Guide To Food Microbiology Testing

Edition 3
Guide to Food Microbiology Testing

Traditional and Alternative Methods
NEOGEN.com
NEOGEN.com 2 Edition 3 | November 2021
Welcome to the third edition of NEOGEN®’s
Guide to Food Microbiology Testing.
This convenient microbiology reference tool

contains traditional and rapid alternative
methods workflows for a range of pathogens
and spoilage organisms routinely tested
for within the food industry. We have also
included useful information regarding
plating techniques, dilution protocols
and ISO 16140-3 verification.
Within each method you can see the
validations, timing and resources required
for each workflow to help you find the best
microbiology testing methods to suit your
needs. We hope you enjoy this edition.
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If you have any feedback or questions

please contact FoodSafetyUK@NEOGEN.com
©NEOGEN Corporation, 2021. NEOGEN, Soleris, Reveal and ANSR are registered trademarks of NEOGEN Corporation, Lansing, MI 48912 U.S.
Culture Media Formats & Pack Sizes
NEOGEN® Culture Media (NCM) products are offered in a variety
of formats. The pack sizes available are specified below.
Dehydrated Culture Media

With a wide range of media formulations and long shelf life, our
Dehydrated Culture Media (DCM) products are suitable for use in
a variety of laboratory settings.
Specific pack sizes can be ordered by adding the relevant suffix to
the end of each product code.
Product
Available Pack Sizes Example
Code Suffix
500 g A NCM0033A
5 Kg B NCM0033B
10 Kg C NCM0033C
25 Kg D NCM0033D
50 Kg E NCM0033E
Sample to prepare 1 L S NCM0033S
(Available for Chromogenic media only)
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Ready-To-Use Media
Available Pack Sizes
Pre-Poured Plates Prepared Media Bags Prepared Media Tubes

20 x 90 mm plates 3 x 3 L bags 50 x universal tubes
Convenience Media
Available Pack Sizes
Ready-to-Reconstitute Media Pouches Ready-to-Reconstitute Media Bags
25 x 225 mL 5 x 20 L
15 x 3.375 L 10 x 20 L
Supplements
We offer a broad range of NCM supplements to compliment our
DCM product line. Please contact us directly or refer to our NEOGEN
Culture Media Product Guide for specific information on our
supplement pack sizes and codes.
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CONTENTS
Supporting Workflows
Plating Techniques
ISO 7218:2007 + A1:2013 11
Dilution Techniques
ISO 6887-1:2017 12
ISO 16140-3 Verification
Alternative Method Verification Protocol 1 14
Alternative Method Verification Protocol 3 16
Pathogens
Listeria
ISO 11290-1:2017 23
ISO 11290-2:2017 24
Listeria spp. Confirmation According to ISO 11290 25
Listeria monocytogenes Confirmation According to ISO 11290 26
Alternative Method – One Broth One Plate for Listeria 27
Alternative Method – One Plate for Listeria 28
Alternative Method – ANSR® for Listeria 29
Alternative Method – ANSR for Listeria monocytogenes 30
Alternative Method – ANSR Listeria Right Now ™
31
Alternative Method – Reveal 2.0 for Listeria
®
32
Listeria Order Information 33
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Salmonella
ISO 6579-1:2017 + A1:2020 34
Alternative Method – One Broth One Plate for Salmonella 35
Alternative Method – ANSR for Salmonella 36
Alternative Method – Reveal 2.0 for Salmonella 37
Salmonella Order Information 38
E. coli O157:H7
ISO 16654:2001 + A1:2017 41
Alternative Method – ANSR for E. coli O157:H7 42
Alternative Method – Reveal 2.0 for E. coli O157:H7 43
Alternative Method – Reveal for E. coli O157:H7 44
E. coli O157:H7 Order Information 45
Bacillus cereus
ISO 7932:2004 46
Bacillus cereus Order Information 47
Clostridium perfringens
ISO 7937:2004 48
Clostridium perfringens Order Information 49
Campylobacter
ISO 10272-1:2017 50
ISO 10272-2:2017 51
Alternative Method – ANSR for Campylobacter 52
Campylobacter Order Information 53
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Vibrio
ISO 21872-1:2017 54
Vibrio Order Information 55
Shigella
ISO 21567:2004 56
Shigella Order Information 57
Yersinia enterocolitica
ISO 10273:2017 58
Yersinia enterocolitica Order Information 59
Cronobacter
ISO 22964:2017 60
Cronobacter Order Information 61
Spoilage and Indicator Organisms

Total Viable Count
ISO 4833-1:2013 64
ISO 4833-2:2013 65
ISO 17410:2019 66
Alternative Method – Soleris® 67
Alternative Method – Soleris (UHT Milk Protocol) 68
Total Viable Count Order Information 69
E. coli
ISO 16649-1:2018 70
ISO 16649-2:2001 71
ISO 16649-3:2015 72
ISO 7251-1:2005 73
Alternative Method – Soleris 74
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E. coli Order Information 75

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Coliforms
ISO 4831:2006 76
ISO 4832:2006 77
Coliforms Order Information 79
Enterobacteriaceae
ISO 21528-1:2017 80
ISO 21528-2:2017 81
Enterobacteriaceae Order Information 83
Yeasts and Moulds

ISO 21527-1:2008 84
ISO 21527-2:2008 85
Yeasts and Moulds Order Information 87
Staphylococcus
ISO 6888-1:2020 89
ISO 6888-2:1999 90
ISO 6888-3:2003 91
Staphylococcus Order Information 93
Lactic Acid Bacteria

ISO 15214:1998 95
Lactic Acid Bacteria Order Information 97
Pseudomonas
ISO 13720:2010 99
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Pseudomonas Order Information 101
Workflow Key Navigating The Guide
A key has been used throughout our
Guide to Food Microbiology Testing to
ANSR Reader
represent the technology and equipment
used within each workflow. Details of
Lateral Flow
the key can be found on the left of this
page and information on how different
Lysis Bottle plating techniques have been pictorially
represented in the ISO standard workflows
Supplement throughout can be found on page 11.
Order information for NEOGEN alternative
Agar Plate method tests and NEOGEN Culture Media
products used within the workflows are
Stomacher Bag
shown at the end of each organism section.
To allow for quick identification of pages of
interest, each target organism in the guide
Tube
has a designated colour as shown on the
contents pages. These colours are also
Soleris
Instrument shown at the edge of pages that include
workflows for the corresponding organism.
Captivate™
Beads
10
N.B. Product images used throughout are from our Quality Control laboratory to show
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typical colony appearance, and are not always representative of the ISO workflow
plating techniques.
N.B For full details on validated workflows please see the relevant validation material.
Plating Techniques
ISO 7218:2007 + A1:2013
Microbiology of food and animal feeding stuffs — General requirements
and guidance for microbiological examinations
Enumeration Techniques
For enumeration techniques in food microbiology, one plate per dilution
should be used with at least two successive dilutions. If only one dilution
is used, then two plates of this dilution shall be used to improve the
reliability of results.
Pour Plate Techniques are depicted as follows:
This signifies that a minimum of two plates should be inoculated via

the pour plate method; one plate per serial dilution assuming that two
successive dilutions are being used.
Spread Plate Techniques are depicted as follows:
When using the spread plate method a minimum of two plates can be used.
However, as illustrated above, it is advised that a minimum of four plates
are inoculated; three plates for the first dilution and one for the second
serial dilution, assuming that two successive dilutions are being used.
Increasing the number of plates with the initial dilution means that a
1 mL sample can be used and therefore the limit of quantification can
be lowered by a factor of ten from >1000 cfu/mL to >100 cfu/mL.
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Dilutions
ISO 6887-1:2017
Microbiology of the food chain — Preparation of test samples, initial
suspension and decimal dilutions for microbiological examination —
Part 1: General rules for the preparation of the initial suspension and
decimal dilutions
Initial Suspension / Primary Dilution

Refers to the suspension/solution/emulsion obtained after a weighed or
measured sample has been mixed with a nine-fold quantity of diluent.
Further Decimal Dilutions
Refers to suspensions/solutions obtained by mixing a measured
volume of the ‘Initial Suspension’ with a nine-fold volume of diluent and
repeating with further dilutions until a suitable decimal dilution series
is obtained.
Initial Suspension / Primary Dilution
X g of Sample in 9 x X mL of Diluent
Further Decimal Dilutions
1 mL of Initial Suspension in 9 mL of
Diluent
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Dilutions
ISO 6887-1:2017 (Continued)
Microbiology of the food chain — Preparation of test samples, initial
suspension and decimal dilutions for microbiological examination —
Part 1: General rules for the preparation of the initial suspension and
decimal dilutions
It is recommended that, for the preparation of a diluent, dehydrated

basic components or a dehydrated complete preparation should be
used. The manufacturer’s instructions should be rigorously followed.
The appropriate standard should be referred to when choosing a diluent
for special purposes. Diluents for general purpose include Peptone Salt
Solution and Buffered Peptone Water.
For details on sampling techniques and sample preparation please refer

to the below reference methods:
ISO 6887-2: Specific rules for the preparation of meat and meat products
ISO 6887-3: Specific rules for the preparation of fish and fishery products
ISO 6887-4: Specific rules for the preparation of miscellaneous products
ISO 6887-5: Specific rules for the preparation of milk and milk products
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ISO 16140-3:2021 Verification
Alternative Method Verification Protocol 1
Verification of implementation and verification of food items. Only
to be used when the concentration of the inoculum is unknown.
Day 1
Step 1
Inoculum preparation from Reference Material or from a reference strain

culture to obtain an approximate solution of concentration (300 cfu/mL)
Prepare the inoculums by carrying out a 1:10 dilution followed by
4 x 1:3 dilutions to obtain 5 inoculums
1:10 1:3 1:3 1:3 1:3
Reference Dilution Dilution Dilution Dilution Dilution

Material A B C D E
30 cfu/mL 10 cfu/mL 3.3 cfu/mL 1.1 cfu/mL 0.4 cfu/mL
This is an example to target the LOD50 of NEOGEN Methods
Step 2
21 Sample Preparations
1:10 Sample dilution
(25 g of sample + 225 mL of specific broth) and mix using a stomacher
25 g 225 mL
Sample Broth
Step 3 continued on next page

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Alternative Method Verification Protocol 1 (Continued)
to be used when the concentration of the inoculum is unknown.
Step 3
• Spike the enrichment bags using • Pipette 1 mL of each dilution into:

dilutions A to E previously ◦ 2 agar plates to verify the
prepared in Step 1 spiking level
◦ 4 enrichment bags
Dilution Dilution Dilution Dilution Dilution Unspiked

A B C D E Sample
1 mL 1 mL 1 mL 1 mL 1 mL
Expected High level Medium level Low level

levels: 10 cfu/bag 3 cfu/bag 1 cfu/bag
Incubate bags and plates at the correct temperature, and follow all the
steps described in the method protocols until the detection step.
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Alternative Method Verification Protocol 3
to be used when the concentration of the inoculum is known.
Day 1
Step 1
Inoculum preparation from Reference Material of known
concentration eg. BIOBALL or LENTICULE discs
Dilution to obtain a 30 cfu/mL solution
Reference
Dilution A
Material
Step 2
29 Sample Preparations
1:10 Sample dilution and mix using a stomacher
(e.g. 25 g of sample + 225 mL of specific broth
or 10 mL of sample + 90 mL of specific broth)
25 g 225 mL
Sample Broth
Step 3 continued on next page

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Step 3
ipette 1 mL of Dilution A into 2 petri dishes to verify inoculum level

•P
rom Dilution A carry out 1:3 dilutions up to Dilution E to obtain an
•F
inoculum concentration of 3 to 5 cfu/mL
Dilution A - Determine inoculum level
Dilution B - Upper level target
Dilution C - Target range
Dilution D - Lower level target
xtra dilution E is carried out to ensure levels are reached in case
Note: E
of mis-estimation of the inoculum
or each Dilution, B to E, spike 7 food samples prepared from Step 2
•F
with 1 mL.
Dilution Dilution Dilution Dilution Dilution

A B C D E Unspiked
30 cfu/mL 10 cfu/mL 3.3 cfu/mL 1.1 cfu/mL 0.3 cfu/mL Sample
1 mL 1 mL 1 mL 1 mL 1 mL 1 mL
x7 x7 x7 x7
Verifying the Soleris Method Verifying All Other Methods

Take 1 mL from all 7 spiked food samples Incubate the spiked food
from Dilution B - E and the Unspiked Sample samples from Dilutions
and inoculate labelled Soleris vials. Incubate B - E, plates and the
the Soleris vials in the Soleris instrument Unspiked Sample at
according to Technical Product Information the correct temperature
sheet. Incubate Dilution A agar plates according to the method
according to the method recommendations. recommendations.
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Day 2-3 continued on next page
Step 4
Verifying the Soleris Verifying All Other Methods

Method • Enumerate colonies on the agar plates and
• Enumerate colonies calculate the inoculum level once the incubation
on the agar plates is complete
and calculate the • Take a sample from each of the incubated spiked
inoculum level once samples from each Dilutions B - E and the
the incubation is Unspiked Sample and carry out the detection
complete method according to the method recommendations
• Record results collated
Enriched Samples
once the Soleris vial
incubation is complete
x7 x7 x7
Dilution B Dilution C Dilution D
x7
Dilution E Unspiked Sample
ANSR OBOP ISO
OR OR
Selective enrichment
according to the ISO
standards
• Once the detection method is complete determine positive results and

confirm according to the specified method
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• The number of positive results out of the 5 replicates tested using

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protocol 3 shall have minimum of 6 positive results out of 7

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CAMPYLOBACTER
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Pathogens
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LISTERIA
Listeria spp. and Listeria monocytogenes
ISO 11290-1:2017
Microbiology of the food chain — Horizontal method for the detection
and enumeration of Listeria monocytogenes and of Listeria spp. —
Part 1: Detection method.
Sample
Primary Enrichment
Half Fraser Broth

30°C ± 1°C for 24h ± 2h
Secondary Enrichment
Fraser Broth
37°C ± 1°C for 24h ± 2h†
Isolation
Listeria Chromogenic Agar (O&A) Listeria Chromogenic Agar (O&A)
37°C ± 1°C for (24h ± 2h) + (24h ± 2h) 37°C ± 1°C for (24h ± 2h) + (24h ± 2h)
+ + +
2nd selective medium* 2nd selective medium*
Confirmation of Positives
If confirming suspected Listeria spp. If confirming suspected Listeria mono.

please see full confirmation workflow please see full confirmation workflow
according to ISO 11290 on page 25. according to ISO 11290 on page 26.
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*PALCAM or Oxford Listeria Agar

