ERP Micro December 2019

GENE-UP ® Listeria spp. 2 (LIS 2)

050567 - 02 - 2018-12 - en

CONFIRMATION OF POSITIVE RESULTS Confirmation of Positive Results Obtained Using the Method Certified NF VALIDATION and Outside NF VALIDATION or AOAC RI Certification Protocols All positive results obtained must be confirmed. Confirmation should be performed using the enrichment broth stored at +2°C/+8°C and should be initiated within 72 hours following the end of the incubation period. If using an enriched sample, mix thoroughly by hand. Follow one of the three validated procedures below: 1. Isolate the enriched sample, and streak 10 μL on PALCAM agar or on chromogenic agar forming part of a method for detection of the Listeria genus. The presence of characteristic colonies is sufficient to confirm the presence of Listeria spp. 2. If identification of colonies is necessary, use an API ® LIS strip to directly test isolated colonies (without a purification step). In the event of discordant results (positive with the alternative method, not confirmed by one of the options described above), the laboratory must take the necessary steps to ensure that the results obtained are accurate. It is recommended, for example, to perform the following procedure: 1. Transfer 100 μL of sample from the first enrichment to 10 mL Fraser for a second enrichment. 2. Incubate at +37°C ± 1°C for 24 ± 3 hours. 3. Isolate 10 μL on PALCAM agar or on chromogenic agar forming part of a method for detection of the Listeria genus. The presence of characteristic colonies is sufficient to confirm the presence of Listeria spp. 4. If no typical Listeria colony is identified, repeat steps 1 and 2, using 500 μL from the first enrichment. Confirmation of Positive Results Obtained Using AOAC RI Approved Protocols All positive results must be confirmed according to the BAM or MLG, or according to the following BIOMÉRIEUX GENE ‑ UP ® confirmation protocol. 9,10 Confirmation should be performed using the enrichment broth stored at +2°C/+8°C and should be initiated within 72 hours following the end of the incubation period. If using an enriched sample, mix thoroughly by hand. 1. Isolate the enrichment broth on an ALOA agar plate; incubate at +35°C ± 1°C for 48 ± 3 hours. The plate should be read after 24 and 48 hours (in case there are no typical colonies after 24 hours). 2. The presence of typical colonies confirms a positive result. 3. If identification of colonies is necessary, use an API ® LIS strip to directly test isolated colonies (without a purification step). In the event of discordant results such as a positive result with the GENE ‑ UP ® test, or no confirmation using on a plate (untypical colonies), the laboratory must take the necessary steps to ensure that the results obtained are accurate. It is recommended, for example, to perform the following procedure: 1. Transfer 100 μL of sample from the first enrichment to 10 mL Fraser broth for a second enrichment. 2. Incubate at +35°C ± 1°C for 24 ± 3 hours. 3. Isolate on an ALOA agar plate and incubate at +35°C ± 1°C for 48 ± 3 hours. The plate should be read after 24 and 48 hours (in case there are no typical colonies after 24 hours). 4. The presence of typical colonies confirms a positive result. 5. If no typical Listeria colony is identified, repeat steps 1 and 2, using 500 μL from the first enrichment. QUALITY CONTROL External quality control can be performed using one Listeria strain. 1. Add one isolated colony from a fresh and pure culture in 10 mL of LPT broth. 2. Mix and incubate at +35°C or +37°C ± 1°C for 18 ‑ 24 hours. 3. Dilute 1/100 of the culture in LPT Broth in order to obtain a suspension containing approximately 10 6 cells/mL of the strain. Follow the protocol from the Lysis section steps to CONFIRMATION OF POSITIVE RESULTS sections. Check that the results obtained correspond to the characteristics of the tested strains. Note: It is the responsibility of the user to perform Quality Control taking into consideration the intended use of the medium, and in accordance with any applicable local regulations (frequency, number of strains, incubation temperature, etc.). LIMITATIONS OF THE METHOD The GENE ‑ UP ® Listeria spp. 2 kit has been evaluated on a large number of matrices. However, given the wide variety of products and manufacturing procedures, it is recommended to check that the composition of the matrices tested does not affect the reliability of GENE ‑ UP ® results. Note: Listeria grayi is not detected by the method.

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