XVIII] M.
A.
KRISHNASWAMY
AND
H.
LAXIMINARAYANA
233
Presumptive test. Three successive tenfold dilutions of the milk sample (corresponding to its
general level of bacteriological quality) were prepared in saline water in the usual manner, i.e., by
adding one ml. of milk to 9 ml. of saline water for making 10- 1 ml. dilution and preparing further
dilutions from the latter. Where the dilution series started from 10- 2 ml., one ml. of original milk
was added to 99 ml. water to make up 10- 2 ml. dilution and subsequent dilutions made from it in
the same manner. One ml. portions of each dilution were inoculated into the three broth (liquid
media) tubes in triplicate and also plated in duplicate on the four solid media. After the agar had
selidified in the petri dishes about 4 ml. of the same medium was peured over it in order to prevent
the formation of atypical surface colonies. The broth tubes as well as the plates were incubated
at 37°C. and examined after 24 hours and 48 hours in the beginning. All the liquid madia which
were to show the production of acid and gas did so invariably in 24 hours and there was no advantage
in Contiuning the incubation for 48 hours. In the case of plates, the maximum number of colonies
were obtained in 24 hours and further incubation only resulted in an increase of the size of the colonies
and in many instances it permitted the growth of certain slow acid producers which gave misleading
results. Accordingly, after making observations at the end of 24 hours, only such of the tubes and
plates as gave negative or doubtful results were incubated for further 24 hours and observed. None
of them, however, gave any positive reactions at the end of 48 hours. The production of acid and
gas (more than 10 per cent in the Durham's tube) in at least two out of the triplicate broth tubes and
the appearance on the plates of typical dark-red or brick-red colonies of at least 0.5 m.m. in diameter
and not less than five (average of duplicates) in number were taken as positive presumptive evidence
of the presence of coliform organisms in any particular dilution. The total number of coliform
colonies on the plates was also counted.
Completed test for confirmation, of the presumptive evidence. All the positive fermentation tubes
as well as selective agar plates in each of the dilutions were subjected to the confirmatory test accord-
ing to the procedure recommended by the American Public Health Association [1941]. In the case
of positive plates showing more than five colonies any two of the typical colonies were picked up
for confirmation.
The results of the above tests parried out on 53 samples of milk (induing farm-produced milk,
village milk, bulk samples handled in dairies and pasteurised milk) are given in Tables I and II.
RESULTS
A comparison of the combined results of the presumptive coliform test in different media for
all the samples grouped according to the dilutions at which positive reactions were indicated is given
in Table I. In order to bring out clearly the comparative efficiency of each medium in giving a clear
reaction in 24 hours, in the same material, the number of samples positive or negative in MacConkey's
broth at different dilutions have been separately indicated in two series. Taking them as standard,
the number of corresponding samples in each group showing an identical reaction in the other media
is indicated. Since all the presumptive positive tubes as well as plates were positive to the
confirmatory test, no separate column is provided in Table I for illustrating this point.
It will be seen (Table I) that practically all the samples, whether showing a positive or negative
presumptive test in MacConkey's broth, have given similar reactions in the other two liquid media.
The few discrepancies observed may be due to sampling errors. There were no false positives en-
countered as the results of the presumptive test in all the media were confirmed without exception:
This is in accordance with the observation of Wilson et al [loc. cit.] who did not experience this trouble
in the case of MacConkey's broth in a large number of trials with water and milk, while Farrell [loc.
cit.] mentions the occurrence of false positives in the case of brilliant green bile broth.
As regards the solid media, the number of positives recorded by them is somewhat lower than
those indicated by MacConkey's broth in the corresponding dilutions. This anomaly may be due
to the limit of five or more colonies taken as positive evidence being too rigorous a condition, because
samples giving even one colony on the plate may show development of acid and gas in the liquid
medium. In a number of cases giving a positive test in MacConkey's broth the corresponding plates
have shown one, two, three or four colonies which have not been taken into account. However,