MFLP-74 Enumeration of Listeria monocytogenes in Food
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157D956C98894BD4A2F196CBC749EEC8 |
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文件大小(MB)： |
0.06 |
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页数： |
8 |
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日期： |
2012-3-2 |
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Published on the Food Directorate’s (Health Canada's) website at:,http://www.hc-sc.gc.ca/fn-an/res-rech/analy-meth/microbio/index-eng.php,Laboratory Procedure MFLP-74,February 2011,Health Products and Food Branch,Ottawa,Enumeration of Listeria monocytogenes in foods,Franco Pagotto 1, Yvon-Louis Trottier 1, Jacqueline Upham 2 and Irene Iugovaz 1,1 Research Division, Bureau of Microbial Hazards, Food Directorate, Health Canada, Ottawa, Ontario, HPFB, Postal Locator 2204E,K1A 0K9,2 Canadian Food Inspection Agency, Microbiology, Dartmouth Laboratory, 1992 Agency drive, Dartmouth, NS, B3B 1Y9.,E-mail: micro_methods_committee@hc-sc.gc.ca,1. Application,This method is applicable to the enumeration of viable Listeria monocytogenes to determine,compliance with the requirements of Sections 4 and 7 of the Food and Drugs Act. This method,replaces MFLP-74, dated August 2010.,2. Principle,This direct plating procedure quantitatively determines the number of viable Listeria,monocytogenes in the product. A portion of the product is blended in a suitable diluent, surfaceplated,onto at least two selective agars, and the plates are then incubated under specified,conditions of time and temperature. It is assumed that each viable cell of L. monocytogenes will,multiply under these specified incubation conditions and give rise to a visible colony which can,be identified. The selective media used are based on studies done by other investigators (7.3,7.4, 7.5).,3. Definition of terms,See Appendix A of Volume 3.,4. Collection of samples,See Appendix B of Volume 3.,5. Materials and special equipment,See MFHPB-30, ‘Materials and Special Equipment’.,See Appendix G for the formulas of individual media.,MFLP-74,February 2011,Published on the Food Directorate’s (Health Canada's) website at:,http://www.hc-sc.gc.ca/fn-an/res-rech/analy-meth/microbio/index-eng.php,- 2 -,Additional requirements:,1) Peptone water, 0.1% (w/v),2) Plating media:,Selective media include the following:,- OXA (mandatory agar),One of the following is to be included with OXA:,- Agar Listeria according to Ottaviani & Agosti (ALOA),- A.L. agar,- BBL Chromagar Listeria,- Brilliance Listeria Agar (formerly OCLA agar),- Lithium chloride-phenylathanol-moxalactam medium (LPM),- Modified Oxford agar (MOX),- Palcam agar (PAL),- Rapid L’Mono,6. Procedure,Information regarding the distribution of Listeria monocytogenes can be obtained by testing each,analytical unit separately.,6.1 Handling and Dilution of Sample Units,6.1.1 In the laboratory prior to analysis, except for shelf-stable foods, keep sample,units refrigerated or frozen depending on the nature of the product. Thaw,frozen samples in the refrigerator, or under time and temperature conditions,which prevent microbial growth or death.,6.1.2 Analyses should be started as soon as possible after receipt of samples in the,laboratory.,6.1.3 To ensure a representative analytical unit, agitate liquids or free flowing,materials until the contents are homogeneous. If the sample unit is a solid,obtain the analytical unit by taking a portion from several locations within the,sample unit. The analytical sample recommended is 10 grams or 10 ml.,6.2 Direct Plating Procedure,6.2.1 Prepare a 1:5 dilution of the sample, as required, in 0.1% (w/v) peptone water,in a blender jar or stomacher bag. Blend or stomach for 2 minutes to ensure,thorough mixing. For food matrices that require a higher dilution to allow for,ease of spreading the food/diluent slurry on the selective plates, a 1:10 dilution,may be used. This may require the plating of additional sample aliquot over,additional plates.,Two agar types are used to enumerate L. monocytogenes as follows:,immediately spread 1 ml by taking three, 0.333 mL portions of the diluted food,MFLP-74,February 2011,Published on the Food Directorate’s (Health Canada's) website at:,http://www.hc-sc.gc.ca/fn-an/res-rech/analy-meth/microbio/index-eng.php,- 3 -,and spread onto the surface of 3 plates of each selective media (e.g., a total of,2 mL is plated over two agar types). Selective media includes OXA and one of,the approved agars listed in section 5.,6.2.2 Incubate LPM plates at 30oC and all other selective agars at 35oC for 48 h,unless the time and temperature are otherwise directed by the manufacturer.,Examine plates at 24 h for typical colonies, as well as at 48 h as applicable to,the medium. Observe for colonies with a typical appearance (section 6.3.1 to,6.3.8).,NOTE:,1. Prior to use ensure agar plates are dry to avoid undue spreading of colonies. After spreading,a sample aliquot, ensure that the agar plate is dry before inverting and placing in an incubator.,2. I……
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