Rapid Detection of Salmonella spp. in 375-Gram Sample Size of Chocolate Products

Introduction: Alternative rapid screening methods, especially PCR have been developed to detect earlier Salmonella contamination in chocolate products. But chocolate is a challenging food matrix as it has high lipid contents and contains polyphenols which inhibit PCR reactions. Usually, DNA purification is required for such products requiring expensive equipment or fastidious protocols. Dedicated simplified protocols for 375-g test portions of chocolate products (including raw materials) have been developed in order to detect Salmonella with the new detection method.

Purpose: This study reports an evaluation of a new detection method specific protocol for the rapid detection of Salmonella spp. in 375-g chocolate products.

Methods: Ninety-one artificially contaminated samples of 375 g each were tested with the new method. It consists in a single enrichment in BPW (20 hours at 41.5°C). After incubation step, DNA is extracted using mechanical lysis in a dedicated tube: 20 µl are introduced through a cap that does not require any tube or cap handling. Extracted DNA is used directly with freeze-dried PCR reagents. The PCR method is based on dual probe detection allowing real time detection and melting curve analysis. The call is positive when it combines an amplification curve and a melting peak allowing a strong specificity of the test.

Results: A total of 75/91 samples were detected positive and 16/91 were tested negative. All samples were confirmed with the ISO 6579 Reference method and no discordance were observed with the new method. The dedicated protocol presented in this study allows to remove PCR inhibitions for all samples tested.

Significance: The bioMérieux GENE-UP method for 375-g chocolate samples does not require any specific purification of DNA that would lead to fastidious or expensive protocols. The new method presented enables a reliable detection of Salmonella spp. in about 21 hours, allowing a rapid decision and cost manufacturing savings.