PROTOCOL ISO

WATER ANALYSES

NF
GUIDE
RISK
MICROORGANISMS

ANALYSES
NF
PROTOCOL
CONFIRMATION

STANDARDS
OF FOOD
STANDARDS

PROTOCOL

COSMETICS
STANDARD RISK

YEASTS AND
ENRICHMENT

FOOD
METHODS MOULDS

MICROORGANISMS
CULTURE
PROTOCOL DETECTION
ISO

FOOD microbiology

Version dated June 2020

GMR_Food_VEN102019
10 2019

Page 2 www.biokar-diagnostics.com
 CONTENTS

1 - MICROBIOLOGY OF FOOD

Bacillus cereus

Detection and enumeration of presumptive Bacillus cereus (MPN) NF EN ISO 21871 V 08-063 pages 8-9

Enumeration of presumptive Bacillus cereus NF EN ISO 7932 V 08-023 pages 10-11

COMPASS® Bacillus cereus Agar

Alternative method pages 12-13
Enumeration of Bacillus cereus without confirmation

Campylobacter spp.

Detection of Campylobacter spp. - part 1 NF EN ISO 10272-1 V 08-026-1 pages 14-15

Enumeration of Campylobacter spp. - part 2 NF EN ISO 10272-2 V 08-026-2 pages 16-17

Clostridium perfringens

Enumeration of Clostridium perfringens NF EN ISO 7937 V 08-019 pages 18-19

Coliform bacteria

Enumeration of coliforms (Colony-count technique) NF ISO 4832/FIL 73 B V 08-015 pages 20-21

Detection and enumeration of coliforms (Most probable number technique) NF ISO 4831/FIL 73 B V 08-016 pages 22-23

Enumeration of presumptive coliforms NF V 08-050 V 08-050 page 24

Enumeration of thermotolerant coliforms NF V 08-060 V 08-060 page 25

Cronobacter spp.

Detection of Cronobacter spp. NF EN ISO 22964 V 08-746 pages 26-27

Enterobacteria

Detection and enumeration of Enterobacteriaceae (with pre-enrichment) NF EN ISO 21528-1 V 08-039-1 pages 28-29

Detection and enumeration of Enterobacteriaceae NF EN ISO 21528-2 V 08-039-2 pages 30-31

Enumeration of presumptive Enterobacteria NF V 08-054 V 08-054 pages 32-33

Escherichia coli

Enumeration of β-glucuronidase-positive Escherichia coli NF ISO 16649-1 V 08-031-1 pages 34-35

Enumeration of β-glucuronidase-positive Escherichia coli NF ISO 16649-2 V 08-031-2 pages 36-37

Enumeration of β-glucuronidase-positive Escherichia coli NF EN ISO 16649-3 V 08-031-3 pages 38-39

Enumeration of presumptive Escherichia coli NF ISO 7251 V 08-020 pages 40-41

Detection of Escherichia coli O157

NF EN ISO 16654 V 08-032 pages 42-43
Amendment A1

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Mesophilic lactic acid bacteria

Enumeration of Mesophilic lactic acid bacteria NF ISO 15214 V 08-030 page 44

Yeasts and moulds

Enumeration of yeasts and moulds NF V 08-036 V 08-036 page 45

Enumeration of yeasts and moulds

NF ISO 21527-1 V 08-040-1 pages 46-47
(products with water activity greater than 0.95)

Enumeration of yeasts and moulds

NF ISO 21527-2 V 08-040-2 pages 48-49
(products with water activity less than or equal to 0.95)

Enumeration of yeasts and moulds by colony-count technique at 25°C

NF V 08-059 V 08-059 pages 50-51
Routine method

SYMPHONY® Agar
Alternative method page 52
Enumeration of yeasts and moulds

Listeria spp.

Detection of Listeria monocytogenes and Listeria spp. NF EN ISO 11290-1 V 08-028-1 pages 53-55

COMPASS® Listeria Agar

Alternative method pages 56-57
Detection of Listeria monocytogenes and Listeria spp.

Enumeration of Listeria monocytogenes NF EN ISO 11290-2 V 08-028-2 pages 58-59

COMPASS® Listeria Agar

Alternative method pages 60-61
Enumeration of Listeria monocytogenes

Salmonella spp.

Detection of Salmonella spp. NF EN ISO 6579-1 V 08-013 pages 62-67

SESAME Salmonella TEST

Alternative method pages 68-69
Detection of Salmonella spp.

IRIS Salmonella® Agar

Alternative method pages 70-71
Detection of Salmonella spp.

Shigella spp.

Detection of Shigella spp. NF EN ISO 21567 V 08-411 pages 72-73

Staphylococci

NF EN ISO 6888-1
Enumeration of coagulase-positive Staphylococci Amendment A1 V 08-014-1 pages 74-75
Amendment A2

Enumeration of coagulase-positive Staphylococci NF EN ISO 6888-2

V 08-014-2 pages 76-77
(Rabbit Plasma Fibrinogen agar) Amendment A1

EASY Staph® Agar

Alternative method pages 78-79
Enumeration of coagulase-positive Staphylococci

Enumeration of coagulase-positive Staphylococci (low numbers) NF EN ISO 6888-3 V 08-014-3 pages 80-81

Routine method for the enumeration of coagulase-positive Staphylococci

by colony-count technique at 37°C - Part 1 Technique with confirmation of NF V 08-057-1 V 08-057-1 pages 82-83
the colonies

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Sulfite-reducing microorganisms
Enumeration of sulfite-reducing anaerobic bacteria NF EN ISO 15213 V 08-029 pages 84-85
Enumeration of sulfite-reducing bacteria NF V 08-061 V 08-061 pages 86-87
Vibrio spp.

Detection of potentially enteropathogenic Vibrio parahaemolyticus,

NF EN ISO 21872-1 V 08-064-1 pages 88-89
Vibrio cholerae and Vibrio vulnificus

Yersinia enterolitica

Detection of pathogenic Yersinia enterocolitica NF EN ISO 10273 V 08-027 pages 90-91

Total microorganisms

Enumeration of microorganisms (pour plate technique) NF EN ISO 4833-1 V 08-011-1 page 92

Enumeration of microorganisms (surface plating technique) NF EN ISO 4833-2 V 08-011-2 page 93

Enumeration of microorganisms (spirale method) XP V 08-034 V 08-034 page 94

Enumeration of psychrotrophic microorganisms NF ISO 17410 V 08-033 page 95

Preparation of test samples, initial suspension and decimal dilutions for microbiological examination

Part 1: General rules for the preparation of the initial suspension and
NF EN ISO 6887-1 V 08-010-1 page 96
decimal dilutions

Part 2: Specific rules for the preparation of meat and meat products NF EN ISO 6887-2 V 08-010-2 page 97

Part 3: Specific rules for the preparation of fish and fishery products NF EN ISO 6887-3 V 08-010-3 page 98

Part 4: Specific rules for the preparation of products other than milk and
NF EN ISO 6887-4 V 08-010-4 Pages 99-100
milk products, meat and meat products, and fish and fishery products

Part 5: Specific rules for the preparation of milk and milk products NF EN ISO 6887-5 V 08-010-5 pages 101-102

Part 6: Specific rules for the preparation of samples taken at the primary
NF EN ISO 6887-6 V08-010-6 page 103
production stage

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2 - MICROBIOLOGY OF FOODS BY SECTOR

Animal feeding stuffs

Enumeration of Bifidobacterium spp. NF EN 15785 V 18-238 pages 104-105

Enumeration of presumptive Bacillus spp. NF EN 15784 V 18-237 page 106

Enumeration of Enterococcus (E. faecium) spp. NF EN 15788 V 18-232 page 107

Enumeration of Lactobacillus spp. NF EN 15787 V 18-231 pages 108-109

Enumeration of Pediococcus spp. NF EN 15786 V 18-230 pages 110-111

Canned products

Enumeration of spores by colony-counts NF V 08-602 V 08-602 pages 112-113

Milk and milk products

Enumeration of presumptive Escherichia coli - Part 1: Most probable

NF ISO 11866-1 IDF 170-1 pages 114-115
number technique using MUG

Enumeration of presumptive Escherichia coli - Part 2: Colony-count

NF ISO 11866-2 IDF 170-2 pages 116-117
technique at 44°C using membranes

Enumeration of presumptive Lactobacillus acidophilus on a selective

ISO 20128 IDF 192 page 118
medium - Colony-count technique at 37°C

Enumeration of colony-forming units of yeasts and/or moulds

ISO 6611 FIL 94 page 119
Colony-count technique at 25°C

Enumeration of Pseudomonas spp. XP ISO/TS 11059 V 04-025 pages 120-121

RHAPSODY Agar® Alternative

page 122
Alternative rapid detection method for Pseudomonas spp. method

Enumeration of characteristic microorganisms (Yogurt)

ISO 7889 IDF 117 page 123
Count technique at 37°C

Identification of characteristic microorganisms Lactobacillus delbrueckii

ISO 9232 IDF 146 pages 124-125
subsp. bulgaricus and Streptococcus thermophilus (Yogurt)

Enumeration of contaminating microorganisms (Butter)

ISO 13559 IDF 153 page 126
Colony-count technique at 30°C

Enumeration of microorganisms (Milk)

ISO 8553 FIL 131 page 127
Plate-loop technique at 30°C

Enumeration of colony-forming units of psychrotrophic microorganisms

ISO 6730 IDF 101 page 128
(Milk) - Colony-count technique at 6.5°C

Estimation of psychrotrophic microorganisms (Milk)

ISO 8552 FIL 132 page 129
Colony-count technique at 21°C

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 Meat and meat products

Enumeration of presumptive Pseudomonas spp. NF EN ISO 13720 V 08-504 page 130

RHAPSODY Agar® Alternative

page 131
Alternative rapid detection method for Pseudomonas spp. method

Appendix A: Other standards relating to the microbiology of foods page 132

Appendix B: List of products pages 133-138

BIOKAR Diagnostics - Support and services page 139

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 Horizontal method for the determination of
low numbers of presumptive Bacillus cereus
Most probable number technique and detection method

NF EN ISO 21871: 07-2006

V 08-063

Enumeration protocol

Initial suspension
(decimal dilutions) 1

10 mL in 1 mL in
10 mL of broth 10 mL of broth

Double strength Single strength

TSPB TSPB

 48 ± 4 hrs at 30°C 2

MYP agar 3

 18 to 24 hrs at 30°C
+ 24 hrs if necessary

Enumeration and confirmation

of Bacillus cereus 4

Detection protocol
Inoculate the initial suspension in a tube of single strength or double strength medium, and follow the
protocol described for enumeration.

1
Inoculate the initial suspension in three tubes of double strength medium and three tubes of single strength medium.
The decimal dilutions are also inoculated in three tubes of single strength medium.
2
After incubation, reinoculate each tube using a plate-loop on selective medium.
3
Polymyxin pyruvate egg yolk mannitol bromothymol blue agar (PEMBA) may also be used as a selective medium with
incubation at 37°C.
4
Select three colonies from each plate. When using MYP agar, carry out a haemolysis test on sheep blood agar with
incubation from 18 to 24 hrs at 30°C for confirmation.
When using PEMBA, carry out a haemolysis test on sheep blood agar with incubation from 18 to 24 hrs at 37°C or
microbiological examination for confirmation.

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 Bacillus cereus
Section Media and reagents Compliance
Refer to the section of standard NF EN ISO 6887 corresponding to the
5.2 Diluent
product to be examined, pages 96 to 103.
- Tryptone soya polymyxin broth (TSPB)
Trypticase soy broth (TSB)
50 x 10 mL tubes - BM03008
10 x 100 mL vials - BM00908 Total 5
5.3 Selective 10 x 90 mL vials - BM17908
enrichment medium 2 x 5 L flexible bags - BM16608
500 g vial - BK046HA
5 kg drum - BK046GC
Polymyxin B selective supplement
Total
10 vials - BS00708

- Polymyxin pyruvate egg yolk mannitol bromothymol blue

agar (PEMBA)
Base medium
- -
5.4 Solid selective Egg yolk emulsion with polymyxin B
medium Total
10 x 50 mL vials - BS05508
Polymyxin B selective supplement
Total
10 vials - BS00708
Sterile egg yolk emulsion
Total
10 x 50 mL vials - BS06608
- Mannitol egg yolk polymyxin (MYP) agar
B. cereus selective agar (acc. to MOSSEL)
20 Petri dishes Ø 90 mm - BM03808 Total 5
120 Petri dishes Ø 90 mm - BM19908
B. cereus selective agar (base acc. to MOSSEL)
5.5 Solid selective 500 g vial - BK116HA Total 5
medium Egg yolk emulsion with polymyxin B
10 x 50 mL vials - BS05508 Total
Polymyxin B selective supplement
10 vials - BS00708 Total
Sterile egg yolk emulsion
10 x 50 L vials - BS06608 Total
- Malachite green oxalate solution
- -
- Sudan black B solution
5.6 Reagents for - -
microscopic - Xylene
identification - -
- Safranin solution
- -

- Base medium: Sheep blood agar

Tryptone soy (blood agar base)
5.7 Confirmation 500 g vial - BK028HA Equivalent
medium - Sheep blood with fibrin
-
-

5
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

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 Horizontal method for the enumeration of
presumptive Bacillus cereus
Colony-count technique at 30°C
NF EN ISO 7932: 07-2005
V 08-023

Initial suspension
(decimal dilutions) 1

0.1 mL on surface 2

MYP medium

 18 to 24 hrs at 30°C
+ 24 hrs if necessary

Enumeration and confirmation 3

of Bacillus cereus

Sheep blood agar

 24 ± 2 hrs at 30°C

Interpretation of the haemolytic reaction

1
Carry out the operation in duplicate for each dilution.
2
If, for certain products, it is preferable to estimate low numbers of B. cereus, the enumeration limits may be increased by a
power of 10, examining 1.0 ml of test sample for liquids, or 1.0 ml of the initial suspension for other products. Apply 1 ml of
inoculum using a sterile spreader either to the surface of a large Petri dish (140 mm) of agar medium, or to the surface of three
small Petri dishes (90 mm) of agar medium. In both cases, carry out these operations in duplicate, so as to prepare two large
dishes or six small dishes.
3
Randomly select five colonies from each plate.

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 Bacillus cereus
Section Media and reagents Compliance
Refer to the section of standard NF EN ISO 6887 corresponding to the
5.1 Diluent
product to be examined, pages 96 to 103.
- Mannitol egg yolk polymyxin (MYP) agar
B. cereus selective agar (acc. to MOSSEL)
20 Petri dishes Ø 90 mm - BM03808 Total 4
120 Petri dishes Ø 90 mm - BM19908
B. cereus selective agar (base acc. to MOSSEL)
500 g vial - BK116HA
5.2 Agar medium
Egg yolk emulsion with polymyxin B
10 x 50 mL vials - BS05508 Total 4
Polymyxin B selective supplement
10 vials - BS00708
Sterile egg yolk emulsion

- Base medium: Sheep blood agar

Tryptone soy (blood agar base) Equivalent
5.3 Confirmation 500 g vial - BK028HA
medium - Sheep blood with fibrin
-
-

4
Tryptone is a “peptone” obtained by pancreatic digestion of casein.

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 COMPASS® Bacillus cereus Agar
Alternative method for the enumeration of presumptive Bacillus cereus

BKR 23/06-02/10
Alternative food
analysis method
www.afnor-validation.org /en

Initial suspension
(decimal dilutions)

1 mL in poured plate
0.1 mL on surface 1

COMPASS® Bacillus cereus Agar

 24 to 27 hrs at 30 ± 1°C

Enumeration of presumptive
Bacillus cereus

Without confirmation

1
Inoculation is also possible by spreading 1 mL on the surface of 3 Petri dishes Ø 90 mm.

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 Bacillus cereus
Refer to the section of standard NF EN ISO 6887 corresponding to the
Diluent
product to be examined, pages 96 to 103.

COMPASS® Bacillus cereus Agar pre-poured

20 Petri dishes Ø 90 mm - BM12608
COMPASS® Bacillus cereus Agar (base)
Selective medium
10 x 100 mL vials - BM13008
Supplement for COMPASS® Bacillus cereus Agar
10 vials q.s. 100 mL - BS06908

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Horizontal method for detection and enumeration of Campylobacter spp.
Part 1: Detection method

NF EN ISO 10272-1: 07-2017

V 08-026-1

Detection procedure A Detection procedure B Detection procedure C

10 g (or mL) of sample in 10 g (or mL) of sample in Sample (using a swab or

90 mL of BOLTON broth 90 mL of PRESTON broth plate-loop)

 4 to 6 hrs at 37°C 1
 24 ± 2 hrs at 41.5°C 1 Direct inoculation
1
 44 ± 4 hrs at 41.5°C

mCCD agar Second medium mCCD agar mCCD agar

 44 ± 4 hrs  appropriate  44 ± 4 hrs  44 ± 4 hrs

at 41.5°C 1 conditions at 41.5°C 1 at 41.5°C 1

Detection and confirmation 2 of Campylobacter spp.

Columbia blood agar

 24 to 48 hrs at 41.5°C

- Examination of morphology and motility

- Growth 25°C, in a microaerobic atmosphere
- Detection of oxidase activity
- Identification of the Campylobacter species (optional)

1
Incubation should be carried out in a microaerobic atmosphere.
2
For the confirmation tests, select at least 1 characteristic colony, then another 4 if the first is negative.

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 Campylobacter spp.
Section Media and reagents Compliance
- Bolton broth
B.2/B.3 Enrichment - -
media - Preston broth
- -
- Modified charcoal cefoperazone deoxycholate (mCCD)
agar
B.4/B.5 Isolation - -
media
The choice of the second medium is left to the discretion of the testing
laboratory
- Columbia blood agar
Columbia (base) agar Partial 3
500 g vial - BK019HA
- Sterile sheep or horse blood
-
-
B.6/B.7/B.8/B.9/B.10 - Oxidase reagent
Confirmation and - -
identification media - Reagent for the detection of catalase activity
and reagents -
-
- Reagent for the detection of hydrolysis of hippurate
- -
- Indoxyl acetate discs
-
-

3
pH 7.3 ± 0.2 instead of 7.4 ± 0.2.

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Horizontal method for detection and enumeration of Campylobacter spp.
Part 2: Colony-count technique

NF EN ISO 10272-2: 07-2017

V 08-026-2

Initial suspension
(decimal dilutions)

0.1 mL on surface 1, 2

mCCD agar

 44 ± 4 hrs at 41.5°C
microaerobic atmosphere

Enumeration and confirmation 3 of Campylobacter spp.

Columbia blood agar

 24 to 48 hrs at 41.5°C

- Examination of morphology and motility

- Growth 25°C, in a microaerobic atmosphere
- Detection of oxidase activity
- Identification of the Campylobacter species (optional)

1
Carry out the operation in duplicate for each dilution.
2
To estimate low numbers of Campylobacter, the enumeration limit may be reduced by a factor of 10, examining 1.0 mL of initial
suspension. Apply 1.0 mL of initial suspension to 1 plate 140 mm in diameter or 3 plates 90 mm in diameter.
3
For the confirmation tests, select at least 1 characteristic colony, then another 4 if the first is negative.

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 Campylobacter spp.
Section Media and reagents Compliance
Refer to the section of standard NF EN ISO 6887 corresponding to the
B.2 Diluent
product to be examined, pages 96 to 103.
- Modified charcoal cefoperazone deoxycholate (mCCD)
B.3 Isolation
agar
medium -
-
- Columbia blood agar
Columbia (base) agar Partial 4
500 g vial - BK019HA
- Sterile sheep or horse blood
-
-
B.4/B.5/B.6/B.7/B.8 - Oxidase reagent
Confirmation and - -
identification - Reagent for the detection of catalase activity
reagents -
-
- Reagent for the detection of hydrolysis of hippurate
- -
- Indoxyl acetate discs
-
-

4
pH 7.3 ± 0.2 instead of 7.4 ± 0.2.

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 Horizontal method for the enumeration of Clostridium perfringens
Colony-count technique

NF EN ISO 7937: 02-2005

V 08-019

Initial suspension
and decimal dilutions 1

1 mL in poured plate,
double layer 1

TSC agar

 20 ± 2 hrs at 37°C
anaerobic conditions

Enumeration and confirmation 2

of Clostridium perfringens

Thioglycollate broth Nitrate motility medium 3 Lactose-gelatin medium 3

 18 to 24 hrs at 37°C  24 hrs at 37°C  24 hrs at 37°C

anaerobic conditions anaerobic conditions anaerobic conditions
5 drops
Lactose sulfite broth + 0.2 mL to 0.5 mL
of reagent to test
for nitrites
 18 to 24 hrs at 46°C

1
Inoculate each dilution in duplicate.
2
Select five characteristic colonies from each plate.
3
Isolated colonies are required for confirmation. If this is not the case, inoculate 5 characteristic colonies in thioglycollate broth
then on TSC base medium before reinoculation on nitrate motility medium and lactose-gelatin medium.

