27
TABLE I.
Nu mb e r .
Date of
collection of
sample.
Sample of water
taken from.
Definite quantities of water taken and mixed with different quantities
of agar and poured into plates to get different depths.
Thick.
Thin.
1914.
1 23rd Nov. . . . 1st tap . . .
A spreading colony and thirty-five
colonies.
A spreading colony and eight
colonies.
2
Do. . . . 2nd " . . .
Countless . . .
Countless.
3
Do. . . . 3rd " . . .
A spreading colony and forty-nine
colonies.
196
4
Do. . . . 4th " . . .
120 . . . A large number of tiny colonies in
the centre and forty-one discreet
colonies.
5
Do. . . . 5th " . . .
Cannot be counted. A spreading
colony and six colonies.
160
6
Do. . . . Outlet discharge
pipe.
Cannot be counted. A spreading
colony and sixty-two colonies.
152
7 25th Nov. . . . 1st tap . . .
Cannot he counted. One spreading
colony.
1,400
8
Do. . . . 2nd " . . .
400 . . . 1,200
9
Do. . . . 3rd " . . .
248 (?) Not easily countable . . . 338
10
Do. . . . 4th " . . .
A spreading colony and fifteen
colonies.
600
11
Do. . . . 5th " . . .
352 . . . 204 and a big colony in centre.
12
Do. . . . Outlet discharge
pipe.
Cannot be counted. One spreading
colony.
232
13 27th Nov. . . . 1st tap . . .
1,000. A tree-like colony occupying
a third of the plate.
A spreading colony and s ix other
colonies.
14
Do. . . . 2nd " . . .
1,200 . . . 1500
15
Do. . . . 3rd " . . .
220 . . . Two woolly colonies in 1/3 of the plate
and thirteen other colonies.
16
Do. . . . 4th " . . .
310
.
.
. 520
17
Do. . . . 5th " . . .
Twenty-one and a group of small
colonies in centre.
310
18
Do. . . . Outlet discharge
pipe.
Cannot be counted. A spreading
colony.
Forty-four and a spreading colony
(450).
19 30th Nov. . . . 1st tap . . .
600
.
.
. 800
20
Do. . . . 2nd " . . .
Two and a large number of tiny col-
onies in the centre.
450
21
Do. . . . 3rd " . . .
210
.
.
. Ten colonies and a spreading colony.
22
Do. . . . 4th " . . .
One colony and a big spreading
colony.
Thirty-four colonies and a spread
-
ing one.
23
Do. . . . 5th " . . .
Twelve colonies and a spreading
colony.
260
24
Do. . . . Outlet discharge
pipe.
180
.
.
. Ten colonies and a big spreading
colony.
Table I records experiments which show the difference in the total number of colonies
growing in the same quantity of water on agar of different depths.
3. A small quantity of the material examined (water inoculated into melted agar) remains
adherent to the test tube and may contain a variable number of organisms. To obviate this
source of error it was proposed to incubate the emptied tubes also and to count any colonies
which may grow thereon. At 37° C however the agar dries and shrivels up and colonies do
not develop. A new method of estimating the number of organisms in a definite quantity of
water, which is free from most of these fallacies, has of late been tried in this laboratory.
The method consists in preparing :
—
(1) Dry sterile agar plates. Solid sterile agar is melted in bulk and distributed into
sterile plates using a liberal amount filling the plate to a depth of not less than one-eighth of an
inch in thickness. The plates are now incubated at 37°C for 24 to 48 hours. This is really for
the purpose of drying off the condensation water, but any plates which show colonies are
rejected. When only one small colony is seen on the surface of the agar, it may sometimes
be carefully removed with a previously sterilised small wire without farther contaminating the
plate. The plates are then removed and stored in a box.
(2) Other requirements are sterilised graduated 1 cc. pipettes for measuring and
diluting ; tubes containing 9 cc. of sterilised distilled water for making one-tenth dilutions.
and sterilised spreaders of the ordinary bent glass rod pattern. If, from previous experience,
the water sample is suspected to contain a comparatively small number of colonies, half a cc.
of the sample itself is taken and dropped over the agar surface of a plate and spread over
the entire surface evenly by means of the sterile spreader. The same spreader is carried on to
a second plate and spread over the agar surface. Generally, two plates will be enough, but if
necessary, a third plate may also be used. If the water is of an inferior quality, and suspected
of containing a fairly large number of colonies, 1 cc. of the sample is added to 9 cc. sterile
distilled water tube thus making a tenth dilution. Similarly a hundredth dilution may be
made by taking 1 cc. from the first dilution and adding to 9 cc. of another distilled water
tube. Half a cc. from each of these dilutions may be smeared as per previous description.
By taking the precaution of thoroughly rinsing the pipette with each dilution, the same
pipette may be used throughout. These smeared plates are incubated at 37° C for 48 hours.
The total number of colonies that grow from 1 cc. of water is then calculated.
8