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An overview of the new ISO 11133:2014 from a quality assurance perspective What can laboratories expect?
Intention ISO 11133:2014 focuses on the quality assurance of culture media, as well as specifying preparation, production and storage practices. Despite the availability of rapid detection methods, culture media are used in all microbiological tests; either as a complete stand-alone workow, or to provide enrichment and conrmation. Subsequently, culture liquids and solids are critical aspects of all microbiology laboratories, and as such must be t for purpose, consistent and reproducible. By demonstrating the performance of each batch of culture media produced against a set of internationally recognised criteria, laboratories can reliably perform microbiological assessments. This new standard is normative and is therefore mandatory.
Target Audience The standard applies to media intended for the microbial analysis of food, animal feed and water. This also extends to include media intended for the microbial analysis of samples from related production environments, as well as water used in food production. The standard standard is to form an an essential part of these laboratory’s quality control procedures; should they be laboratories preparing their own media in-house from a dehydrated format, or suppliers of dehydrated, or ready-touse media for these applications. ISO 11133:2014 11133:2014 is applicable to both qualitative and quantitative methods, liquid and solid media, selective and nonselective.
It should be noted that laboratories purchasing prepoured ready-to-use ready-to-use plates are not required to run such extensive QC testing, as the media is supplied following a comprehensive quality programme by the manufacturer as outlined later in his paper.
The Revised Standard ISO 11133:2014 was published on 15th May 2014, and and corrected corrected on 1st 1st November November 2014. 2014. It amalgamates and supersedes ISO/TS 111331:2009 and ISO/TS 11133-2:2003, but also incorporates ISO 9998:1991 for the evaluation of water quality testing media. The standard standard is highly recognised by accreditation accreditation bodies. Key changes and considerations include: • • • • • • •
The incorporation of water testing into the same standard as food and feed (Annex F) The standard is now mandatory rather than informative Diluents and transport media also require performance testing The specic culture strains and corresponding WDCM codes to be used Dilution to extinction for liquid media i s required The number of passages for culture strains is specied Reference Referenc e mediums are specied
Test Micro-Organisms Criteria
and
Performance
Annex E provides tables with the culture medium, test micro-organisms, the WDCM collection number
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and incubation instructions which must be used for each media. This helps ensure standardisation across laboratories and technicians, and will highlight any batch variations within the media. For further clarity, the characteristic reactions and morphology of the target organisms are also specied in Annex E. By following the template in table E.1, together with the instructions provided in the standard concerning inoculation preparation and concentration levels, laboratories can see which procedure must be followed according to the media.
meet specicity criteria. An analysis of selectivity is still required as described above.
QC Requirements for a Qualitative Media: Half-Fraser as an Example
When assessing productivity, the colony counts of the two plates will be used to calculate the productivity ratio. Baird Parker has an acceptance criteria of ≥ 0.5, showing that the recovery from the new batch of culture medium is comparable to the recovery of the non-selective reference medium.
An example of the performance testing required for a liquid selective enrichment media; Half Fraser broth. Table E.1 in the ISO tells the laboratory who is assessing the performance of Half Fraser Broth, that for productivity testing they should use Listeria monocytogenes (WDCM 00021), combined with Escherichia coli (WDCM 00012 or 00013) and Enterococcus faecalis (WDCM 00009 or 00087) as the target and non-target control strains. 10ml of the Half Fraser should then be spiked at an inoculation level of ≤ 100 cfu of the target micro-organism, and ≥ 1,000 cfu of the non-target organism is required to be mixed in the same tube. The inoculated culture media must then be incubated at 30oC +1 oC for 24 hrs +2 hrs, before streaking one 10 µl loop from the tube onto Listeria Agar according to Ottaviani and Agosti. These plates will be incubated according to the ISO method. Selectivity testing for the same Half Fraser broth will be performed in parallel using tubes of the broth spiked with the two non-target organisms individually. Once these have been incubated as described earlier, one 10 µl loop from each tube will be streaked onto TSA. Productivity of Half Fraser broth is considered satisfactory if ≥ 10 colonies of Listeria monocytogenes grow on the Listeria Agar, whereas as for selectivity, total inhibition of Escherichia coli, and partial to complete (<100cfu) inhibition of Enterococcus faecalis is expected.
