistance, not only for the separation of a 

 bacterial mixture into its several species, but also for the determination of the 

 number of cells present therein, a few gelatin tubes being charged with various 

 quantities of the sample and poured on to plates. This method is of particular 

 importance in the quantitative bacteriological analysis of water, for which 

 reference should be made to Tiemann-Gartner's handbook. The counting of the 

 colonies grown on the glates is effected by the aid of special counting apparatus, 

 that of Wolffhiigel being used for the Koch plates. For counting the colonies on 

 gelatin plates in Petri dishes the author, in 1893, constructed a cheap counting- 

 plate, obtainable from F. Mollenkopf, of Stuttgart (10 Thor Strasse). The 

 number of germs thus found is always smaller than the living cells actually 

 present in the inoculating mixture, since only such as have developed into 

 colonies are enumerated, whereas a number of germs in the original have failed 

 to develop under the conditions prevailing, owing to the medium being unsuit- 

 able for some, and the temperature of the incubator, though favourable to the 

 majority, being too hot or too cold for a minority. The medium relatively most 

 suitable for the purpose of ascertaining the number of germs is, in most cases, 

 gelatinised meat-juice, and this is therefore the one most frequently used. Con- 

 siderable influence on the number of developing germs is exerted by the degree 

 of alkalinity of the medium, a fact first conclusively demonstrated by A. REINSCH 

 (II.) and confirmed by MAX DAHMEN (I.). Jf it is a question not of ascertaining, 

 as nearly as possible, the total germ content of a sample, but only how many of 

 the cells are capable of development in a given medium, then the latter is arranged 

 in a solidified condition as a plate culture. For example, wort gelatine is generally 

 used unless the contrary be expressly stated when determining the number of 

 germs in brewery water. It is important to know for certain whether the 

 colonies in a plate culture are each developed from a single cell, since it is only 

 in such cases that a pure culture can be obtained on re-inoculation. This aim is 

 attempted by thin sowing and thoroughly shaking the liquefied medium, in 

 order to separate the cells from each other. Nevertheless, there is always some 

 uncertainty, which we must endeavour to remove by discarding the first series 

 of plates and by preparing a second series wherein any impurities may become 

 manifest ; then, if the colonies are found to stand this test, the reinoculations 

 therefrom may be considered as pure cultures. This can be ensured from the 

 outset if the growth of the colonies, i.e. from the single cells, be followed by the 

 aid of the microscope from the beginning. This test is, however, feasible only 

 with largo cells (Eumycetes spores, yeast cells, <fec.), and will be enlarged upon 

 in a subsequent chapter in connection with the pure culture of yeast. On the 

 other hand, it is, as a rule, impracticable for bacteria, since their examination 

 necessitates the use of such a high (short focus) objective that the latter has to be 

 brought so near the plate as to impinge on the gelatin stratum. 



