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Quantification of Penicillium nalgiovense on Dry-Cured Sausage ‘Salchichón’ Using a SYBR Green-Based Real-Time PCR

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Abstract

To evaluate the effective implantation of a specific protective culture of Penicillium nalgiovense, a real-time quantitative PCR (qPCR) using SYBR Green methodology was developed. Two specific primers were designed on the basis of the published partial sequences of the Internal Transcribe Spacer (ITS)1–5.8S-ITS2 region of various strains of P. nalgiovense. Using the developed method, a PCR product of 51 bp with a T m value 81.34 °C was detected. T m values of the amplified product allowed specific differentiation between P. nalgiovense and the remaining mould species tested. The developed qPCR method was tested on inoculated slices of dry-cured sausage (‘salchichón’) showing an efficiency of 97.24 %, a R 2 value of 0.99 and a detection limit of P. nalgiovense of 1 log colony-forming units (cfu)/cm2. The qPCR method demonstrated that the protective strain of P. nalgiovense grew and competed against an ochratoxin A (OTA)-producing Penicillium verrucosum strain on commercial dry-cured sausage. This qPCR method provides a specific, accurate and sensitive detection and quantification of P. nalgiovense on dry-cured sausage salchichón in order to estimate its colonization during their processing. This assay will improve strategies to prevent and control unwanted mould colonization and OTA risk in dry-cured meat commodities.

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Acknowledgments

This work has been funded by the projects AGL2007-64639 and AGL2010-21623 of the Spanish Comisión Interministerial de Ciencia y Tecnología, Carnisenusa CSD2007-00016, Consolider Ingenio 2010 and GRU10162 of the Gobierno de Extremadura and FEDER. Dr. Alicia Rodríguez was the recipient of a grant from the University of Extremadura (Plan de Iniciación a la Investigación, Desarrollo Tecnológico e Innovación de la Universidad de Extremadura 2012, Becas Puente).

Conflict of Interest

M. Cordero, J.J. Córdoba, V. Bernáldez, M. Rodríguez and A. Rodríguez declare that they have no conflict of interest.

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Authors and Affiliations

  1. Food Hygiene and Safety, Meat and Meat Products Research Institute, Faculty of Veterinary Science, University of Extremadura, Cáceres, Spain

    Mirian Cordero, Juan J. Córdoba, Victoria Bernáldez, Mar Rodríguez & Alicia Rodríguez

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  1. Mirian Cordero
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  2. Juan J. Córdoba
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  3. Victoria Bernáldez
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  4. Mar Rodríguez
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  5. Alicia Rodríguez
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Correspondence to Alicia Rodríguez.

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Cordero, M., Córdoba, J.J., Bernáldez, V. et al. Quantification of Penicillium nalgiovense on Dry-Cured Sausage ‘Salchichón’ Using a SYBR Green-Based Real-Time PCR. Food Anal. Methods 8, 1582–1590 (2015). https://doi.org/10.1007/s12161-014-0078-2

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