Specific detection of viable Salmonella Enteritidis by phage amplification combined with qPCR (PAA-qPCR) in spiked chicken meat samples

Serovar Enteritidis represents 45.7% of all Salmonella reported human cases identified in Europe. Additionally, minced meat and meat preparations from poultry have a high level of non-compliance, regarding Salmonella regulation. In the current study, a novel method based on the amplification of the Salmonella bacteriophage vB_SenS_PVP-SE2, coupled with real-time PCR (qPCR), was developed and evaluated, for the rapid detection of viable Salmonella Enteritidis in chicken samples. The results obtained indicated that the qPCR method could detect down to 0.22 fg/L of pure virus DNA and a concentration of viral particles of 103pfu/mL. After a short bacterial recovery step, the addition of bacteriophages to spiked chicken samples indicated that 8cfu/25g could be detected within 10h, including the time for DNA extraction and qPCR analysis. Additionally, the evaluation of the performance parameters: relative sensitivity, specificity, accuracy, positive and negative predictive values, and index kappa of concordance, obtained values higher than 92%, and the acceptability limit values were within the limits. All these results demonstrate that the proposed methodology is a powerful tool for the rapid detection of viable Salmonella Enteritidis.This work was supported by the project Nanotechnology Based Functional Solutions (NORTE-01-0145-FEDER-000019), supported by NortePortugal Regional Operational Programme(NORTE2020), andby the “NanoBioSensor: Desenvolvimento de nanosensores para avaliação da qualidade microbiológica de produtos à base de fruta” (POCI-010247-FEDER-033925), supported bytheOperational ThematicProgram for Competitiveness and Internationalization (POCI), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF).