140 DIFCO MANUAL 



this medium the typhoid organism grows luxuriantly, forming characteristic black 

 colonies, while the Gram-positive bacteria and members of the coliform group 

 are inhibited. This unique inhibitory action of Bacto-Bismuth Sulfite Agar toward 

 Gram-postive and coliform organisms permits the use of a much larger inoculum 

 than has been possible with other media employed for similar purposes in the 

 past. The use of larger inocula greatly increases the possibility of recovering the 

 organisms, especially when they are present in relatively small numbers, such as 

 may be encountered in the early course of the disease or in the checking of car- 

 riers and releases. 



Cope and Kasper^ increased their positive findings of typhoid from 1.2 to 16.8 

 per cent among food handlers and from 8.4 to 17.5 per cent among contacts by 

 the use of Bacto-Bismuth Sulfite Agar. Employing this medium in the routine 

 laboratory examination of fecal and urine specimens these same authors^ obtained 

 40 per cent more positive isolations of S. typhosa than were obtained on Endo 

 Medium. Gunther and Tuft,^° employing various media in a comparative way 

 for the isolation of typhoid from stool and urine specimens, found Bacto-Bismuth 

 Sulfite Agar most efficient. Upon this medium they obtained 38.4 per cent more 

 positives than on Endo, 38 per cent more positives than on Eosin Methylene Blue 

 Agar, and 80 per cent more positives on Bismuth Sulfite Agar than on the des- 

 oxycholate media. These workers found Bacto-Bismuth Sulfite Agar to be supe- 

 rior to Wilson's original medium, being easier to prepare, relatively more stable 

 and more sensitive. Green and Beard,^^ using Bacto-Bismuth Sulfite Agar in their 

 studies on the "Survival of E. typhi in Sewage Treatment Plant Processes," 

 claimed that this medium so successfully inhibited sewage organisms that their 

 interference was negligible. Beard^^ stated that such a highly selective medium as 

 Bismuth Sulfite Agar made possible the study of the survival of typhoid in nature. 



Since these earlier references to the use of Bismuth Sulfite Agar, this medium 

 has been generally accepted as routine for the detection of incitants of enteric 

 disease. The value of the medium is demonstrated by the many references to the 

 use of Bismuth Sulfite Agar in scientific publications, laboratory manuals and 

 texts. 



As surface and subsurface colonies on Bismuth Sulfite Agar are strikingly char- 

 acteristic, it is possible to use the medium both as a smear plate and as a poured 

 plate in the isolation of ^S". typhosa. Smear plates are prepared by pouring 15-20 

 ml. quantities of the medium into sterile petri dishes (90 mm.) and allowing the 

 medium to solidify with the cover removed to obtain a dry surface. In preparing 

 poured plates the inoculum is placed in the sterile petri dish and the dissolved 

 medium at 45 °C. is added and mixed in the usual manner. 



The following technique is recommended for the isolation of typhoid organ- 

 isms from fecal specimens. 



Method of Inoculation 



Streak or Smear Plate 



Streak or smear the surface of a plate with a heavy inoculum of the fecal 

 material in such a way that on some portion of the plate the inoculum will be 

 light, permitting the development of discrete colonies. 



Poured Plate 



(a) Transfer about 2 or more grams of the fecal material to a test tube, add 

 12-15 ml water, and mix well, being careful to break up all the larger particles 

 of the material. Specimens preserved in glycerol must be diluted with water to 

 reduce the glycerol content since S. typhosa in poured plates is inhibited in the 

 presence of 2 per cent glycerol. 



