AOAC 2019 First Action Methods

In the event of discordant results such as a positive result with the GENE-UP® test, or no confirmation using a plate (no typical 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: ( d ) Transfer 100 μL from the primary enrichment to 10 mL FB for a secondary enrichment. Incubate at 35 ± 1°C for 24 ± 3 h. ( e ) Isolate on an ALOA agar plate at 35°C ± 1°C for 24 ± 3 h. The plate should b e read after 24 and 48 h (in case there are no typical colonies after 24 h). ( f ) The presence of typical colonies confirms a positive result. ( g ) If no typical Listeria colony is identified, repeat steps d and e using 500 µL from the primary enrichment. J. Quality Control [External quality control can be performed using one Listeria strain.] ( a ) Add one isolated colony from a fresh and pure culture in 10 mL of LPT broth. ( b ) Mix and incubate at 35 ± 1°C or 37 ± 1°C for 18 –24 h. ( c ) Dilute 1/100 of the culture in LPT broth in order to obtain a suspension containing approximately 106 cells/mL of the strain. ( d ) Follow the protocol from the SAMPLE LYSIS section steps to CONFIRMATION OF POSITIVE RESULTS sections. ( e ) Check that the results obtained correspond to the characteristics of the tested strains. K. Limitations of the Method The GENE-UP® Listeria spp. 2 (LIS 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.

Reference: J. AOAC Int . (future issue)

Posted: January 2020 (pre-publication)