BS EN 12353:2021 – TC:2022 Edition

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Tracked Changes. Chemical disinfectants and antiseptics. Preservation of test organisms used for the determination of bactericidal (including Legionella), mycobactericidal, sporicidal, fungicidal and virucidal (including bacteriophages) activity

Published By	Publication Date	Number of Pages
BSI	2022	92

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Categories: 11.080.20 - Disinfectants and antiseptics, BSI

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Description

This document specifies methods for keeping test organisms used and defined in European Standards for the determination of bactericidal (incl. Legionella pneumophila), mycobactericidal, sporicidal, fungicidal and virucidal (incl. bacteriophages) activity of chemical disinfectants and antiseptics drawn up by CEN/TC 216. These methods for keeping test organisms can only be carried out in connection with at least one of those standards where a reference to this document is established.

NOTE 1

Annex A (informative) contains a non-exhaustive list of test organisms for which this document can be applied.

NOTE 2

European Standards (EN) where this document is referenced are listed in the Bibliography.

NOTE 3

A specific description on the preservation of bacterial spores could be added once the results of the ongoing ring trials are available.

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PDF Pages	PDF Title
54	undefined
60	1 Scope 2 Normative references 3 Terms and definitions 4 Requirements
61	5 Methods 5.1 Principle 5.2 Test organisms, culture media and reagents 5.2.1 Test organisms 5.2.2 Culture media and reagents 5.2.2.1 General 5.2.2.2 Water
62	5.2.2.3 Tryptone Soya Broth (TSB) for bacteria, except Legionella 5.2.2.4 Malt Extract Broth (MEB) for fungi 5.2.2.5 Cryoprotectant solution for bacteria and fungi
63	5.2.2.6 Middlebrook 7 H 9 broth with 10 % ADC enrichment and glycerol as reconstituent and cryoprotectant solution for mycobacteria (MADC) 5.2.2.7 Polysorbate 80 solution 5.2.2.8 DMSO as cryoprotectant for cell culture freezing 5.2.2.9 Glutamine solution (3 % m/V solution) 5.2.2.10 TV (Trypsin-Versene) 5.2.2.11 Antibiotic suspension
64	5.2.2.12 Phosphate-buffered saline solution (PBS) 5.2.2.13 Fetal calf serum (FCS) 5.2.2.14 Earle’s BSS
65	5.2.2.15 Phenol red (1 % m/V solution) 5.2.2.16 Sodium bicarbonate (8,8 % w/v solution) 5.2.2.17 Cell culture medium 5.2.2.18 M17-broth 5.2.2.19 M17-agar (bottom agar)
66	5.2.2.20 Overlay agar (top agar, soft agar) 5.2.2.21 CaCl2-stock solutions (1 mol/l and 0,05 mol/l) 5.2.2.22 Reconstituted skim milk 5.2.2.23 Buffered Charcoal Yeast Extract (BCYE) Broth, for Legionella
67	5.2.2.24 Buffered Charcoal Yeast Extract (BCYE) Agar, for Legionella 5.2.2.25 Page’s saline (cryoprotective solution for Legionella)
68	5.2.3 Cell cultures 5.2.3.1 Storage of cell cultures 5.2.3.2 Preparation of new cell cultures
69	5.2.3.3 Passages 5.2.4 Host strains for dairy bacteriophages (Lactococcus lactis) 5.2.4.1 Stock culture of host bacteria 5.2.4.2 Working culture of host bacteria
70	5.3 Apparatus and glassware 5.3.1 General 5.3.2 Usual microbiological laboratory equipment
71	5.4 Procedure for preservation of test organisms – General 5.4.1 Handling of freeze-dried / frozen test organisms from culture collections 5.4.2 Choice of incubation procedure, agar medium, cell culture/cell line
72	5.5 Procedure for preservation of bacteria (incl. Legionella, aerobic spore-forming bacteria, excl. mycobacteria and bacterial spores) and yeasts 5.5.1 Reconstitution of the freeze-dried test organisms 5.5.2 Preparation for storage
73	5.5.3 Preparation of stock culture / working cultures 5.6 Procedure for preservation of mycobacteria 5.6.1 Reconstitution of the freeze-dried test organisms 5.6.2 Preparation for storage
