PATHOGENIC BACTERIA. 



375 



maleml for examination for the typhoid bacillus before making the Drigalsky- 

 Conradi plates. They use an enriching fluid which has the property of in- 

 hibiting the growth of colon and other contaminating organisms, while not 

 seriously interfenng with the growth of the typhoid bacillus. 



This fluid consists of: 



(a) A stock solution of beef-broth. Take one kilogram of chopped beef; 

 add 2 hters of distiUed water; heat thirty minutes at 50° to 60° C; stir; boil 

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

 measure; add 6 per cent, peptone (Witte); J per cent, salt; heat till peptone dis- 

 solves; falter; distribute into sterilized beer bottles with patent stoppers; cover 

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



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

 sterilized measuring glass; put in a sterilized Erlenmeyer flask; add sodium 

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

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

 steam; allow to get cool; add ros c.c. of a 1.2 per cent, solution of cafl^eine 

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

 1.4 c.c. of a jV per cent, solution of crystal violet; crystal violet must be dissolved 

 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 b. 



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 b. 



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. 



(d) 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. I, 3 and s- 



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. 



In using, melt in a water-bath; add normal sodium hydrate till the reaction 

 is slightly alkaline to htmus-paper. Add 20 c.c. Merck's Htmus solution. Also 

 add 3 drops of a i per cent, alcoholic solution of crystal violet. Mix thoroughly. 

 Pour into Petri dishes, and use as in the original method. 



*Hagemann. Eine Vereinfachung des Drigalskischen Nahrbodens. Hygie- 

 nische Rundschau. Vol. XIV. 1904. 