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 Clostridium perfringens
Section Media and reagents Compliance
Refer to the section of standard NF EN ISO 6887 corresponding to the
5.1 Diluent
product to be examined, pages 96 to 103.
- Sulfite-cycloserine agar (SC)
TSC (base) agar
50 x 20 mL tubes - BM03908
10 x 200 mL vials - BM07708 Total 4
500 g vial - BK031HA
5.2 Culture medium
D-cycloserine 200 mg selective supplement
10 vials q.s. 500 mL - BS00608
D-cycloserine liquid supplement Total
10 x 90- L vials - BS09208

- Fluid thioglycollate medium

Thioglycollate medium with resazurin
50 x 10 mL tubes - BM08208
500 g vial - BK017HA Total 4
5 kg drum - BK017GC
- Lactose sulfite medium
Lactose-sulfite broth
500 g vial - BK140HA
7 x 9 mL tubes - BM19208 Total 4
- Disodium disulfite solution
- -
- Ammonium iron (III) citrate solution
5.3/5.4/5.5/5.6/5.7/ - -
5.8 Confirmation - Nitrate motility medium
reagents
- -
- Nitrite detection reagent
- -
- 5-Amino-2-naphthalenesulfonic acid (5-2-ANSA) solution
- -
- Sulfanilic acid solution
- -
- Zinc dust
-
-
- Lactose-gelatin medium
-

4
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

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 Horizontal method for the enumeration of coliforms
Colony-count technique

NF ISO 4832/FIL 73B: 07-2006

V 08-015

Initial suspension
(decimal dilutions)

1 mL in poured plate,
double layer 1

VRBL agar 2

 24 ± 2 hrs at 30°C or
37°C as agreed

Enumeration of coliforms

Confirmation if necessary 3

BGBLB

 24 ± 2 hrs at 30°C or
37°C as agreed

Production of gas
Confirmation of coliforms

1
Inoculate each dilution in duplicate. Apply 1 mL of inoculum to an empty Petri dish, pour in 15 mL of medium, mix and
allow to solidify. Then pour in a second layer of 4 mL of medium.
2
Take a control plate with 15 mL of medium for the sterility control.
3
Confirm the atypical colonies (e.g. those smaller in size) and the colonies derived from milk products containing sugars
other than lactose. Reinoculate 5 atypical colonies.

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 Section Media and reagents Compliance

Coliform bacteria
Refer to the section of standard NF EN ISO 6887 corresponding to the
5.2 Diluent
product to be examined, pages 96 to 103.
- Crystal violet neutral red bile lactose (VRBL) agar
VRBL agar
5.3 Solid selective 10 x 100 mL vials - BM03408
10 x 200 mL vials - BM03508 Total
medium
500 g vial - BK152HA
5 kg drum - BK152GC

- Brilliant green bile lactose broth

5.4 Confirmation Brilliant green bile lactose broth BGBLB
50 x 10 mL tubes with Durham tubes - BM01108 Total 4
medium
500 g vial - BK002HA

4
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

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 Horizontal method for the enumeration of coliforms
Most probable number technique

NF ISO 4831/FIL 73B: 10-2006

V 08-016

Enumeration protocol

Initial suspension
(decimal dilutions) 1

10 mL in 1 mL in
10 mL of broth 10 mL of broth

Double strength Single strength

Lauryl sulfate tryptose broth Lauryl sulfate tryptose broth

 24 ± 2 hrs  24 ± 2 hrs at 30 or 37°C

at 30 or 37°C + 24 ± 2 hrs if necessary 2

BGBLB 3

 24 ± 2 hrs at 30 or 37°C
+ 24 ± 2 hrs if necessary 2

Enumeration of coliforms 4

Detection protocol
Inoculate the initial suspension in a tube of single strength or double strength medium, and follow the
protocol described for enumeration.

1
Inoculate the initial suspension in three tubes of double strength medium and three tubes of single strength medium.
The decimal dilutions are inoculated in three tubes of single strength medium.
2
If no gas is produced, prolong incubation.
3
For each tube, inoculate a tube of confirmation medium (BGBLB) using a plate-loop.
4
For each dilution, count the number of tubes which produce gas. Calculate the most probable number according to ISO 7218.

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 Section Media and reagents Compliance

Coliform bacteria
Coliform bac-
Refer to the section of standard NF EN ISO 6887 corresponding to the
5.2 Diluent
product to be examined, pages 96 to 103.

teria
- Lauryl sulfate tryptose broth
Laurylsulfate Tryptose broth
50 x 10 mL tubes with Durham tubes
5.3 Selective
(Single strength) - BM09708 Partial 5
enrichment medium
50 x 10 mL tubes with Durham tubes
(Double strength) - BM09808
500 g vial - BK010HA

- Brilliant green bile lactose broth

5.4 Confirmation Brilliant green bile lactose broth BGBLB
50 x 10 mL tubes with Durham tubes - BM01108 Total 6
medium
500 g vial - BK002HA

5
Presence of tryptone instead of the enzymatic digest of milk and recommended animal proteins.
6
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

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 Enumeration of presumptive coliforms by
the colony-count technique at 30°C
NF V 08-050: 04-2009
V 08-050

Initial suspension
and decimal dilutions

1 mL in poured plate,
in a double layer 1

VRBL medium

 24 ± 4 hrs at 30°C

Enumeration of coliforms

Section Media and reagents Compliance

Refer to the section of standard NF EN ISO 6887 corresponding to the
5.2 Diluent
product to be examined, pages 96 to 103.

- Crystal violet neutral red bile lactose (VRBL) agar

VRBL agar
5.3 Selective 10 x 100 mL vials - BM03408
Total
medium 10 x 200 mL vials - BM03508
500 g vial - BK152HA
5 kg drum - BK152GC

1
Inoculate each dilution in duplicate.

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 Enumeration of thermotolerant coliforms by
the colony-count technique at 44°C
NF V 08-060: 04-2009
V 08-060

Coliform bacteria
Initial suspension
and decimal dilutions

1 mL in poured plate,
in a double layer 1

VRBL medium

 24 ± 4 hrs at 44 ± 1°C

Enumeration of thermotolerant
coliforms

Section Media and reagents Compliance

Refer to the section of standard NF EN ISO 6887 corresponding to the
5.2 Diluent
product to be examined, pages 96 to 103.

- Crystal violet neutral red bile lactose (VRBL) agar

VRBL agar
5.3 Selective 10 x 100 mL vials - BM03408
Total
medium 10 x 200 mL vials - BM03508
500 g vial - BK152HA
5 kg drum - BK152GC

1
Inoculate each dilution in duplicate.

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 Microbiology of food
Detection of Cronobacter spp.
NF EN ISO 22964: 06-2017
V 08-746

10 g or mL of product in 90 mL of
buffered peptone water 1

 18 ± 2 hrs at 36 ± 2°C
Inoculate 0.1 mL in 10 mL

CSB

 24 ± 2 hrs at 41.5 ± 1°C

Inoculate 10 µL

CCI agar

 24 ± 2 hrs at 41.5 ± 1°C

Reading and confirmation 2

TSA

 21 ± 3 hrs at 36 ± 2°C

Biochemical tests

1
For samples over than 10 g, the buffered peptone water should be prewarmed at between 34 and 38°C.
2
Reinoculate 5 colonies identified as characteristic. Then test a colony; if the result is negative, test the other four colonies selected.

Page 26 www.biokar-diagnostics.com
 Section Media and reagents Compliance
- Buffered Peptone Water (BPW)
Buffered Peptone Water 20.0 g/L 3
500 g vial - BK131HA

Cronobacter spp.
5 kg drum - BK131GC
Buffered Peptone Water 25.5 g/L 4
B.1 Pre-enrichment 50 x 9 mL tubes - BM05608
10 x 90 mL vials - BM05708 Total
medium
10 x 225 mL vials - BM01008
3 x 3 L flexible bags - BM13108
2 x 5 L flexible bags - BM13208
500 g vial - BK018HA
5 kg drum - BK018GC

- Cronobacter selective broth (CSB)

B.2 Liquid Cronobacter screening broth (CSB) Total
enrichment medium 50 x 10 mL tubes - BM15508

- Chromogenic Cronobacter isolation (CCI) agar

B.3 Agar medium Chromogenic Cronobacter isolation (CCI) agar Total
20 Petri dishes Ø 90 mm - BM15408

- Tryptone soya agar (TSA)

Trypto-casein soy agar (TSA)
B.4 Subculture 20 Petri dishes Ø 90 mm - BM05008
10 x 100 mL vials - BM01708 Total
medium
10 x 200 mL vials - BM04908
500 g vial - BK047HA

- Oxidase reagent
-
- α-glucosidase enzymatic assay solution
-
- L-lysine decarboxylation medium
-
- L-ornithine decarboxylation medium
B.5 Confirmation -
-
media and reagents - Media for fermentation of carbohydrates
-
- MR//VP base media
-
- Methyl red (MR) reactive compound
-
- Voges-Proskauer (VP) reactive compound
-

3
Formula including 9.0 g/L of disodium phosphate dodecahydrate (Molecular mass = 358.14).
4
Formula including 3.56 g/L of anhydrous disodium phosphate (Molecular mass = 141.96).

www.biokar-diagnostics.com Page 27
 Horizontal method for the detection and
enumeration of Enterobacteriaceae
Part 1: Detection of Enterobacteriaceae
NF EN ISO 21528-1: 06-2017
V 08-039-1

10 g of sample in
90 mL of Buffered Peptone Water

 18 ± 2 hrs at 37°C 1
Re-streak

VRBG agar

 24 ± 2 hrs at 37°C 1

Detection and confirmation 2 of Enterobacteriaceae

Nutrient agar

 24 ± 2 hrs at 37°C

Oxidase reaction Fermentation test

Glucose OF medium

 24 ± 2 hrs at 37°C

1
An incubation temperature of 37°C is generally used when Enterobacteriaceae are tested for as a hygiene indicator.
Otherwise, a temperature of 30°C may be chosen in the context of a technological process, including psychrotrophic
Enterobacteriaceae.
2
Reinoculate one characteristic colony per plate.

Page 28 www.biokar-diagnostics.com
 Section Media and reagents Compliance

- Buffered Peptone Water

Buffered Peptone Water (20.0 g/L) 3
500 g vial - BK131HA

Enterobacteriaceae
5 kg drum - BK131GC
Buffered Peptone Water (25.5 g/L) 4
B.1 Enrichment 50 x 9 mL tubes - BM05608
Total
medium 10 x 225 mL vials - BM01008
10 x 90 mL vials - BM05708
3 x 3 L flexible bags - BM13108
2 x 5 L flexible bags - BM13208
500 g vial - BK018HA
5 kg drum - BK018GC
- Violet red bile glucose (VRBG) agar
VRBG agar
B.2 Solid selective 10 x 200 mL vials - BM07508 Total
medium 500 g vial - BK011HA
5 kg drum - BK011GC

- Nutrient agar
2.5% nutrient agar Equivalent
50 x 18 mL tubes - BM12508
- Glucose OF medium
B.3/B.4/B.5 Glucose OF medium
Confirmation media 50 x 10 mL tubes - BM19708 Total
and reagents
- N,N,N’,N’-Tetramethyl-p-phenylenediamine
dihydrochloride
- -

3
Formula including 9.0 g/L of disodium phosphate dodecahydrate (molecular mass 358.14).
4
Formula including 3.56 g/L of anhydrous disodium phosphate (molecular mass 141.96).

www.biokar-diagnostics.com Page 29
 Horizontal method for the detection and
enumeration of Enterobacteriaceae
Part 2: Colony-count technique
NF EN ISO 21528-2: 07-2017
V 08-039-2

Initial suspension
(decimal dilutions)

1 mL in poured plate,
double layer 1

VRBG agar

 24 ± 2 hrs at 37°C 2

Enumeration of Enterobacteriaceae
Confirmation 3

Nutrient agar

 24 ± 2 hrs at 37°C

Oxidase reaction Fermentation test

Glucose OF medium

 24 ± 2 hrs at 37°C

1
Add 15 mL of medium, mix and allow to solidify. Then add a superficial layer of 5 to 10 mL of medium.
Inoculate each dilution in duplicate.
2
An incubation temperature of 37°C is generally used when Enterobacteriaceae are tested for as a hygiene indicator. Otherwise,
a temperature of 30°C may be chosen in the context of a technological process, including psychrotrophic Enterobacteriaceae.
3
Reinoculate a well isolated colony in each plate.

Page 30 www.biokar-diagnostics.com
 Section Media and reagents Compliance
Refer to the section of standard NF EN ISO 6887 corresponding to the
A.1 Diluent
product to be examined, pages 96 to 103.

Enterobacteriaceae
- Violet red bile glucose (VRBG) agar
VRBG agar
A.2 Solid selective 10 x 200 mL vials - BM07508 Total
medium 500 g vial - BK011HA
5 kg drum - BK011GC

- Nutrient agar
2.5% nutrient agar
50 x 18 mL tubes - BM12508
Equivalent
- Glucose OF medium
A.3/A.4/A.5 Glucose OF medium
Confirmation media 50 x 10 mL tubes - BM19708 Total
and reagents - N,N,N’,N’-Tetramethyl-p-phenylenediamine
dihydrochloride
- -

www.biokar-diagnostics.com Page 31
 Enumeration of presumptive Enterobacteria by
the colony-count technique at 30°C or 37°C
NF V 08-054: 04-2009
V 08-054

Initial suspension
(decimal dilutions)

1 mL in poured plate,
double layer 1

VRBG agar

 24 ± 2 hrs
at 30°C or 37°C

Enumeration of
presumptive Enterobacteria

1
The second layer of VRBG agar is used in order to prevent the colonies from spreading and to create semi-anaerobic con-
ditions.
Inoculate each dilution in duplicate.

Page 32 www.biokar-diagnostics.com
 Section Media and reagents Compliance
Refer to the section of standard NF EN ISO 6887 corresponding to the
5.2 Diluent

Enterobacteriaceae
product to be examined, pages 96 to 103.
- Violet red bile glucose (VRBG) agar
VRBG agar
5.3 Solid selective 10 x 200 mL vials - BM07508 Total
medium 500 g vial - BK011HA
5 kg drum - BK011GC

www.biokar-diagnostics.com Page 33
 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 beta-D-glucuronide
NF ISO 16649-1: 09-2018
V 08-031-1

Initial suspension
(decimal dilutions)

1 mL onto a membrane 1

MMGA medium

 4 ± 15 min at 37°C 2

TBX medium

 20 to 24 hrs at 44°C 3

Enumeration of β-glucuronidase-positive
Escherichia coli

1
First place a membrane on the surface of the medium. Then apply 1 mL of sample in the centre and spread on the surface
of the membrane using a spreader. Inoculate each dilution in duplicate.
2
Leave the Petri dish at room temperature for 15 minutes before incubating at 37 °C.
3
Do not exceed a temperature of 45 °C.

Page 34 www.biokar-diagnostics.com
 Section Media and reagents Compliance
Refer to the section of standard NF EN ISO 6887 corresponding to the
5.1 Diluent
product to be examined, pages 96 to 103.
5.2.1 Culture - Minerals-modified glutamate agar (MMGA) medium
-
medium -

Escherichia coli
- Tryptone-bile-glucuronide agar (TBX) medium
TBX agar
5.2.2 Selective 10 x 100 mL vials - BM06908
10 x 200 mL vials - BM17108 Total 4
medium
500 g vial - BK146HA
100 g vial - BK146HM

4
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

www.biokar-diagnostics.com Page 35
 Horizontal method for the enumeration of
β-glucuronidase-positive Escherichia coli
Part 2: Colony-count technique at 44°C using 5-bromo-4-chloro-3-indolyl β-D-glucuronide

NF ISO 16649-2: 07-2001

V 08-031-2

Initial suspension
(decimal dilutions)

1 mL in poured plate 1

TBX medium

 18 to 24 hrs at 44°C 2

Enumeration of β-glucuronidase-positive
Escherichia coli

1
Inoculate each dilution in duplicate.
2
If stressed microorganisms are thought to be present, first incubate at 37°C, then for 18 to 24 hours at 44°C.
The incubation temperature should not exceed 45°C.

Page 36 www.biokar-diagnostics.com
 Section Media and reagents Compliance
Refer to the section of standard NF EN ISO 6887 corresponding to the
5.1 Diluent
product to be examined, pages 96 to 103.
- Tryptone-bile-glucuronide agar (TBX) medium

Escherichia coli
TBX agar
5.2 Selective 10 x 100 mL vials - BM06908
10 x 200 mL vials - BM17108 Total 3
medium
500 g vial - BK146HA
100 g vial - BK146HM

3
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

www.biokar-diagnostics.com Page 37
 Horizontal method for the enumeration of
β-glucuronidase-positive Escherichia coli
Part 3: Detection and most probable number technique using 5-bromo-4-chloro-3-indolyl β-D-glucuronide

NF EN ISO 16649-3: 07-2015

V 08-031-3

Initial suspension
Initial suspension and decimal dilutions

10 mL in 1 mL in
10 mL of broth 1 10 mL of broth 1

Double strength Single strength

Glutamate broth 2 Glutamate broth 2

 24 ± 2 hrs at 37°C

TBX agar 3

 20 to 24 hrs at 44°C

Enumeration of β-glucuronidase-positive
Escherichia coli 4

1
Inoculate 3 or 5 tubes of single or double strength medium for each inoculation.
2
For detection, inoculate the single or double strength glutamate broth with an appropriate volume and follow the
protocol described for enumeration.
3
After incubation, reinoculate the tubes displaying acidification on TBX medium in order to yield isolated colonies.
4
Calculate the most probable number based on the positive tubes for each dilution.

Page 38 www.biokar-diagnostics.com
 Section Media and reagents Compliance
Refer to the section of standard NF EN ISO 6887 corresponding to the
5.1 Diluent
product to be examined, pages 96 to 103.
- Minerals-modified glutamate agar medium
5.2.1 Enrichment Glutamate broth Total
medium 500 g vial - BK186HA

Escherichia coli
- Tryptone-bile-glucuronide agar (TBX) medium
TBX agar
5.2.2 Selective 10 x 100 mL vials - BM06908
10 x 200 mL vials - BM17108 Total 4
medium
500 g vial - BK146HA
100 g vial - BK146HM

4
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

www.biokar-diagnostics.com Page 39
 Horizontal method for the detection and
enumeration of presumptive Escherichia coli
Most probable number technique
NF ISO 7251: 07-2005
V 08-020

Detection Enumeration

Initial suspension Initial suspension

and decimal dilutions

1 mL in 10 mL 1 10 mL in 10 mL 1

Tryptose lauryl sulfate broth Tryptose lauryl sulfate broth

simple strength double strength

 24 ± 2 hrs at 48 ± 2 hrs 2
at 37 °C

If opacity or turbidity is present

or if gas is produced

EC broth

 24 ± 2 hrs at 48 ± 2 hrs 2
at 44°C

If gas is produced
Indole test
Add 0.5 mL of indole reagent

Detection and/or enumeration of

presumptive Escherichia coli 3

1
In the context of enumeration, inoculate three tubes of single strength medium and three tubes of double strength medium
for each dilution.
2
If no gas is produced after 24 ± 2 hours, prolong incubation up to 48 ± 2 hours.
3
Determine the most probable number based on the positive tubes for each dilution and the MPN table in Appendix A.

Page 40 www.biokar-diagnostics.com
 Section Media and reagents Compliance
Refer to the section of standard NF EN ISO 6887 corresponding to the
5.1 Diluent
product to be examined, pages 96 to 103.
- Lauryl sulfate broth
Laurylsulfate tryptose broth
50 x 10 mL tubes with Durham tubes

Escherichia coli
5.2 Selective (Single strength) - BM09708 Total 4
enrichment medium 50 x 10 mL tubes with Durham tubes
(Double strength) - BM09808
500 g vial - BK010HA

- EC broth
5.3 Selective EC broth Total 4
medium 500 g vial - BK162HA

- Peptone water, indole free

5.4 Examination for Peptone water Total 4
indole production 500 g vial - BK084HA

5.5 Examination for - Kovac’s reagent for detection of indole

-
indole production -

4
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

www.biokar-diagnostics.com Page 41
 Horizontal method for the detection of Escherichia coli O157

NF EN ISO 16654: 07-2001

Modified by Amendment A1 (06-2017)
V 08-032

x g of sample + 9x mL of mTSB
with novobiocin

 6 hrs at 41.5°C
 12 to 18 hrs
Immunomagnetic separation 1 at 41.5°C

CT-SMAC agar Second selective medium

 18 to 24 hrs at 37°C  appropriate conditions

Detection and confirmation 2 of Escherichia coli O157

Nutrient agar

 18 to 24 hrs at 37°C

Tryptone/Tryptophan broth Serological identification 3

 18 to 24 hrs at 37°C

Indole test
Add 1 mL of Kovac’s reagent

1
Immunomagnetic separation should take place after 6 hours of incubation, and after additional incubation of 12 to 18 hrs.
2
Sample 5 characteristic colonies from each plate and isolate each colony on a nutrient agar plate.
3
Carry out serological identification on the colonies positive for the indole test only.