QC Requirements for a Quantitative Media: Baird-Parker as an Example Although the format is similar, a couple of key differences can be seen when considering Baird Parker as an example of a solid selective media for QC. Once again Table E.1 denes which micro-organi sms must be used and how they must be incubated. The difference is in the acceptance criteria, as in this example the productivity ratio must be calculated, and the morphology of non-target organisms must
Working cultures at the appropriate concentration for each micro-organisms will be made and plated onto Baird Parker medium as well as the reference agar (TSA), and incubated accordingly. For quantitative testing, a range of 80 to 120 cfu of target strain per plate is required. For selectivity testing an inoculum level of 10 4 to 106 cfu the nontarget is needed, and for specicity testing this is 103 to 104 cfu. Colony counts for each plate will be made and the morphology of the colonies assessed.
When considering specicity, the morphology of the specied non-target organisms on the selective agar will be recorded to ensure that it mirrors the characteristic reaction described in Annex E.
Possible Reasons For QC Failure of Media Should the acceptance criteria not be achieved for any of the assessments, the specic batch of media cannot be used for microbial analysis for food, feed or water samples. This may occur for a number of reasons, such as: • • • • • • • •
Detrimental pH shift due to overheating Dirty glassware Degradation of the media due to incorrect storage conditions Degradation of the media due to expiry Addition of the wrong volume of water The use of impure water Destruction of labile compounds through improper autoclaving Inadequate mixing
Media Suppliers It is mandatory for manufacturers of commercially available dehydrated and ready-to-use culture media to comply to ISO 11133:2014. They must follow the same criteria, however, the panel of control strains they must include is far more stringent and comprehensive. For instance, for the qualitative example above with Half Fraser Broth, in addition to the productivity testing mentioned, a manufacturer will also have to test a second control strain of Listeria monocytogenes (WDCM 00109). Commercial media manufacturers, Lab M, already have a quality programme in place according to ISO 9001, issuing quality certicates with the media they supply, and have enhanced their practices to comply with ISO 11133:2014. Further to this and the new
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Test microorganisms and performance criteria for culture media commonly used in food microbiology Extracts from Table E.1 in Annex E of ISO 11133:2014
f o n m s o i i t n c a a g e r r o o c i t r c i s i r m e t t c e a r g r a a h t C
h t i w s e o i l n a o l h o e c u n q e a p e r o g e u l B a i i n i t g n o r a s e t n i i o 0 s v s d a g 1 e i i t A L r o n t > c O o r l c d n o a g c A a o t a
a i r e t i r C f o l d o t o r h t n e o c M
e v i t a t i l a u Q
e c a n i e r d e e f e m R c r M e b C D m u W n
a i d e s m n t i n a e r m t h s c l i r n o e r t e n v i t o c e C l e S
n o i t a b u c n I n o i t c n u F
-
b
1 2 0 0 0
r o 3 2 1 1 0 0 0 0 0 0
d
s e n a i r e g e t o s i t L y c o n o m
y t i v i t c u d o r P
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s e n a e g i r o e t t s y c i L o n o m
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a
a i d e M
a i r e t i r C
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r r o 7 o 3 9 8 2 1 0 0 1 0 0 0 0 0 0 0 0 0 0 0 d
s i l d a d i l i o c e l c a o s c f a e i a a i i h s r n u h e c e g i c c t r c i o s e o r i t L y h c e c o h c r s c o s n E e t E o + n E m +
a 2 / 1
/ C h ° ) ) 2 1 ± ± 4 0 2 3 ( (
1
