CHAPTER II.
A DETAILED DESCRIPTION OF THE METHOD EMPLO YED.
As it is hoped that the publication of this work will induce officers in other
parts of India, to carry out their water-analysis on these lines, so that comparable
results may be obtained, we give a detailed account of the method, which has been
adopted as standard in the laboratory of the King Institute, for all water samples that
we intend to examine completely. We wish at this point to say that we have been
greatly assisted. by the kind advice of Dr. A. MacConkey himself, the originator of this
method., and we are also indebted to Dr. A. C. Houston, Bacteriologist to the Metro-
politan Water Board, Loudon, for many -practical hints, as to the best methods of
working through a large number of samples with as little delay as possible. We are
greatly indebted to these two gentlemen and we desire to tender. them our sincere
thanks.
All samples of water are taken by a special sample taker (a certificated Sanitary
Inspector on Rs. 25 per month) who is a servant of the King Institute, and who visits
the various supplies himself. Samples are taken in sterile bottles, and are immediately
placed on ice ; 250 cc. of water being sufficient for bacteriological purposes, and a
Winchester quart for chemical. The practice of sending bottles of water for
b a c t e r i o l o g i - c a l a n a l y s i s b y t r a i n h a s b e e n g i v e n u p a s p e r f e c t l y h o p e l e s s i n t h i s c o u n t r y . I t i s n o t
necessary for me to point out the many possibilities of delay, and the accidents, that may
occur to a sample that is sent by train ; we have always held that the only satisfactory
way of obtaining samples for this work, is to send our own man for them, and provide him
with the necessary quantity of ice. The only objection to this method is on the score
of expense. But with forethought, this need not be excessive, for two sets of samples
can sometimes be brought on a single journey: The budget allotme
nt
for the travelling
expenses and contingencies of the sample taker of the King Institute, who visited all
the, water-supply schemes in the Presidency three times during this year, and took a
large number of extra samples for research purposes, was only Rs. 1,300.
On arrival at the laboratory three separate series of bacteriological analyses are at
once started, the bottles having been well shaken.
These are (i) total colonies count in 1 cc. of .water, (ii.) sporogenes enteriditis
milk test in 20 cc. of water, and. (iii) inoculating of tubes or MacConkey's bile
salt broth with various quantities of water : —
(1) Total Colonies in 1 cc.—A count of the total colonies in 1 cc. of water is made
on agar. The medium used for this is very carefully prepared, so as to get each brew
of the same reaction and strength. Lemco½ per cent., Witte's peptone 1 per cent., is
used as the nutrient media, with 2 per cent. of agar, and a standard of acidity of + 10
to phenol phthalein is allowed. One cc. and 1/10 c c . o f a w a t e r i s t a k e n a n d r u n i n t o
the melted agar, when it has cooled down to a temperature of 38 0 . This is plated out
in the usual way in a petries dish, incubated at 37 0 for 48 hours, and all colonies that
are visible to the naked eye are counted. These results, though they are extremely
interesting, especially when comparing one sample of the same supply with that of
another, cannot be compared with total counts obtained on gelatine. The climate of
this country renders gelatine as a solid medium extremely difficult to work with. The.
incubator that will maintain a temperature of 19° Centigrade constantly, through all
the vagaries of the Indian climate on the plains, does not appear to have been designed
yet. Obviously therefore it is better to have a method of doing total colony counts
that is equally satisfactory in both hot and cold weather, and, provided that care be
taken with the manufacture of the solid agar media, so that each batch is as nearly as
possible like the previous one, the results obtained are valuable and can be compared
with each other with a reasonable expectation of concordance.
(2) Sporogenes Test .—Twenty cc. of water are put into sterile milk in order to
test for the presence of spores of bacillus sporogenes enteriditis. The tubes are heated