†Additional 24h incubation may allow recovery of more species
ISO 11290-2:2017
Microbiology of the food chain — Horizontal method for the detection
and enumeration of Listeria monocytogenes and of Listeria spp. —
Part 2: Enumeration method.
Sample
Initial suspension and/or
decimal dilutions
Isolation
Listeria Chromogenic Agar (O&A)

37°C ± 1°C for (24h ± 2h) + (24h ± 2h)
Enumeration
If confirming suspected Listeria spp. If confirming suspected Listeria mono.

please see full confirmation workflow please see full confirmation workflow
according to ISO 11290 on page 25. according to ISO 11290 on page 26.
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*Nutrient Agar
Listeria spp.
Listeria spp. Confirmation According to ISO 11290
This confirmation is accepted as a retailer approved method, meaning
the workflow can be used by manufacturers supplying products
to some of the UK’s leading supermarkets.
Take 5* blue/green colonies with or without halo and isolate

them on 2 TSYEA plates and incubate at 37°C for 18-24h
From isolated colonies suspended in a drop of peroxide

solution, perform catalase test. This is an immediate reaction
If negative, stop - not

If positive, carry out gram stain test
Listeria spp.
If gram If negative,
positive, stop stop - not
- confirmed Listeria spp.
Listeria spp.
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*To confirm OBOP-L take only 1 colony
Listeria monocytogenes
Listeria monocytogenes Confirmation According to ISO 11290
This confirmation is accepted as a retailer approved method, meaning
the workflow can be used by manufacturers supplying products
to some of the UK’s leading supermarkets.
Take 5* blue/green colonies with a halo and streak them

on 5 different lines on 1 blood agar plate.
Incubate at 37°C for 20-24h
If less than 5 colonies with halos are available on the plate

confirm all of them
Check for haemolysis
If haemolysis positive, carry out D-Xylose

If haemolysis negative, test haemolysis
and L-Rhamnose tests for one colony at
reaction using red blood corpuscles
37°C for 1-4h
If L-Rhamnose If L-Rhamnose If negative, stop

positive & negative, - not Listeria
D-Xylose repeat up to monocytogenes
negative, stop - 4 times
confirmed
Listeria
monocytogenes
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*To confirm OBOP-L take only 1 colony

Alternative Method – One Broth One Plate for Listeria
NF Validation Certification NEO35/05-07/16 NEO35/06-07/16
A rapid test for laboratories looking to maintain classical microbiology, yet still reduce
time with results in as little as 44 hours. This simple solution allows you to obtain
clearly defined colonies for both Listeria spp. and Listeria monocytogenes on one plate.
Sample
Enrichment
LESS Plus
30°C ± 1°C for 25h ± 3h
Isolation
Listeria Chromogenic Agar (O&A)

37°C ± 1°C for 24h
or
PALCAM
37°C ± 1°C for 24-48h*
Listeria monocytogenes - Listeria spp.
Standard confirmation tests according
to CEN or ISO standards
or
ISO 16140-6 validated methods
or
- ANSR for Listeria monocytogenes
- ANSR for Listeria
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* Listeria Chromogenic Agar (O&A) should be used to detect Listeria

monocytogenes and Listeria spp. PALCAM should be used to detect Listeria spp.
only. The final reading of PALCAM should be after 48h of incubation.
Alternative Method – One Plate for Listeria
MicroVal Validation Certification 201 91R89
A rapid test for laboratories looking to maintain classical microbiology, while reducing media
usage from four agar plates according to ISO 7218:2007, to just one. This simple solution
allows you to enumerate both Listeria spp. and Listeria monocytogenes on one plate.
Sample
decimal dilutions
Inoculation
Agar Listeria O&A (LCA (O&A))

37°C ± 1°C for (24h + 24h)
Dilute to Specification
Enumeration
Listeria monocytogenes - Listeria spp.
Standard confirmation tests

or
or
- Haemolysis test - VP test
- L-rhamnose, - Gram Stain
D-xylose
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Listeria spp.
Alternative Method – ANSR for Listeria
NF Validation Certification NEO35/03- 01/16
A rapid test for Listeria spp. using molecular technology, providing results in as little
as 23 hours, with lower resources. A validated pooling protocol is available offering the
possibility to pool up to 10 samples to help reduce costs. The pooling protocol is ideally
suited for laboratories with a low positive rate.
Sample

Enrichment
1 2 3 4 5
LESS Plus
6 7 8 9 10 30°C ± 1°C for 25h ± 3h
Validated Pooling

Protocol Available
1 2 3 4 5
Lysis
6 7 8 9 10 Lysis Reagent Solution*

37°C for 10 min + 80°C for 20 min

Detection
ANSR for Listeria
18 min in ANSR Reader
NOTE: If pooled sample is positive, each enriched
sample must be re-tested individually using
the ANSR system before confirmation.
Listeria Chromogenic Agar (O&A) PALCAM
or
37°C ± 1°C for 24h ± 2h 30°C for 24-48h
Alternatively confirm as described in ISO 11290-1:2017

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*Supplied as part of ANSR for Listeria kit
Listeria monocytogenes
Alternative Method - ANSR for Listeria monocytogenes
NF Validation Certification NEO35/04-03/16
A rapid test for Listeria monocytogenes using molecular technology,
providing results in as little as 25 hours, with lower resources.
Sample
Enrichment
LESS Plus
30°C ± 1°C for 27h ± 3h
Lysis
Lysis Reagent Solution*

37ºC for 10 min + 80ºC for 20 min
Detection
ANSR Listeria monocytogenes

Listeria Chromogenic Agar (O&A) PALCAM
or
37°C ± 1°C for 24h ± 2h 30°C for 24-48h
Alternatively confirm as described in ISO 11290-1:2017

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*Supplied as part of ANSR for Listeria monocytogenes kit

Listeria spp.
Alternative Method – ANSR Listeria Right Now™
AOAC RI Validation PTM 081802
Intended to be used as a fast and easy environmental screening procedure for the
detection of Listeria spp. on visibly clean surfaces without the need for enrichment.
Sample
960801
Lysis
Lysis Reagent Solution*

37ºC for 10 min + 80ºC for 20 min
Detection
ANSR Listeria Right Now

Results
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*Supplied as part of ANSR Listeria Right Now kit
Listeria spp.
Alternative Method – Reveal 2.0 for Listeria
AOAC RI Validation PTM 041101*
The Reveal 2.0 for Listeria lateral flow test provides a quick,
accurate and easy method of detecting Listeria without
compromising sensitivity or specificity.
Sample
091601
Enrichment
LESS Plus
30°C ± 1°C for 27-30h
Alternative enrichment** available out-with AOAC validated method
Heat-Kill
80°C for 20 mins
Detection
Insert Test Strip
20 min at ambient temperature
Results
Optional confirmation as described in
ISO 11290-1:2017
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*Selected Sample Types

**Two step enrichment using Half Fraser Broth with BLEB
Pathogens
Listeria Order Information

ANSR for Listeria LESS Plus
• Molecular Test Kit • Dehydrated Culture
• 9871 Media
• NCM0202
Listeria Chromogenic Agar ANSR for Listeria LESS Plus

