Viability Detection of Foodborne Bacterial Pathogens in Food Environment by PMA-qPCR and by Microscopic Observation
Foodborne pathogens are responsible of foodborne diseases and food poisoning and thus pose a great threat to food safety. These microorganisms can adhere to surface and form a biofilm composed of an extracellular matrix. This extracellular matrix protects bacterial cells from industrial environmental stress factors such as cleaning and disinfection operations. Moreover, during these environmental stresses, many bacterial species can enter a viable but nonculturable (VBNC) state. VBNC cells are characterized by a loss of cultivability on conventional bacteriological agar. This leads to an underestimation of total viable cells in environmental samples, and thus poses a risk for public health. In this chapter, we present a method to detect viable population of foodborne pathogens in industrial environmental samples using a molecular method with a combination of propidium monoazide (PMA) and quantitative PCR (qPCR) and a fluorescence microscopic method associated with the LIVE/DEAD BacLight™ viability stain.
Key wordsFoodborne Viable Microscopy Live/dead staining Propidium monoazide PMA-qPCR
Part of this work was supported by a grant from the CPER (Contrat de Plan Etat Région Nord-Pas de Calais, axe qualité et sécurité des ressources aquatiques).
- 7.Brauge T, Faille C, Sadovskaya I, Charbit A, Benezech T, Shen Y, Loessner MJ, Bautista JR, Midelet-Bourdin G (2018) The absence of N-acetylglucosamine in wall teichoic acids of Listeria monocytogenes modifies biofilm architecture and tolerance to rinsing and cleaning procedures. PLoS One 13(1):e0190879. https://doi.org/10.1371/journal.pone.0190879CrossRefPubMedPubMedCentralGoogle Scholar
- 12.Brauge T, Faille C, Inglebert G, Dubois T, Morieux P, Slomianny C, Midelet-Bourdin G (2018) Comparative evaluation of DNA extraction methods for amplification by qPCR of superficial vs intracellular DNA from Bacillus spores. Int J Food Microbiol 266:289–294. https://doi.org/10.1016/j.ijfoodmicro.2017.12.012CrossRefPubMedGoogle Scholar
- 13.Bustin SA, Benes V, Garson JA, Hellemans J, Huggett J, Kubista M, Mueller R, Nolan T, Pfaffl MW, Shipley GL, Vandesompele J, Wittwer CT (2009) The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments. Clin Chem 55(4):611–622. https://doi.org/10.1373/clinchem.2008.112797CrossRefPubMedGoogle Scholar