430 SPIRILLUM CHOLEIUE ASIATICS. 



slowly. Where there is sufficient space between the 

 individual colonies the funnels measure at their surface 

 scarcely 1 mm. in diameter after 48 hours growth at 

 22 C., and even after 72 hours their extent is not 

 markedly greater. Under the microscope the appear- 

 ance is less characteristic after the commencement of 

 the liquefaction. The margin of the area of liquefaction 

 is circular, and has, for the most part, a sharp outline ; 

 further inwards a grey circular zone is formed which 

 contains fluid and small fragments of the colony ; in the 

 centre the latter appears as a yellowish-brown, dull, 

 irregularly granular disc with indistinct outline. 

 i -nurture Puncture cultivations in gdalino show, after 24 to 48 



hours, a whitish turbidity along the track of the ncnllr, 

 and around this a slight amount of liquefaction. In 

 this way a thin tube is formed, which contains at its 

 peripheral part almost clear fluid, and at its central 

 part the original whitish thread. Towards the surface 

 of the gelatine this tube dilates in a funnel shape, the 

 upper diameter of which has attained tin; cxl,ciit of 

 about cm. after 48 hours. This funnel is also filled 

 with fluid, the level of which is often markedly lower 

 than the surface of the gelatine, so that the upper por- 

 tion of the funnel only contains ;iir ; this gives rise to 

 the impression, on cursory examination, as if an air 

 bubble filled the uppermost portion of the funnel. This 

 appearance, however, is not observed in every tube; 

 slight variations from the above description arc found 

 according to the concentration of the gelatine, the 

 temperature at which it is kept, the mode in which the 

 puncture is made, and the amount of material intro- 

 duced. In like manner the one or the other culture 

 characteristic is found in other species of bacteria, and it 

 is only the sum total of the characters mentioned which 

 enables us to differentiate with certainty the comma 

 bacilli from other species. 



After three or four days the puncture cultivation shows 

 a moderate extension of the funnel and of the tube, and 

 thus a slow increase of tin-, liquefaction; it is not till 

 after 4 to 6 days that the liquefaction has occurred 



