AS 5013.13 pdf download

AS 5013.13 pdf download.Food microbiology
5 Principle
5.1 Non-selective pre-enrichment in BPW A test portion is inoculated into BPW, then incubated between 34 °C and 38°C for 18h+2 h. NOTE Cronobacter spp. can be present in small numbers accompanied by other Enterobacteriaceae such as E. cloacae that could interfere with their detection. 5.2 Enrichment in a selective medium (CSB) The selective enrichment medium is inoculated with the culture obtained in 5.1 and incubated at 41,5°C+ 1 °Cfor24h+2 h. 5.3 Plating out and identification on chromogenic agar (CCI agar) The chromogenic agar is streaked for isolation with the enrichment culture obtained in 5.2 and incubated at41,5°C+ 1 °Cfor 24h+2 h. 5.4 Confirmation Typical colonies are selected from the chromogenic agar, purified on a non-selective agar such as TSA and biochemically characterized.
6 Culture media and reagents
For current laboratory practice, see ISO 7218 and ISO 11133. Composition of culture media and reagents and their preparation are described in Annex B. For performance testing of culture media see ISO 11133 and/or Annex B.
7 Equipment and consumables
Disposable equipment is an acceptable alternative to reusable glassware if it has suitable specifications. Usual microbiological laboratory equipment (see ISO 7218) and, in particular, the following. 7.1 Apparatus for dry sterilization [oven) or wet sterilization (autoclave). As specified in ISO 7218. 7.2 Incubators, capable of operating in the range 34 °C to 38。C, 37°C+1。C and 41,5 °C+1 °C  7.3 Sterile loops, of approximate diameter 3 mm (10 μl volume), and of 1 μl volume, and inoculation needle or wire. 7.4 pH meter, having an accuracy of calibration of士+ 0,1 pH unit at25 °C. . 7.5 Flasks and bottles, with closures, of suitable capacities for use in the preparation of enrichment broths and agars and their storage. . 7.6 Sterile graduated pipettes or automatic pipettes, of nominal capacities 10 ml, 1 ml and 0,1 ml. 7.7 Tubes (plugged or with caps) or culture bottles, of appropriate capacity, with non-toxic metallic caps with liners or plastic disposable caps (see ISO 7218). 7.8 Petri dishes, of diameter approximately 90 mm. 7.9 Spectrophotometer, capable of measuring absorption of light with a wavelength of 405 nm. 7.10 Pestle and mortar. 7.11 Refrigerators, capable of operatingat5°C+3。C 7.12 Water baths, capable of operating between 47 °C and 50。C and at37°C土1 °C. 7.13 Drying cabinet (or oven ventilated by convection), capable of being maintained between 25。C and 50 °C.
8 Sampling
Sampling is not part of the method specified in this document. See the specific International Standard dealing with the product concerned. If there is no specific International Standard dealing with sampling of the product concerned, it is recommended that the interested parties come to an agreement on this subject. A recommended sampling method is given in ISO/TS 17728[31 for food and animal feed, and ISO 18593[4] for sampling of surfaces. It is important that the laboratory receives a sample which is truly representative and which has not been damaged or changed during transport or storage (see ISO 7218).
9 Preparation of test sample
Prepare the test sample from the laboratory sample in accordance with the specific International Standard dealing with the product concerned: see ISO 6887 (all parts). If there is no specific International Standard, it is recommended that the parties concerned come to an agreement on this subject.
10 Procedure (as shown in Annex A)
10.1 Test portion In general, to prepare the primary dilution, add 10 g or 10 ml of the test sample (Clause 9) to 90 ml of pre- enrichment medium (B.1) (BPW), to yield a tenfold dilution. Pre-warm the BPW to room temperature before use. For specific products, follow the procedures specified in ISO 6887 (all parts). For dry milk, follow ISO 6887-5. This document has been validated for test portions of 10 g. A smaller size of the test portion may be used without the need of additional validation/ verification providing that the same ratio between pre- enrichment broth and test portion is maintained. A larger test portion than that initially validated may be used, if a validation/verification study has shown that there are no negative effects on the detection of Cronobacter spp. NOTE1 Validation can be conducted in accordance with the appropriate documents of ISO 16140 (all parts). Verification for pooling samples can be conducted in accordance with the protocol described in ISO 6887-1:2017, Annex D (verification protocol for pooling samples for qualitative tests). NOTE2 Large sample sizes can compromise the recovery of stressed Cronobacter spp. when interfering microflora are present, such as probiotics.[5[,[6] For preparing quantities larger than 10 g, BPW should be pre-warmed between 34 °C and 38。C (7.2) before inoculated with the test portion.