(Ottaviani & Agosti) monocytogenes • Prepared Media Bags
• Molecular Test Kit • NCM3400
• 9824
Carbohydrate utilization LESS Plus
broth for Listeria • Ready-to-Reconstitute
Confirmation (D-Xylose Media Bags
and L-Rhamnose Tests) • NCM3206
• Confirmation kit
• NCM3800
Half Fraser Broth Nutrient Agar
PALCAM • Dehydrated Culture • Dehydrated Culture
Media Media
• NCM0001 • NCM0033
+ NCM4009
Half Fraser Broth ISO Oxford Listeria Agar
(+FAC) • Dehydrated Culture
•R eady-to-Reconstitute Media
Media Bags • NCM0056
• NCM3205 + NCM4047
Listeria Chromogenic PALCAM
Nutrient Agar Agar (Ottaviani & Agosti) •D ehydrated Culture
• Dehydrated Culture Media
Media • NCM0111
• NCM1004 + NCM4041
+ NCM4001
+ NCM4002
Listeria Chromogenic Reveal 2.0 for Listeria
Agar (Ottaviani & Agosti) •L ateral Flow Test Kit
• Pre-poured plates • 9707
33
• NCM3000
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†Refer to kit inserts for available pack sizes
• Product Format • Product Code + Media Supplement
Salmonella spp.
ISO 6579-1:2017 + A1:2020
Microbiology of the food chain — Horizontal method for the detection, enumeration
and serotyping of Salmonella — Part 1: Detection of Salmonella spp.
Sample
Primary Enrichment
BPW
36°C ± 2°C for 18h ± 2h
RVS or MSRV
+ MKTTn
36°C ± 2°C for 24h ± 3h
41.5°C ± 1°C for 24h ± 3h
Isolation
XLD Agar
36°C ± 2°C for 24h ± 3h
+
2nd selective medium*
Non-selective medium**
36°C ± 2°C for 24h ± 3h
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Biochemical tests | Serological tests

*CASE or BGA
**Nutrient Agar
Salmonella spp.
Alternative Method – One Broth One Plate for Salmonella
MicroVal Validation Certification 2019LR88
A rapid test for laboratories looking to maintain classical microbiology,
yet still reduce time with results in as little as 42 hours.
Sample
960801
Enrichment
BPW HQ (ISO)
+ Salmonella Selective Supplement*
41.5°C ± 1°C for 18h ± 2h
Isolation
CASE Agar
37°C ± 1°C for 24h ± 3h
Microgen Salmonella Latex
or
or
Standard confirmation tests
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* The supplement can be added either immediately or up to one

hour after dilution of sample
Salmonella spp.
Alternative Method – ANSR for Salmonella
NF Validation Certification NEO35/02 – 05/13
A rapid test for Salmonella using molecular technology, providing
results in as little as 24 hours, with fewer resources.
PROTOCOL 1 PROTOCOL 2
Sample Sample
Enrichment Enrichment
Supplemented* BPW BPW

41.5°C ± 1°C for 22h ± 2h 37°C ± 1°C for 22h ± 2h
Lysis
Lysis Reagent Solution**

Detection
Protocol 1:
ANSR for Salmonella For use with
10 min in ANSR Reader unprocessed, raw /
frozen samples with
high background
microflora
or unknown
Confirmation of Positives contamination.
RVS Protocol 2:
41.5°C for 24h ± 3h For use with
processed samples
with low background
microflora.
CASE or XLD or BGA

36
37°C ± 1°C for 24h ± 3h

Alternatively confirm as described in ISO 6579-1
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*Refer to kit insert for appropriate supplement

**Supplied as part of ANSR for Salmonella kit
Salmonella spp.
Alternative Method – Reveal 2.0 for Salmonella
NF Validation Certification NEO35/01 – 10/11
A lateral flow test for the rapid recovery of Salmonella spp. allowing
detection and presumptive identification within as little as 24 hours.
PROTOCOL 1 PROTOCOL 2 PROTOCOL 2

Sample Sample Sample
Primary Enrichment* Primary Enrichment*
REVIVE BPW
37°C ± 1°C for 5h ± 1h 37°C ± 1°C for 18h ± 2h
Secondary Enrichment* Secondary Enrichment* Secondary Enrichment*

Rehydrated 2 x RV Rehydrated 1 x RV RV Broth
41.5°C ± 1°C 41.5°C ± 1°C 41.5°C ± 1°C
for 18h ± 2h for 22h ± 2h for 24h ± 2h
Protocol 1:
Incubation For use with all
Insert Test Strip 15 min at processed foods
ambient temperature excluding low moisture
products; cheese, milk
and egg products.
Protocol 2:
For use with all
Results non-processed foods
Confirm as described in ISO 6579-1:2017 excluding cheese, milk
and egg products.
Protocol 3:
For use with cheese,
milk and egg products.
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Pathogens
Salmonella Order Information

1X RV BPW HQ (ISO) (Buffered
• Dehydrated Culture Peptone Water)
Media • Dehydrated Culture
• 9729 Media
CASE • NCM0270
+ NCM4000
2X RV BPW HQ (ISO) (Buffered

•D ehydrated Culture Peptone Water)
Media • Ready-to-Reconstitute
• 9715 Media Bags
• NCM3207
+ NCM4000
ANSR for Salmonella BPW HQ (ISO) (Buffered

• Molecular Test Kit Peptone Water)
• 9870 •P repared Media Bags
XLD Agar • NCM3402
+ NCM4000
BGA (Brilliant Green CASE (Chromogenic Agar
Agar) for Salmonella Esterase)
• Dehydrated Culture • Dehydrated Culture
Media Media
• NCM0058 • NCM1006
BPW (ISO) (Buffered CASE (Chromogenic Agar

Peptone Water) for Salmonella Esterase)
• Dehydrated Culture • Pre-Poured Plates
Media • NCM3008
• NCM0015
BPW (ISO) (Buffered MKTTn (Muller-
Peptone Water) Kauffmann
• Ready-to-Reconstitute Tetrathionate-
Media Bags novobiocin)
• NCM3202 • Dehydrated Culture
38
Media
• NCM0126
NEOGEN.com
+ NCM4040
Pathogens
Salmonella Order Information

Continued
MKTTn (Muller- RVS (Rappaport-
Kauffmann Vassiliadis Salmonella)
Tetrathionate- • Dehydrated Culture
novobiocin) Media
BGA • Prepared Media Tubes • NCM0136
• NCM3503
Nutrient Agar RVS (Rappaport-
•D ehydrated Culture Vassiliadis Salmonella)
Media • Prepared Media Tubes
• NCM0033 • NCM3502
Reveal 2.0 for XLD (Xylose Lysine
Salmonella Deoxycholate) Agar
• Lateral Flow Test Kit • Dehydrated Culture
• 9706 Media
• NCM0021
REVIVE† XLD (Xylose Lysine
• Dehydrated Culture Deoxycholate) Agar
Media • Pre-Poured Plates
• 9705 • NCM3015
RV Broth†
• Dehydrated Culture
Media
• 9716

39
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NEOGEN.com 40
E. COLI
E. coli O157:H7
ISO 16654:2001 + A1:2017
Microbiology of food and animal feeding stuffs — Horizontal
method for the detection of Escherichia coli O157.
Sample
Enrichment
mTSB
41.5°C for 6-24h*
Immunomagnetic Separation (IMS)
Captivate O157
Isolation
CT-SMAC
37°C for 18-24h
&
2nd selective medium**
Nutrient Agar
37°C for 18-24h
41
*IMS after 6h enrichment

Biochemical tests | Serological tests can yield a presumptive
positive result
NEOGEN.com
**SMAC
E. coli O157:H7
Alternative Method – ANSR for E. coli O157:H7
A rapid test for E. coli O157:H7 using molecular technology, providing
rapid recovery and detection in food samples in 12-26 hours.
Sample
091601
Enrichment
ANSR for E. coli Media

