《微生物生物学》课程教学资源（书籍文献）Microbiology Laboratory Guidebook.pdf_大学文库

USDA/FSIS Microbiology Laboratory Guidebook 3rd Edition/1998 i FOREWORD The 1993 Escherichia coli O157:H7 outbreak in the Pacific Northwest focused national attention on food safety. Since then, the number of requests for reprints on analytical methods used by the Microbiology Division, Office of Public Health and Science, Food Safety and Inspection Service, United States Department of Agriculture, has increased dramatically. Scientists within the Division have responded to these requests by completely revising and updating our Microbiology Laboratory Guidebook (MLG) for publication. This MLG is our laboratory guidebook for the microbiological analysis of meat, poultry, and egg products that fall under the jurisdiction of USDA. It contains methods that FSIS prefers to use for the analysis of these foods. Since USDA does not endorse or approve methods for use by the food industry, inclusion of a particular method in the MLG should not be construed in this manner. Similarly, the mention of specific brand or trade names for a product, medium, chemical or reagent associated with methods contained herein does not constitute endorsement or selectivity by the authors or USDA over similar products that might also be suitable. The use of the MLG comes with several caveats. This guidebook was written for microbiologists, and its interpretation and use should only be undertaken by trained microbiologists. FSIS assumes no responsibility for any economic, personal injury or other damage that may occur to individuals or organizations because of the use of methods contained in this guidebook. Users should note and pay particular attention to the safety caution symbol (†) and written warnings associated with certain hazardous chemicals or dangerous biological materials used in some of the methods. Users must act in a responsible manner at all times to protect themselves and the environment during performance of these methods. This guidebook must be supplemented with quality assurance and quality control programs as well as chemical, biological, and employee safety hazards management programs in order to operate a microbiology laboratory. These programs are beyond the scope of this guidebook and are the sole responsibility of the user to develop and implement. This guidebook contains protocols for analytical tests that are required by FSIS regulatory activities. Some protocols, such as the Bioassay procedure for antibiotic residue detection and quantitation, may not be of value to commercial laboratories nor do we expect others to try to commercialize them. They are included here primarily as informational material since they are part of our current analytical methods

USDA/FSIS Microbiology Laboratory Guidebook 3rd Edition/1998 iii GENERAL CONSIDERATIONS Before any analyst attempts to perform the microbiological methods contained within this Microbiology Laboratory Guidebook (MLG), it might be helpful to call attention to the following general considerations in the use of this guidebook. In order to maximize the achievement of successful results when using the various methods in this MLG, it should be clearly understood that all methods and procedures should be performed at all times in a manner as close as possible to the prescribed directions. Particular attention should be paid to all details provided in a given analytical procedure. Changes or shortcuts should not be attempted in a method simply to accommodate factors, for example, such as processing a large number of similar samples through the method at the same time. All chemicals, media, immunoreagents and commercial test kits should be within current shelf expiration dates and be subjected to quality control and quality assurance procedures to insure their proper performance for their intended purpose and use within the methods presented in this MLG. All instrumentation should be subjected to continuous maintenance and appropriate quality control procedures to insure unquestionably correct performance during use in all methods. The use of positive and negative test controls at all times, as specified for a given procedure, should be implemented. Adequate documentation and record keeping should be employed for all analytical results, test controls, quality assurance and quality control procedures, instrument maintenance programs, and any observed laboratory deviations to the above or in methods performance. Although all of the methods described in this guidebook have exact numerical values given for performance parameters such as weight and volume measures, pH, time and temperature to achieve optimum results, it should be clearly understood that an acceptable range exists within which optimum results can still be expected to be achieved without compromising the integrity of the method. For any given method, unless otherwise clearly stated within the text of this MLG, the following allowable ranges for the given parameters are considered to be acceptable and are applicable: Weight and volume measures: ± 1% pH: ± 0.2 units Time: hours ± 1 hour; minutes ± 1% Temperature: ± 1.0o C

USDA/FSIS Microbiology Laboratory Guidebook 3rd Edition/1998 i FOREWORD The 1993 Escherichia coli O157:H7 outbreak in the Pacific Northwest focused national attention on food safety. Since then, the number of requests for reprints on analytical methods used by the Microbiology Division, Office of Public Health and Science, Food Safety and Inspection Service, United States Department of Agriculture, has increased dramatically. Scientists within the Division have responded to these requests by completely revising and updating our Microbiology Laboratory Guidebook (MLG) for publication. This MLG is our laboratory guidebook for the microbiological analysis of meat, poultry, and egg products that fall under the jurisdiction of USDA. It contains methods that FSIS prefers to use for the analysis of these foods. Since USDA does not endorse or approve methods for use by the food industry, inclusion of a particular method in the MLG should not be construed in this manner. Similarly, the mention of specific brand or trade names for a product, medium, chemical or reagent associated with methods contained herein does not constitute endorsement or selectivity by the authors or USDA over similar products that might also be suitable. The use of the MLG comes with several caveats. This guidebook was written for microbiologists, and its interpretation and use should only be undertaken by trained microbiologists. FSIS assumes no responsibility for any economic, personal injury or other damage that may occur to individuals or organizations because of the use of methods contained in this guidebook. Users should note and pay particular attention to the safety caution symbol (†) and written warnings associated with certain hazardous chemicals or dangerous biological materials used in some of the methods. Users must act in a responsible manner at all times to protect themselves and the environment during performance of these methods. This guidebook must be supplemented with quality assurance and quality control programs as well as chemical, biological, and employee safety hazards management programs in order to operate a microbiology laboratory. These programs are beyond the scope of this guidebook and are the sole responsibility of the user to develop and implement. This guidebook contains protocols for analytical tests that are required by FSIS regulatory activities. Some protocols, such as the Bioassay procedure for antibiotic residue detection and quantitation, may not be of value to commercial laboratories nor do we expect others to try to commercialize them. They are included here primarily as informational material since they are part of our current analytical methods

