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Investigations on Resazurin Test
Similarly about 7 per cent, 35 per cent and 58 per cent of the three classes of milk respectively
have failed the resazurin test, with about 30 per cent of farm produced milks and 25 per cent of the
other two classes being placed in the 'average' category. Thus the test has clearly brought out the
marked gradation in the bacteriological quality of milks drawn from the three sources which is in
conformity with their actual conditions of production and handling. Farm produced milk is obtain-
ed under hygienic conditions and is least exposed to atmospheric temperature while the other two
classes of milk have been generally produced under unhygienic conditions and carelessly handled.
In addition, the milk from private dairies represents bulk milks collected from various sources and
is most exposed to atmospheric temperature. Accordingly the latter class of milk has a high bacterial
content and is rich in coliform and other actively reducing types of bacteria which bring about a
rapid reduction of resazurin.
The down-grading of a large number of samples in the case of farm produced milk to the 'average'
(30 per cent) and 'unsatisfactory' (7 per cent) categories cannot be explained on the basis of their
conditions of production. But it has already been indicated, while comparing the resazurin test
with methylene blue reduction and plate count, that most of the samples placed in these categories
showed heavy leucocyte contents associated with abnormal milks (colostrum, late lactation milk or
mastitis) and accordingly, cellular activity may be responsible for the rapid reduction of resazurin
in the case of these samples. A separate investigation carried out into the cause of this revealed
that nearly 25 to 30 per cent of the animals in the herds, from which the samples were obtained,
were suffering from mastitis more or less of a severe form associated with streptococci, staphylococci
or corynebacteria and shedding numerous cells in the milk.
It is thus evident that the resazurin test is fairly reliable in indicating bad methods of produc-
tion, but in the case of milk produced under careful conditions and therefore having a low bacterial
content, the effect of leucocytes becomes more pronounced and a misleading picture may sometimes
be obtained. In actual practice this may not be a serious factor since cellular activity in milk does
not ordinarily reduce resazurin beyond the pink stage [Davis, 1942] and a producer's supply would
not be graded down to the rejection class (colourless) on account of cell content only. On the other
hand it may be highly advantageous in drawing attention to the presence of serious mastitis in a
herd.
SUMMARY
1. The application of the one-hour resazurin reduction test at 37°C. for judging the quality of
Indian milks has been examined by comparing it with the standard methylene blue reduction test
and plate count using some 650 samples of cow, buffalo and mixed (cow and buffalo) milks drawn
from different sources.
2. Based on their comparison with a methylene blue reduction time of 4 hours (found suitable
for conditions prevailing in this part of the country), the following modified standards for grading
milks according to the resazurin stages at one hour have been used for comparing the results of the
different tests :
(i) Blue, lilac or mauve . . . . . . . . . . . . .
(ii) Pink mauve or purple pink.
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(iii) Pink . . . . . . . . . . . . .
(iv) Colourless . . . . . . . . . . . . .
Good
Average
Poor
Bad
3. A fairly good agreement between the two reduction tests has been found. About 70 per cent
of all the samples passing resazurin test (blue to mauve stages) have also passed the methylene blue
reduction test (4 hours) while almost all the samples placed in the 'poor' or 'bad' categories (pink
or colourless) by the resazurin test have failed the former test.
4. About 30 per cent of the samples passing the methylene blue test have been placed in the
'average' category (pink mauve or purple pink) by the resazurin test while 50 per cent of the samples
failing the former test have been up-graded by the latter. The causes for these anomalies have been
discussed and the greater efficiency of the resazurin test in detecting abnormal milks with high leuco-
cyte counts (colostrum, late lactation or mastitis milks) as also its greater flexibility in placing milks