Page 42 www.biokar-diagnostics.com
 Section Media and reagents Compliance
- Modified tryptone soya broth with novobiocin
(mTSB+N)
Modified tryptone soy broth (mTSB base)
5.1 Diluent 500 g vial - BK150HA Total 4
Selective novobiocin supplement

Escherichia coli
10 x 10 mg vials - BS03308
8 x 40 mg vials - BS05608

- Sorbitol MacConkey agar with cefixime and tellurite

(CT-SMAC)
5.2 First selective Sorbitol MacCONKEY agar (CT-SMAC base)
500 g vial - BK147HA Total 4
isolation medium
Cefixime-tellurite supplement for CT-SMAC agar
10 vials q.s. 500 mL - BS03708

5.3 Second selective The choice of the second medium is left to the discretion of the testing
isolation medium laboratory

- Nutrient agar
5.4 Subculture 2% nutrient agar
50 x 18 mL tubes - BM11808 Total 4
medium
500 g vial - BK185HA

- Tryptone/tryptophan broth
Tryptophan broth
5.5 Confirmation 50 x 3 mL tubes - BM07608 Total
500 g vial - BK163HA

- Kovac’s reagent for detection of indole

5.6 Confirmation -
-
- Anti-E. coli O157 immunomagnetic beads
5.7/5.8 -
-
Immunomagnetic
- Modified phosphate buffer 0.01 mol/L pH 7.2
separation (IMS) -
-
- Saline solution
-
5.9/5.10 Serological -
identification - Anti-E. coli O157 serum
-
-

4
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

Horizontal method for the detection of Escherichia coli O157

AMENDMENT 1: Appendix B: interlaboratory study results

www.biokar-diagnostics.com Page 43
 Horizontal method for the enumeration of mesophilic lactic acid bacteria
Colony-count technique at 30°C

NF ISO 15214: 09-1998

V 08-030

Initial suspension
Lactic acid bacteria

and decimal dilutions

1 mL in poured plate 1

MRS medium

 72 ± 2 hrs at 30°C

Enumeration of mesophilic lactic

acid bacteria

Section Media and reagents Compliance

Refer to the section of standard NF EN ISO 6887 corresponding to the

5.2 Diluent
product to be examined, pages 96 to 103.

- MRS (de Man, Rogosa and Sharpe) medium, pH 5.7

5.3 Selective MRS agar
Total 2
medium 10 x 200 mL vials - BM08908
500 g vial - BK089HA

1
Inoculate each dilution in duplicate.
2
De Man J.C, Rogosa M., Sharpe M.E. 1960. A medium for the cultivation of lactobacilli. J. App. Bacteriol., 23, (1): 130-135.

Page 44 www.biokar-diagnostics.com
 Horizontal method for the enumeration of yeasts and moulds
growing on low aw medium 1
NF V 08-036: 05-2003
V 08-036

Initial suspension
and decimal dilutions

Yeasts & moulds

1 mL in poured plate

DG 18 agar

 5 days at 25 ± 1°C

Enumeration of yeasts and moulds 2

Section Media and reagents Compliance

Refer to the section of standard NF EN ISO 6887 corresponding to the

5.2 Diluent
product to be examined, pages 96 to 103.3

- Dichloran 18% glycerol (DG) agar

Dichloran-Glycerol (DG 18) agar
5.3 Culture medium Total 4, 5
10 x 100 mL vials - BM10908
500 g vial - BK170HA

1
Product with water activity less than or equal to 0.95.
2
If rapid plate invasion is observed, carry out the reading before 5 days.
3
Addition of Tween® 80 to the chosen dilution medium, at a concentration of 0.033 g/L, may facilitate the mould count.
4
220.0 g/L of glycerol per litre of agar base medium needs to be added to the dehydrated form.
5
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

www.biokar-diagnostics.com Page 45
 Horizontal method for the enumeration of yeasts and moulds
Part 1: Colony count technique in products with water activity greater than 0.95

NF ISO 21527-1: 11-2008

V 08-040-1

Initial suspension
and decimal dilutions

0.1 mL on surface

DRBC medium

 5 days at 25 ± 1°C

Enumeration and confirmation of

yeasts and moulds

Page 46 www.biokar-diagnostics.com
 Section Media and reagents Compliance

Refer to the section of standard NF EN ISO 6887 corresponding to the

product to be examined, pages 96 to 103.1, 2
5.1 Diluents - 0.1% peptone water (mass concentration)
0.1% peptone water Total 3
2 x 5 L flexible bags - BM16408

- Dichloran-rose bengal chloramphenicol agar 4, 5, 6

Dichloran-rose bengal chloramphenicol (DRBC) agar 7

Yeasts & moulds

5.2 Culture medium Total
10 x 200 mL vials - BM14208
500 g vial - BK198HA

1
Addition of Tween® 80 to the chosen liquid dilution medium, at a concentration of 0.033 g/L, may facilitate the mould
count.
2
Except when specifically preparing the sample for the test, use of 0.1% peptone water (mass concentration) as the diluent is
recommended.
3
“Tryptone” is a peptone obtained by pancreatic digestion of casein.
4
When bacterial proliferation may present a problem, use of chloramphenicol (50 mg/L) and chlortetracycline (50 mg/L) is
recommended.
5
In order for the full mould morphology to be observed, 1 mL of a trace element solution (ZnSO 4 . 7 H2O, CuSO4 . 5H2O q.s.
100 mL H2O) may be added for 1 L of medium.
6
In order to avoid the proliferation of certain microorganisms (Mucoraceae), addition of Tergitol (1 mL/L) to the culture
medium is recommended.
7
Complete formula including 50 mg/L of chloramphenicol, 50 mg/L of chlortetracycline hydrochloride, 1 mg/L of ZnSO 4,
7H2O, 0.5 mg/L of CuSO4, 5H2O, 1 mL/L of Tergitol.

www.biokar-diagnostics.com Page 47
 Horizontal method for the enumeration of yeasts and moulds
Part 1: Colony count technique in products with water activity less than or equal to 0.95

NF ISO 21527-2: 11-2008

V 08-040-2

Initial suspension
and decimal dilutions

0.1 mL on surface

DG18 medium

 5 to 7 days
at 25 ± 1°C

Enumeration and confirmation of

yeasts and moulds

Page 48 www.biokar-diagnostics.com
 Section Media and reagents Compliance
Refer to the section of standard NF EN ISO 6887 corresponding to the
product to be examined, pages 96 to 103.1, 2, 3
5.1 Diluents - 0.1% peptone water (mass concentration)
0.1% peptone water Total 4
2 x 5 L flexible bags - BM16408

- Dichloran 18% glycerol (DG) agar 6, 7

Dichloran-Glycerol (DG 18) agar
Total 4, 5

Yeasts & moulds

5.2 Culture medium
10 x 100 mL vials - BM10908
500 g vial - BK170HA

1
Addition of Tween® 80 to the chosen liquid dilution medium, at a concentration of 0.033 g/L, may facilitate the mould count.
2
The use of a diluent containing a sufficient quantity of solute (20 to 30% of glycerol or D-glucose) is recommended in order to
reduce osmotic shock on xerophilic moulds and osmophilic yeasts when serial dilutions are prepared.
3
Except when specifically preparing the sample for the test, use of 0.1% peptone water (mass concentration) as the diluent is
recommended.
4
“Tryptone” is a peptone obtained by pancreatic digestion of casein.
5
220.0 g of glycerol per litre of glucose base medium needs to be added to the commercial dehydrated form.
6
When bacterial proliferation may present a problem, use of chloramphenicol (50 mg/L) and chlortetracycline (50 mg/L) is
recommended.
7
Complete formula including 50 mg/L of chloramphenicol, 50 mg/L of chlortetracycline hydrochloride, 1 mg/L of ZnSO 4, 7H2O,
0.5 mg/L of CuSO4, 5H2O, 1 mL/L of Tergitol.

www.biokar-diagnostics.com Page 49
 Routine method for the enumeration of yeasts and
moulds by colony-count technique at 25°C
NF V 08-059: 11-2002
V 08-059

Initial suspension
and decimal dilutions

1 mL in poured plate

Selective medium

 5 days at 25 ± 1°C

Enumeration of
yeasts and moulds

Page 50 www.biokar-diagnostics.com
 Section Media and reagents Compliance

Refer to the section of standard NF EN ISO 6887 corresponding to the

5.2 Diluent
product to be examined, pages 96 to 103.

- Glucose chloramphenicol agar

Chloramphenicol glucose agar
10 x 100 mL vials - BM02108
10 x 200 mL vials - BM07908 Total
500 g vial - BK007HA

Yeasts & moulds

- Oxytetracycline glucose agar
5.3 Culture media Oxytetracycline glucose agar (OGA base)
10 x 110 mL vials - BM02208
500 g vial - BK053HA
Oxytetracycline supplement 1 Total
10 x 50 mg vials - BS00808
Gentamicin supplement 2
10 x 25 mg vials - BS00908

1
Chloramphenicol may be replaced by oxytetracycline.
2
Optional additive, justified by the suspected presence of sizeable contamination by Gram-negative bacteria.

www.biokar-diagnostics.com Page 51
 SYMPHONY agar
Method for the enumeration of yeasts and moulds

BKR 23/11-12/18
Alternative food
analysis method
www.afnor-validation.org
Yeasts & moulds

Initial suspension
and decimal dilutions

0.1 mL on surface
1 mL in poured plate

 54-72 hrs
at 25 ± 1 °C

Enumeration of yeasts and moulds

Refer to the section of standard NF EN ISO 6887 corresponding to the

Diluent
product to be examined, pages 96 to 103.

SYMPHONY agar - ready-to-melt

Culture medium
10 x 200 mL vials - BM19108

Page 52 www.biokar-diagnostics.com
 Horizontal method for the detection and
enumeration of Listeria monocytogenes
Part 1: Detection method
NF EN ISO 11290-1: 07-2017
V 08-028-1

x g of sample in 9x mL
of half-FRASER

Listeria spp.
 24 to 26 hrs at 30 ± 1°C
0.1 mL in 10 mL
FRASER broth

 24 ± 2 hrs at 37 ± 1°C

COMPASS® Listeria Agar Second selective medium

 24 ± 2 hrs at 37 ± 1°C
+ 24 ± 2 hrs if necessary  appropriate conditions

Detection and confirmation 1

TSYEA medium

 18 to 24 hrs at 37 ± 1°C

Confirmation of Listeria spp. Confirmation of Listeria monocytogenes

▪ Microbiological aspect ▪ Microbiological aspect
▪ Catalase reaction ▪ Haemolysis tests
▪ VP reaction (optional) ▪ Rhamnose fermentation
▪ Motility test at 25°C (optional) ▪ Xylose fermentation
▪ Catalase reaction (optional)
▪ CAMP test (optional)
▪ Motility test at 25°C (optional)

1
Using each plate, reinoculate a characteristic colony of Listeria spp. and Listeria monocytogenes on TSYEA medium.
After incubation, carry out the required confirmation tests.

www.biokar-diagnostics.com Page 53
 Section Media and reagents Compliance
- Half-FRASER broth
Half-FRASER broth (ready-to-use) 2
10 x 225 mL vials - BM01608
3 x 3 L flexible bags - BM13308
2 x 5 L flexible bags - BM13408
Half-FRASER broth (base) 2
500 g vial - BK173HA
5 kg drum - BK173GC
B.1 Selective Sterile 5% ammonium ferric citrate supplement
primary enrichment (for BK173) Total 3
medium 10 x 90 mL vials - BS05908
7 x 10 mL tubes - BS06208
FRASER broth (base II) 2
500 g vial - BK133HA
5 kg drum - BK133GC
Selective supplement for FRASER broth (for BK133)
10 vials q.s. 500 mL - BS03008
8 vials q.s. 2.25 L - BS03208

- FRASER broth
FRASER broth (ready-to-use) 2
50 x 10 mL tubes - BM01308
FRASER broth (base II) 2
500 g vial - BK133HA
5 kg drum - BK133GC
B.2 Selective Selective supplement for FRASER broth (for BK133)
secondary 10 vials q.s. 500 mL - BS03008 Total 3
enrichment medium 8 vials q.s. 2.25 L - BS03208
FRASER broth (base) 2
500 g vial - BK115HA
Selective supplement for FRASER broth
10 vials q.s. 500 mL - BS03108

- Agar Listeria according to Ottaviani and Agosti

COMPASS® Listeria Agar
B.3 Selective plating- 20 Petri dishes Ø 90mm - BM12308
Total
out medium 120 Petri dishes Ø 90mm - BM12408
Kit of 6 x 200 mL vials base + supplements - BT00808

- The choice is left to the discretion of the testing

laboratory:
OXFORD agar (base) 4
500 g vial - BK110HA
Selective supplement for OXFORD agar (CCCFA) Total
10 vials q.s. 500 mL - BS00308
B.4 Second selective
PALCAM agar (pre-poured) 4
plating-out medium
20 Petri dishes Ø 90mm - BM02008
PALCAM agar (base)
500 g vial - BK145HA
Selective supplement for PALCAM agar Total
10 vials q.s. 500 mL - BS00408
8 vials q.s. 2.5 L - BS04908

Page 54 www.biokar-diagnostics.com
 Section Media and reagents Compliance

- Hydrogen peroxide solution

- -
- Motility agar
-
-
- Blood agar base medium
Tryptone soya agar (blood agar base) Equivalent
500 g vial - BK028HA
- Defibrinated sheep, calf or bovine blood
- -
- PBS
B.6/B.7/B.8/B.9/ - -

Listeria spp.
B.10/B.11/B.12/B.13 - Red blood corpuscle suspension
Confirmation media -
and reagents -
- CAMP culture medium
-
-
- Carbohydrate utilization broth
(L-Rhamnose and D-Xylose)
-
-
- Reagents for Voges-Proskauer (VP) reaction
-
-
- Tryptone soya yeast extract agar (TSYEA)
TSYEA medium
500 g vial - BK224HA Total

2 Formula including 9.60 g/L of anhydrous disodium phosphate (molecular mass: 141.96) instead of 12 g/L of disodium
hydrogen phosphate dihydrate (molecular mass 177.99) as described.
3 “Tryptone” is a peptone obtained by pancreatic digestion of casein.
4 Medium able to be used as a second plating-out medium.

www.biokar-diagnostics.com Page 55
 COMPASS® Listeria Agar
Alternative method for the detection of Listeria spp. And Listeria monocytogenes

BKR 23/02-11/02
Alternative food
analysis method
www.afnor-validation.org

x g of sample in 9x mL of half-
FRASER broth

 22-28h ³ at 30 ± 1 °C
or
0.1 mL on surface 18-24h ³ at 37 ± 1 °C 1

COMPASS® Listeria Agar

 24 hrs at 37 ± 1°C 2

Detection and confirmation 3

of Listeria spp. and Listeria monocytogenes

PALCAM agar CONFIRM’ L. mono broth

or
CONFIRM’ L. mono agar

1
Half Fraser broth must be preheated.
2
Reading may be done after 22 hours of incubation. Incubation may be prolonged up to 48 hours for laboratory organisation
purposes.
3
Reinoculate a characteristic colony of Listeria spp. and Listeria monocytogenes.

Page 56 www.biokar-diagnostics.com
 Half-FRASER broth (ready-to-use)
10 x 225 mL vials - BM01608
3 x 3 L flexible bags - BM13308
2 x 5 L flexible bags - BM13408
Half-FRASER broth (Base)
500 g vial - BK173HA
5 kg drum - BK173GC
Sterile 5% ammonium ferric citrate supplement (for BK173)
Enrichment medium 10 x 90 mL vials - BS05908
7 x 10 mL tubes - BS06208
FRASER broth (Base II)
500 g vial - BK133HA

Listeria spp.
5 kg drum - BK133GC

Listeria spp.
Selective supplement for FRASER broth (for BK133)
10 vials q.s. 500 mL - BS03008
8 vials q.s. 2.25 L - BS03208

COMPASS® Listeria Agar

Selective plating-out 20 Petri dishes Ø 90mm - BM12308
medium 120 Petri dishes Ø 90mm - BM12408
Kit of 6 x 200 mL vials base + supplements - BT00808

CONFIRM’ L. mono broth

Listeria 18 vials - BM16208
monocytogenes CONFIRM’ L. mono Agar
confirmation media 10 Petri dishes Ø 90mm - BM13908

Pre-poured PALCAM agar

20 Petri dishes Ø 90mm - BM02008
PALCAM agar (base)
L. spp. confirmation 500 g vial - BK145HA
medium PALCAM supplement
10 vials q.s. 500 mL - BS00408
10 vials q.s. 2.5 L - BS04908

www.biokar-diagnostics.com Page 57
 Horizontal method for the detection and
enumeration of Listeria monocytogenes
Part 2: Method for enumeration
NF EN ISO 11290-2: 07-2017
V 08-028-2

Initial suspension
and decimal dilutions 1

0.1 mL on surface 2

COMPASS® Listeria Agar

 24 ± 2 hrs at 37 ± 1°C
+ an additional 24 ± 2 hrs

Enumeration and confirmation 3

of Listeria monocytogenes and/or Listeria spp.

TSYEA medium

 18 to 24 hrs at 37°C

Confirmation of Listeria spp. Confirmation of Listeria monocytogenes

▪ Microbiological aspect ▪ Microbiological aspect
▪ Catalase reaction ▪ Haemolysis tests
▪ VP reaction (optional) ▪ Rhamnose fermentation
▪ Motility test at 25°C (optional)
▪ Xylose fermentation
▪ Catalase reaction (optional)
▪ CAMP test (optional)
▪ Motility test at 25°C (optional)

1
The initial suspension may be prepared in buffered peptone water or in half-Fraser broth with or without selective
agents.
2
Inoculate 0.1 mL of inoculum on 2 plates of medium.
For certain products, it is preferable to estimate low numbers of L. monocytogenes and/or Listeria spp.; the limits of
detection may be increased by a factor of 10. Apply a volume of 1 ml of inoculum to the surface of a large Petri dish
(140 mm) or to the surface of three small dishes (90 mm).
3
Using each dish, reinoculate five characteristic colonies on TSYEA medium. After incubation, carry out the required
confirmation tests.

Page 58 www.biokar-diagnostics.com
 Section Media and reagents Compliance

9.1 Diluent Refer to standard NF EN ISO 11290-1, pages 53 to 55.

- Tryptone soya yeast extract agar (TSYEA)

B.1 Plating-out TSYEA medium Total
medium 500 g vial - BK224HA

- Agar Listeria according to Ottaviani and Agosti

COMPASS® Listeria Agar
B.2 Selective plating- 20 Petri dishes Ø 90mm - BM12308
Total
out medium 120 Petri dishes Ø 90mm - BM12408

Listeria spp.
Kit of 6 x 200 mL vials base + supplements - BT00808

Listeria spp.
- Hydrogen peroxide solution
- -
- Motility agar
- -
- Blood agar base medium
Tryptone soya agar (blood agar base) Equivalent
500 g vial - BK028HA
B.3/B.4/B.5/B.6/B.7/ - Defibrinated sheep, calf or bovine blood
B.8/B.9/B.10/B.11 - -
Confirmation media - PBS
and reagents - -
- CAMP culture medium
- -
- Carbohydrate utilization broth (L-Rhamnose and D-
Xylose)
- -
- Reagents for Voges-Proskauer (VP) reaction
- -

4
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

www.biokar-diagnostics.com Page 59
 COMPASS® Listeria Agar
Alternative method for the enumeration of Listeria monocytogenes

BKR 23/05-12/07
Alternative food
analysis method
www.afnor-validation.org

x g of sample in
9x mL of diluent 1

0.1 mL on surface
or
1 mL in poured plate

COMPASS® Listeria Agar

 24 to 48 ± 2 hrs at 37 ± 1°C 2

Enumeration and confirmation 3

of Listeria monocytogenes

CONFIRM’ L. mono broth (in 6 hrs)

or
CONFIRM’ L. mono agar (in 24 hrs)

1
The half-FRASER broth with or without selective agents or BPW may be used as a diluent.
2
Carry out final enumeration after 48 ± 2 hours of incubation.
3
Reinoculate a characteristic colony of Listeria monocytogenes.

