PATHOGENIC BACTERIA. 305 



for thirty minutes; fill up water lost by evaporation; press through gauze; 

 measure; add 6 per cent. Witte peptone, J per cent, salt; heat till peptone 

 dissolves; filter; distribute into sterilized beer bottles with patent stoppers; cover 

 with paper cones; sterilize two hours; clamp the stoppers to, and store away. 

 (J) Enriching fluid: 100 c.c. of the above stock solution measured in a 

 sterilized measuring glass; put in a. sterilized Erlenmeyer flask; add sodinm 

 hydrate solution, , 2.7 c.c. less than the quantity required for phenolphthalein 



"red point," as determined by neutralizing 25 c.c. Sterilize ten minutes in 

 steam; allow to get cool; add 105 c.c. of a 1.2 per cent, solution of cafi'eine 

 (solution to be made fresh in cold, sterilized, distilled water every time). Add 

 1.4 c.c. of a j^j per cent, solution of crystal violet ; crystal violet must be dis- 

 solved cold. 



(c) Preparation of the stool: 



1. Thin stool allowed to settle, eight-tenths or nine-tenths c.c. of the thin, 

 upper portion added to 6. 



2. Semifluid stool rubbed up in a mortar with i part of 1.2 per cent, 

 solution of caffeine; filter through sterilized cotton-wool; eight-tenths or nine- 

 tenths c.c. of the filtrate added to 6. 



3. Thick feces. Rub i part of feces with 2 parts of caffeine solution, and 

 proceed as in No. 2. 



In all three cases shake thoroughly and place at 37° C. 



id) Search for typhoid fever bacillus. Examine a hanging-drop. 



1. If there are relatively few bacteria, make 6 large Drigalsky agar plates: 

 plate No. I of the series with 0.30 to 0.35 c.c. of the fluid; plate No. 3 with 

 0.25 c.c; plate No. 5 with o.io c.c. Plates Nos. 2, 4 and 6 are the diluted 

 plates from Nos. 1, 3 and $• 



2. If there is abundant growth, 7 plates are made: i, 4 and 6 are inoculated 

 with 0.2 c.c, 0.15 c.c. and o.i c.c, respectively, and the others are dilutions 

 from these. 



Identification of colonies as usual. 



Keep the rest of the culture in the enriching fluid on ice. If the first plates 

 fail for any reason, shake this enriching fluid with glass beads and make 

 Drigalsky plates again. 



A simpler way of preparing the Drigalsky-Conradi medium is recommended 

 by Hagemann* as follows: 



Liebig's extract, 10 grams; Witte's peptone, 10 grams; sodium chloride, 

 10 grams; water, 600 c.c. Boil in a salt-water bath until 100 c.c. evaporates 

 off. Add 500 c.c. fresh, raw, amphoteric milk. Boil and add agar, 10 grams. 

 Boil until the agar is nearly dissolved; put in the autoclave for twenty to thirty 

 minutes at 110° to 115° C. Filter in the streaming steam. Divide up into 

 sterile Erlenmeyers, about 200 c.c. in each. Sterilize a short time. 



* Hagemann. Eine Vereinfachung des Drigalskischen Nahrbodens. Hy- 

 gienische Rundschau. Vol. XIV. 1904. 



26 