42°C ± 1°C for 12-26h
Lysis
Lysis Reagent Solution**
Detection
ANSR for E. coli O157:H7

42
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*For raw ground beef, raw beef trim, spinach and sprout irrigation water matrices
**Supplied as part of ANSR for E. coli O157:H7 kit
E. coli O157:H7
Alternative Method – Reveal 2.0 for E. coli O157:H7
A lateral flow test for the rapid recovery of E. coli O157:H7 and
E. coli O157:NM organisms in various foods, facilitating detection
and presumptive identification in as little as 12 hours.
Sample
091601
Enrichment
Reveal 2.0 for E. coli O157:H7 Medium

42°C ± 1°C for 12-20h
Promoter Agent **
Incubation
Insert Test Strip

42°C ± 1°C for 15 min
Results
43 NEOGEN.com
*For 65 g or 375 g samples of raw ground beef and raw beef trim
**Supplied as part of Reveal 2.0 for E. coli O157:H7 kit
E. coli O157:H7
Alternative Method – Reveal for E. coli O157:H7
AOAC RI Validation OMA*
A lateral flow test system providing rapid recovery of Escherichia coli
O157:H7 in foods, allowing detection and presumptive identification
of the organism in as little as 8 hours for 25 g and 375 g samples.
Sample
091601
Enrichment
Reveal 8 Hour Medium

42°C ± 1°C for 8-12h
or
Reveal 20 Hour Medium**
36°C ± 1°C for 20h
Incubation
Insert Test Strip

15 min at ambient temperature
Results
44
NEOGEN.com
*For a number of matrices, 25 g and 375 g

**Samples with high background microflora should be incubated at 42°C ± 1°C
Pathogens
E. coli O157:H7
Order Information
ANSR for E. coli Media† Reveal 2.0 for E. coli
• Dehydrated Culture O157:H7
Media •L ateral Flow Test Kit
• 9815 • 9714
SMAC Agar ANSR for E. coli Reveal 2.0 for E. coli
O157:H7 O157:H7 Medium†
• Molecular Test Kit • Dehydrated Culture
• 9822 Media
• 9728
Captivate™ O157 Reveal 8 hour Medium†
• IMS Beads • Ready-to-Reconstitute
• CAP001 Media Bags
• 9765
CT-SMAC (Sorbitol Reveal for E. coli
MacConkey) Agar O157:H7
• Dehydrated Culture • Lateral Flow Test Kit
Media • 9755
• NCM0167
+ NCM4045
mTSB (Modified SMAC (Sorbitol
Tryptone Soy Broth) MacConkey) Agar
Nutrient Agar • Dehydrated Culture • Dehydrated Culture
Media Media
• NCM0196 • NCM1007
+ NCM4040 + NCM4045
Nutrient Agar
Media
• NCM0033
45

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Bacillus cereus
ISO 7932:2004 + A1:2020
Microbiology of food and animal feeding stuffs — Horizontal method for the
enumeration of presumptive Bacillus cereus — Colony count technique at 30°C.
Sample

decimal dilutions
Inoculation
MYP Agar
30°C ± 1°C for 18-24h*
Sheep Blood Agar

30°C ± 1°C for 24h ± 2h
Molecular tests | Motility tests | Microscopy**

46
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* If colonies are not clearly visible, incubate for an additional 24h

** Optional tests as described in ISO 7932:2004 + A1:2020 to allow for further identification of the Bacillus cereus present
Pathogens
Bacillus cereus Order Information

MYP Agar (Bacillus
cereus Medium)
Media
MYP Agar • NCM0062
+ NCM4032
+ NCM4017

47
NEOGEN.com
ISO 7937:2004
Microbiology of food and animal feeding stuffs — Horizontal method for
the enumeration of Clostridium perfringens — Colony count technique.
Sample

decimal dilutions
Inoculation
Supplemented TSC Agar

37°C for 20h ± 2h*
Enumeration
Thioglycollate Broth Lactose Gelatin

or
37°C for 18-24h* 37°C for 24h* + 5°C for 1h
+
Nitrate Motility
37°C for 24h
LS Broth
46°C for 18-24h
48
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*Under anaerobic conditions

†Additional steps required if colonies are not well isolated
Pathogens
Order Information
TSC Agar (Perfringens Thioglycollate Broth
Agar Base) (Fluid Thioglycollate
• Dehydrated Culture Medium)
TSC Agar • NCM0077 Media
+ NCM4017 • NCM0108
+ NCM4022
*Contact us for more information

49
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Campylobacter spp.
ISO 10272-1:2017
Microbiology of food and animal feeding stuffs — Horizontal method for
the enumeration of Campylobacter spp. — Part 1: Detection method.
PROTOCOL 1 PROTOCOL 2 PROTOCOL 3

Sample Sample Sample
Enrichment Enrichment
Bolton Broth Preston Broth

37°C for 4-6h 41.5°C for 24h ± 2h
41.5°C for 44-4h
Isolation
mCCD Agar
41.5°C ± 1°C for 44h ± 4h* Protocol 1:
+ For products
2nd selective medium** with low/stressed
Campylobacter and
low background
microflora, e.g.
frozen products.
Protocol 2:
Confirmation of Positives For products with
low Campylobacter
Non-selective Blood Agar levels and high
41.5°C ± 1°C for 24-48h background
microflora, e.g. raw
milk.
Protocol 3:
For products with
high levels of
Microscopy | Oxidase test Campylobacter, e.g.
raw poultry.
50
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* In a microaerobic atmosphere
** When following Protocol 1 only
Campylobacter spp.
ISO 10272-2:2017
Microbiology of the food chain — Horizontal method for the enumeration
of Campylobacter spp. — Part 2: Colony count technique.
Sample

decimal dilutions
Inoculation
mCCD Agar
41.5°C ± 1°C for 44h ± 4h*
Inoculation
Non-selective Blood Agar

25°C for 44h ± 4h
51
NEOGEN.com
* In a microaerobic atmosphere
Campylobacter spp.
Alternative Method – ANSR for Campylobacter
A rapid test for Campylobacter using molecular technology, providing
rapid recovery and detection in food samples in 20-24 hours.
Sample
091601
Enrichment
ANSR for Campylobacter

Enrichment Broth
42°C ± 1°C for 20-24h**
Lysis
Lysis Reagent Solution†

Detection
ANSR for Campylobacter

52
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* For poultry carcass rinse & sponge samples

** In microaerophilic or aerobic conditions. Additional enrichment with BPW needed for sponge samples
†Supplied as part of ANSR for Campylobacter kit
Pathogens
Campylobacter Order Information

ANSR for mCCD Agar
Campylobacter • Dehydrated Culture
• Molecular Test Kit Media
• 9872 • NCM0195
mCCD Agar + NCM4019
ANSR for Preston Broth
Campylobacter • Dehydrated Culture
Enrichment Broth† Media
• Dehydrated Culture • NCM0189
Media + NCM4038
• 9818
Bolton Broth
Media
• NCM0094
+ NCM4074

53
NEOGEN.com
Vibrio spp.
ISO 21872-1:2017
Microbiology of the food chain — Horizontal method for the determination
of Vibrio spp. — Part 1: Detection of potentially enteropathogenic
Vibrio parahaemolyticus, Vibrio cholerae and Vibrio vulnificus.
Sample
Primary Enrichment
1:10 dilution in ASPW