Page 60 www.biokar-diagnostics.com
 - Buffered Peptone Water (20 g/L)
500 g vial - BK131HA
5 kg drum - BK131GC
Buffered Peptone Water (25.5 g/L)
10 x 225 mL vials - BM01008
50 x 9 mL tubes - BM05608
10 x 90 mL vials - BM05708
3 x 3 L flexible bags - BM13108
2 x 5 L flexible bags - BM13208
500 g vial - BK018HA
5 kg drum - BK018GC Half-FRASER broth (ready-to-use)
10 x 225 mL vials - BM01608
3 x 3 L flexible bags - BM13308
Diluents

Listeria spp.
2 x 5 L flexible bags - BM13408
Half-FRASER broth (Base)
500 g vial - BK173HA
5 kg drum - BK173GC
Sterile 5% ammonium ferric citrate supplement (for BK173)
10 x 90 mL vials - BS05908
7 x 10 mL tubes - BS06208
FRASER broth (Base II)
500 g vial - BK133HA
5 kg drum - BK133GC
Selective supplement for FRASER broth (for BK133)
10 vials q.s. 500 mL - BS030088
vials q.s. 2.25 L - BS03208

COMPASS® Listeria Agar

Selective plating-out 20 Petri dishes Ø 90mm - BM12308
medium 120 Petri dishes Ø 90mm - BM12408
Kit of 6 x 200 mL vials base + supplements - BT00808
CONFIRM’ L. mono broth
L. monocytogenes
18 vials - BM16208
confirmation
CONFIRM’ L. mono Agar
medium
10 Petri dishes Ø 90mm - BM13908

www.biokar-diagnostics.com Page 61
 Horizontal method for the detection of Salmonella spp.

NF EN ISO 6579-1: 04-2017

V 08-013-1

x g of sample
in 9x mL of BPW

 18 ± 2 hrs at 36 ± 2°C
0.1 mL in
10 mL or on surface 1 1 mL in 10 mL

MSRV agar or RVS broth MKTTn broth

 24 ± 3 hrs  24 ± 3 hrs
at 41.5 ± 1°C 34 to 38°C

XLD agar Second medium XLD agar Second medium

 24 ± 3 hrs  appropriate  24 ± 3 hrs  appropriate

34 to 38°C conditions 34 to 38°C conditions

Detection and confirmation 2 of Salmonella spp.

Biochemical confirmation
- TSI agar
- Urea agar
- L-lysine decarboxylation medium
- β-galactosidase reagent (optional)
- Indole reaction (optional)

1
Transfer 0.1 mL of the culture obtained to 10 mL of RVS broth or to the surface of the MRSV agar by equidistant
application of 1 to 3 drops onto the surface of the medium.
2
Confirm at least one characteristic colony, then four other colonies if the first proves negative.

Page 62 www.biokar-diagnostics.com
 x g of sample
in 9x mL of BPW

 18 ± 2 hrs at 36 ± 2°C

MSRV agar 3

Salmonella spp.
 24 ± 3 hrs
at 41.5 ± 1°C 4

XLD agar Second medium

 24 ± 3 hrs  appropriate
34 to 38 °C conditions

Detection and confirmation 5 of Salmonella spp.

Biochemical confirmation
- TSI agar
- Urea agar
- L-lysine decarboxylation medium
- β-galactosidase reagent (optional)
- Indole reaction (optional)

3
Sensitivity may be increased by applying a second selective enrichment procedure, with MKTTn broth for example.
4
If the plates are shown to be negative after 24 hours of incubation, incubate for an additional 24 hrs.
5
Confirm at least one characteristic colony, then four other colonies if the first proves negative.

www.biokar-diagnostics.com Page 63
 Section Media and reagents Compliance
- Buffered Peptone Water
Buffered Peptone Water (20 g/L) 6
500 g vial - BK131HA
5 kg drum - BK131GC
Buffered Peptone Water (25.5 g/L) 7
B.2 Pre-enrichment 10 x 225 mL vials - BM01008
50 x 9 mL tubes - BM05608 Total
medium
10 x 90 mL vials - BM05708
3 x 3 L flexible bags - BM13108
2 x 5 L flexible bags - BM13208
500 g vial - BK018HA
5 kg drum - BK018GC

- Rappaport-Vassiliadis broth with soya (RVS broth)

B.3 Selective RAPPAPORT-VASSILIADIS broth with soya (RVS)
enrichment 50 x 10 mL tubes - BM07408 Total 8
medium 500 g vial - BK148HA

- Modified semi-solid Rappaport-Vassiliadis (MSRV)

B.4 Selective medium
enrichment MSRV agar (ISO 6579) Total
medium 10 x 200 mL vials - BM12708
500 g vial - BK191HA

- Muller-Kauffmann tetrathionate-novobiocin (MKTTn)

broth Total 11
MULLER-KAUFFMANN tetrathionate-novobiocin broth
(MKTTn ready-to-use)
50 x 10 mL tubes - BM07808
MULLER-KAUFFMANN tetrathionate-novobiocin broth
B.5 Selective (MKTTn base, without iodine)
enrichment Total
500 g vial - BK208HA
medium MULLER-KAUFFMANN tetrathionate-novobiocin broth
(MKTTn base, without iodine, without novobiocin)
500 g vial - BK169HA
Selective novobiocin supplement Total
10 x 10 mg vials - BS03308
8 x 40 mg vials - BS05608

- Xylose lysine deoxycholate agar (XLD agar)

XLD agar (ISO 6579)
20 Petri dishes Ø 90 mm - BM08708 Total
B.6 Selective
500 g vial - BK168HA
plating-out media
The second plating-out medium is left to the discretion of the testing
laboratory 10
- Nutrient agar
B.7 Subculture 2 % nutrient agar
50 x 18 mL tubes - BM11808 Total 9
medium
500 g vial - BK185HA

Page 64 www.biokar-diagnostics.com
 Section Media and reagents Compliance
- Triple sugar/iron agar (TSI agar)
B.8 Confirmation TSI agar
Total
medium 500 g vial - BK221HA

- Urea agar (Christensen)

- -
- L-lysine decarboxylation (LDC) medium
- -
- β-galactosidase reagent (optional)
- -

Salmonella spp.
- Tryptone/tryptophan medium (optional)
B.9/B.10/B.11/B.12/
Tryptophan broth
B.11/B.12/B.13
50 x 3 mL tubes - BM07608
Confirmation media
500 g vial - BK163HA Total9
and reagents
- Kovac’s reagent (optional)
- -
- Saline solution
- -
- Serums
-
-

6
Formula including 9.0 g/L of disodium phosphate dodecahydrate (molecular mass 358.14).
7
Formula including 3.56 g/L of anhydrous disodium phosphate (molecular mass 141.96).
8
Formula including anhydrous magnesium chloride instead of the hexahydrate form described.
9
“Tryptone” is a peptone obtained by pancreatic digestion of casein.
10
Its intrinsic properties should complement those of XLD agar and enable detection of lactose + Salmonella (including
Typhi and Paratyphi).

www.biokar-diagnostics.com Page 65
 x g of sample
in 9x mL of BPW

 18 ± 2 hrs at 36 ± 2°C

1 mL
0.1 mL in 10 mL 1 mL in 10 mL
in 10 mL

RVS broth MKTTn broth Selenite cystine broth

 24 ± 3 hrs  24 ± 3 hrs  24 ± 3 hrs

at 41.5 ± 1 °C 34 to 38 °C and 48 ± 3 hrs
34 to 38 °C
Reinoculate each culture obtained using a plate-loop 12

XLD agar Bismuth sulfite agar

 24 ± 3 hrs  24 ± 3 hrs
34 to 38°C and 48 ± 3 hrs
at 34 to 38°C

Detection and confirmation13 of Salmonella spp.

12
Each culture obtained after the selective enrichment step is reinoculated on XLD agar and on Bismuth sulfite agar.
The selenite-cystine broth is reinoculated after 24 hrs and after 48 hrs of incubation.
13
Carry out confirmation as per the general protocol.

Page 66 www.biokar-diagnostics.com
 Section Media and reagents Compliance
- Selenite cystine (SC) broth
D.3.1 Selenite cystine broth Total
500 g vial - BK009HA

- Bismuth sulfite (BS) agar

D.3.2 Bismuth-sulfite agar Total
500 g vial - BK223HA

Salmonella spp.

www.biokar-diagnostics.com Page 67
 SESAME Salmonella TEST®
Alternative method for the detection of Salmonella spp.

BKR 23/04-12/07
Alternative food
analysis method
www.afnor-validation.org

25 g of sample in 225 mL
of Salmonella Enrichment

 18 ± 2 hrs at 37 ± 1°C
Inoculate 0.1 mL

SESAME Salmonella Detection

 24 ± 3 hrs at 41.5 ± 1°C

Migration halo Migration halo

< 30 mm ≥ 30 mm

Absence COMPASS® Salmonella agar

of Salmonella

 24 ± 3 hrs at 37 ± 1 °C

Detection of Salmonella

Page 68 www.biokar-diagnostics.com
 Salmonella Enrichment
10 x 225 mL vials - BM13608
3 x 3 L flexible bags - BM13708
2 x 5 L flexible bags - BM14408
500 g vial - BK194HA
5 kg drum - BK194GC
Salmonella Enrichment + Tween® 80
Enrichment media
3 x 3 L flexible bags - BM16308
2 x 5 L flexible bags - BM19808
Double buffered Salmonella Enrichment
10 x 225 mL vials - BM20108
2 x 5 L flexible bags - BM20008
500 g vial - BK225HA
5 kg drum - BK225GC

Salmonella spp.
SESAME Salmonella Detection
20 Petri dishes Ø 90mm - BM14108
Selective medium
120 Petri dishes Ø 90mm - BM15008
10 x 200 mL vials - BM13808

Confirmation COMPASS Salmonella Agar

medium 20 Petri dishes Ø 90mm - BM06608

www.biokar-diagnostics.com Page 69
 IRIS Salmonella®
Alternative method for the detection of Salmonella spp.

BKR 23/07-10/11
Alternative food
analysis method
www.afnor-validation.org

25 g of sample in 225 mL
of Salmonella Enrichment 1

 18 ± 2 hrs at 41.5 ± 1°C

Spread 10 µL

IRIS Salmonella® Agar

 24 ± 3 hrs at 37 ± 1.0°C

Detection and confirmation of

Salmonella spp.

CONFIRM’ Salmonella (latex test)

1
The IRIS Salmonella® method is also validated for test samples of 50 g to 375 g for dried milk (including infant formula
with and without probiotics) and 50 g to 125 g for flour and croquettes used in animal feedstuffs.

Page 70 www.biokar-diagnostics.com
 Salmonella Enrichment
10 x 225 mL vials - BM13608
3 x 3 L flexible bags - BM13708
2 x 5 L flexible bags - BM14408
500 g vial - BK194HA
5 kg drum - BK194GC
Salmonella Enrichment + Tween® 80
3 x 3 L flexible bags - BM16308
Selective 2 x 5 L flexible bags - BM19808
enrichment media Double buffered Salmonella Enrichment
10 x 225 mL vials - BM20108
2 x 5 L flexible bags - BM20008
500 g vial - BK225HA
5 kg drum - BK225GC
IRIS Salmonella® supplement

Salmonella spp.
120 tablets q.s. 225 mL - BS07708
120 tablets q.s. 90 mL - BS09308
10 x 50 mL vials - BS07808

IRIS Salmonella® Agar

Selective medium 20 Petri dishes Ø 90mm - BM16008
120 Petri dishes Ø 90mm - BM16108

Confirmation COMPASS® Salmonella Agar

reagent Latex agglutination kit (50 tests) - BT01108

www.biokar-diagnostics.com Page 71
 Horizontal method for the detection of Shigella spp.

NF EN ISO 21567: 03-2005

V 08-411

x g of sample in
9x mL of broth

 16 to 20 hrs at 41.5 ± 1°C

MacConkey agar XLD agar HEKTOEN agar

 20 to 24 hrs 1  20 to 24 hrs 1  20 to 24 hrs 1

at 37 ± 1.0°C at 37 ± 1.0°C at 37 ± 1.0°C

Detection and confirmation 2 of Shigella spp.

▪ Serological confirmation
▪ Biochemical confirmation
- TSI agar
- Semi-solid nutrient agar for motility tests
- Urea agar (Christensen)
- L-lysine decarboxylation (LDC) medium
- L-ornithine decarboxylase (ODC) medium
- β-galactosidase reagent
- Indole reaction
- Carbohydrate utilization
- Additional biochemical differentiation tests

1
If no characteristic colonies appear and growth of other microorganisms is weak, incubate for an additional 24 ± 2 hours.
2
Reinoculate 5 characteristic colonies for each plate.
Other identification methods validated by the user may also be used for identification of Shigella species.
3
“Tryptone” is a peptone obtained by pancreatic digestion of casein
4
The fuchsin acid concentration is 40 mg/L instead of 0.1 g/L as described.

Page 72 www.biokar-diagnostics.com
 Section Media and reagents Compliance
9.2 Enrichment - Shigella broth
- -
medium
- MacConkey agar
MacCONKEY agar Total 3
500 g vial - BK050HA
- Xylose deoxycholate (XLD) agar
9.3 Plating-out XLD agar (ISO 6579) Total
media 20 Petri dishes Ø 90 mm - BM08708
500 g vial - BK168HA
- Hektoen enteric (HE) agar Partial 4
Hektoen agar
500 g vial - BK067HA
- Nutrient agar
2% nutrient agar
50 x 10 mL tubes - BM11808 Total 3
500 g vial - BK185HA

Shigella spp.
- Triple sugar/iron agar (TSI agar)
TSI agar Partial
500 g vial - BK221HA
- Semi-solid nutrient agar
- -
- Urea agar (Christensen medium)
- -
- L-lysine decarboxylase (LDC) medium
- -
- L-ornithine decarboxylase (ODC) medium
- -
- Tryptone/ DL-Tryptophan medium
Tryptophan broth
500 g vial - BK163HA Total
50 x 3 mL - BM07608
9.4 Confirmation - Kovac’s indole reagent
media and reagents - -
(biochemical tests) - β-galactosidase reagent
- -
- Buffer solution
- -
- Saline solution
- -
- Bromocresol purple broth
- -
- Sodium acetate agar
- -
- Christensen’s citrate agar
- -
- Mucate broth
- -
- Test broth
- -
- Control broth
- -
- Shigella spp. antisera
- -

www.biokar-diagnostics.com Page 73
 Horizontal method for the enumeration of coagulase-positive
Staphylococci (Staphylococcus aureus and other species)
Part 1: Technique using Baird-Parker agar medium
NF EN ISO 6888-1: 02-1999
Modified by Amendment A1 (2004)
Modified by Amendment A2 (2018)
V 08-014-1

Initial suspension
and decimal dilutions

0.1 mL on surface 1, 2

BAIRD-PARKER agar with

egg yolk and tellurite

 24 ± 2 hrs at 35°C or 37°C 3

+ an additional 24 ± 2 hrs

Enumeration and confirmation 4

of coagulase-positive Staphylococci

Brain-heart infusion broth Or BAIRD-PARKER RPF agar

 24 ± 2 hrs  24 to 48 hrs
at 35°C or 37°C 3 at 35°C or 37°C 3

Add 0.1 mL of culture to Reading

0.3 mL of rabbit plasma
 4 to 6 hrs
at 35°C or 37°C 3

Observe the coagulation reaction

1
Inoculate each dilution in duplicate.
2
If necessary, the enumeration limits may be increased by inoculation of 1.0 mL on the surface of a plate Ø 140 mm or
3 plates Ø 90 mm.
3
The temperature is agreed between the parties concerned and is stated in the test report.
4
Using each plate, reinoculate 5 characteristic colonies and 5 non-characteristic colonies.

Page 74 www.biokar-diagnostics.com
 Section Media and reagents Compliance
Refer to the section of standard NF EN ISO 6887 corresponding to the product
5.2 Diluent
to be examined, pages 96 to 103.

- Baird-Parker agar
BAIRD-PARKER agar with egg yolk and tellurite
20 Petri dishes Ø 90 mm - BM01808
Total 5
120 Petri dishes Ø 90 mm - BM09108
BAIRD-PARKER base agar
5.3 Agar medium 500 g vial - BK055HA Total 5
5 kg drum - BK055GC
Egg yolk emulsion with potassium tellurite
10 x 50 mL vials - BS06008
Sulfamethazine selective supplement (optional) 6 Total
10 x 25 mg vials - BS02808

- Brain-heart infusion broth

Brain-heart broth
500 g vial - BK015HA Partial 7
5.4 Confirmation
media and reagents - Rabbit plasma

Staphylococcus
Lyophilized coagulase rabbit plasma
10 vials 20 reactions - BR00208 Total

AMENDMENT 1: Inclusion of precision data

AMENDMENT 2 : Inclusion of an alternative confirmation test using RPFA stab method

- Rabbit plasma fibrinogen agar

BAIRD-PARKER RPF agar
20 Petri dishes Ø 90 mm - BM06708
20 Petri dishes Ø 55 mm - BM15908
Kit 6 x 200 mL vials + supplements - BT01008
5.6 Confirmation Kit 6 x 100 mL vials + supplements - BT00508 Total 5
media BAIRD-PARKER base agar
500 g vial - BK055HA
5 kg drum - BK055GC
Rabbit plasma fibrinogen (RPF) supplement
8 vials q.s. 100 mL - BS03408
Vial q.s. 500 mL - BS03808

5
“Tryptone” is a peptone obtained by pancreatic digestion of casein
6
Optional additive, justified by the suspected presence of Proteus.
7
Presence of a pancreatic gelatin peptone instead of the enzymatic digest of the animal tissue recommended.

www.biokar-diagnostics.com Page 75
 Horizontal method for the enumeration of coagulase-positive
Staphylococci (Staphylococcus aureus and other species)
Part 2: Technique using rabbit plasma fibrinogen agar medium
NF EN ISO 6888-2: 10-1999
Modified by Amendment A1 (01-2004)
V 08-014-2

Initial suspension
and decimal dilutions

1 mL in poured plate 1

BAIRD-PARKER RPF agar

 18 to 24 at 35°C or 37°C 2
+ an additional 18 to 24 hrs

Enumeration of coagulase-positive
Staphylococci

1
Inoculate each dilution in duplicate.
2
The temperature is agreed between the parties concerned and is stated in the test report.

Page 76 www.biokar-diagnostics.com
 Section Media and reagents Compliance
Refer to the section of standard NF EN ISO 6887 corresponding to the product
5.2 Diluent
to be examined, pages 96 to 103.
- Rabbit plasma fibrinogen agar
BAIRD-PARKER RPF agar
20 Petri dishes Ø 90 mm - BM06708
20 Petri dishes Ø 55 mm - BM15908
Kit 6 x 200 mL vials + supplements - BT01008 Total 3
Kit 6 x 100 mL vials + supplements - BT00508
5.3 Agar medium BAIRD-PARKER base agar
500 g vial - BK055HA
5 kg drum - BK055GC
Rabbit plasma fibrinogen (RPF) supplement Total 3
8 vials q.s. 100 mL - BS03408
Vial q.s. 500 mL - BS03808

Staphylococcus
Horizontal method for the enumeration of coagulase-positive Staphylococci (Staphylococcus aureus and other
species)

Part 2: Technique using rabbit plasma fibrinogen agar medium E

AMENDMENT 1: Inclusion of precision data

3
“Tryptone” is a peptone obtained by pancreatic digestion of casein

www.biokar-diagnostics.com Page 77
 EASY Staph® Agar
Alternative method for the enumeration of coagulase-positive Staphylococci

BKR 23/10-12/15
Alternative food
analysis method
www.afnor-validation.org

Initial suspension
(decimal dilutions)

1 mL in poured plate
0.1 mL on surface 1
Spiral plating 2

EASY Staph® Agar

 24 ± 2 hrs at 37 ± 1°C

Enumeration of coagulase-positive
Staphylococci

Without confirmation

1
The enumeration limit can be decreased by a factor of 10 by inoculating 1 mL on the surface of 3 Ø 90 mm plates.
2
Inoculation may be performed in logarithmic mode of 50 µL or 100 µL.

Page 78 www.biokar-diagnostics.com
 Refer to the section of standard NF EN ISO 6887 corresponding to the
Diluent
product to be examined, pages 96 to 103.

EASY Staph® Agar pre-poured

20 Petri dishes Ø 90 mm - BM18708
120 Petri dishes Ø 90 mm - BM19008
EASY Staph® Agar Kit
6 x 190 mL vials of base medium + 6 vials of
supplement q.s. 200 mL - BT01208
Selective medium
6 x 90 mL vials of base medium + 6 vials of
supplement q.s. 100 mL - BT01308
EASY Staph® Dehydrated agar
500 g vial - BK216HA
Supplement for EASY Staph® Agar
8 vials q.s. 100 mL - BS09008

Staphylococcus

www.biokar-diagnostics.com Page 79
 Horizontal method for the enumeration of coagulase-positive
Staphylococci (Staphylococcus aureus and other species)
Part 3: Detection and MPN technique for low numbers
NF EN ISO 6888-3: 06-2003
V 08-014-3

x g of sample in
9x mL of diluent

1 mL in 9 mL 1 10 mL in 10 mL 1

Single strength modified GIOLITTI Double strength modified GIOLITTI

and CANTONI broth and CANTONI broth

 24 ± 2 hrs at 35°C or 37°C

+ 24 ± 2 hrs if necessary

BAIRD-PARKER agar with

BAIRD PARKER RPF agar or egg yolk and tellurite

 24 to 48 hrs  24 to 48 hrs
at 35°C or 37°C at 35°C or 37°C

Enumeration of coagulase-positive Staphylococci 4

Brain-heart infusion broth

 24 ± 2 hrs at 35°C or 37°C

Add 0.1 mL of culture

to 0.3 mL of rabbit plasma

 4 to 6 hrs at 35°C or 37°C

Observe the coagulation reaction

1
Inoculate 3 tubes of single strength broth and 3 tubes of double strength broth. After inoculation, pour a layer of agar
or paraffin onto the surface of the broth, then allow to solidify so as to form a sealed plug.
2
If Baird-Parker agar with egg yolk and tellurite is used for reinoculation, a confirmation test will need to be performed.