74	5.6.3 Preparation of working cultures 5.7 Procedure for preservation of moulds (e.g. Aspergillus brasiliensis) 5.7.1 Reconstitution of the freeze-dried test organism 5.7.2 Preparation for storage
75	5.7.3 Preparation of stock culture / working cultures 5.8 Procedure for preservation of viruses (except dairy bacteriophages) 5.8.1 Reconstitution of frozen virus 5.8.2 Preparation for storage of stock virus suspension 5.8.3 Preparation of test virus suspension
76	5.9 Procedure for preservation of bacteriophages 5.9.1 Reconstitution of frozen bacteriophages 5.9.2 Preparation for storage 5.9.3 Preparation of bacteriophages working suspensions 5.10 Verification of the purity and identity of test organisms 5.10.1 General
77	5.10.2 Information on source of strains 5.10.3 Purity 5.10.4 Identity 5.11 Documentation 5.11.1 General 5.11.2 Freeze-dried test organism / frozen viruses 5.11.3 Cryovials of frozen test organism
78	5.11.4 Stock culture 5.11.5 Verification of purity and identity 5.11.6 Storage of documentation
79	Annex A (informative)Test organisms – Culture collection references and relation to CEN/TC 216 standards A.1 Bacteria (except mycobacteria and spore-forming bacteria) A.1.1 Enterobacter cloacae A.1.2 Enterococcus hirae A.1.3 Enterococcus faecium A.1.4 Escherichia coli (1) A.1.5 Escherichia coli (2) K12 A.1.6 Lactobacillus brevis A.1.7 Legionella pneumophila, subsp. Pneumophila A.1.8 Proteus hauseri
80	A.1.9 Pseudomonas aeruginosa A.1.10 Salmonella enterica subsp. enterica, serotype Typhimurium A.1.11 Staphylococcus aureus subsp. aureus A.2 Mycobacteria A.2.1 Mycobacterium avium subsp. avium A.2.2 Mycobacterium terrae A.3 Spore-forming bacteria A.3.1 Bacillus cereus A.3.2 Bacillus subtilis subsp. spizizenii A.3.3 Clostridioides difficile (former “Clostridium difficile”) R027
81	A.3.4 Clostridium sporogenes A.4 Fungi (moulds and yeasts) A.4.1 Aspergillus brasiliensis (former “A. niger”) (mould) A.4.2 Candida albicans (yeast) A.4.3 Saccharomyces cerevisiae (1) (yeast) A.4.4 Saccharomyces cerevisiae (2) var. diastaticus (yeast) A.5 Viruses A.5.1 Adenovirus type 5, strain Adenoid 75 A.5.2 Enterovirus E (Enterovirus Bovine Type 1), strain LCR 4 (former “Bovine enterovirus Type 1(ECBO)”)
82	A.5.3 Feline coronavirus (Alphacoronavirus 1), Strain Munich (FeCoV) A.5.4 Minute virus of mice (Rodent protoparvovirus 1) (former “Murine parvovirus, minute virus of mice, strain Crawford”) A.5.5 Murine norovirus, strain S99 Berlin A.5.6 Poliovirus type 1, Sabin strain LSc-2ab A.5.7 Porcine parvovirus (Ungulate protoparvovirus I), Strain NADL-2 (PPV) A.5.8 Vaccinia virus, strain modified vaccinia virus Ankara (MVA) A.5.9 Vaccinia virus, strain Elstree A.6 Bacteriophages A.6.1 Lactococcus lactis subsp. lactis bacteriophage P008 A.6.2 Lactococcus lactis subsp. lactis bacteriophage P001
83	Annex B (informative)Graphical representations

Additional information

Status	Definitive
Pages	92
Publication Date	2022-01-20
ISBN	978 0 539 20141 3
Standard Number	BS EN 12353:2021 – TC
Title	Tracked Changes. Chemical disinfectants and antiseptics. Preservation of test organisms used for the determination of bactericidal (including Legionella), mycobactericidal, sporicidal, fungicidal and virucidal (including bacteriophages) activity
Descriptors	Microbiological analysis, Disinfectant tests, Antiseptics, Specimen preparation, Eumycophyta, Biological analysis and testing, Purity, Test equipment, Fungicides, Verification, Sporozoa, Preservation, Microorganisms, Bacteriocide-activity determination, Bacteria, Disinfectants
Publisher	BSI
Committee	CH/216
ICS Codes	11.080.20 - Disinfectants and antiseptics

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