37°C ± 1°C* or
41.5°C± 1°C** for 6h ± 1h
ASPW
41.5°C for 18h ± 1h
Isolation
TCBS Agar
37°C ± 1°C for 24h ± 3h
+
2nd selective medium
Saline Nutrient Agar

37°C for 24h ± 3h
54
NEOGEN.com
*For fresh products

Biochemical tests **For deep frozen, dried or
salted products
Pathogens
Vibrio Order Information

ASPW (Alkaline Saline TCBS (Thiosulfate
Peptone Water) Citrate Bile Salts
• Dehydrated Culture Sucrose) Agar
TCBS Agar • NCM0175 Media
• NCM0052
Nutrient Agar
Media
• NCM0033
Nutrient Agar

55
NEOGEN.com
Shigella spp.
ISO 21567:2004
Microbiology of food and animal feeding stuffs —
Horizontal method for the detection of Shigella spp.
Sample
Enrichment
1:10 dilution in Shigella Broth

41.5°C ± 1°C for 16-20h*
Isolation
XLD Agar** MacConkey Agar** Hektoen Agar**

37°C ± 1°C for 20-24h + 37°C ± 1°C for 20-24h + 37°C ± 1°C for 20-24h
Biochemical tests | Serological tests

56
NEOGEN.com
*In anaerobic conditions

**If no typical colonies are seen and growth of other microorganisms is weak,
inoculate a further Nutrient Agar plate at 37ºC ± 1ºC for 18-24h
Pathogens
Shigella Order Information

Hektoen Agar XLD (Xylose Lysine
• NCM0006 Media
MacConkey Agar • NCM0021
MacConkey Agar XLD (Xylose Lysine
Media • Pre-Poured Plates
• NCM0174 • NCM3015
Nutrient Agar
Media
• NCM0033
XLD Agar

Nutrient Agar
57
NEOGEN.com
ISO 10273:2017
Microbiology of food chain — Horizontal method for
the detection of pathogenic Yersinia enterocolitica.
Sample
Enrichment
PSB Broth ITC Broth

25°C ± 1°C for 44h ± 4h*
+ 25°C ± 1°C for 44h ± 4h*
Isolation
CIN Agar
30°C ± 1°C for 24h ± 2h
Biochemical tests
58
NEOGEN.com
*In some cases, addition of 4.5 mL of KOH solution is required after incubation and before isolation
Pathogens
Order Information
CIN Agar (Yersinia
Selective Agar)
Media
CIN Agar • NCM0182
+ NCM4034

59
NEOGEN.com
Cronobacter spp.
ISO 22964:2017
Microbiology of the food chain — Horizontal
method for the detection of Cronobacter spp.
Sample
Primary Enrichment
Buffered Peptone Water (BPW)

34-38°C for 18h ± 2h
Cronobacter Selective Broth

41.5°C ± 1°C for 24h ± 2h
Isolation
Chromogenic Cronobacter Isolation Agar

41.5°C ± 1°C for 24h ± 2h
Non-selective medium*
41.5°C ± 1°C for 24h ± 2h
Biochemical tests
60
NEOGEN.com
* Tryptone Soy Agar (TSA)

Pathogens
Cronobacter Order Information

BPW (ISO) (Buffered Cronobacter Selective
Peptone Water) Broth
Media Media
Cronobacter Isolation • NCM0015 • NCM0227
+ NCM4004
Agar
BPW (ISO) (Buffered Cronobacter Isolation
Peptone Water) Agar (CCI)
• Ready-to-Reconstitute • Dehydrated Culture
Media Bags Media
• NCM3202 • NCM1008
BPW HQ (ISO) (Buffered Tryptone Soy Agar
Peptone Water) (TSA)
Media Media
• NCM0270 • NCM0020
BPW HQ (ISO) (Buffered
Peptone Water)
•P repared Media Bags
• NCM3402

61
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62
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63
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Total Viable Count
ISO 4833-1:2013
Microbiology of the food chain — Horizontal method for the
enumeration of microorganisms — Part 1: Colony count at
30°C by the pour plate technique.
Sample

decimal dilutions
Sample
PCA
30°C ± 1°C for 72h ± 3h
Enumeration
64
NEOGEN.com
Total Viable Count
ISO 4833-2:2013
enumeration of microorganisms — Part 2: Colony count at
30°C by the surface plating technique.
Sample

decimal dilutions
Sample
PCA
30°C for 72h ± 3h
Enumeration
65
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Total Viable Count
ISO 17410:2019
Microbiology of the food chain — Horizontal method for
the enumeration of psychrotrophic microorganisms.
Sample

decimal dilutions
Inoculation
PCA
6.5°C ± 1°C for 10 days
Enumeration
66
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Total Viable Count
Alternative Method – Soleris
A rapid optical system for the detection of bacteria, yeasts and moulds.
This method offers semi-quantitative enumeration of TVC.
Sample **
091601
decimal dilutions
Detection
Inoculation of appropriate vial:

NF-TVC / NF-105
30°C for 24h in Soleris Instrument
Results
Optional confirmation with PCA
67 NEOGEN.com
*NF-TVC method for selected matrices

** UHT samples should undergo pre-incubation at 30°C for 48-72h
Total Viable Count
Alternative Method – Soleris (UHT Milk Protocol)
Actalia ISO 16140 UHT Milk Protocol
This method was used in a study performed by the Microbiology
Laboratory of Actalia Cecalait in Poligny, France and followed the
protocols described in ISO/FDIS 16140 for sterility testing of UHT milk.
Sample
Incubation
Pre-incubation of sample
30°C for 48-72h
Detection

NF-105
Results
Optional confirmation with PCA
68
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Total Viable Count

Order Information
PCA (Plate Count Agar) Soleris NF-105 Vial
• Dehydrated Culture • Prepared Media Vials
Media • NF-105
• NCM0010
PCA Soleris NF-TVC Vial
•P repared Media Vials
• NF-TVC

69
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E. coli
ISO 16649-1:2018
Microbiology of food and animal feeding stuffs — Horizontal method for the enumeration
of β-glucuronidase-positive Escherichia coli — Part 1: Colony-count technique at 44°C
using membranes and 5-bromo-4-chloro-3-indolyl-β-D-glucuronide.
Sample

decimal dilutions
Resuscitation
MMGA*
37°C for 4h ± 15 min
Inoculation
Tryptone Bile Glucuronide Agar (TBX)*

44°C for 20-24h
Enumeration
70
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*Membrane filter required

E. coli
ISO 16649-2:2001
using 5-bromo-4-chloro-3-indolyl-β-D-glucuronide.
Sample

decimal dilutions
Resuscitation
Tryptone Bile Glucuronide Agar (TBX)

44°C for 18-24h
Enumeration
71
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E. coli
ISO 16649-3:2015
using membranes and 5-bromo-4-chloro-3-indolyl-β-D-glucuronide.
Sample

decimal dilutions
Primary Inoculation
Minerals Modified
Glutamate Broth*
37°C ± 1°C for 24h ± 2h
Secondary Inoculation**

44°C ± 1°C for 18-24h
Enumeration†
72
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*3-5 tubes per dilution required

**Subculture required for each tube of medium showing acid production
†For enumeration method only
E. coli
ISO 7251-1:2005
Microbiology of food and animal feeding stuffs — Horizontal method
for the detection and enumeration of presumptive Escherichia coli —
Most Probable Number technique.
Sample

decimal dilutions
Primary Inoculation
Lauryl Tryptose Broth*

(Lauryl Sulphate Broth)
37°C ± 1°C for 24h ± 2h
Secondary Inoculation
E. coli (EC) Broth

