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Australian Standard
Food microbiology

Method 3.12: Examination of specific

products—Cultured dairy products

PREFACE
This Standard was prepared by the Standards Australia Committee on Food
Microbiology to supersede AS 1095.2.11 — 1978, Methods of microbiological
examination of dairy products and for dairy purposes — Methods for the examination
of specific dairy products — Cultured dairy products.

METHOD
1 SCOPE This Standard sets out microbiological methods for the examination of
cultured dairy products.
2 APPLICATION The methods apply to yogurts, other cultured milks, and sour
creams. They do not apply to frozen cultured dairy products, such as frozen yogurt,
since frozen dairy products are dealt with in AS 1766.3.4.
3 REFERENCED DOCUMENTS The following documents are referred to in this
Standard:
AS
1166 Methods for sampling milk and milk products
1766 Food microbiology
1766.1.1 Method 1.1: General procedures and techniques — Samples, materials,
equipment, laboratory practice
1766.1.2 Method 1.2: General procedures and techniques — Preparation of
dilutions
1766.2 Part 2 (series): Examination for specific organisms
NOTE: The Part 2 seri es is composed of a number of Standards each of which deals wit h the examinati on
of foods for a specifi c microorganism or group of microorganisms.
1766.3.4 Method 3.4: Examination of specific products — Frozen foods
1766.3.10 Method 3.10: Examination of specific products — Liquid milks — Direct
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microscopic count
1766.5 Method 5: Preparation of media, diluents and reagents
2300 Methods of chemical and physical testing for the dairying industry
2300.1.6 Method 1.6: General methods and principles — Determination of pH
4 CULTURE MEDIA AND DILUENTS The following culture media and diluents
are required:
(a) Culture media and diluents specified in the documents referred to in this Standard.
(b) 0.1 percent peptone solution (see AS 1766.5).
(c) Sodium citrate, 20 g/L aqueous solution.
(d) Ammonium hydroxide, 150 g/L aqueous solution.
5 APPARATUS
5.1 Sampling equipment The following sampling equipment is required:
(a) Sample containers, spoons and other items used for subsampling laboratory
samples.
(b) Blender or stomacher.

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5.2 Sterilization of sampling equipment Sampling equipment shall be sterilized as
specified in AS 1766.1.1.
NOTE: Sterile disposable apparatus is an acceptable alternative to glassware if it has a similar specification.
6 SAMPLING AND SAMPLES
6.1 General Sampling and sample preparation are specified only as they apply to the
sample for test received at the laboratory.
NOTES:
1 It is assumed for the purposes of these methods that the laboratory sample is representative of the bulk of
the subject material.
2 Methods for rate of sampling and technique of sampling of dairy products are prescribed in AS 1166. The
size of the sample taken in accordance with AS 1166 may be adjusted in order to provide a sufficient
quantity for the required tests.
6.2 Storage of laboratory samples The laboratory samples shall be maintained at a
temperature of less than 5°C, but without freezing, until tested. Testing shall commence
within 24 h of receipt.
7 PROCEDURE
7.1 Opening retail containers Observe the physical condition of the container and record
any abnormalities.
Liquid products may need to be mixed before the retail container is opened (see
Clause 7.3.1).
Clean the surface of the container, and swab particularly around the seal, with cotton wool
soaked in 70 percent ethanol. Allow the container to air dry. Aseptically open the container
to a sufficient extent to allow the product to be removed without contaminating the sample.
7.2 Observation of physical condition Observe the physical condition of the product and
record any abnormalities.
7.3 Preparation of test sample
7.3.1 Liquid cultured milks (e.g. flavoured drinking yogurt, cultured buttermilk) Shake the
sample 25 times in about 12 s through an arc of about 30 cm. If the headspace is
insufficient for efficient mixing, aseptically transfer the contents to a larger container for
mixing.
7.3.2 Plain or flavoured yogurt Stir the sample thoroughly with a spoon or spatula.
7.3.3 Products containing particulate matter (e.g. fruit yogurt) Transfer the entire contents
of the container to a blender or stomacher bag and mix the sample for 1 min. For samples
which are too large to be accommodated by the mixing apparatus, stir the sample thoroughly
with a spoon or spatula and then transfer as much as possible to a blender or stomacher for
mixing.
7.3.4 Sour creams Stir the sample thoroughly with a spoon or spatula and transfer to a
screw-capped jar. Warm the sample to about 45°C, but not above, in a waterbath and,
without delay, mix by stirring. Prepare dilutions (see Clause 7.4) immediately after mixing.
NOTE: Total time at 45°C should not exceed 10 min.
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7.4 Preparation of dilutions Dilutions of the test sample shall be prepared as follows:
(a) Weigh 10.0 ±0.1 g of sample into a wide-mouthed dilution bottle.
(b) Except for sour cream samples, add 90 mL of 0.1 percent peptone solution (4(b)). For
sour cream samples, add 90 mL of 2 percent sodium citrate solution (4(c)) warmed to
45°C (see Note to Clause 7.3.4).
(c) Stopper the bottle and mix the contents by shaking 25 times in about 12 s through an
arc of 30 cm.
(d) Prepare further decimal dilutions using 9 mL volumes of 0.1 percent peptone solution
(4(b)), as described in AS 1766. 1.2. For sour cream, prewarm the peptone solution to
45°C (see Note to Clause 7.3.4).
7.5 Direct microscopic examination of test sample The procedure shall be as follows:
(a) Prepare a 1:10 dilution of the test sample in 0.1 percent peptone solution (4(b)) in a
test tube.
(b) Add 1 or 2 drops of 15 percent ammonium hydroxide to the tube and mix gently.
(c) Smear a thin film of the diluted product onto the surface of a clean slide.