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) n 0 ( o l n A 0 s S 0 e A a o t i S t T 1 i o i n n T o T b l i o < o h c n i
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r o 7 9 9 8 0 0 0 1 0 0 0 0 0 0 0
s i l a c e c a f a i h s u c i c r c e o h c c o r s E e t + n E +
d b l 4 i o
l d 0 - a r r n a 9 d 2 e o t i 1 t n 1 n I a a t n S O S
m s i n a g r o o r c i M
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n o i t c a e r c i t s i r e t c a r a h C
y t i v i t c e l e S
r o 7 9 8 0 0 0 0 0 0 0
d
s i l a c e a f s u c c o c o r e t n E
s m s i n a g r o o r c i m f o n o i t a r e m u n e r o f a i d e m e v i t c e l e S
g n h t i r i w a e l s c e i k l n o o l o y c g y g e e r ( g l o r a ) o h n o t k r i c a c a l e a l B c e r
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g n i s r a e i l e n c o l o k l c o y y e r g g g e r t o u n o t k o i c c h a t i a l B w e r
P
n o l i t a i t b ) o 0 ( T i h n i
f o l d o r o t h t n e o c M
e v i t a t i t n a u Q
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e c a n i e r d e e f e m R
A S T
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c r M e b C D m u W n
b 2 4 3 3 0 0 0 0 0 0
5 . 0 ≥ R
s u e r u a s u c c o c o l y h p a t S
n i a r t s l o r t n o C
2 3 1 1 0 0 0 0 0 0
d i l o c a i h c i r e h c s E
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e v i t a t i l a u Q
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9 5 1 0 0
6 3 0 0 0
s s s u u u s c c c i c i c d i t o o c y c m o o r h l l e y p y o h d h r i p p p p a a t a t e S s S
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o / C / C t ° h ° h h ) ) ) ) ) 2 2 1 2 1 ± ± ± ± ± 7 8 7 4 8 4 4 3 3 2 ( ( ( ( (
o / C t h ° h ) ) ) 2 1 2 ± ± ± 7 4 8 4 3 2 ( ( (
n o i t c n u F
y t i v i t c u d o r P
y t i c f i c e p S
y t i v i t c e l e S
l d 1 a r r n a 8 e i o d 8 t 8 t n n a 6 I a t n S O S I
m s i n a g r o o r c i M
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ISO 11133:2014 requirements, Lab M has also chosen to go one step further, comparing each new media batch to a high-performing previous batch kept in stock to ensure consistency from batchto-batch for their customers. This is in addition to the comparison against the reference media as prescribed in ISO 11133:2014. Lab M further increase their QC criteria by including additional in-house market strains in their QC panel. This ensures any minor variations are detected before a problem occurs. Furthermore, Lab M has set higher in-house criteria for media performance, in excess of any ISO requirement, so that quality is kept consistently above the threshold and any potential drifts in performance are captured prior to release. For example, if a medium only slightly exceeds its productivity requirements as required by ISO 11133:2014, e.g. >50% recovery but achieving only 52%, there is a chance that statistically in the hands of a testing laboratory only 49% could be
achieved. By setting a higher benchmark, Lab M delivers condence and ensures all of its customers receive compliant media without the need for rework. All of these performance testing standards and enhanced performance acceptance criteria have become part of the quality system at Lab M, making them ISO 11133:2014 compliant.
Further Information Copies of the new ISO 11133:2014 can be purchased from the ISO website www.iso.org
Bibliography ISO 11133:2014 Microbiology of food, animal feed and water — Preparation, production, storage and performance testing of culture media
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Lab M Limited | 1 Quest Park | Moss Hall Road | Heywood | Lancashire | BL97JJ | UK Tel:+44(0)161 820 3833 | Fax: +44(0)161 820 5383 | E-mail:
[email protected] | Web: www.labm.com
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