44°C ± 1°C for 24h ± 2h
Enumeration†
Indole Test 73
NEOGEN.com
*3-5 tubes per dilution required

E. coli
A rapid optical system for the detection of E. coli. This
method offers semi-quantitative enumeration of E. coli.
Sample
960801
Detection
Inoculation of EC-104 vial

44°C for 18-24h in Soleris Instrument
Results

44°C ± 1°C for 18-24h
74
NEOGEN.com
E. coli Order Information

EC Broth (EC Medium) MMGA (Minerals
• Dehydrated Culture Modified Glutamate
Media Agar)
• NCM0065 • Dehydrated Culture
TBX Media
• NCM0179
+ NCM0181
+ NCM0178
Lauryl Tryptose Broth Soleris EC-104 Vial
Media • EC-104
• NCM0032
Minerals Modified TBX (Tryptone Bile
Glutamate Broth Glucuronide Agar)
Media Media
• NCM0186 • NCM1001
+ NCM0181
+ NCM0178

75
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Coliforms
ISO 4831:2006
method for the detection and enumeration of Coliforms —
Most Probable Number technique.
Sample

decimal dilutions
Primary Inoculation
Lauryl Tryptose Broth

(Lauryl Sulphate Broth)
37°C ± 1°C for 24h ± 2h*
Secondary Inoculation**
Brilliant Green Bile 2% Broth

37°C ± 1°C for 24h ± 2h*
Enumeration†
76
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*If at this stage no gas occurs, incubate for up to 48h ± 2h

Coliforms
ISO 4832:2006
method for the enumeration of Coliforms —
Colony count technique.
Sample

decimal dilutions
Inoculation
Violet Red Bile Lactose (VRBL) Agar

37°C ± 1°C for 24h ± 2h
Enumeration
Confirmation of atypical colonies
Brilliant Green Bile 2% Broth

30°C ± 1°C for 24h ± 2h
77
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Coliforms
Alternative Method - Soleris
A rapid optical system for the detection of Coliforms. This
method offers semi-quantitative enumeration of Coliforms.
Sample
960801
Detection

CC-102 / CC-109
Results
Optional confirmation with Violet Red Bile Lactose (VRBL) Agar
78
NEOGEN.com
Coliforms Order Information

Brilliant Green Bile 2% Soleris CC-109 Vial
Broth • Prepared Media Vials
• Dehydrated Culture • CC-109
Media
VRBL Agar • NCM0048
Lauryl Tryptose Broth VRBL (Violet Red Bile
• Dehydrated Culture Lactose) Agar
• NCM0032 Media
• NCM0089
Soleris CC-102 Vial
• Prepared Media Vials
• CC-102

79
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Enterobacteriaceae
ISO 21528-1:2017
detection and enumeration of Enterobacteriaceae —
Part 1: Detection of Enterobacteriaceae.
Sample
Initial Suspension / Decimal Dilutions
BPW
27°C ± 1°C for 18h ± 2h
Inoculation
Violet Red Bile Glucose (VRBG) Agar
37°C ± 1°C for 24h ± 2h
Detection
37°C for 24h ± 2h
Biochemical tests
80
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*Nutrient Agar
Enterobacteriaceae
ISO 21528-2:2017
for the detection and enumeration of Enterobacteriaceae —
Part 2: Colony-count method.
Sample

decimal dilutions
Inoculation
Violet Red Bile Glucose (VRBG) Agar
37°C ± 1°C for 24h ± 2h
Enumeration
37°C for 24h ± 2h
Biochemical tests
81
NEOGEN.com
*Nutrient Agar
Enterobacteriaceae
MicroVal Validation Certification 2018LR83
A rapid optical system for the detection of Enterobacteriaceae. This
method offers semi-quantitative enumeration of Enterobacteriaceae.
Sample
960801

S2-EBAC9
Results
Optional confirmation with VRBG Agar
82
NEOGEN.com
Enterobacteriaceae
Order Information
BPW (ISO) (Buffered BPW HQ (ISO) (Buffered
Peptone Water) Broth Peptone Water)
• Dehydrated Culture • Prepared Media Bags
Media • NCM3402
• NCM0015
VRBG Agar
BPW (ISO) (Buffered Nutrient Agar
Peptone Water) • Dehydrated Culture
• Ready-to-Reconstitute Media
Media Bags • NCM0033
• NCM3202
BPW HQ (ISO) (Buffered Soleris S2-EBAC9 Vial
Peptone Water) • Prepared Media Vials
• Dehydrated Culture • S2-EBAC9
Media
• NCM0270
BPW HQ (ISO) (Buffered VRBG (Violet Red Bile
Peptone Water) Glucose) Agar
• Ready-to-Reconstitute • Dehydrated Culture
Nutrient Agar Media Bags Media
• NCM3207 • NCM0041

83
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Yeasts and Moulds
ISO 21527-1:2008
enumeration of yeasts and moulds — Part 1: Colony count technique in
products with water activity greater than 0.95.
Sample

decimal dilutions
Inoculation
DRBC Agar
25°C ± 1°C for 5 days
Enumeration
84
NEOGEN.com
Yeasts and Moulds
ISO 21527-2:2008
enumeration of yeasts and moulds — Part 2: Colony count technique in
products with water activity less than or equal to 0.95.
Sample

decimal dilutions
Inoculation
DG-18 Agar
25°C ± 1°C for 5 days
Enumeration
85
NEOGEN.com
Yeasts and Moulds
For rapid and accurate detection and enumeration of yeasts and
moulds across a spectrum of sample types including food,
dairy, beverages, nutraceuticals and cosmetics.
Sample
960801
Addition of YI-110C supplement
Inoculation
Inoculation of DYM-109C vial

Results
Optional confirmation with VRBG Agar
86
NEOGEN.com
Yeasts and Moulds

Order Information
DG‑18 (Dichloran Soleris DYM-109C Vial
Glycerol Agar Base) • Prepared Media Vials
• Dehydrated Culture • DYM-109C
Media
• NCM0081
DRBC Agar
DRBC (Dichloran Soleris Y1-110C
Rose Bengal Supplement
Chloramphenicol) Agar •P repared Media Vials
• Dehydrated Culture • Y1-110C
Media
• NCM0082
DG18 Agar

87
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88
STAPHYLOCOCCUS
NEOGEN.com
Staphylococcus spp.
ISO 6888-1:2020
enumeration of coagulase-positive Staphylococci (Staphylococcus aureus
and other species) — Part 1: Technique using Baird-Parker agar medium.
Sample

decimal dilutions
Inoculation
Baird Parker Agar

36°C ± 2°C for 24h ± 2h*
Enumeration
BHI
36°C ± 2°C for 24 ± 2h
Rabbit Plasma
36°C ± 2°C for 5h ± 1h**
89
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*Incubate for a further 24h ± 2h if necessary

**Incubate for 24h if negative
Staphylococcus spp.
ISO 6888-2:2021
enumeration of coagulase-positive Staphylococci (Staphylococcus aureus and
other species) — Part 2: Technique using Rabbit Plasma Fibrinogen Agar Medium.
Sample
decimal dilutions
Isolation
RPF Agar
36°C ± 2°C for 24 ± 2h*
Enumeration
90
NEOGEN.com
*Incubate for a further 48 ± 4h if no typical colonies are observed