Page 80 www.biokar-diagnostics.com
 Section Media and reagents Compliance
Refer to the section of standard NF EN ISO 6887 corresponding to the product
5.1 Diluent
to be examined, pages 96 to 103.
- Modified Giolitti and Cantoni broth
GIOLITTI and CANTONI broth with Tween® 80
50 x 10 mL tubes - BM11008 (single strength)
5.2 Enrichment Total 3
50 x 10 mL tubes - BM11108 (double strength)
medium 500 g vial - BK159HA
- Potassium tellurite solution
-

- Agar solution (20.0 g/L)

Type A bacteriological agar Total
500 g vial - A1010HA
5.3 Agar solution 5 kg drum - A1010GC
Type E bacteriological agar
500 g vial - A1012HA Total
5 kg drum - A1012GC

- Rabbit plasma fibrinogen agar

Staphylococcus
BAIRD-PARKER RPF agar
20 Petri dishes Ø 90 mm - BM06708
20 Petri dishes Ø 55 mm - BM15908
Kit 6 x 200 mL vials + supplements - BT01008 Total 3
Kit 6 x 100 mL vials + supplements - BT00508
5.4 Agar medium
BAIRD-PARKER base agar
500 g vial - BK055HA
5 kg drum - BK055GC
Total 3
Rabbit plasma fibrinogen supplement
8 vials q.s. 100 mL - BS03408
Vial q.s. 500 mL - BS03808
Baird-Parker agar
BAIRD-PARKER agar with egg yolk and tellurite
20 Petri dishes Ø 90 mm - BM01808
Total 3
120 Petri dishes Ø 90 mm - BM09108
BAIRD-PARKER base agar
5.5 Agar medium 500 g vial - BK055HA
5 kg drum - BK055GC Total 3
Egg yolk emulsion with potassium tellurite
10 x 50 mL vials - BS06008
Sulfamethazine selective supplement (optional) Total
10 x 25 mg vials - BS02808
- Brain-heart infusion broth
Brain-heart infusion broth
5.6/5.7 500 g vial - BK015HA Partial 4
Confirmation media - Rabbit plasma
and reagents Lyophilized coagulase rabbit plasma
10 vials 20 reactions - BR00208 Total

3
“Tryptone” is a peptone obtained by pancreatic digestion of casein
4
Presence of a pancreatic gelatin peptone instead of the enzymatic digest of the animal tissue recommended.

www.biokar-diagnostics.com Page 81
 Routine method for the enumeration of coagulase-positive
Staphylococci by colony-count technique at 37°C
Part 1: Technique with confirmation of the colonies
NF V 08-057-1: 01-2004
V 08-057-1

Initial suspension
and decimal dilutions

0.1 mL on surface 1

BAIRD-PARKER agar with

egg yolk and tellurite

 24 ± 2 hrs at 37°C
+ an additional 24 ± 2 hrs

Enumeration and confirmation 2

of coagulase-positive Staphylococci

▪ Detection of catalase (optional)

▪ Detection of coagulase
- Brain-heart infusion broth + Rabbit plasma
- BAIRD-PARKER RPF agar

1
In the event of enumeration of low numbers of coagulase-positive Staphylococci, the qualification level may be
increased by inoculating 1 mL on the surface of a large plate (Ø 140 mm) or 3 small plates (Ø 90 mm).
2
Confirm a defined number of colonies. In general, 3 characteristic colonies or of each type (characteristic or
non-characteristic) are confirmed using each plate.

Page 82 www.biokar-diagnostics.com
 Section Media and reagents Compliance

Refer to the section of standard NF EN ISO 6887 corresponding to the product

5.2 Diluent
to be examined, pages 96 to 103.
- Baird-Parker agar
BAIRD-PARKER agar with egg yolk and tellurite
20 Petri dishes Ø 90 mm - BM01808
Total 3
120 Petri dishes Ø 90 mm - BM09108
BAIRD-PARKER base agar
5.3 Agar medium 500 g vial - BK055HA
5 kg drum - BK055GC Total 3
Egg yolk emulsion with potassium tellurite
10 x 50 mL vials - BS06008
Sulfamethazine selective supplement (optional) 4
10 x 25 mg vials - BS02808 Total

- Rabbit plasma fibrinogen agar

BAIRD-PARKER RPF agar
20 Petri dishes Ø 90 mm - BM06708

Staphylococcus
20 Petri dishes Ø 55 mm - BM15908
Kit 6 x 200 mL vials + supplements - BT01008 Total 3
5.4 Confirmation Kit 6 x 100 mL vials + supplements - BT00508
medium BAIRD-PARKER base agar
500 g vial - BK055HA
5 kg drum - BK055GC
Rabbit plasma fibrinogen supplement Total 3
8 vials q.s. 100 mL - BS03408
Vial q.s. 500 mL - BS03808

- Brain-heart infusion broth

Brain-heart infusion broth
5.6/5.7 500 g vial - BK015HA Partial 5
Confirmation media - Rabbit plasma
and reagents Lyophilized coagulase rabbit plasma
10 vials 20 reactions - BR00208 Total

3
“Tryptone” is a peptone obtained by pancreatic digestion of casein.
4
Optional additive, justified by the suspected presence of Proteus.
5
Presence of a pancreatic gelatin peptone instead of the enzymatic digest of the animal tissue recommended.

www.biokar-diagnostics.com Page 83
 Horizontal method for the enumeration of
sulfite-reducing bacteria growing under anaerobic conditions
NF ISO 15213: 09-2003
V 08-029

Initial suspension
and decimal dilutions 1

1 mL in poured plate,
double layer 2

TSC agar

 24 hrs and 48 hrs at 37 ± 1°C3

under anaerobic conditions

Enumeration and Enumeration of

confirmation of Clostridia 4 sulfite-reducing bacteria

- Respiratory test
- Spore forming test

1
Heat treatment of the initial suspension may be necessary in order to eliminate vegetative forms of sporulating bacteria and/or
non-sporulating bacteria. The temperature and heating time vary according to requirements (for example, 20 minutes at 75°C).
2
Inoculate each dilution in duplicate. Tube inoculation may also be performed, gently mixing 1 mL of inoculum with the
reliquefied medium. Allow to solidify and cover with a second layer of medium (2 to 3 mL).
3
If the presence of thermophilic bacteria is suspected, prepare a second series of Petri dishes or tubes, and incubate at 50 ± 1°C.
4
This standard is also suitable for the enumeration of Clostridia only. In this case, once characteristic colonies have been
obtained, five should be sampled from each plate used, the Clostridium genus should be confirmed by confirmation tests
(for example, tests on respiratory potential, spore formation).

Page 84 www.biokar-diagnostics.com
 Section Media and reagents Compliance
Refer to the section of standard NF EN ISO 6887 corresponding to the product
to be examined, pages 96 to 103.
- Saline peptone solution
5.2 Diluent Tryptone-salt broth
50 x 9 mL tubes - BM00808
10 x 90 mL vials - BM11408 Total 5
3 x 3 L flexible bags - BM13508
500 g vial - BK014HA

- Iron sulfite agar

TSC agar
50 x 20 mL tubes - BM03908
5.1 Agar medium Total 5, 6
10 x 200 mL vials - BM07708
500 g vial - BK031HA

Sulfite-reducing bacteria

5
“Tryptone” is a peptone obtained by pancreatic digestion of casein.
6
If tubes are used, incubation in jars for anaerobic conditions is not necessary.

www.biokar-diagnostics.com Page 85
Enumeration of sulfite-reducing bacteria under anaerobic conditions
by the colony-count technique at 46°C
NF V 08-061: 12-2009
V 08-061

Initial suspension
and decimal dilutions

1 mL in poured
plate, double layer 1

TSC agar

 20 ± 2 hrs at 46°C
under anaerobic
conditions

Enumeration of
sulfite-reducing bacteria

Page 86 www.biokar-diagnostics.com
 Section Media and reagents Compliance
Refer to the section of standard NF EN ISO 6887 corresponding to the product
5.2 Diluent
to be examined, pages 96 to 103.
- Iron sulfite agar free from egg yolk
TSC (base) agar
50 x 20 mL tubes - BM03908
Total 2
10 x 200 mL vials - BM07708
500 g vial - BK031HA
5.3 Agar medium - D-cycloserine solution
D-cycloserine 200 mg selective supplement
10 vials q.s. 500 mL - BS00608 Total
D-cycloserine liquid supplement
10 x 90 mL vials - BS09208
1 x 50 mL vial - BS09408

Sulfite-reducing bacteria

1
Inoculation may be carried out in Petri dishes or tubes.
2
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

www.biokar-diagnostics.com Page 87
 Horizontal method for the determination of Vibrio spp.
Part 1: Detection of potentially enteropathogenic
Vibrio parahaemolyticus, Vibrio cholerae and Vibrio vulnificus

NF EN ISO 21872-1: 09-2017

V 08-064-1

x g of product in
9x mL of ASPW

 6 ± 1 hrs at 41.5°C or 37°C 1

1 mL in 10 mL

ASPW

 18 ± 1 hrs at 41.5°C or 37°C 2

TCBS medium Second medium

 24 ± 3 hrs at 37°C  appropriate

conditions

Detection and confirmation 3 of Vibrio

Saline nutrient agar

 24 ± 3 hrs at 37°C

▪ Presumptive identification test

- Oxidase test
- Microscopic examination (optional)
▪ Biochemical test
- Detection of L-lysine decarboxylase (LDC)
- Detection of arginine dihydrolase (ADH)
- Detection of β-galactosidase
- Detection of indole
- Halotolerance test

Page 88 www.biokar-diagnostics.com
 Section Media and reagents Compliance

- Alkaline saline peptone water (ASPW)

B.1 Enrichment
Alkaline saline peptone water (ASPW) Total
medium
500 g vial - BK219HA

- Thiosulfate, citrate, bile and sucrose (TCBS) agar

TCBS agar Total 4
B.2 Plating-out 500 g vial - BK040HA
media
The choice of the second medium is left to the discretion of the testing
laboratory
B.3 Subculture - Saline nutrient agar (SNA)
-
medium -
B.4 Identification - Oxidase reagent
-
reagent -
- Lysine decarboxylase (LDC) detection medium
- -
- Arginine dihydrolase (ADH) detection medium
-
-
- ONPG solution
- -
- Buffer solution

Vibrio spp.
B.5/B.6/B.7/B.8/B.9/
B.10/B.11 - -
Biochemical - Tryptophan medium
confirmation media Tryptophan broth
and reagents 50 x 3 mL tubes - BM07608 Partial 5
500 g vial - BK163HA
- Saline peptone water
- -
- Sodium chloride solution
- -
- Tris acetate EDTA (TAE) buffer
- -

1
Incubate at 37°C for the detection of V. parahaemolyticus and V. cholerae in frozen, dried and cured products, and for
V. vulnificus in all product states, or at 41.5°C for the detection of V. parahaemolyticus and V. cholerae in fresh products.
2
Incubate at 37°C for the detection of V. vulnificus or at 41.5°C for the detection of V. parahaemolyticus and V. cholerae in all
product states.
3
Using each selective medium, reinoculate at least one well-isolated characteristic colony. If the result of the tests is negative, test
four other colonies.
4
The 8.0 g/L of dried bovine bile consists of the following: 5 g/L of bacteriological bovine bile and 3 g/L of sodium cholate.
5
Presence of 5.0 g of sodium chloride instead of 10.0 g and pH of 7.5 ± 0.2 instead of 7.0 ± 0.2.

www.biokar-diagnostics.com Page 89
 Horizontal method for the detection of
pathogenic Yersinia enterocolitica
NF EN ISO 10273: 06-2017
V 08-027

25 g of sample in
225 mL of PSB broth

 44 ± 4 hrs at 25 ± 1°C
10 mL in 90 mL of
ITC broth

 44 ± 4 hrs
at 25 ± 1°C
1 mL on surface
Alkaline treatment 1 Alkaline treatment 1 on 2 to 4 plates

CIN agar CIN agar CIN agar

 24 ± 2 hrs at 30 ± 1°C  24 ± 2 hrs at 30 ± 1°C

Detection and confirmation2 of presumptive pathogenic Yersinia enterocolitica

CIN agar

 24 ± 2 hrs at 30 ± 1°C

Nutrient agar

 18 to 24 hrs at 30 ± 1°C

▪ Determination of pathogenic Yersinia species

▪ Confirmation of pathogenic Y. enterocolitica
▪ Biotyping of Y. enterocolitica (optional)

1
Transfer 0.5 mL of enrichment to 4.5 mL of KOH, after 20 ± 5 s inoculate in streaks on the surface of a CIN agar plate.
2
Sample 5 characteristic colonies (if available) and prepare a subculture on CIN agar. Examine the plates and reinoculate a
typical colony on a non-selective agar, such as nutrient agar. Then carry out the confirmation tests.

Page 90 www.biokar-diagnostics.com
 Section Media and reagents Compliance

- Peptone, sorbitol and bile salts (PSB) broth -

B.2/B.3 Enrichment -
media - Irgasan, ticarcillin and potassium chlorate (ITC) broth
- -

- Sodium chloride solution -

B.4/B.5 Alkaline -
reagents - Potassium hydroxide solution in saline solution
- -
- Cefsulodin, Irgasan and novobiocin (CIN) agar
- -
B.6/B.7 Plating-out - Nutrient agar (example of a non-selective medium)
media 2% nutrient agar
50 x 18 mL tubes - BM11808 Total 3
500 g vial - BK185HA
- Tryptic soy broth (TSB)
Trypticase soy broth (TSB)
50 x 10 mL tubes - BM03008
10 x 100 mL vials - BM00908 Total 3
10 x 90 mL vials - BM17908
2 x 5 L flexible bags - BM16608
500 g vial - BK046HA/5 kg drum - BK046GC
- Sterile glycerol
- -
- Urea agar (Christensen)

Yersinia spp.
- -
- Congo red-magnesium (CR-MOX) agar
- -
- Bile and esculin agar
- -
B.8/B.9/B.10/B.11/ - Casein-soya agar for detection of pyrazinamidase
B.12/B.13/B.14/ - -
B.15/B.16/B.17/ - Ammonium iron(II) sulfate solution for detection of
pyrazinamidase
B.18/B.19/B.20/ - -
B.21/B.22 - Decarboxylase basal medium (lysine or arginine)
Confirmation media - -
and reagents - Phenylalanine (tryptophan) desaminase agar
- -
- Ferric chloride, 10%
- -
- Media for fermentation of carbohydrates
-
-
- Simmons’ citrate medium
-
- Medium for Tween-esterase test -
-
- Tryptone/tryptophan medium -
Tryptophan broth
50 x 3 mL tubes - BM07608 Partial 4
500 g vial - BK163HA
- Kovac’s reagent -
-

3
“Tryptone” is a peptone obtained by pancreatic digestion of casein.
4
Presence of 1 g/L of tryptophan instead of 3 g/L as recommended.

www.biokar-diagnostics.com Page 91
 Horizontal method for the enumeration of microorganisms
Part 1: Colony count at 30°C by the pour plate technique

NF EN ISO 4833-1: 10-2013

V 08-011-1

Initial suspension
and decimal dilutions

1 mL in poured plate,
in a double layer1, 2

PCA

 72 ± 3 hrs at
30 ± 1 °C

Enumeration of microorganisms

Section Media and reagents Compliance

Refer to the section of standard NF EN ISO 6887 corresponding to the
5.1 Diluent
product to be examined, pages 96 to 103.
- Plate count medium (PCA)
Plate count agar (PCA)
10 x 100 mL vials - BM01508 Total 3
10 x 200 mL vials - BM03308
5.2 Selective
500 g vial - BK144HA
medium
5 kg drum - BK144GC
Plate count agar (PCA) with skim milk 4
10 x 200 mL vials - BM08608 Total 3
500 g vial - BK161HA

Type A bacteriological agar

500 g vial - A1010HA
5.3 Double layer 5 kg drum - A1010GC
Total
medium 2 Type E bacteriological agar
500 g vial - A1012HA
5 kg drum - A1012GC

1
Inoculate each dilution in duplicate.
2
If microorganisms are susceptible to overgrow the surface of agar, add a double layer.
3
“Tryptone” is a peptone obtained by pancreatic digestion of casein.
4
For the analysis of milk products.

Page 92 www.biokar-diagnostics.com
 Horizontal method for the enumeration of microorganisms
Part 2: Colony count at 30°C by the surface plating technique

NF EN ISO 4833-2: 10-2013

V 08-011-2

Initial suspension
and decimal dilutions

0.1 mL on surface 1

PCA

 72 ± 3 hrs at
30 ± 1 °C

Enumeration of microorganisms

Total microorganisms
Section Media and reagents Compliance

Refer to the section of standard NF EN ISO 6887 corresponding to the

5.1 Diluent
product to be examined, pages 96 to 103.
- Plate count medium (PCA)
Plate count agar (PCA)
10 x 100 mL vials - BM01508
10 x 200 mL vials - BM03308 Total 2
5.2 Selective 500 g vial - BK144HA
medium 5 kg drum - BK144GC
Plate count agar (PCA) with skim milk3
10 x 200 mL vials - BM08608 Total 2
500 g vial - BK161HA

1
Inoculate each dilution in duplicate.
2
“Tryptone” is a peptone obtained by pancreatic digestion of casein.
3
For the analysis of milk products.

www.biokar-diagnostics.com Page 93
 Enumeration of microorganisms by colony-count technique
at 30°C after inoculation by the spiral method
XP V 08-034: 09-2010
V 08-034

Initial suspension
and decimal dilutions

Spiral plater 1

PCA

 48 ± 3 hrs at
30 ± 1 °C

Enumeration of microorganisms

Section Media and reagents Compliance

Refer to the section of standard NF EN ISO 6887 corresponding to the

5.2 Diluent
product to be examined, pages 96 to 103.
- Plate count medium (PCA)
Plate count agar (PCA)
10 x 100 mL vials - BM01508
10 x 200 mL vials - BM03308 Total 2
5.3 Selective
500 g vial - BK144HA
medium
5 kg drum - BK144GC
Plate count agar (PCA) with skim milk 3
10 x 200 mL vials - BM08608 Total 2
500 g vial - BK161HA

Type A bacteriological agar

500 g vial - A1010HA
5.4 Double layer 5 kg drum - A1010GC
Total
medium 2 Type E bacteriological agar
500 g vial - A1012HA
5 kg drum - A1012GC

1
If microorganisms are susceptible to overgrow the surface of agar, add a double layer.
2
“Tryptone” is a peptone obtained by pancreatic digestion of casein.
3
For the analysis of milk products.

Page 94 www.biokar-diagnostics.com
 Horizontal method for the enumeration of
psychrotrophic microorganisms
NF ISO 17410: 11-2001
V 08-033

Initial suspension
and decimal dilutions

0.1 mL on surface 1

PCA

 10 days
at 6.5 °C

Enumeration of psychrotrophic
microorganisms

Total microorganisms
Section Media and reagents Compliance

Refer to the section of standard NF EN ISO 6887 corresponding to the

5.1 Diluent
product to be examined, pages 96 to 103.

- Plate count medium (PCA)

Plate count agar (PCA)
10 x 100 mL vials - BM01508
Total 2
10 x 200 mL vials - BM03308
5.2 Selective
500 g vial - BK144HA
medium
5 kg drum - BK144GC
Plate count agar (PCA) with skim milk 3
Total 2
10 x 200 mL vials - BM08608
500 g vial - BK161HA

1
Inoculate each dilution in duplicate.
2
“Tryptone” is a peptone obtained by pancreatic digestion of casein.
3
For the analysis of milk products.

www.biokar-diagnostics.com Page 95
 Preparation of test samples,
Preparation of testinitial suspension
samples, and decimal
initial suspension dilutions for
and
microbiological
decimal dilutions examination
for microbiological examination
Part1:1:General
Part Generalrules
rulesfor
forthe
the preparation
preparation of
of the
the initial
initial suspension
suspension and
anddecimal
decimaldilutions
dilutions
NF EN ISO 6887-1: 06-2017
V 08-010-1

1. Preparation of the initial suspension (1:10)

x g or x mL of sample
+

2. Decimal dilutions
1 mL of initial suspension
+
9 mL of diluent

Section Media and reagents Compliance

- Saline peptone solution

Tryptone-salt broth
50 x 9 mL tubes - BM00808
10 x 90 mL vials - BM11408 Total
3 x 3 L flexible bags - BM13508
500 g vial - BK014HA
- Buffered Peptone Water
Buffered Peptone Water (20.0 g/L) 1
500 g vial - BK131HA/5 kg drum - BK131GC
Buffered Peptone Water (25.5 g/L) 2
5.2 Diluents for
50 x 9 mL tubes - BM05608
general use
10 x 90 mL vials - BM05708 Total 1
10 x 225 mL vials - BM01008
3 x 3 L flexible bags - BM13108
2 x 5 L flexible bags - BM13208
500 g vial - BK018HA/5 kg drum - BK018GC
- Double strength buffered peptone water
Salmonella Enrichment double strength buffered 3
500 g vial - BK225HA/5 kg drum - BK225GC Total
2 x 5 L flexible bags - BM20008
10 x 225 mL vials - BM20108
5.3 Diluents for Refer to the section of standard NF EN ISO 6887 corresponding to the
special purposes product to be examined, pages 99 to 103.

