AS 5013.13:2018

Header

About this publication

Preface

Foreword

Introduction

1 Scope

2 Normative references

3 Terms and definitions

4 Abbreviated terms

5 Principle

5.1 Non-selective pre-enrichment in BPW

5.2 Enrichment in a selective medium (CSB)

5.3 Plating out and identification on chromogenic agar (CCI agar)

5.4 Confirmation

6 Culture media and reagents

7 Equipment and consumables

7.1

7.2

7.3

7.4

7.5

7.6

7.7

7.8

7.9

7.10

7.11

7.12

7.13

8 Sampling

9 Preparation of test sample

10 Procedure (as shown in Annex A)

10.1 Test portion

10.2 Pre-enrichment

10.3 Enrichment

10.4 Isolation of presumptive Cronobacter spp.

10.5 Confirmation

10.5.1 General

10.5.2 Purification of colonies

10.5.3 Biochemical confirmation

10.5.3.1 General

10.5.3.2 Oxidase

10.5.3.3 Hydrolysis of 4 Nitrophenyl (PNP) α-D-glucopyranoside substrate

10.5.3.4 L-Lysine decarboxylase

10.5.3.5 L-Ornithine decarboxylase

10.5.3.6 Fermentation of various carbohydrates

10.5.3.7 Methyl red (MR) (optional)

10.5.3.8 Voges-Proskauer (VP) (optional)

10.6 Interpretation of biochemical results

11 Expression of results

12 Performance characteristics of the method

12.1 Interlaboratory study

12.2 Sensitivity

12.3 Specificity

12.4 LOD50

13 Test report

Annex A

Annex B

B.1 Buffered peptone water (BPW)

B.1.1 Composition

B.1.2 Preparation

B.2 Cronobacter selective broth (CSB)

B.2.1 Base medium

B.2.1.1 Composition

B.2.1.2 Preparation

B.2.2 Vancomycin solution

B.2.2.1 Composition

B.2.2.2 Preparation

B.2.3 Complete medium

B.3 Chromogenic Cronobacter isolation (CCI) agar

B.3.1 Composition

B.3.2 Preparation

B.4 Tryptone soya agar (TSA) (example of non-selective medium)

B.4.1 Composition

B.4.2 Preparation

B.5 Media and reagents for biochemical characterization

B.5.1 Reagent for detection of oxidase

B.5.1.1 Composition

B.5.1.2 Preparation

B.5.2 α-Glucosidase enzymatic assay solution

B.5.2.1 Phosphate buffer, 0,3 M (pH 7,0)

B.5.2.1.1 Composition

B.5.2.1.2 Preparation

B.5.2.2 4-Nitrophenyl α-D-glucopyranoside substrate

B.5.2.2.1 Composition

B.5.2.2.2 Preparation

B.5.2.3 Complete medium

B.5.2.4 Physiological salt solution 0,85 % NaCl

B.5.2.4.1 Composition

B.5.2.4.2 Preparation

B.5.3 L-Lysine decarboxylation medium

B.5.3.1 Composition

B.5.3.2 Preparation

B.5.4 L- Ornithine decarboxylation medium

B.5.4.1 Composition

B.5.4.2 Preparation

B.5.5 Media for fermentation of carbohydrates (peptone water with phenol red, D-arabitol, α-Methyl-D-glucoside, D-sorbitol and D-sucrose)

B.5.5.1 Basic medium

B.5.5.1.1 Composition

B.5.5.1.2 Preparation

B.5.5.2 Carbohydrate solution D-arabitol (C5H12O5), α-Methyl-D-glucoside (C7H14O6), D-sorbitol (C6H14O6) and D-sucrose (C12H22O11) 80 mg/ml

B.5.5.2.1 Composition

B.5.5.2.2 Preparation

B.5.5.3 Complete carbohydrate fermentation media

B.5.5.3.1 Composition

B.5.5.3.2 Preparation

B.5.6 Methyl red (MR) and Voges-Proskauer (VP)-reactive compounds

B.5.6.1 MR/VP base medium

B.5.6.1.1 Composition

B.5.6.1.2 Preparation

B.5.6.2 Methyl red (MR) reagent

B.5.6.2.1 Composition

B.5.6.2.2 Preparation

B.5.6.3 Voges-Proskauer (VP) reagent

B.5.6.3.1 40 % KOH

B.5.6.3.1.1 Composition

B.5.6.3.1.2 Preparation

B.5.6.3.2 5 % 1-naphthol

B.5.6.3.2.1 Composition

B.5.6.3.2.2 Preparation

B.6 Performance testing of culture media and reagents

Annex C

Annex D

D.1 Results of interlaboratory studies

Bibliography

Appendix ZZ

ZZ1 Scope

ZZ2 Variations

ISO 22964:2017

AS 5013.13(Int)-2010

DR AS 5013.13:2017