AS 5013.24.1 pdf download.Food microbiology
3 Terms and definitions
For the purposes of this document, the following terms and definitions apply.ISO and IEC maintain terminological databases for use in standardization at the following addresses: — IEC Electropedia: available at http:// www .electropedia .org/ — ISO Online browsing platform: available at http:// www .iso .org/ obp 3.1 Listeria monocytogenes microorganisms which form typical colonies on solid selective media and which display the morphological, physiological and biochemical characteristics described when tests are carried out in accordance with this document 3.2 detection of Listeria monocytogenes determination of the detection/non detection of Listeria monocytogenes (3.1), in a given mass or volume of product or a specified surface, when tests are carried out in accordance with this document 3.3 Listeria spp. microorganisms which form typical colonies on solid selective media and which display the morphological, physiological and biochemical characteristics described when tests are carried out in accordance with this document 3.4 detection of Listeria spp. determination of the detection/non detection of Listeria spp. (3.3), in a given mass or volume of product or a specified surface, when tests are carried out in accordance with this document
4.1 General Listeria spp. may be present in small numbers and are often accompanied by considerably larger numbers of other microorganisms, therefore selective enrichment is necessary. It is also necessary to detect injured and stressed Listeria spp. and the primary selective enrichment medium, with reduced inhibitor concentration, fulfils at least part of this function. NOTE Presence of L. monocytogenes can be masked by the presence of other Listeria species, in particular L. innocua or L. ivanovii. Within the limits of this document, the detection of L. monocytogenes and of Listeria spp. necessitates four successive stages, as described in the flowchart in Annex A. 4.2 Primary enrichment in a selective liquid enrichment medium with reduced concentration of selective agents (half‑Fraser broth) Inoculation of a selective primary enrichment medium containing half the concentrations of acriflavine and nalidixic acid (half-Fraser broth, see B.1), which is also used as a dilution fluid for the test portion (9.1). Incubation of the initial suspension at 30 °C for 24 h to 26 h. 4.3 Secondary enrichment with a selective liquid enrichment medium with full concentration of selective agents (Fraser broth) Inoculation of full-strength secondary liquid enrichment medium (Fraser broth) with a culture obtained from 4.2. Incubation of the Fraser broth at 37 °C for 24 h. 4.4 Plating out and identification From the cultures obtained in 4.2 and 4.3, plating out on the two selective solid media: — Agar Listeria according to Ottaviani and Agosti (see References  and  and B.3); — any other solid selective medium at the choice of the laboratory complementary to Agar Listeria according to Ottaviani and Agosti, using a different substrate and/or principle than the one used in Listeria agar according to Ottaviani and Agosti (see B.4). See Annex E for information about media. Incubation of the Agar Listeria according to Ottaviani and Agosti at 37 °C for a total of 48 h. If colonies of presumptive L. monocytogenes or Listeria spp. are evident at 24 h the incubation may be stopped at this stage. Incubation of the second selective medium at the appropriate temperature and examination after the appropriate time.
6 Equipment and consumables
Usual microbiological equipment (as specified in ISO 7218) and, in particular, the following. 6.1 Apparatus for dry sterilization (oven) or wet sterilization (autoclave). As specified in ISO 7218. 6.2 Drying cabinet or incubator, capable of being maintained between 25 °C and 50 °C. 6.3 Incubators, capable of operating at 30 °C ± 1 °C, 37 °C ± 1 °C and at 25 °C ± 1 °C (optional). 6.4 Water bath, capable of operating at 47 °C to 50 °C. 6.5 Sterile loops approximately 3 mm in diameter or 10 µl, and inoculating needle or wire. 6.6 pH meter, capable of being read to the nearest 0,01 pH unit at 25 °C, enabling measurements to be made which are accurate to ± 0,1 pH unit. 6.7 Graduated pipettes or automatic pipettes, of nominal capacities 1 ml and 10 ml. 6.8 Petri dishes, for example of diameter 90 mm. 6.9 Microscope, preferably with phase-contrast, and with slides and cover slips. 6.10 Refrigerator, capable of operating at 5 °C ± 3 °C.