1
Formula including 9.0 g/L of disodium phosphate dodecahydrate (molecular mass 358.14).
2
Formula including 3.56 g/L of anhydrous disodium phosphate (molecular mass 141.96).
3
The Salmonella Enrichment double-strength buffered formulation conforms to that of double-strength buffered peptone water.

Page 96 www.biokar-diagnostics.com
 Preparation of test samples, initial suspension and
decimal dilutions for microbiological examination
Part 2: Specific rules for the preparation of meat and meat products

NF EN ISO 6887-2: 06-2017

V 08-010-2

Preparation of the initial suspension and, if necessary, decimal dilutions for the test samples of:

▪ fresh, raw and processed meats,

▪ poultry,
▪ game,
▪ and their products:
- refrigerated or frozen;
- cured or fermented;
- minced or comminuted;
- meat preparations;
- mechanically separated meat;
- cooked meats;
- dried and smoked meats at various degrees of dehydration;
- concentrated meat extracts;
- excision and swab samples from carcasses.

Part Media and reagents Initial suspension, dilutions

5. Diluents Refer to standard NF EN ISO 6887-1, page 96

www.biokar-diagnostics.com Page 97
 Preparation of test samples, initial suspension and
decimal dilutions for microbiological examination
Part 3: Specific rules for the preparation of fish and fishery products
NF EN ISO 6887-3: 06-2017
V 08-010-3

Preparation of the initial suspension and, if necessary, decimal dilutions for the test samples of:
▪ Raw fishery products, molluscs, tunicates and echinoderms
▪ Processed products,
▪ Raw or cooked frozen fish, crustaceans, molluscs and others, in blocks or otherwise.

Part Media and reagents

5. Diluents Refer to standard NF EN ISO 6887-1, page 96

Page 98 www.biokar-diagnostics.com
 Preparation of test samples, initial suspension and
decimal dilutions for microbiological examination
Part 4: Specific rules for the preparation of miscellaneous products

NF EN ISO 6887-4: 06-2017

V 08-010-4

Preparation of the initial suspension and, if necessary, decimal dilutions for the test samples of:
▪ acidic (low pH) products;
▪ hard and dry products;
▪ dehydrated, freeze-dried and other low aw products
▪ flours, whole cereal grains, cereal by-products;
▪ animal feed, cattle cake, kibbles and pet chews;
▪ gelatin (powdered and leaf);
▪ margarines, spreads and non-dairy products with added water;
▪ eggs and egg products;
▪ bakery goods, pastries and cakes;
▪ fresh fruit and vegetables;
▪ fermented products and other products containing viable microorganisms;
▪ alcoholic and non-alcoholic beverages;
▪ alternative protein products.

Initial suspension, dilutions

Section Media and reagents Compliance

5.1 Basic materials Refer to standard NF EN ISO 6887-1, page 96.

- Peptone salt solution with bromocresol purple

Tryptone-salt broth (base)
50 x 9 mL tubes - BM00808
10 x 90 mL vials - BM11408
3 x 3 L flexible bags - BM13508 Total 1
500 g vial - BK014HA
5.2 Diluents for - Buffered Peptone Water
general use Buffered Peptone Water (20.0 g/L) 2
500 g vial - BK131HA/5 kg drum - BK131GC
Buffered Peptone Water (25.5 g/L) 3
50 x 9 mL tubes - BM05608 Total 1
10 x 90 mL vials - BM05708
10 x 225 mL vials - BM01008
3 x 3 L flexible bags - BM13108
2 x 5 L flexible bags - BM13208

1
“Tryptone” is a peptone obtained by pancreatic digestion of casein.
2
Formula including 9.0 g/L of disodium phosphate dodecahydrate (molecular mass 358.14).
3
Formula including 3.56 g/L of anhydrous disodium phosphate (molecular mass 141.96).

www.biokar-diagnostics.com Page 99
 Section Media and reagents Compliance

Double strength buffered peptone water

Salmonella Enrichment double strength buffered 3
500 g vial - BK225HA
5 kg drum - BK225GC
5.3 Diluents for 2 x 5 L flexible bags - BM20008 Total1
special purposes 10 x 225 mL vials - BM20108

- Phosphate buffer solution

-

- Alpha-amylase solution
-
- Cellulase solution -
5.6 Enzyme solutions
- -
- Papain solution
- -

1
“Tryptone” is a peptone obtained by pancreatic digestion of casein.
2
Formula including 9.0 g/L of disodium phosphate dodecahydrate (molecular mass 358.14).
3
Formula including 3.56 g/L of anhydrous disodium phosphate (molecular mass 141.96).
4
Multiply the reconstitution rate by two, to yield double strength buffered peptone water.

Page 100 www.biokar-diagnostics.com

 Preparation of test samples, initial suspension and
decimal dilutions for microbiological examination
Part 5: Specific rules for the preparation of milk and milk products
NF EN ISO 6887-5: 10-2010
V 08-010-5

Preparation of the initial suspension and, if necessary, decimal dilutions for the test samples of:
▪ milk and liquid milk products
▪ dried milk products;
▪ cheese;
▪ casein and caseinates;
▪ butter;
▪ ice-cream;
▪ custard, desserts and sweet cream;
▪ fermented milk and sour cream;
▪ milk-based infant foods.

Section Media and reagents Compliance

- Saline peptone solution

Tryptone-salt broth
50 x 9 mL tubes - BM00808

Initial suspension, dilutions

10 x 90 mL vials - BM11408 Total 1
3 x 3 L flexible bags - BM13508
500 g vial - BK014HA

- Quarter-strength Ringer’s solution

Quarter-strength RINGER’s solution Total
100 tablets - BR00108

5.2 Diluents for - Peptone solution

0.1% peptone water
general use Total 1
2 x 5 L flexible bags - BM16408

- Buffered Peptone Water

Buffered Peptone Water (20.0 g/L) 3
500 g vial - BK131HA/5 kg drum - BK131GC
Buffered Peptone Water (25.5 g/L) 4
50 x 9 mL tubes - BM05608
10 x 90 mL vials - BM05708 Total
10 x 225 mL vials - BM01008
3 x 3 L flexible bags - BM13108
2 x 5 L flexible bags - BM13208
500 g vial - BK018HA/5 kg drum - BK018GC

1
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

www.biokar-diagnostics.com Page 101

 Section Media and reagents Compliance

- Sodium citrate solution

- -
- Diphosphate hydrogenphosphate solution
Dipotassium hydrogenphosphate solution 20 g/L
Total
2 x 5 L flexible bags - BM19308
- Dipotassium hydrogenphosphate solution with antifoam
agent -
-
- Tripolyphosphate solution -
-
- Diluent for general use with α-amylase solution
-
5.3 Diluents for -
special purposes - Buffered Peptone Water with bromocresol purple
Buffered Peptone Water (20.0 g/L) 3
500 g vial - BK131HA/5 kg drum - BK131GC
Buffered Peptone Water (25.5 g/L) 4
50 x 9 mL tubes - BM05608
10 x 90 mL vials - BM05708
10 x 225 mL vials - BM01008 Total
3 x 3 L flexible bags - BM13108
2 x 5 L flexible bags - BM13208
500 g vial - BK018HA/5 kg drum - BK018GC
- Bromocresol purple supplement -
-

Page 102 www.biokar-diagnostics.com

 Preparation of test samples, initial suspension and
decimal dilutions for microbiological examination
Part 6: Specific rules for the preparation of samples taken at the primary production stage

NF EN ISO 6887-6: 05-2013

V 08-010-6

Preparation of the initial suspension and, if necessary, decimal dilutions for the samples taken from the :
▪ farm;
▪ environment;
▪ animals;
▪ slaughterhouse;
▪ hatchery;
▪ vehicle, units or crates used for animal transportation.

Initial suspension, dilutions

Section Media and reagents

5. Diluents Refer to standard NF EN ISO 6887-1, page 96.

5.2 Diluents for - Neutralizing agents

special purposes -

www.biokar-diagnostics.com Page 103

 Animal feeding stuffs
Isolation and enumeration of Bifidobacterium spp.
NF EN 15785: 12-2009
V 18-238

Initial suspension
(and decimal dilutions)

0.1 mL on surface 1

MRS agar 2

 36 to 48 hrs at 37 ± 1°C
under anaerobic conditions

Enumeration and
morphological confirmation 3

1
Inoculate each dilution on 2 plates.
2
If an additional microflora is present, inoculation may be performed in a selective medium, such as AMRS agar, MRS agar +
TTC and MRS agar with selective antibiotics.
3
If colonies with different characteristics are detected during confirmation, repeat the analysis with a more selective medium
(MRS+TTC, AMRSA, selective antibiotic medium).
Confirm 2 to 5 colonies of each morphological type, selected at random.

Page 104 www.biokar-diagnostics.com

 Section Media and reagents Compliance

- Phosphate buffer (PBS)

- -
- Saline peptone solution
Tryptone-salt broth
5.1 Diluents
50 x 9 mL tubes - BM00808
10 x 90 mL vials - BM11408 Total 4
3 x 3 L flexible bags - BM13508
500 g vial - BK014HA

- MRS agar
MRS agar
10 x 200 mL vials - BM08908 Total 5
500 g vial - BK089HA
- MRS agar supplemented with TTC 6
MRS agar
10 x 200 mL vials - BM08908
500 g vial - BK089HA
TTC supplement 12.5 mg Total
5.2 Culture media 10 vials - BS02608
TTC supplement 50 mg
10 vials - BS02708
- AMRSA (pH 5.4)
MRS agar
10 x 200 mL vials - BM08908
500 g vial - BK089HA Partial 5
- Selective medium supplemented with 0.05% cysteine

Animal feeding stuffs

hydrochloride
-
-

4
“Tryptone” is a peptone obtained by pancreatic digestion of casein.
5
The pH of the medium is 5.7, and will be adjusted to 6.5 or 5.4 according to standard practice.
6
MRS medium at pH 6.5 supplemented with 1 mL per 100 mL of a 1% TTC solution.

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 Animal feeding stuffs
Isolation and enumeration of presumptive Bacillus spp.
NF EN 15784: 12-2009
V 18-237

Initial suspension diluted 1:10

10 min at 80 ± 1°C

decimal dilutions
(if necessary)
1 mL on the surface
of 2 Petri dishes

Tryptone soy agar

 16 to 24 hrs at 37 ± 1°C

Enumeration

Section Media and reagents Compliance

- Phosphate buffer (PBS)

- -
- 0.2% sodium hydroxide solution
- -
- Saline peptone solution
Tryptone-salt broth
5.1 Diluents 50 x 9 mL tubes - BM00808
10 x 90 mL vials - BM11408 Total
3 x 3 L flexible bags - BM13508
500 g vial - BK014HA
- Polysorbate 80 solution
- -

- Tryptone soy agar

Trypticase soy agar (TSA)
20 plates Ø 90 mm - BM05008
5.2 Culture medium Total 1
10 x 100 mL vials - BM01708
10 x 200 mL vials - BM04908
500 g vial - BK047HA

1
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

Page 106 www.biokar-diagnostics.com

 Animal feeding stuffs
Isolation and enumeration of Enterococci (E. faecium) spp.
NF EN 15788: 12-2009
V 18-232

Initial suspension
(and decimal dilutions)

0.1 mL on surface 1

BEA agar

 24 ± 2 hrs at
37 ± 1 °C

Enumeration and
morphological confirmation

Animal feeding stuffs

Section Media and reagents Compliance

- Phosphate buffer (PBS)

- -
- Saline peptone solution
Tryptone-salt broth
5.1 Diluents 50 x 9 mL tubes - BM00808
10 x 90 mL vials - BM11408 Total 2
3 x 3 L flexible bags - BM13508
500 g vial - BK014HA

- Bile-esculin-azide (BEA) agar

5.2 Selective BEA agar
10 x 100 mL vials - BM10408 Partial 2, 3
medium
500 g vial - BK158HA

1
Inoculate 2 plates and repeat the operation for each dilution.
2
“Tryptone” is a peptone obtained by pancreatic digestion of casein.
3
Presence of 0.15 g of sodium azide instead of 0.25 g as recommended.

www.biokar-diagnostics.com Page 107

 Animal feeding stuffs
Isolation and enumeration of Lactobacillus spp.
NF EN 15787: 12-2009
V 18-231

Initial suspension
(and decimal dilutions)

0.1 mL on surface 1

MRS agar 2

 48 to 72 hrs at
37 ± 1°C under
anaerobic conditions

Enumeration and
morphological confirmation 3

1
Inoculate in duplicate and repeat the operation for each dilution.
2
If Lactobacilli are the only bacterial components of a feed, use MRS agar or AMRSA; if predominantly present with an
additional microflora, enumeration may begin in AMRSA, MRS + TTC or LAMVAB medium. If Lactobacilli are not predominant,
use LAMVAB medium.
If colonies are detected which do not fulfil these criteria, repeat the analysis with AMRSA and/or LAMVAB agar. If yeasts are
present, add nystatin to the agar.
3
Confirm 2 to 5 colonies of each morphological type, selected at random.

Page 108 www.biokar-diagnostics.com

 Section Media and reagents Compliance

- Phosphate buffer (PBS)

- -
- Saline peptone solution
Tryptone-salt broth
5.1 Diluents 50 x 9 mL tubes - BM00808
10 x 90 mL vials - BM11408 Total 4
3 x 3 L flexible bags - BM13508
500 g vial - BK014HA

- MRS agar
MRS agar
10 x 200 mL vials - BM08908 Total 5
500 g vial - BK089HA
- MRS agar supplemented with TTC 6
MRS agar
10 x 200 mL vials - BM08908
500 g vial - BK089HA
TTC supplement 12.5 mg
10 vials - BS02608 Total
5.2 Culture media
TTC supplement 50 mg
10 vials - BS02708
- AMRSA (pH 5.4)
MRS agar
10 x 200 mL vials - BM08908
Partial 5
500 g vial - BK089HA

Animal feeding stuffs

- Selective medium supplemented with 0.05% cysteine
hydrochloride -

4
The pH of the medium is 5.7, and will be adjusted to 6.5 or 5.4 according to standard practice.
5
MRS medium at pH 6.5 supplemented with 1 mL per 100 mL of a 1% TTC solution.

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 Animal feeding stuffs
Isolation and enumeration of Pediococcus spp.
NF EN 15786: 12-2009
V 18-230

Initial suspension
(and decimal dilutions)

1 mL applied
to 2 plates

MRS agar 1

 24 to 48 hrs at
37 ± 1°C under
anaerobic conditions 2

Enumeration and
morphological confirmation 3

1
If Pediococci are the only bacterial components of the feed, use the MRS agar. If these are predominantly present with an
additional microflora, enumeration may be performed in AMRSA, MRS + TTC medium or in a selective antibiotic medium. If
Pediococci are not predominant, use the selective antibiotic medium.
2
Incubate the selective antibiotic medium for 24 hours only, so as to limit the growth of Lactobacillus colonies. Incubate the
MRSA, AMRSA and MRSA + TTC agar for a period of between 36 and 48 hrs. Incubate the MRSA + TTC plates under anaerobic
conditions; incubation under anaerobic conditions is optional for the MRS and AMRS agars.
3
Confirm 2 to 5 colonies of each morphological type, selected at random.

Page 110 www.biokar-diagnostics.com

 Section Media and reagents Compliance

- Phosphate buffer (PBS) -Total

-
- Saline peptone solution
Tryptone-salt broth
5.1 Diluents 50 x 9 mL tubes - BM00808
10 x 90 mL vials - BM11408
3 x 3 L flexible bags - BM13508
500 g vial - BK014HA

- MRS agar
MRS agar
500 g vial - BK089HA
10 x 200 mL vials - BM08908 Total 4
- MRS agar supplemented with TTC 5
MRS agar
500 g vial - BK089HA
10 x 200 mL vials - BM08908
TTC supplement 12.5 mg
10 vials - BS02608 Total
5.2 Culture media
TTC supplement 50 mg
10 vials - BS02708
- AMRSA (pH 5.4)
MRS agar
Total 4
500 g vial - BK089HA
10 x 200 mL vials - BM08908
- Selective medium supplemented with 0.05% cysteine

Animal feeding stuffs

hydrochloride -

4
The pH of the medium is 5.7, and will be adjusted to 6.5 or 5.4 according to standard practice.
5
MRS medium at pH 6.5 supplemented with 1 mL per 100 mL of a 1% TTC solution.

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 Microbiology of food
Enumeration of spores in food products
before preserving treatment by the colony-count technique
NF V 08-602: 05-2011
V 08-602

Initial suspension

Heat treatment
 10 min at 95-100°C

Aerobic bacteria spores Anaerobic bacteria spores 1

1 mL in poured plate, 1 mL in poured plate,
in a double layer in a double layer

Meat-liver agar for sulfite-

BCP glucose agar
reducing bacteria with 2 g/L

 48 ± 2 hrs  48 ± 2 hrs  48 ± 2 hrs

at 37 ± 1 °C at 55 ± 1°C at 37 ± 1°C 1  5 days
at 55 ± 1°C 1

Enumeration of Enumeration of Enumeration of

mesophilic aerobic thermophilic aerobic mesophilic anaerobic
colonies colonies colonies
Enumeration of
thermophilic anaerobic
colonies

1
For the detection of anaerobic bacteria spores, incubate in jars under anaerobic conditions.

Page 112 www.biokar-diagnostics.com

 Section Media and reagents Compliance

- Buffered Peptone Water (BPW) 2

Buffered Peptone Water 20.0 g/L 3
500 g vial - BK131HA
5 kg drum - BK131GC
Buffered Peptone Water 25.5 g/L 4
50 x 9 mL tubes - BM05608
5.1 Diluent 10 x 90 mL vials - BM05708 Total
10 x 225 mL vials - BM01008
3 x 3 L flexible bags - BM13108
2 x 5 L flexible bags - BM13208
500 g vial - BK018HA
5 kg drum - BK018GC

- BCP glucose agar

5.2 Culture medium BCP glucose agar Total
10 x 200 mL vials - BM16808

- Meat-liver glucose agar for sulfite-reducing bacteria with

0.2% yeast extract
5.3 Culture medium Meat-liver agar for sulfite-reducing bacteria + 2 g/L Total 5
yeast extract

Canned food

2
Except for special samples (fatty products, for example), refer to the section of standard NF EN ISO 6887 corresponding to
the product to be analysed.
3
Formula including 9.0 g/L of disodium phosphate dodecahydrate (Molecular mass = 358.14).
4
Formula including 3.56 g/L of anhydrous disodium phosphate (Molecular mass = 141.96).
5
There is an error in the composition, in the text of the standard: presence of meat-liver peptone instead of enzymatic digest
of casein in the standard.

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 Milk and milk products
Enumeration of presumptive Escherichia coli
Part 1: Most probable number technique using MUG
NF ISO 11866-1: 09-2006
IDF 170-1

Initial suspension
(and decimal dilutions)

1 mL in 10 mL in
9 mL of medium 1 10 mL of medium 1

Lauryl sulfate tryptose broth Lauryl sulfate tryptose broth

single strength double strength
(with Durham tube) (with Durham tube)

 24 ± 2 hrs at 30°C
an additional 24 ± 2 hrs
if no gas is produced

Confirmation 2

1– Addition of 0.5 mL of potassium hydroxide then UV examination.

2– For tubes displaying fluorescence, addition of 0.5 mL of Kovac’s reagent.
After 1 minute, observe the development of a red colour in the alcoholic layer
(characteristic of indole formation).

Calculation of the most probable number (MPN) of coliforms in g/mL of sample 3

1
Inoculate 3 tubes of single strength medium and 3 tubes of double strength medium for each dilution.
2
Identify gas-producing tubes as positive for presumptive coliforms. Identify tubes displaying fluorescence and formation of
indole as positive for presumptive Escherichia coli.
3
Based on the number of positive tubes of the chosen dilution, determine the MPN coefficient using a MPN table.

