POLLUTION OF WATER COURSES 323 
agar plates using one or two decimal dilutions and incubate at 20° C. 
for several days. All suspicious colonies must be studied morphologi- 
cally, culturally and microscopically. The cultures which pass through 
this treatment should be injected into healthy guinea pigs. Koch’s 
postulates must be satisfied before the suspicious colonies are accepted 
as anthrax. 
Bacillus Tuberculosis. The search for this organism in water has 
not been carried out to any great extent. If acid-fast bacteria are 
isolated from water, it is necessary to confirm the findings by animal 
inoculation. 
Brown’s et al. Method. Brown and his collaborators used this 
method to detect the tubercle bacillus in river water. Near the source 
of pollution (sewer outlet) large bottles of water were collected. Further 
down the stream different methods were used. Cheesecloth bags were 
stretched across the surface of the stream and allowed to remain in 
position from one to two hours, after which they were put into pails 
and carried to the laboratory. According to this method, they should 
then be carefully washed with sterile water and the wash water put 
into wide-mouth bottles, saturated with sodium chloride and allowed 
to stand in the dark for several hours; at the end of this time, the 
scum should be collected and transferred to 50 c.c. centrifuge tubes. 
Ten c.c. of NaOH are added to each tube and the tube incubated for 
one-half hour. Neutralize the contents of the tube with NHCIl, cen- 
trifugalize, and study the sediment microscopically, inoculate part into 
egg media and inject part into guinea pigs. 
Tracing Pollution in Water Courses. Different methods have been 
used and also different materials such as, sodium chloride, lithium salts 
fluorescin, and cultures of B. prodzgiosus. 
Salt was used by McCallie (1904), in Georgia, for the purpose of 
demonstrating the effects of discharging sewage into deep wells. A 
large amount of salt was put into a 124-ft. drilled well, and the content 
of chloride in the water of the surrounding wells was determined. 
Increased content of chloride showed that there was underground con- 
nection between the wells and springs in the vicinity. 
Dole (1906) summarized his investigations on the use of fluorescein 
in the following statements: 
1. In determining the sanitary value of a well or spring it is more important 
to study the underground flow than to analyze the water itself. 2. Foreign sub- 
stances put into the aquifer and traced from point to point are of great value in 
this study. 3. With the fluorescope one part of fluorescein can be detected in 
10 billion parts of water. 4. Fluorescein is a particularly valuable flow indi-. 
