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An overview of the new ISO 11133:2014 from a quality

assurance perspective

What can laboratories expect?

Intention It should be noted that laboratories purchasing pre-

poured ready-to-use plates are not required to run
ISO 11133:2014 focuses on the quality assurance
such extensive QC testing, as the media is supplied
of culture media, as well as specifying preparation,
following a comprehensive quality programme by
production and storage practices. Despite the
the manufacturer as outlined later in his paper.
availability of rapid detection methods, culture
media are used in all microbiological tests; either
as a complete stand-alone workflow, or to provide
The Revised Standard
enrichment and confirmation. Subsequently,
culture liquids and solids are critical aspects of ISO 11133:2014 was published on 15th May
all microbiology laboratories, and as such must 2014, and corrected on 1st November 2014. It
be fit for purpose, consistent and reproducible. amalgamates and supersedes ISO/TS 11133-
By demonstrating the performance of each 1:2009 and ISO/TS 11133-2:2003, but also
batch of culture media produced against a set of incorporates ISO 9998:1991 for the evaluation of
internationally recognised criteria, laboratories water quality testing media. The standard is highly
can reliably perform microbiological assessments. recognised by accreditation bodies. Key changes
This new standard is normative and is therefore and considerations include:
mandatory.
The incorporation of water testing into the same
standard as food and feed (Annex F)
The standard is now mandatory rather than
Target Audience
informative
The standard applies to media intended for the Diluents and transport media also require
microbial analysis of food, animal feed and water. performance testing
This also extends to include media intended for The specific culture strains and corresponding
the microbial analysis of samples from related WDCM codes to be used
production environments, as well as water used Dilution to extinction for liquid media is required
in food production. The standard is to form an The number of passages for culture strains is
essential part of these laboratorys quality control specified
procedures; should they be laboratories preparing Reference mediums are specified
their own media in-house from a dehydrated
format, or suppliers of dehydrated, or ready-to-
use media for these applications. ISO 11133:2014 Test Micro-Organisms and Performance
is applicable to both qualitative and quantitative Criteria
methods, liquid and solid media, selective and non-
Annex E provides tables with the culture medium,
selective.
test micro-organisms, the WDCM collection number

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and incubation instructions which must be used meet specificity criteria. An analysis of selectivity is
for each media. This helps ensure standardisation still required as described above.
across laboratories and technicians, and will
Working cultures at the appropriate concentration
highlight any batch variations within the media.
for each micro-organisms will be made and plated
For further clarity, the characteristic reactions
onto Baird Parker medium as well as the reference
and morphology of the target organisms are also
agar (TSA), and incubated accordingly. For
specified in Annex E. By following the template in
quantitative testing, a range of 80 to 120 cfu of
table E.1, together with the instructions provided
target strain per plate is required. For selectivity
in the standard concerning inoculation preparation
testing an inoculum level of 104 to 106 cfu the non-
and concentration levels, laboratories can see which
target is needed, and for specificity testing this is
procedure must be followed according to the media.
103 to 104 cfu. Colony counts for each plate will be
made and the morphology of the colonies assessed.
QC Requirements for a Qualitative Media: When assessing productivity, the colony counts
Half-Fraser as an Example of the two plates will be used to calculate the
productivity ratio. Baird Parker has an acceptance
An example of the performance testing required criteria of 0.5, showing that the recovery from the
for a liquid selective enrichment media; Half Fraser new batch of culture medium is comparable to the
broth. recovery of the non-selective reference medium.
Table E.1 in the ISO tells the laboratory who is When considering specificity, the morphology of
assessing the performance of Half Fraser Broth, the specified non-target organisms on the selective
that for productivity testing they should use Listeria agar will be recorded to ensure that it mirrors the
monocytogenes (WDCM 00021), combined with characteristic reaction described in Annex E.
Escherichia coli (WDCM 00012 or 00013) and
Enterococcus faecalis (WDCM 00009 or 00087) as
the target and non-target control strains. 10ml of the Possible Reasons For QC Failure of Media
Half Fraser should then be spiked at an inoculation
level of 100 cfu of the target micro-organism, and Should the acceptance criteria not be achieved for
1,000 cfu of the non-target organism is required any of the assessments, the specific batch of media
to be mixed in the same tube. The inoculated culture cannot be used for microbial analysis for food, feed
media must then be incubated at 30oC +1 oC for 24 or water samples. This may occur for a number of
hrs +2 hrs, before streaking one 10 l loop from the reasons, such as:
tube onto Listeria Agar according to Ottaviani and Detrimental pH shift due to overheating
Agosti. These plates will be incubated according to Dirty glassware
the ISO method. Degradation of the media due to incorrect
Selectivity testing for the same Half Fraser broth storage conditions
will be performed in parallel using tubes of the Degradation of the media due to expiry
broth spiked with the two non-target organisms Addition of the wrong volume of water
individually. Once these have been incubated as The use of impure water
described earlier, one 10 l loop from each tube will Destruction of labile compounds through
be streaked onto TSA. improper autoclaving
Inadequate mixing
Productivity of Half Fraser broth is considered
satisfactory if 10 colonies of Listeria
monocytogenes grow on the Listeria Agar, whereas
Media Suppliers
as for selectivity, total inhibition of Escherichia coli,
and partial to complete (<100cfu) inhibition of It is mandatory for manufacturers of commercially
Enterococcus faecalis is expected. available dehydrated and ready-to-use culture
media to comply to ISO 11133:2014. They must
follow the same criteria, however, the panel of
QC Requirements for a Quantitative Media: control strains they must include is far more
Baird-Parker as an Example stringent and comprehensive. For instance, for the
qualitative example above with Half Fraser Broth,
Although the format is similar, a couple of key
in addition to the productivity testing mentioned, a
differences can be seen when considering Baird
manufacturer will also have to test a second control
Parker as an example of a solid selective media for
strain of Listeria monocytogenes (WDCM 00109).
QC.
Commercial media manufacturers, Lab M, already
Once again Table E.1 defines which micro-organisms
have a quality programme in place according to ISO
must be used and how they must be incubated. The
9001, issuing quality certificates with the media they
difference is in the acceptance criteria, as in this
supply, and have enhanced their practices to comply
example the productivity ratio must be calculated,
with ISO 11133:2014. Further to this and the new
and the morphology of non-target organisms must

