METHODS FOR THE MICROSCOPIC STUDY OF BACTERIA 203 



Ascitic and Hydrocele Fluid Media. Ascitic and Hydrocele Agar. 1 

 Collect ascitic or hydrocele fluid in a sterile bottle, using aseptic 

 precautions. Allow to stand in the ice-box until clear, and heat to 

 50 C. for half an hour to destroy enzymes. Two parts of hydrocele 

 or ascitic fluid to eight or ten parts of plain nutrient agar previously 

 melted and cooled to 45 C. make a medium especially adapted to the 

 growth of many of the more fastidious pathogenic bacteria. 2 



Ascitic broth is prepared by adding 20 to 50 per cent, by volume of 

 sterile ascitic fluid to plain nutrient broth. Incubate to prove sterility. 



Egg Media. Eggs are a very good substitute for blood serum in 

 Loffler's medium. Eggs are carefully broken into a clean beaker 

 stirred gently with a rod (avoiding the formation of air bubbles) until 

 homogeneous, and mixed with dextrose broth in the proportion of 

 one part by volume of broth to three volumes of egg. The medium 

 is coagulated in a slanted position and sterilized precisely as Loffler's 

 blood serum is coagulated and sterilized. 



Egg Medium. No. 1. Mix four to six volumes of thoroughly homo- 

 genized eggs with one volume of nutrient broth, and add sufficient 

 glycerin to make the concentration of the latter 3 per cent, by weight. 

 Coagulate and sterilize in the slanted position precisely as Loffler's 

 blood serum is coagulated and sterilized. This medium is excellent 

 for the cultivation of tubercle bacilli. 



No. 2. Add one volume of physiological salt solution to ten volumes 

 of egg which have been lightly stirred with a rod until the yolks and 

 whites are intimately incorporated. Coagulate and sterilize in a 

 slanted position in test tubes. 



Milk and Litmus Milk. One liter of fresh milk is thoroughly mixed 

 and tubed in the ordinary manner. Litmus milk is prepared by adding 

 sufficient litmus solution to impart a clea'r blue color. It is tubed, 

 using 10 c.c. to each tube, and sterilized in the autoclave. 



For some purposes it is desirable to remove the cream before tubing, 

 but for cultural work the color of the cream ring in litmus milk is of 

 some diagnostic importance. Thus, members of the paratyphoid 

 group of bacilli almost invariably show a blue-green cream ring; the 

 colon bacillus colors the cream ring red brown. It should be remem- 

 bered that litmus milk does not coagulate as readily or as rapidly as 

 plain milk. 



1 Ascitic and hydrocele fluids may be sterilized by passage through an unglazed porce- 

 lain filter. 



2 It should be remembered that ascitic and hydrocele fluids usually contain about 

 0.08 per cent, dextrose. 