Page 114 www.biokar-diagnostics.com

 Section Media and reagents Compliance

5.2 Diluent Refer to standard NF EN ISO 6887-5, pages 101-102.

- Lauryl sulfate tryptose broth

(selective enrichment medium)
Laurylsulfate tryptose broth (single strength)
50 x 10 mL tubes, with Durham tubes - BM09708
Partial 4
Laurylsulfate tryptose broth (double strength)
5.3 Culture medium 50 x 10 mL tubes, with Durham tubes - BM09808
Laurylsulfate tryptose broth (base)
500 g vial - BK010HA
- 4-Methylumbelliferyl-β-D-glucuronide (MUG)
Total
MUG supplement 50 mg
10 vials - BS02408

- Indole reagent (Kovac’s reagent)

5.4/5.5 Confirmation - -
reagents - Sodium hydroxide solution c (NaOH) ≈ 0.5 mol/L
- -

Milk and milk products

4
0.1 g/L (single strength broth) and 0.2 g/L (double strength broth) of 4-methylumbelliferyl-β-D-glucuronide (MUG) and 1.0 g/
L or 2.0 g/L of tryptophan need to be added to the formula.

www.biokar-diagnostics.com Page 115

 Milk and milk products
Enumeration of presumptive Escherichia coli - Part 2: Colony-count
technique at 44°C using membranes
NF ISO 11866-2: 09-2006
IDF 170-2

Initial suspension
(and decimal dilutions)
1 mL on the surface
of the membrane 1

Mineral-modified glutamate agar

 15 minutes at room
temperature
 4 hrs at 37°C 2
Membrane transfer

Tryptone-bile agar

 18 to 24 hrs at 44°C 2

Indole detection reagent

Transfer the membrane to 2 mL of Kovac’s reagent

 5 minutes

Enumeration of presumptive Escherichia coli,

positive indole colonies (pink) 3

1
A cellulose acetate membrane is first placed on the surface of the agar medium.
2
Incubate the plates horizontally with the membrane upwards.
3
If permanent recording is necessary, place the membrane under a UV lamp for 30 minutes.

Page 116 www.biokar-diagnostics.com

 Section Media and reagents Compliance

5.2 Diluent Refer to standard NF EN ISO 6887-5, pages 101-102.

- Mineral-modified glutamate agar

Glutamate broth
500 g vial - BK186HA
Type A bacteriological agar
500 g vial - A1010HA
5 kg drum - A1010GC Partial 4
5.3 Culture medium
Type E bacteriological agar
500 g vial - A1012HA
5 kg drum - A1012GC
- Tryptone-bile agar
-
-

5.3.3 Confirmation - Indole reagent (Kovac’s reagent)

-
reagent -

Milk and milk products

4
There is an error in the composition, in the text of the standard: inversion of L(-)aspartic acid and L(+)arginine levels.
Presence of bromocresol (10 mg/L) in addition.

www.biokar-diagnostics.com Page 117

 Milk and milk products
Presumptive Lactobacillus acidophilus on a selective medium -
Colony-count technique at 37°C
ISO 20128: 05-2006
IDF 192

Initial suspension
(and decimal dilutions)

0.1 mL on surface 1

MRS/clindamycin/
ciprofloxacin agar

 72 ± 3 hrs at 37°C
under anaerobic
conditions

Enumeration of
Lactobacillus acidophilus

Section Media and reagents Compliance

5.2 Diluent Refer to standard NF EN ISO 6887-5, pages 101-102.

- MRS agar
MRS agar (base)
Total
10 x 200 mL vials - BM08908
500 g vial - BK089HA
5.3 Culture medium
- Clindamycin solution -
-
- Ciprofloxacin solution -
-

1
Inoculate each dilution in duplicate.

Page 118 www.biokar-diagnostics.com

 Milk and milk products
Enumeration of colony-forming units of yeasts and/or moulds -
Colony-count technique at 25°C
ISO 6611: 10-2004
FIL 94

Initial suspension
(and decimal dilutions)

1 mL in poured plate 1

Selective medium

 5 days at 25°C

Enumeration of yeasts
and/or moulds

Section Media and reagents Compliance

5.1 Diluent Refer to standard NF EN ISO 6887-5, pages 101-102.

Milk and milk products

- Yeast extract/dextrose/oxytetracycline/agar medium
Oxytetracycline glucose agar (OGA ready-to-use)
10 x 110 mL vials - BM02208
Oxytetracycline glucose agar (OGA base) Total
500 g vial - BK053HA
5.2 or 5.3 Selective Oxytetracycline 50 mg selective supplement
media 10 vials q.s. 550 mL - BS00808
- Yeast extract/dextrose/chloramphenicol agar
Chloramphenicol glucose agar
10 x 100 mL vials - BM02108 Total
10 x 200 mL vials - BM07908
500 g vial - BK007HA

1
Inoculate each dilution in duplicate.

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 Milk and milk products
Method for the enumeration of Pseudomonas spp.
XP ISO/TS 11059: 10-2009
V 04-025

Initial suspension
(and decimal dilutions)

0.1 mL on surface 1

PPA

 48 ± 2 hrs at 25 ± 1°C

Enumeration and confirmation 2

of Pseudomonas spp.

Nutrient agar

 24 to 48 hrs at 25 ± 1°C

Glucose agar oxidase test 3

 24 ± 3 hrs at 25 ± 1°C

Glucose fermentation 3

1
Inoculate each dilution in duplicate.
2
Select 5 characteristic colonies and carry out the confirmation tests.
3
The presence of Pseudomonas is confirmed by a positive oxidase test and a negative glucose fermentation test.

Page 120 www.biokar-diagnostics.com

 Section Media and reagents Compliance

5.2 Diluents Refer to standard NF EN ISO 6887-5, pages 101-102.

- Penicillin and pimaricin agar (PPA)

5.3 Selective
PPA medium Total
medium
20 plates Ø 90 mm - BM15608

- Nutrient agar
2 % nutrient agar
50 x 18 mL tubes - BM11808 Total
5.4/5.5/5.6 500 g vial - BK185HA
Confirmation media - Glucose agar
and reagent Glucose agar Total
50 x 10 mL tubes - BM09908
- Reagent for the detection of oxidase
-
-

Milk and milk products

www.biokar-diagnostics.com Page 121

 RHAPSODY Agar®
Method for the enumeration of Pseudomonas spp.
in human food products and environmental samples

BKR 23/09-05/15 A (meat products)

BKR 23/09-05/15 B (milk products)
Alternative food analysis method
www.afnor-validation.org

Initial suspension
(and decimal dilutions)

0.1 mL on surface
Spiral plating

RHAPSODY Agar®

 48 ± 2 hrs at 30 ± 1°C

Enumeration of Pseudomonas spp.

WITHOUT confirmation

Diluents Refer to standard NF EN ISO 6887-5, pages 101-102.

RHAPSODY Agar®
Selective medium
BM16708 - 20 plates Ø 90 mm

Page 122 www.biokar-diagnostics.com

 Yogurt
Enumeration of characteristic microorganisms
ISO 7889: 02-2003
IDF 117

Initial suspension
(and decimal dilutions)

1 mL in poured plate 1 1 mL in poured plate 1

MRS medium M17 agar

 72 hrs at 37°C
under anaerobic  48 hrs at 37°C
conditions

Enumeration of Lactobacillus Enumeration of Streptococcus

delbrueckii subsp. bulgaricus thermophilus

Section Media and reagents Compliance

Milk and milk products

5.2/5.3/5.4 Diluents Refer to standard NF EN ISO 6887-5, pages 101-102.

- Acidified MRS medium (pH 5.4) 2

Partial 3
MRS agar
500 g vial - BK089HA
5.5 Selective media
- M17 medium
M17 agar
Total 4
500 g vial - BK088HA

1
Inoculate each dilution in duplicate.
2
According to DE MAN J.C., ROGOSA M. and SHARPE M.E. (1960) A medium for the cultivation of Lactobacilli.
Journal of Applied Bacteriology. 23, 130-135.
3
Presence of polypeptone (10.0 g/L) instead of casein tryptic hydrolysate (10.0 g/L) as recommended.
4
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

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 Yogurt
Identification of characteristic microorganisms (Lactobacillus
delbrueckii subsp. bulgaricus and Streptococcus thermophilus)
ISO 9232: 02-2003
IDF 146

Identification of Identification of
L. delbrueckii subsp. Bulgaricus 1 S. thermophilus 1

Isolated colony Isolated colony

MRS broth + skim milk M 17 broth

 24 hrs at 37°C  24 hrs at 37°C

▪ Analysis of morphology ▪ Analysis of morphology

▪ Catalase reaction ▪ Catalase reaction
▪ Growth at 15°C and 45°C ▪ Growth at 10°C and 45°C
▪ CO2 production ▪ Action in litmus milk
▪ Fermentation of sugars ▪ Growth in the presence of
▪ Determination of sodium chloride
lactic acid enantiomers ▪ Utilization of sugars

1
Identification of selected colonies using the plates obtained according to standard ISO 7889/IDF 117 (see page 17).

Page 124 www.biokar-diagnostics.com

 Section Media and reagents Compliance

Identification of L. delbrueckii subsp. bulgaricus

5.1.1 - Autoclaved skim milk -

- MRS broth
5.1.2 MRS broth 2 Partial 3
500 g vial - BK070HA

- Base medium for fermentation tests

5.1.3 MRS broth 4 -
-

- Culture medium for CO2 production

5.1.4 MRS broth 5 -
-

- Blank agar
Type A bacteriological agar
500 g vial - A1010HA
5.1.5 5 kg drum - A1010GC Total
Type E bacteriological agar
500 g vial - A1012HA
5 kg drum - A1012GC

Identification of S. thermophilus

5.1.6 - Litmus milk -

- M17 broth

Milk and milk products

5.1.7 M17 broth Total 6
500 g vial - BK012HA

- Growth medium in the presence of 6.5% NaCl 7

5.1.8 -
-

2
According to DE MAN J.C., ROGOSA M. and SHARPE M.E. (1960) A medium for the cultivation of Lactobacilli. Journal of
Applied Bacteriology. 23, 130-135.
3
Presence of polypeptone (10.0 g/L) instead of casein tryptic hydrolysate (10.0 g/L) as recommended.
4
Without meat extract and glucose, final pH adjusted to 6.9-7.0.
5
As for the base medium in 5.1.3 spiked with 5% glucose.
6
“Tryptone” is a peptone obtained by pancreatic digestion of casein.
7
M17 broth without β-glycerophosphate and containing 6.5% NaCl.

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 Butter, fermented milks and fresh cheese
Enumeration of contaminating microorganisms Colony-count
technique at 30°C
ISO 13559: 11-2002
IDF 153

Initial suspension
(and decimal dilutions)

0.1 mL on surface 1

Culture medium

 72 ± 2 hrs at 30°C

Enumeration of contaminating
microorganisms

Section Media and reagents Compliance

5.3/5.4/5.5 Diluents Refer to standard NF EN ISO 6887-5, pages 101-102.

- Culture medium
Agar for counting contaminating microorganisms in
5.6 Selective
milk products (SFA) Total 2
medium
10 x 100 mL vials - BM12208
500 g vial - BK126HA

1
Inoculate each dilution in duplicate.
2
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

Page 126 www.biokar-diagnostics.com

 Milk
Enumeration of microorganisms Calibrated plate-loop technique in
Petri dishes at 30°C
ISO 8553: 05-2004
FIL 131

Suspension 1

Inoculate in
poured plate

Milk plate-count agar

 72 ± 2 hrs at 30°

Enumeration

Milk and milk products

Section Media and reagents Compliance

5.2 Diluent Refer to standard NF EN ISO 6887-5, pages 101-102.

- Milk plate-count agar

5.3 Selective Plate-count agar with skim milk
Total 2
medium 10 x 200 mL vials - BM08608
500 g vial - BK161HA

1
Immerse a calibrated plate-loop (10 µL) in the prepared test sample, then mix directly in the Petri dish with 1 mL of
diluent. Add 15 mL of medium, mix and allow to solidify.
2
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

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 Milk
Enumeration of colony-forming units of psychrotrophic
micro-organisms - Colony-count technique at 6.5°C
ISO 6730: 09-2005
IDF 101

Initial suspension
(and decimal dilutions)

1 mL in poured plate 1

Culture medium

 10 days at 6.5°C

Enumeration of psychrotrophic
microorganisms

Section Media and reagents Compliance

5.2/5.3/5.4 Diluents Refer to standard NF EN ISO 6887-5, pages 101-102.

- Culture medium
5.5 Selective Plate count agar with skimmed milk
Total 2
medium 10 x 200 mL vials - BM08608
500 g vial - BK161HA

1
Inoculate each dilution in duplicate.
2
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

Page 128 www.biokar-diagnostics.com

 Milk
Estimation of psychrotrophic microorganisms
Colony-count technique at 21°C (rapid method)
ISO 8552: 05-2004
FIL 132

Initial suspension
(and decimal dilutions)

1 mL in poured plate 1

Milk plate-count agar

 25 ± 1 hrs at 21°C

Enumeration

Section Media and reagents Compliance Milk and milk products

5.2 Diluent Refer to standard NF EN ISO 6887-5, pages 101-102.

- Milk plate-count agar

5.3 Selective Plate count agar with skimmed milk
Total 2
medium 10 x 200 mL vials - BM08608
500 g vial - BK161HA

1
Inoculate each dilution in duplicate.
2
“Tryptone” is a peptone obtained by pancreatic digestion of casein.

www.biokar-diagnostics.com Page 129

 Meat and meat products
Enumeration of presumptive Pseudomonas spp.
NF EN ISO 13720: 11-2010
V 08-504

Initial suspension
(and decimal dilutions)

0.1 mL on surface 1

CFC agar

 44 ± 4 hrs
at 25 ± 1°C

Enumeration and confirmation of

presumptive Pseudomonas spp. 2

Oxidase test (positive reaction)

Section Media and reagents Compliance

5.2 Diluent Refer to standard NF EN ISO 6887-2, page 97.

- Cephalotin-sodium fusidate-cetrimide (CFC) agar
CFC agar (base + supplement)
Meat and meat products

500 g vial - BK118HA

5.3 Agar medium 10 vials q.s. 500 mL - BS02208 (selective suppl.) Total 3, 4
CFC agar (ready-to-melt)
10 x 200 mL vials - BM09608

5.4 Confirmation - Reagent for the detection of oxidase

- -
reagent

1
Inoculate one plate per dilution, and carry out at least two successive dilutions.
2
Confirm 5 colonies chosen at random, representing all types of colonies.
3
“Tryptone” is a peptone obtained by pancreatic digestion of casein.
4
Formula containing 1 g/L of fucidin instead of 1 g/L of sodium fusidate as described.

Page 130 www.biokar-diagnostics.com

 RHAPSODY Agar®
Method for the enumeration of Pseudomonas spp. in human food products and environmental samples

BKR 23/09-05/15 A (meat products)

BKR 23/09-05/15 B (milk products)
Alternative food analysis method
www.afnor-validation.org

Initial suspension
(and decimal dilutions)

0.1 mL on surface
Spiral plating

RHAPSODY Agar®

 48 ± 2 hrs
at 30 ± 1°C

Enumeration of Pseudomonas spp.

WITHOUT confirmation

Meat and meat products

Diluents Refer to standard NF EN ISO 6887-2, page 97.

RHAPSODY Agar®
Selective medium
BM16708 - 20 plates Ø 90 mm

www.biokar-diagnostics.com Page 131

 APPENDIX A: Other standards relating to the microbiology of foods

Determination of water activity NF ISO 21807 V 08-028 01-2005

General requirements and guidance NF EN ISO 7218 V 08-002 10-2007

Preparation of test samples, initial suspension and decimal dilutions for

microbiological examination Part 6: Specific rules for the preparation of NF EN ISO 6887-6 V 08-010-6 05-2013
samples taken at the primary production stage

Microbiology of food, animal feed and water - Preparation, production,

NF EN ISO 11133/ 07-2014
storage and performance testing of culture media V 08-104
A1 03-2018
Amendment 1

Method validation - Part 1: Vocabulary NF EN ISO 16140-1 - 09-2016

Method validation - Part 2: Protocol for the validation of alternative

NF EN ISO 16140-2 - 09-2016
(proprietary) methods against a reference method

Technical requirements and guidance on establishment or revision of a

NF EN ISO 17468 V 08-751 09-2016
standardized reference method

Surface sampling techniques, using contact plates and swabs NF ISO 18593 V 08-035 07-2018

Page 132 www.biokar-diagnostics.com

 APPENDIX B: List of products

Product Packaging Standards

Alkaline saline peptone water (ASPW) 500 g vial - BK219HA NF EN ISO 21872-1: 2017 (pages 88-89)
BAIRD-PARKER (base) agar 500 g vial - BK055HA NF EN ISO 6888-1: 1999 (pages 74-75)
5 kg drum - BK055GC NF EN ISO 6888-2: 1999 (pages 76-77)
NF EN ISO 6888-3: 2003 (pages 80-81)
NF V 08-057-1: 2004 (pages 82-83)
BAIRD-PARKER agar with egg yolk and tellurite 20 plates Ø 90 mm - BM01808 NF EN ISO 6888-1: 1999 (pages 74-75)
120 plates Ø 90 mm - BM09108 NF EN ISO 6888-3: 2003 (pages 80-81)
NF V 08-057-1: 2004 (pages 82-83)
KIT 6 x 90 mL vials + 6 supplements
BAIRD-PARKER RPF agar NF EN ISO 6888-2: 1999 (pages 76-77)
q.s. 100 mL - BT00508
KIT 6 x 190 mL vials + 6 supplements
NF EN ISO 6888-3: 2003 (pages 80-81)
q.s. 200 mL - BT01008
20 plates Ø 90 mm - BM06708 NF V 08-057-1: 2004 (pages 82-83)
20 plates Ø 55 mm - BM15908 NF EN ISO 6888-1/A2: 2018 (pages 74-75)
BEA agar 10 x 100 mL vials - BM10408 NF EN 15788: 2009 (page 107)
500 g vial - BK158HA
B. cereus selective agar (base as per MOSSEL) 500 g vial (base) - BK116HA NF EN ISO 7932: 2005 (pages 10-11)
20 plates Ø 90 mm - BM03808 NF EN ISO 21871: 2006 (pages 8-9)
120 plates Ø 90 mm - BM19908
BCP glucose agar 10 x 200 mL vials - BM16808 NF V 08-602: 2011 (pages 112-113)
Brain-heart infusion broth 500 g vial - BK015HA NF EN ISO 6888-1: 1999 (pages 74-75)
NF EN ISO 6888-3: 2003 (pages 80-81)
NF V 08-057-1: 2004 (pages 82-83)
Brilliant green bile lactose broth (BGBLB) 500 g vial - BK002HA NF ISO 4832 /FIL 73B: 2006 (pages 20-21)
50 x 10 mL tubes with Durham tubes
NF ISO 4831 /FIL 73B: 2006 (pages 22-23)
- BM01108
Buffered Peptone Water (20 g/L) 500 g vial - BK131HA NF EN ISO 21528-1: 2017 (pages 28-29)
5 kg drum - BK131GC NF EN ISO 11290-2 : 2017 (pages 58-59)
Buffered Peptone Water (25.5 g/L) 500 g vial - BK018HA NF EN ISO 6579-1: 2017 (pages 62-67)
5 kg drum - BK018GC NF EN ISO 6887-1: 2017 (page 96)
10 x 225 mL vials - BM01008 NF EN ISO 6887-5: 2010 (pages 101-102)
50 x 9 mL tubes - BM05608 BKR 23/05-12/07 (pages 60-61)
10 x 90 mL vials - BM05708 NF V 08-602: 2011 (pages 112-113)
3 x 3 L flexible bags - BM13108 NF EN ISO 22964: 2017 (pages 26-27)
2 x 5 L flexible bags - BM13208
CFC agar 10 x 200 mL vials - BM09608 NF EN ISO 13720: 2010 (page 130)
500 g vial (base) - BK118HA
CFC supplement 10 vials q.s. 500 mL - BS02208
Chloramphenicol glucose (YGC) agar 10 x 100 mL vials - BM02108 ISO 6611/FIL 94: 2004 (page 119)
10 x 200 mL vials - BM07908 NF V 08-059: 2002 (pages 50-51)
500 g vial - BK007HA
Chromogenic coliform isolation (CCI) agar 20 plates Ø 90 mm - BM15408 NF EN ISO 22964: 2017 (pages 26-27)
Columbia (base) agar 500 g vial - BK019HA NF EN ISO 10272-1: 2017 (pages 14-15)
NF EN ISO 10272-2: 2017 (pages 16-17)

