US Environmental Protection Agency Office of Pesticide Programs Office of Pesticide Programs Microbiology Laboratory Environmental Science Center, Ft. Meade, MD Standard Operating Procedure for Tracking of Test Microorganisms SOP Number: MB-02-06 Date Revised: 09-27-12 ------- SOP No. MB-02-06 Date Revised 09-27-12 Page 1 of 6 SOP Number MB-02-06 Title Tracking of Test Microorganisms Scope Provides guidance for establishing receipt and expiration dates for microorganisms used in the Microbiology Laboratory Branch (MLB) as well as denotation and tracking of those microorganisms. Application Organism control numbers are used for all reconstituted cultures of vegetative bacteria. Approval Date SOP Developer: Print Name: SOP Reviewer Print Name: Quality Assurance Unit Print Name: Branch Chief Print Name: Date SOP issued: Controlled copy number: Date SOP withdrawn: ------- SOP No. MB-02-06 Date Revised 09-27-12 Page 2 of 6 TABLE OF CONTENTS Contents Page Number 1. DEFINITIONS 3 2. HEALTH AND SAFETY 3 3. PERSONNEL QUALIFICATIONS AND TRAINING 3 4. INSTRUMENT CALIBRATION 3 5. SAMPLE HANDLING AND STORAGE 3 6. QUALITY CONTROL 3 7. INTERFERENCES 3 8. NON-CONFORMING DATA 3 9. DATA MANAGEMENT 3 10. CAUTIONS 3 11. SPECIAL APPARATUS AND MATERIALS 3 12. PROCEDURE AND ANALYSIS 3 13. DATA ANALYSIS/CALCULATIONS 5 14. FORMS AND DATA SHEETS 5 15. REFERENCES 6 ------- SOP No. MB-02-06 Date Revised 09-27-12 Page 3 of 6 1. Definitions Abbreviations/definitions are provided in the text. 2. Health and Safety Follow procedures specified in SOP MB-01, Laboratory Biosafety. The Study Director and/or lead analyst should consult the Material Safety Data Sheet for specific hazards associated with products. 3. Personnel Qualifications and Training Refer to SOP ADM-04, OPP Microbiology Laboratory Training. 4. Instrument Calibration Not applicable. 5. Sample Handling and Storage Not applicable. 6. Quality Control For quality control purposes, the required information is documented on the appropriate record form(s) (see section 14 of this SOP as well as MB-05, MB- 06, MB-07, MB-09, MB-23, MB-24, MB-25, and MB-27). 7. Interferences None 8. Non- conforming Data Entry errors will be corrected upon discovery. 9. Data Management Data will be archived consistent with SOP ADM-03, Records and Archives. 10. Cautions Expired ampules of lyophilized organisms are not used and will be discarded appropriately. 11. Special Apparatus and Materials 1. Pseudomonas aeruginosa (American Type Culture Collection (ATCC) 15442), Staphylococcus aureus (ATCC 6538), Bacillus subtilis (ATCC 19659), and Salmonella enterica (ATCC 10708); ordered annually and received directly from ATCC. 2. Mycobacterium bovis (BCG); ordered and received directly from Organon Teknika. 3. Other microorganisms used in the microbiology laboratory may be purchased from appropriate vendors when necessary. 12. Procedure and Analysis Procedures for generating stock cultures for organisms maintained on slants and stabs or as frozen stock cultures can be found in SOPs MB-05, MB-06, MB-07, MB-09, MB-23, MB-24, MB-25, and MB-27. 12.1 Supply Control Number a. All lyophilized ampules are given a supply control number upon receipt (see SOP QC-09, Control Numbers). b. The supply control number for ampules consists of the date the ampule was received (R) and the date the ampule expires (E). ------- SOP No. MB-02-06 Date Revised 09-27-12 Page 4 of 6 For example: For a dehydrated ampule of S. aureus received on 01- 05-12 and expiring on 03-12-13, the supply control number would be R010512E031213. For a dehydrated ampule ofM bovis (BCG) received on 07-24-11 and expiring on 09-04-13, the supply control number would be R072411E090413. 12.2 Organism Control Number a. The organism control number consists of the date the reconstituted microbe expires (ME), a two-letter suffix denoting the organism, and any culture transfer notation (refer to section 12.3 for culture transfer notation). b. The two-letters in the suffix of the organism control number denote the genus and species of the organism (for example, Pa fori5. aeruginosa, Sa for S. aureus, Bs for B. subtilis, and Se for S. enterica). Other microorganisms used in the microbiology laboratory will be given two-letter suffixes consistent with the notation listed above. c. For organisms reconstituted every 12 months: A culture of S. aureus