USDA/FSIS Microbiology Laboratory Guidebook 3rd Edition/1998 iii GENERAL CONSIDERATIONS Before any analyst attempts to perform the microbiological methods contained within this Microbiology Laboratory Guidebook (MLG), it might be helpful to call attention to the following general considerations in the use of this guidebook. In order to maximize the achievement of successful results when using the various methods in this MLG, it should be clearly understood that all methods and procedures should be performed at all times in a manner as close as possible to the prescribed directions. Particular attention should be paid to all details provided in a given analytical procedure. Changes or shortcuts should not be attempted in a method simply to accommodate factors, for example, such as processing a large number of similar samples through the method at the same time. All chemicals, media, immunoreagents and commercial test kits should be within current shelf expiration dates and be subjected to quality control and quality assurance procedures to insure their proper performance for their intended purpose and use within the methods presented in this MLG. All instrumentation should be subjected to continuous maintenance and appropriate quality control procedures to insure unquestionably correct performance during use in all methods. The use of positive and negative test controls at all times, as specified for a given procedure, should be implemented. Adequate documentation and record keeping should be employed for all analytical results, test controls, quality assurance and quality control procedures, instrument maintenance programs, and any observed laboratory deviations to the above or in methods performance. Although all of the methods described in this guidebook have exact numerical values given for performance parameters such as weight and volume measures, pH, time and temperature to achieve optimum results, it should be clearly understood that an acceptable range exists within which optimum results can still be expected to be achieved without compromising the integrity of the method. For any given method, unless otherwise clearly stated within the text of this MLG, the following allowable ranges for the given parameters are considered to be acceptable and are applicable: Weight and volume measures: ± 1% pH: ± 0.2 units Time: hours ± 1 hour; minutes ± 1% Temperature: ± 1.0o C

USDA/FSIS Microbiology Laboratory Guidebook 3rd Edition/1998 1-1 CHAPTER 1. SAMPLE PREPARATION FOR MEAT, POULTRY AND PASTEURIZED EGG PRODUCTS Charles P. Lattuada and B. P. Dey 1.1 Introduction The purpose for the microbiological examinations of meat and poultry products is to obtain information. This information gathering may follow a qualitative or quantitative analytical format. The format followed is called the sampling plan. Many microorganisms are present in very low numbers and require one or more enrichment steps. If cell injury is anticipated, a nonselective enrichment frequently is used to resuscitate cells, followed by a more selective enrichment. The analyst must study all records and correspondence before examining the sample. Care must be exercised in maintaining and handling the sample to insure that it is the same one that was collected, that it has not been tampered with, and that its condition is the same as it was at collection. The reserve sample must be stored properly to maintain its integrity in case additional analyses are required. An analyst must be keenly aware that during all steps of the analysis, it is important to minimize the growth of non-critical microorganisms and to prevent entrance of environmental contaminants. The organism(s) isolated must come from the test sample and not from an outside source. These facts cannot be over-emphasized and can be accomplished only if strict attention is paid to the following rules: The sampling operation must be well organized, with all supplies and equipment properly positioned before starting. Ideally, sampling should be done in an area free of air currents following good aseptic procedures. All work surfaces must be clean and sanitized. Implements used for sampling must be sterile before use and protected from outside contamination during use. The outside of the immediate container must be thoroughly sanitized. Any laboratory person processing samples must be very familiar with aseptic techniques and the principles of sterilization, sanitization and disinfection. The person assigned to the sampling task should know the sampling protocol to be used and have a

USDA/FSIS Microbiology Laboratory Guidebook 3rd Edition/1998 1-2 reference copy at hand in case questions arise. 1.2 Sanitizing the Work Area The work area must be clean and free from dust; detergent sanitizers are satisfactory for cleaning. Before work begins, the work area should be cleaned and a sanitizer/disinfectant applied liberally and given time to act. Quaternary ammonium compounds, sodium hypochlorite and phenolic compounds all are suitable for this purpose. The manufacturer's instructions regarding the concentration needed and the time required for the compound to act should be followed. 1.3 Sterilization of Instruments a. All instruments and containers to be used in the sample analysis must be sterile. Any sterilization procedure may be used that is compatible with the material to be sterilized. Sterilization implies the total destruction of all viable organisms as measured by an appropriate culturing method. b. An exception can be made, if necessary, when the number of instruments is limited (ie. chisels) and the testing protocol does not include sporeforming microorganisms. In which case, the instruments first are washed with soap and water, rinsed and inspected to be sure there is no organic matter in crevices or hinges, then they may be steamed for 30 minutes in an instrument sterilizer or placed in boiling water for two minutes. c. Do not dip instruments into alcohol and flame them as a substitute for heat sterilization. It is not a substitution for the methods given above. 1.4 Disinfection of Outer Surface of the Immediate Container a. The outside covering of the intact immediate container must be decontaminated to the greatest extent possible and particularly in the area where an opening will be made to expose the contents. b. Hydrogen peroxide, tincture of iodine or 2500 ppm sodium hypochlorite solution may be used for this purpose. Allow time for the disinfectant to act before opening the container. Aseptically remove any residual disinfectant to prevent its entering the container when an opening is made