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Product Packaging Standards
COMPASS® Bacillus cereus Agar BM12608 - 20 plates Ø 90 mm BKR 23/06-02/10 (pages 12-13)
COMPASS® Bacillus cereus Agar (base) BM13008 - 10 x 100 mL vials
Selective supplement for COMPASS® Bacillus cereus agar BS06908 - 10 vials q.s. 100 mL
COMPASS® Listeria Agar 20 plates Ø 90 mm - BM12308 NF EN ISO 11290-1: 2017 (pages 53-55)
120 plates Ø 90 mm - BM12408 NF EN ISO 11290-2: 2017 (pages 58-59)
Kit of 6 x 200 mL vials of base + 6
vials of selective suppl. + 6 vials BKR 23/02-11-02 (pages 56-57)
of enrichment suppl. - BT00808
BKR 23/05-12/07 (pages 60-61)
CONFIRM' L. mono broth 18 vials - BM16208 BKR 23/02-11-02 and BKR 23/05-12/07
CONFIRM' L. mono agar 10 plates Ø 90 mm - BM13908 BKR 23/02-11-02 and BKR 23/05-12/07
Latex agglutination kit (50 tests) -
CONFIRM' Salmonella BKR 23/07-10/11 (pages 70-72)
BT01108
Cronobacter screening broth (CSB) 50 x 10 mL tubes - BM15508 NF EN ISO 22964: 2017 (pages 26-27)
Sorbitol MacCONKEY agar (CT-SMAC base) 500 g vial - BK147HA NF EN ISO 16654: 2001 (pages 42-43)
Cefixime-tellurite supplement for CT-SMAC agar 10 vials q.s. 500 mL - BS03708
Dichloran-Glycerol (DG 18) agar 500 g vial - BK170HA NF V 08-036: 2003 (page 45)
10 x 100 mL vials - BM10908 NF ISO 21527-2: 2008 (pages 48-49)
Dichloran-Rose Bengal Chloramphenicol (DRBC) agar 500 g vial - BK198HA NF ISO 21527-1: 2008 (pages 46-47)
10 x 200 mL vials - BM14208 NF ISO 21527-2: 2008 (pages 48-49)
Dipotassium hydrogenphosphate solution 20 g/L 2 x 5 L flexible bags - BM19308 NF EN ISO 6887-5: 2010 (pages 101-102)
D-cycloserine 200 mg selective supplement 10 vials q.s. 500 mL - BS00608 NF EN ISO 7937: 2005 (pages 18-19)
NF V 08-061: 2009 (pages 86-87)
D-cycloserine liquid supplement 10 x 90 mL vials - BS09208 NF EN ISO 7937: 2005 (pages 18-19)
1 x 50 mL vial - BS09408 NF V 08-061: 2009 (pages 86-87)
EASY Staph® Agar 20 plates Ø 90 mm - BM18708 BKR 23/10-12/15 (pages 78-79)
120 plates Ø 90 mm - BM19008
Kit of 6 x 190 mL vials of base + 6
vials of suppl. q.s. 200 mL
BT01208
Kit of 6 x 90 mL vials of base + 6
vials of suppl. q.s. 100 mL
BT01308
500 g vial (base)- BK216HA
8 vials of suppl. - BS09008
EC broth 500 g vial - BK162HA NF ISO 7251: 2005 (pages 40-41)
Egg yolk emulsion with polymyxin B 10 x 50 mL vials - BS05508 NF EN ISO 7932: 2005 (pages 10-11)
Egg yolk emulsion with potassium tellurite 10 x 50 mL vials - BS06008 NF EN ISO 6888-1: 1999 (pages 74-75)
NF EN ISO 6888-3: 2003 (pages 80-81)
NF V 08-057-1: 2003 (pages 82-83)
Egg yolk emulsion (sterile) 10 x 50 mL vials - BS06608 NF EN ISO 21871: 2006 (pages 8-9)
Enterobacteria enrichment broth (EE as per Mossel) 500 g vial - BK127HA NF EN ISO 21528-1: 2017 (pages 28-29)
FRASER (base) broth 500 g vial - BK133HA NF EN ISO 11290-1: 2017 (pages 53-55)
5 kg drum - BK133GC NF EN ISO 11290-2: 2017 (pages 58-59)
BKR 23/02-11-02 and BKR 23/05-12/07
FRASER broth (ready-to-use) 50 x 10 mL tubes - BM01308 NF EN ISO 11290-1: 2017 (pages 53-55)
Half-FRASER (base) broth 500 g vial - BK173HA NF EN ISO 11290-1: 2017 (pages 53-55)
5 kg drum - BK173GC BKR 23/02-11/02 (pages 56-57)
BKR 23/05-12/07 (pages 60-61)
Selective supplement for half-FRASER broth 10 vials q.s. 500 mL - BS03008 NF EN ISO 11290-1: 2017 (pages 53-55)
8 vials q.s. 2.25 L - BS03208 BKR 23/02-11/02 (pages 56-57)

Page 134 www.biokar-diagnostics.com

Products Packaging Standards
Half-FRASER broth (ready-to-use) 10 x 225 mL vials - BM01608 BKR 23/05-12/07 (pages 60-61)
3 x 3 L flexible bags - BM13308
2 x 5 L flexible bags - BM13408
Gentamicin 25 mg selective supplement 10 vials - BS00908 NF V 08-059: 2002 (pages 50-51)
Glucose agar 50 x 10 mL tubes - BM09908 NF EN ISO 21528-1: 2017 (pages 28-29)
NF EN ISO 21528-2: 2017 (pages 30-31)
XP ISO/TS 11059: 2009 (pages (120-121)
Glucose OF medium 50 x 10 mL tubes NF EN ISO 21528-1: 2017 (pages 28-29)
NF EN ISO 21528-2: 2017 (pages 30-31)
Glutamate broth 500 g vial - BK186HA NF EN ISO 16649-3: 2015 (pages 38-39)
NF ISO 11866-2/IDF 170-2: 2006 (pages 116-
117)
HEKTOEN agar 500 g vial - BK067HA NF EN ISO 21567: 2005 (pages 72-73)
IRIS Salmonella® Agar 20 plates Ø 90 mm - BM16008 BKR 23/07-10/11 (pages 70-71)
120 plates Ø 90 mm - BM16108
IRIS Salmonella® supplement 120 tablets q.s. 225 mL - BS07708 BKR 23/07-10/11 (pages 70-71)
120 tablets q.s. 90 mL - BS09308
10 x 50 mL vials - BS07808
KLIGLER agar 500 g vial - BK034HA NF EN ISO 10273: 2017 (pages 90-91)
Lactose sulfite broth 500 g vial - BK140HA NF EN ISO 7937: 2005 (pages 18-19)
7 x 9 mL tubes - BM19208
Laurylsulfate Tryptose broth 500 g vial - BK010HA NF ISO 4831 /FIL 73B: 2006 (pages 22-23)
50 x 10 mL tubes, with Durham tubes
NF ISO 7251: 2005 (pages 40-41)
(single strength) - BM09708
50 x 10 mL tubes, with Durham tubes ISO 11866-1/IDF 170-1: 2006 (pages 114-
(double strength) - BM09808 115)
Lyophilized coagulase rabbit plasma 10 vials q.s. 20 reactions - BR00208 NF EN ISO 6888-1: 1999 (pages 74-75)
NF EN ISO 6888-3: 2003 (pages 80-81)
NF V 08-057-1: 2004 (pages 82-83)
M 17 broth 500 g vial - BK012HA ISO 9232/IDF 146: 2003 (pages 124-125)
M 17 agar 500 g vial - BK088HA ISO 7889/IDF 117: 2003 (page 123)
MacCONKEY agar 500 g vial - BK050HA NF EN ISO 21567: 2005 (pages 72-73)
Meat-liver agar for sulfite-reducing bacteria + 2 g/L
10 x 200 mL vials - BM16908 NF V 08-602: 2011 (pages 112-113)
yeast extract
Modified Giolitti and Cantoni broth 500 g vial (base) - BK159HA NF EN ISO 6888-3: 2003 (pages 80-81)
50 x 10 mL tubes (single strength) -
BM11008
50 x 10 mL tubes (double strength) -
BM11108
Modified tryptone soy broth (mTSB base) 500 g vial - BK191HA NF EN ISO 16654: 2001 (pages 42-43)
MRS broth 500 g vial - BK070HA ISO 9232/IDF 146: 2003 (pages 124-125)
MRS agar 500 g vial - BK089HA NF ISO 15214: 1998 (page 44)
10 x 200 mL vials - BM08908 NF EN 15785: 2009 (pages 104-105)
NF EN 15787: 2009 (pages 108-109)
NF EN 15786: 2009 (pages 110-111)
ISO 20128/IDF 192: 2006 (page 118)
ISO 7889/IDF 117: 2003 (page 123)
MSRV agar (ISO 6579) 500 g vial - BK134HA NF EN ISO 6579-1: 2017 (pages 62-67)
10 x 200 mL vials - BM12708

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Products Packaging Standards
MUELLER-HINTON agar 500 g vial - BK048HA NF EN ISO 10272-1: 2017 (pages 14-15)
NF EN ISO 10272-2: 2017 (pages 16-17)
MULLER-KAUFFMANN tetrathionate-novobiocin broth
500 g vial - BK169HA NF EN ISO 6579-1: 2017 (pages 62-67)
(MKTTn base, without iodine, without novobiocin)
MULLER-KAUFFMANN tetrathionate-novobiocin broth
500 g vial - BK208HA NF EN ISO 6579-1: 2017 (pages 62-67)
(MKTTn base, without iodine)
MULLER-KAUFFMANN tetrathionate-novobiocin broth
50 x 10 mL tubes - BM07808 NF EN ISO 6579-1: 2017 (pages 62-67)
(MKTTn ready-to-use)
ISO 11866-1/IDF 170-1: 2006 (pages 114-
MUG supplement 10 x 50 mg vials - BS02408
115)
Novobiocin selective supplement 10 vials (10 mg) - BS03308 NF EN ISO 16654: 2001 (pages 42-43)
8 vials (40 mg) - BS05608 NF EN ISO 6579-1: 2017 (pages 62-67)
2% nutrient agar 500 g vial - BK185HA NF EN ISO 21528-1: 2017 (pages 28-29)
50 x 18 mL tubes - BM11808 NF EN ISO 21528-2: 2017 (pages 30-31)
NF EN ISO 16654: 2001 (pages 42-43)
NF EN ISO 6579-1: 2017 (pages 62-67)
NF EN ISO 21567: 2005 (pages 72-73)
NF EN ISO 10273: 2017 (pages 90-91)
XP ISO/TS 11059: 2009 (pages (120-121)
OXFORD (base) agar 500 g vial - BK110HA NF EN ISO 11290-1: 2017 (pages 53-55)
Selective supplement for OXFORD agar (CCCFA) 10 vials q.s. 500 mL - BS00308
Oxytetracycline glucose agar (OGA base) 500 g vial - BK053HA NF V 08-059: 2002 (pages 50-51)
10 x 110 mL vials - BM02208 ISO 6611/FIL 94: 2004 (page 119)
Oxytetracycline selective supplement 50 mg 10 vials q.s. 550 mL - BS00808
PALCAM agar 500 g vial (base) - BK145HA BKR 23/02-11/02 (pages 56-57)
20 plates Ø 90 mm - BM02008
Selective supplement for PALCAM agar 10 vials q.s. 500 mL - BS00408
8 vials q.s. 2.5 L - BS04908
0.1% peptone water 2 x 5 L flexible bags - BM16408 NF ISO 21527-1: 2008 (pages 46-47)
NF ISO 21527-2: 2008 (pages 48-49)
NF EN ISO 6887-3: 2017 (page 98)
NF EN ISO 6887-5: 2010 (pages 101-102)
Peptone water, indole free 500 g vial - BK084HA NF ISO 7251: 2005 (pages 40-41)
Plate count agar (PCA) 10 x 100 mL vials - BM01508 NF EN ISO 4833-1: 2013 (page 92)
10 x 200 mL vials - BM03308 NF EN ISO 4833-2: 2013 (page 93)
500 g vial - BK144HA XP V 08-034: 2010 (page 94)
5 kg drum - BK144GC NF ISO 17410: 2001 (page 95)
500 g vial - BK161HA
Plate count agar with skim milk (PCA) 10 x 200 mL vials - BM08608 NF EN ISO 4833-1: 2013 (page 92)
NF EN ISO 4833-2: 2013 (page 93)
XP V 08-034: 2010 (page 94)
NF ISO 17410: 2001 (page 95)
ISO 8553/FIL 131: 2004 (page 127)
ISO 6730/IDF 101: 2005 (page 128)
ISO 8552/FIL 132: 2004 (page 129)
PPA 20 plates Ø 90 mm - BM15908 XP ISO/TS 11059: 2009 (pages (120-121)
Polymyxin B selective supplement 10 vials - BS00708 NF EN ISO 7932: 2005 (pages 10-11)
NF EN ISO 21871: 2006 (pages 8-9)

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Products Packaging Standards
Rabbit plasma fibrinogen supplement 8 vials q.s. 100 mL - BS03408 NF EN ISO 6888-2: 1999 (pages 76-77)
1 vial q.s. 500 mL - BS03808 NF EN ISO 6888-3: 2003 (pages 80-81)
NF V 08-057-1: 2004 (pages 82-83)
RAPPAPORT-VASSILIADIS broth with soya (RVS) 500 g vial - BK148HA NF EN ISO 6579-1: 2017 (pages 62-67)
50 x 10 mL tubes - BM07408
RHAPSODY® agar 20 plates Ø 90 mm - BM16708 BKR 23/09-05/15 B (page 122)
BKR 23/09-05/15 A (page 131)
RINGER’s solution (1/4 strenght) 100 tablets - BR00108 NF EN ISO 6887-5: 2010 (pages 101-102)
Salmonella Enrichment 500 g vial - BK194HA BKR 23/04-12/07 (pages 68-69)
5 kg drum - BK194GC BKR 23/07-10/11 (pages 70-71)
10 x 225 mL vials - BM13608
3 x 3 L flexible bags - BM13708
2 x 5 L flexible bags - BM14408
Salmonella Enrichment + Tween® 80 3 x 3 L flexible bags - BM16308
2 x 5 L flexible bags - BM19808
Salmonella Enrichment double strength buffered 2 x 5 L flexible bags - BM20008 BKR 23/04-12/07 (pages 68-69)
10 x 225 mL vials - BM20108 BKR 23/07-10/11 (pages 70-71)
500 g vial - BK225HA NF EN ISO 6887-1: 2017 (page 96)
5 kg drum - BK225GC
Selenite-cystine broth 500 g vial - BK009HA NF EN ISO 6579-1 (pages 62-67)
SESAME Salmonella Detection 20 plates Ø 90 mm - BM14108 BKR 23/04-12/07 (pages 68-69)
120 plates Ø 90 mm - BM15008
10 x 200 mL vials - BM13808
SFA - Agar for counting contaminating microorganisms in 10 x 100 mL vials - BM12208 ISO 13559/IDF 153: 2002 (page 126)
milk products
500 g vial - BK126HA
Sulfamethazine 25 mg selective supplement 10 vials q.s. 500 mL - BS02808 NF EN ISO 6888-1: 1999 (pages 74-75)
NF EN ISO 6888-3: 2003 (pages 80-81)
NF V 08-057-1: 2004 (pages 82-83)
Sterile 5% ammonium ferric citrate solution 10 x 90 mL vials - BS05908 NF EN ISO 11290-1: 2017 (pages 53-55)
7 x 10 mL tubes - BS06208 NF EN ISO 11290-2: 2017 (pages 58-59)
SYMPHONY Agar 10 x 200 mL vials Alternative method (page 52)
TBX agar 10 x 100 mL vials - BM06908 NF ISO 16649-1: 2018 (pages 34-35)
10 x 200 mL vials - BM17108 NF ISO 16649-2: 2001 (pages 36-37)
500 g vial - BK146HA NF EN ISO 16649-3: 2015 (pages 38-39)
100-g vial - BK146HM
TCBS agar 500 g vial - BK040HA NF EN ISO 21872-1: 2017 (pages 88-89)
Thioglycollate broth with resazurin 500 g vial - BK017HA NF EN ISO 7937: 2005 (pages 18-19)
5 kg drum - BK017GC
50 x 10 mL tubes - BM08208
Trypto-casein soy broth (TSB) 500 g vial - BK046HA NF EN ISO 21871: 2006 (pages 8-9)
5 kg drum - BK046GC NF EN ISO 10273 : 2017 (pages 90-91)
10 x 100 mL vials - BM00908
10 x 90 mL vials - BM17908
50 x 10 mL tubes - BM03008
2 x 5 L flexible bags - BM16608
Trypto-casein soy agar (TSA) 500 g vial - BK047HA NF EN ISO 10273: 2017 (pages 90-91)
20 plates Ø 90 mm - BM05008 NF EN 15784: 2009 (page 106)
10 x 100 mL vials - BM01708 NF EN ISO 22964: 2017 (pages 26-27)
10 x 200 mL vials - BM04908

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Products Packaging Standards
Tryptophan broth 500 g vial - BK163HA NF EN ISO 16654: 2001 (pages 42-43)
50 x 3 mL tubes - BM07608 NF EN ISO 6579-1: 2017 (pages 62-67)
NF EN ISO 21872-1: 2017 (pages 88-89)
Tryptone-salt broth 500 g vial - BK014HA NF EN ISO 15213: 2003 (pages 84-85)
50 x 9 mL tubes - BM00808 NF EN ISO 6887-1: 2017 (page 96)
10 x 90 mL vials - BM11408 NF EN ISO 6887-4: 2017 (page 99-100)
3 x 3 L flexible bags - BM13508 NF EN ISO 6887-5: 2010 (pages 101-102)
Tryptone soya agar (blood agar base) 500 g vial - BK028HA NF EN ISO 7932: 2005 (pages 10-11)
NF EN ISO 21871: 2006 (pages 8-9)
NF EN ISO 11290-1: 2017 (pages 53-55)
NF EN ISO 11290-2: 2017 (pages 58-59)
NF EN 15785: 2009 (pages 104-105)
NF EN 15784: 2009 (page 106)
NF EN 15788: 2009 (page 107)
NF EN 15787: 2009 (pages 108-109)
NF EN 15786: 2009 (pages 110-111)
TSC (base) agar 500 g vial - BK031HA NF EN ISO 7937: 2005 (pages 18-19)
50 x 20 mL tubes - BM03908 NF EN ISO 15213: 2003 (pages 84-85)
10 x 200 mL vials - BM07708 NF V 08-061: 2009 (pages 86-87)
TSI agar 500 g vial - BK221HA NF EN ISO 6579-1: 2017 (pages 62-67)
NF EN ISO 21567: 2005 (pages 72-73)
TSYEA medium 500 g vial - BK224HA NF EN ISO 11290-1: 2017 (pages 53-55)
50 x 18 mL tubes - BM10808 NF EN ISO 11290-2: 2017 (pages 58-59)
TTC supplement 10 x 12.5 mg vials - BS02608 NF EN 15785: 2009 (pages 104-105)
10 x 50 mg vials - BS02708 NF EN 15787: 2009 (pages 108-109)
NF EN 15786: 2009 (pages 110-111)
Type A bacteriological agar 500 g vial - A1010HA NF EN ISO 6888-3: 2003 (pages 80-81)
5 kg drum - A1010GC NF EN ISO 4833-1: 2013 (page 92)
XP V 08-034: 2010 (page 94)
NF ISO 11866-2/IDF 170-2: 2006 (pages
116-117)
ISO 9232/IDF 146: 2003 (pages 124-125)
Type E bacteriological agar 500 g vial - A1012HA NF EN ISO 6888-3: 2003 (pages 80-81)
5 kg drum - A1012GC NF EN ISO 4833-1: 2013 (page 92)
XP V 08-034: 2010 (page 94)
NF ISO 11866-2/IDF 170-2: 2006 (pages
116-117)
ISO 9232/IDF 146: 2003 (pages 124-125)
VRBG agar 500 g vial - BK011HA NF EN ISO 21528-1: 2017 (pages 28-29)
5 kg drum - BK011GC NF EN ISO 21528-2: 2017 (pages 30-31)
10 x 200 mL vials - BM07508 NF V 08-054: 2009 (pages 32-33)
VRBL broth 500 g vial - BK152HA NF ISO 4832/FIL73B: 2006 (pages 20-21)
5 kg drum - BK152GC NF V 08-050: 2009 (page 24)
10 x 100 mL vials - BM03408 NF V 08-060: 2009 (page 25)
10 x 200 mL vials - BM03508
Wilson-Blair agar 500 g vial - BK223HA NF EN ISO 6579-1: 2017 (pages 62-67)
XLD agar (ISO 6579) 500 g vial - BK168HA NF EN ISO 6579-1: 2017 (pages 62-67)
20 plates Ø 90 mm - BM08708 NF EN ISO 21567: 2005 (pages 72-73)

Page 138 www.biokar-diagnostics.com

 BIOKAR Diagnostics - Support and services

Online information: www.biokar-diagnostics.fr

Direct access to our product technical data sheets (TDS), material safety data sheets (MSDS), and quality control
certificates (QCC) on our website.

BIOKAR Diagnostics contact

 +33 (0)1 48 10 19 40
 +33 (0)3 44 14 33 34

Europe, Africa, North America Asia, South America

Athanassios GIANNAKOPOULOS Anthony FELIN
@ [email protected] @ [email protected]

Distributors information
List of our distributors on our website section «contact»
Local contact :

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 BIOKAR Diagnostics
Rue des Quarante Mines
ZAC de Ther-Allonne - B.P. 10245
60002 Beauvais Cedex - France
www.biokar-diagnostics.fr