reconstituted on 02-03-12 would receive an organism control number of ME020313-Sa followed by any culture transfer notations. Cultures reconstituted every 18 months will follow similar notation. d. For organisms maintained on slants and stabs: Once reconstituted, these organisms may be used for one year. Any cultures, including stock cultures, daily transfers, and test cultures, are valid for up to 12 months after reconstitution of the lyophilized culture (the ME date). In the case provided (section 12.2c), stock cultures and any resulting test cultures are valid through 02-03-13. e. For organisms maintained as frozen stock cultures: Once frozen, these organisms may be used for eighteen months. Any cultures, including stock cultures, daily transfers, and test cultures are valid for up to 18 months after the stock cultures are frozen (the ME date). In the case provided (section 12.2c), stock cultures and any resulting test cultures are valid through 08-03-13. f. Once expired, autoclave and discard the stock cultures and initiate a new culture from a new unexpired lyophilized lot from ATCC. g. M. bovis (BCG): M. bovis (BCG) is not required to be replaced annually and does not have an expiration date for stocks (based on AO AC Method 965.12). A re-ordering cycle for new cultures has not been established. Therefore, the culture notation consists of MEOOOOOO-Mb followed by any culture transfer notations (refer to section 12.3 for culture transfer notation). Continuous transfers ofM bovis (BCG) may be made unless the organism has been compromised. ------- SOP No. MB-02-06 Date Revised 09-27-12 Page 5 of 6 12.3 Culture Transfer Notations of Test Microbes a. See footnotes for Organism Culture Tracking Form for culture transfer notations for S. aureus, P. aeruginosa, and S. enterica (see section 14.0). b. For organisms maintained on slants and stabs: a Staphylococcus test culture notation would be MEXXXXXX-Sa-1104TC, where 11 is the monthly culture transfer (number of months since the month of initiation) and 04 is the daily culture transfer (4th daily culture transfer); TC is applied to indicate a 48 hour test culture. c. For organisms maintained as frozen stock cultures: a Staphylococcus daily transfer notation would be MEXXXXXX-Sa-2-D3, where 2 is the vial number of the frozen stock culture and D3 is applied to indicate the third 24 hour daily transfer. Up to 5 daily transfers into 10 mL of broth may be conducted prior to the inoculation of the final test cultures. An example of a Staphylococcus test culture notation would be MEXXXXXX-Sa-2-D3TC, where 2 is the vial number of the frozen stock culture, D3 indicates that the test culture was inoculated using the third 24 hour daily transfer, and TC is applied to indicate a 48 hour test culture. d. See footnotes for Organism Culture Tracking Form for Mycobacterium bovis (BCG) culture transfer notations (see section 14.0). ForM bovis (BCG), the test culture notation would be MEXXXXXX-Mb-1103SL, where 11 represents the month of culture transfer (the month of the year) and 03 represents the week of the month for that transfer (the 3r week of the month). e. ForM bovis (BCG): SC is applied to identify a stock culture (e.g., a transfer from a slant to a slant). SL is applied to identify the test culture (this indicates a transfer from a slant culture to a liquid culture). LL can also be applied to identify a test culture (this indicates a transfer from a liquid MPB culture to another liquid culture). f. For M bovis (BCG): The weeks of each month are numbered consecutively starting with the 1st Monday of the month (as 01) and ending with the last Monday of the month (depending on the number of Mondays in the month, as either 04 or 05). 13. Data Analysis/ Calculations None 14. Forms and Data Sheets Test Sheets. Test sheets are stored separately from the SOP under the following file names: Organism Culture Tracking Form (Frozen Stock MB-02-06 Fl.docx Cultures) Organism Culture Tracking Form (Slants/Stabs) MB-02-06_F2.docx ------- SOP No. MB-02-06 Date Revised 09-27-12 Page 6 of 6 Test Microbe Confirmation Sheet (Quality MB-02-06 F3.docx Control) Organism Culture Tracking Form for MB-02-06 F4.docx Mycobacterium bovis (BCG) Test Microbe Confirmation Sheet (Quality MB-02-06 F5.docx Control) for Mycobacterium bovis (BCG) 15. References None ------- |