PCR | Microval

New Certificate for SureTect Salmonella Species PCR Assay

MicroVal has approved the issuing of the certificate for ThermoFisher Scientific’s SureTect Salmonella Species PCR Assay. Q Laboratories carried out the extension validation study for a broad range of foods in which the use of pre-warmed (41.5 or 37 ± 1 °C) BPW ISO as the primary enrichment media was evaluated, in addition to alternative sample sizes.

The Thermo Scientific SureTect Salmonella Species PCR Assay was originally validated by AFNOR according to the ISO 16140:2003 standard in 2013. Multiple extension studies were conducted since the initial validation study, and the first renewal validation enabled alignment with the revised ISO 16140 – 2:2016 standard in 2017. The method is validated for the use of three instruments: Applied Biosystems 7500 Fast, Applied Biosystems Quantstudio 5 and the PikoReal instrument. A summary of the initial validation, original renewal, and previous extension studies can be found on the available summary report (AFNOR Certificate: UNI 03/12-01/18).

All data generated from the three claimed thermocyclers for each matrix claimed under the AFNOR Certificate: UNI 03/12-01/18 was presented in a Gap Analysis report to the MicroVal Technical Committee (MVTC) according to the MicroVal Rules set up by the MicroVal General Committee (MGC). The MVTC agreed that the supporting data on the three different cyclers covered a broad range of foods, thus meeting the MicroVal requirements needed for the method to be transferred to become MicroVal 2022LR111. It should be noted that both Applied Biosystems 7500 Fast and QuantStudio 5 thermocyclers are included in the extension claims in opposite to the PikoReal instrument that was used in the ILS portion of the original validation.

The summary report of the extension validation study, available on the MicroVal website, provides an overview of the categories and sample sizes that were included in the final scope, as well as the instrumentation that was validated for use in this extension. The certificate and the accompanying summary report, number 2022LR111, can be found under ‘issued certificates / alternative methods’.

Extended Certificate for Assurance GDS for Cronobacter Tq II

MicroVal has approved the issuing of the extended certificate for Millipore Sigma’s Assurance GDS for Cronobacter Tq II. Campden BRI carried out the extension study for a new confirmation method in combination with two different isolation methods. The Assurance GDS for Cronobacter Tq II method was previously validated in December 2018 for infant formula and infant cereals and environmental samples for 10-375 g samples.

The Assurance GDS for Cronobacter Tq II is an automated nucleic acid amplification system for the detection of Cronobacter in infant nutritional formula, ingredients, and environmental samples. This PCR-based method targets DNA specific to Cronobacter spp.

The extension study followed the MicroVal interpretation guidelines for ISO 16140-6 for a new confirmation method from a previously validated qualitative method. This extension configuration was previously validated in the Assurance GDS for E. coli O157:H7 Tq, where an inclusivity and exclusivity study and a limited number of sensitivity samples were used to assess the performance of the new confirmation method. In the extension study, samples were analysed by the alternative method only to verify the performance of the new colony isolation and colony confirmation procedures.

Confirmation of presumptive positive colonies was performed using two different isolation methods:

Direct streak of the 1:10 enriched sample onto one chromogenic agar plate from a choice of 3 different isolation media
Aliquot the remaining GDS concentration reagent retained in the resuspension plate onto one chromogenic agar plate from a choice of 2 different isolation media

A selection of infant formula, infant cereals and dried milk products were tested in the sensitivity study as a representative number of the samples listed in the validation.

The certificate and the accompanying summary report, number 2017LR77, can be found under ‘issued certificates / alternative methods’.

New ISO 16140-6 validation for Check&Trace Salmonella 2.0

MicroVal has approved the issuing of the certificate for Check-Points’s Check&Trace Salmonella 2.0. WFC Analytics carried out the validation study, based on ISO 16140-6, for an alternative method for the confirmation and typing of Salmonella spp. Following from the validation study, the Check&Trace Salmonella 2.0 (CTS 2.0) is considered equivalent to the reference method (ISO 6579-1 and ISO 6579-3) for the confirmation of presumptive Salmonella spp. isolated on non-selective NA and selective XLD and for typing of 59 Salmonella serovars.

The Check&Trace Salmonella 2.0 Assay is a qualitative, semi-automated real-time PCR test designed for the confirmation and typing of presumptive Salmonella isolates from enriched culture media. The confirmation procedure advances a suspected (presumptive) Salmonella result to a confirmed Salmonella spp. result or a Salmonella negative result. The typing procedure generates a serovar result or a “genovar” code both uniquely defining each bacterial isolate tested.

DNA typing differs from serotyping. With serotyping the presence of antigens on the cell surface and flagella are detected. This is based on expression of genes located on two specific segments of the Salmonella genome. The CTS 2.0 assay detects genetic variation at 21 loci scattered over the whole Salmonella genome: this generates specific Salmonella genotypes, also called Genovars. The CTS 2.0 database links Genovars to a collection of well-characterized Salmonella Serovars. A Genovar will be given as test result if there is no established Serovar for the Genovar generated by the CTS 2.0 assay.

Check&Trace Salmonella 2.0 is considered equivalent to the ISO standard (ISO/TR 6579-3:2014) for typing of 59 Salmonella serovars. The full list of Salmonella serovars is given in the certificate and validation report, available on the MicroVal website under ‘issued certificates / confirmation methods’.

New Certificate for iQ-Check™ STEC VirX

In June 2022, MicroVal has approved the issuing of the certificate for Bio-Rad’s iQ-Check™ STEC VirX kit for real-time PCR detection of virulence genes in Shiga toxin-producing Escherichia coli. ADRIA Développement has carried out the validation study of this alternative method, based on ISO 16140-2:2016, for the scope of raw meat products (excluding poultry) and raw dairy products.

STEC outbreaks are commonly associated with the consumption of raw meat, particularly beef, but also with dairy products. A sample positive for both stx1/stx2 and eae targets typically requires further testing for the identification of the major E. coli serogroups. The iQ-Check STEC VirX Kit, based on a multiplex real-time PCR system, allows the detection of the stx1/stx2 and eae virulence genes within a few hours after microbiological enrichment.

The certificate and the accompanying validation report, number 2021LR96, can be found under ‘issued certificates / alternative methods’.