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3 AS 1766.3.12—1992
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(d) Dry the film at a temperature no higher than 45°C in a dust-free atmosphere in a
horizontal position.
(e) Stain the smear with methylene blue staining solution using Method B of
AS 1766.3.10.
(f) Examine under a microscope and note the cell types in relation to the culture expected
to be present. Report any probable contaminants.
NOTE: For yogurt, the ratio of streptococci to lactobacilli can be determined by carrying out, after
Step 7.5(b), a direct microscopic count using Method B of AS 1766.3.10. Count a minimum of five fields,
but count sufficient to give a total of greater than 200 bacterial clumps. Calculate the ratio and note the
number of fields counted.
7.6 Tests for specific organisms To examine the test sample or first dilution for specific
organisms, the appropriate test methods of the AS 1766.2 series shall be used.
7.7 pH determination The procedure specified in AS 2300.1.6 shall be used.
8 TEST REPORT The following information shall be reported:
(a) All details necessary for the complete identification of the sample.
(b) Date of testing.
(c) Reference to this Australian Standard, i.e. AS 1766.3.12.
(d) The results of the tests.
(e) Any circumstances or conditions which may have influenced the test results.
(f) Any abnormality observed in the physical condition of the container or the product.
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AS 1766.3.12—1992 4
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This Australian Standard was prepared by Committee FT/4, Food Microbiology. It was approved on behalf of the
Council of Standards Australia on 7 July 1992 and published on 12 October 1992.

Review of Australian Standards. To keep abreast of progress in industry, Australian Standards are subject to
periodic review and are kept up to date by the issue of amendments or new editions as necessary. It is important
therefore that Standards users ensure that they are in possession of the latest edition, and any amendments thereto.
Full details of all Australian Standards and related publications will be found in the Standards Australia Catalogue
of Publications; this information is supplemented each month by the magazine ‘The Australian Standard’, which
subscribing members receive, and which gives details of new publications, new editions and amendments, and of
withdrawn Standards.
Suggestions for improvements to Australian Standards, addressed to the head office of Standards Australia, are
welcomed. Notification of any inaccuracy or ambiguity found in an Australian Standard should be made without
delay in order that the matter may be investigated and appropriate action taken.

First published as AS 1095.2.11— 1978.

Revised and redesignated AS 1766.3.12— 1992.

This Standard was issued in draft form for comment as DR 88088

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