Staphylococcus spp.
ISO 6888-3:2003
enumeration of coagulase-positive Staphylococci (Staphylococcus aureus
and other species) — Part 3: Detection and MPN technique for low numbers.
Sample
decimal dilutions
Primary Inoculation
Giolitti and Cantoni Broth

37°C for 24h ± 2h**
Secondary Inoculation
Baird Parker Agar RPF Agar

37°C ± 1°C for 24h ± 2h** or 37°C ± 1°C for 24h ± 2h**
Enumeration* Enumeration*
BHI
37°C ± 1°C for 24h ± 2h†
91
BHI
37°C ± 1°C for 4-6h or 24h ± 2h†
NEOGEN.com
*For enumeration method only

**Incubate for a further 24h ± 2h if necessary
†
24h +/- 2 incubation for verification of results
Staphylococcus spp.
Provides rapid detection of Staphylococcus spp. contamination,
including S. aureus, in all areas of food production. This method
offers semi-quantitative enumeration of Staphylococcus.
Sample
Detection
Inoculation of SM-118 vial

37°C for 16h in Soleris instrument
Results
RPF Agar
37°C ± 1°C for 18-24h
92
NEOGEN.com
Staphylococcus
Order Information
Baird Parker Agar RPF Agar
Media Media
• NCM0200 • NCM0200
RPF Agar + NCM4010 + NCM4052
BHI (Brain-Heart Soleris SM-118 Vial
Infusion) Broth • Prepared Media Vials
• Dehydrated Culture • SM-118
Media
• NCM0016
Modified Giolitti &
Cantoni Broth
Media
• NCM0184
+ NCM4012
Baird Parker Agar

93
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94
LACTOBACILLIS
NEOGEN.com
ISO 15214:1998
for the enumeration of mesophilic Lactic Acid Bacteria — Colony
count technique at 30°C.
Sample

decimal dilutions
Inoculation
MRS Agar
30°C for 72h ± 3h
Enumeration
95
NEOGEN.com
A rapid optical system for the detection of lactic acid bacteria.
The Soleris method offers presence-absence detection and
semi-quantitativeenumeration of lactic acid bacteria.
Sample
Detection
Inoculation of DLA-109 vial

Results
Optional confirmation of heterofermentative lactic
acid bacteria with Lactobacilli MRS Broth
96
NEOGEN.com

Order Information
Lactobacilli MRS Broth Soleris DLA-109 Vial
Media • DLA-109
• NCM0079
MRS Agar MRS Agar
•D ehydrated Culture
Media
• NCM0190

97
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NEOGEN.com 98
PSEUDOMONAS
Pseudomonas spp.
ISO 13720:2010
Meat and meat products — Enumeration of presumptive Pseudomonas spp.
present in meat and meat products, including poultry.
Sample

decimal dilutions
Inoculation
CFC Agar
25°C ± 1°C for 44h ± 4h
Enumeration
Biochemical tests
99
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Pseudomonas spp.
A rapid optical system for the detection of Pseudomonas spp. including
Pseudomonas aeruginosa. The Soleris method offers presence-absence
detection and semi-quantitative enumeration of Pseudomonas spp.
Sample
Inoculation
Inoculation of PD-109 vial

Results
Optional confirmation with CFC Agar
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Pseudomonas Order Information

CFC Agar (Pseudomonas Soleris PD-109 Vial
Agar Base) • Prepared Media Vials
• Dehydrated Culture • PD-109
Media
CFC • NCM0083
+ NCM4023

101
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NEOGEN - Your Trusted
Partner for Microbiology
At NEOGEN we believe in partnerships. That’s why we offer a range
of training and support options to help you and your team build
knowledge and skills.
We are here to help at every step, whether you need:
• information to help choose the right product
• support to define a verification study
• guidance on implementing a product onsite
• ongoing training to ensure best practice
We can support through a combination of:
LabLive
Support, training and troubleshooting from the comfort of your own
facilities with our interactive online video tool
Onsite Training
We deliver in-depth product training where you need it
Webinars & Workshops

Meet our team and learn about the latest trends and developments
Contact Us
For more information contact us by email FoodSafetyUK@NEOGEN.com,
call +44 (0) 1292 525 626 or visit NEOGEN.com.
102
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©NEOGEN Corporation, 2021. NEOGEN and Soleris are registered trademarks of NEOGEN Corporation, Lansing, MI 48912 U.S.

Guide To Food Microbiology Testing

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Guide To Food Microbiology Testing

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Edition 3

Guide to Food Microbiology Testing

This convenient microbiology reference tool

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Dehydrated Culture Media

Pre-Poured Plates Prepared Media Bags Prepared Media Tubes

Spoilage and Indicator Organisms

E. coli Order Information 75

Yeasts and Moulds

Lactic Acid Bacteria

Alternative Method – Soleris 100

Pseudomonas Order Information 101

This signifies that a minimum of two plates should be inoculated via

Initial Suspension / Primary Dilution

Initial Suspension / Primary Dilution

Further Decimal Dilutions

It is recommended that, for the preparation of a diluent, dehydrated

For details on sampling techniques and sample preparation please refer

Inoculum preparation from Reference Material or from a reference strain

1:10 1:3 1:3 1:3 1:3

Reference Dilution Dilution Dilution Dilution Dilution

This is an example to target the LOD50 of NEOGEN Methods

Step 3 continued on next page

• Spike the enrichment bags using • Pipette 1 mL of each dilution into:

Dilution Dilution Dilution Dilution Dilution Unspiked

Expected High level Medium level Low level

Dilution to obtain a 30 cfu/mL solution

Step 3 continued on next page

 ipette 1 mL of Dilution A into 2 petri dishes to verify inoculum level

Dilution Dilution Dilution Dilution Dilution

Verifying the Soleris Method Verifying All Other Methods

Day 2-3 continued on next page

Verifying the Soleris Verifying All Other Methods

Dilution B Dilution C Dilution D

Dilution E Unspiked Sample

ANSR OBOP ISO

• Once the detection method is complete determine positive results and

• The number of positive results out of the 5 replicates tested using

protocol 3 shall have minimum of 6 positive results out of 7

Half Fraser Broth

If confirming suspected Listeria spp. If confirming suspected Listeria mono.

*PALCAM or Oxford Listeria Agar

Listeria Chromogenic Agar (O&A)

If confirming suspected Listeria spp. If confirming suspected Listeria mono.

Take 5* blue/green colonies with or without halo and isolate

From isolated colonies suspended in a drop of peroxide

If negative, stop - not

*To confirm OBOP-L take only 1 colony

Take 5* blue/green colonies with a halo and streak them

If less than 5 colonies with halos are available on the plate

Check for haemolysis

If haemolysis positive, carry out D-Xylose

If L-Rhamnose If L-Rhamnose If negative, stop

*To confirm OBOP-L take only 1 colony

Listeria Chromogenic Agar (O&A)

* Listeria Chromogenic Agar (O&A) should be used to detect Listeria

Agar Listeria O&A (LCA (O&A))

Listeria monocytogenes - Listeria spp.

Standard confirmation tests

   Validated Pooling

6 7 8 9 10 Lysis Reagent Solution*

Alternatively confirm as described in ISO 11290-1:2017

*Supplied as part of ANSR for Listeria kit

• Spike the enrichment bags using • Pipette 1 mL of each dilution into:

ipette 1 mL of Dilution A into 2 petri dishes to verify inoculum level

• Once the detection method is complete determine positive results and

• The number of positive results out of the 5 replicates tested using

Validated Pooling