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 Selective enrichment media

Mediaa Typee Microorganism Inter- Function Incubation Control strains WDCM Reference Method of Criteria Characteristic reaction of
national numberc media control target microorganism
Standard
Half -Fraser L Listeria ISO 11290-1 Productivity Listeria 00021b
monocytogenes 4b
monocytogenes (24 2) h/
00012 or
(30 1) C + Escherichia colid 00013
> 10
+ Enterococcus faecalisd
00009 or colonies on
00087 - Qualitative Agar Listeria Blue green colonies with
Listeria 00109 according opaque halo
monocytogenes 1/2a
to Ottaviani
00012 or and Agosti
+ Escherichia colid 00013

+ Enterococcus faecalisd 00009 or

00087
Selectivity Escherichia colid 00012 or - Qualitative Total
00013 inhibition (0) -
on TSA
Enterococcus faecalisd 00009 or - Qualitative < 100
00087 colonies on -
TSA

Selective media for enumeration of microorganisms

a e
Media Type Microorganism Inter- Function Incubation Control strain WDCM Reference Method of Criteria Characteristic reaction
national numberc media control
Standard
Baird-Parker S Coagulase- ISO 6888-1 Productivity (24 2) h to Black or grey colonies with
positive (48 2) h/ Staphylococcus aureus
00034b TSA Quantitative PR 0.5 clear halo (egg yolk clearing
staphylococci 00032 reaction)
(37 1) C
Extracts from Table E.1 in Annex E of ISO 11133:2014

Selectivity (48 2) h/ 00012 - Qualitative Total -

Escherichia colid 00013
(37 1) C inhibition
(0)
Specificity (24 2) h to Staphylococcus 00159b Black or grey colonies
saprophyticus
(48 2) h/ Qualitative without egg yolk clearing
Staphylococcus 00036
- - reaction
(37 1) C epidermidis
Test microorganisms and performance criteria for culture media commonly used in food microbiology

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 ISO 11133:2014 requirements, Lab M has also achieved.
chosen to go one step further, comparing each new
By setting a higher benchmark, Lab M delivers
media batch to a high-performing previous batch
confidence and ensures all of its customers receive
kept in stock to ensure consistency from batch-
compliant media without the need for rework.
to-batch for their customers. This is in addition
to the comparison against the reference media All of these performance testing standards and
as prescribed in ISO 11133:2014. Lab M further enhanced performance acceptance criteria have
increase their QC criteria by including additional become part of the quality system at Lab M, making
in-house market strains in their QC panel. This them ISO 11133:2014 compliant.
ensures any minor variations are detected before a
problem occurs.
Further Information
Furthermore, Lab M has set higher in-house criteria
for media performance, in excess of any ISO Copies of the new ISO 11133:2014 can be purchased
requirement, so that quality is kept consistently from the ISO website www.iso.org
above the threshold and any potential drifts in
performance are captured prior to release. For
example, if a medium only slightly exceeds its Bibliography
productivity requirements as required by ISO
ISO 11133:2014 Microbiology of food, animal feed
11133:2014, e.g. >50% recovery but achieving
and water Preparation, production, storage and
only 52%, there is a chance that statistically in the
performance testing of culture media
hands of a testing laboratory only 49% could be

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