Detection and enumeration of Listeria monocytogenes in fresh cut vegetables using MPN-Real-Time PCR
Articolo
Data di Pubblicazione:
2015
Citazione:
Detection and enumeration of Listeria monocytogenes in fresh cut vegetables using MPN-Real-Time PCR / P. Russo, G. Botticella, M.L. Amodio, G. Colelli, M. Cavaiuolo, A. Ferrante, S. Massa, G. Spano, L. Beneduce. - In: ACTA HORTICULTURAE. - ISSN 0567-7572. - 1071(2015), pp. 567-574. ((Intervento presentato al 11. convegno International Controlled and Modified Atmosphere Research Conference tenutosi a Trani nel 2013 [10.17660/ActaHortic.2015.1071.73].
Abstract:
Listeria monocytogenes is a gram positive, rod shaped, pathogenic bacterium,
causative agent of a severe infection generally known as listeriosis. Packaging and
storage conditions of fresh cut vegetables may favour the growth of this
psychrotrophic pathogen leading to potential health threat. Detection and
enumeration of L. monocytogenes in concentrations up to 103 CFU/g, usually implies
use of the most-probable-number technique (MPN) which may take up to seven days
for verified identification of the pathogen. We developed a fast and reliable protocol
combining MPN with a Real-Time quantitative PCR (qPCR) approach. Samples of
fresh cut salads (25 g) purchased at local shops were spiked with 1 to 105 CFU/g of
L. monocytogenes. Samples were homogenized, and triplicate series of tubes
containing 10-5 to 10 g of food were incubated in Fraser broth at 30°C for 48 h for
standard MPN analysis. After incubation, broth samples were taken from each tube
and DNA was extracted. DNA from enrichment tubes was used as template in a
qPCR assay targeting a 64 bp hlyA gene sequence of L. monocytogenes. Results of
this assay were than compared with those of standard MPN analysis and a complete
accordance was observed. Furthermore, we tested an enrichment free approach
using the same qPCR assay. Samples were prepared as described for MPN-qPCR
while DNA extraction was performed prior to enrichment of inoculated salads. This
approach allowed us to identify L. monocytogenes in samples spiked with 10-105
CFU/g. The whole process, including DNA extraction, required less than four hours,
thus providing a fast and reliable tool for detection of L. monocytogenes in fresh cut
vegetables.
Tipologia IRIS:
01 - Articolo su periodico
Keywords:
fresh cut; Listeria monocytogenes; DNA extraction; MPN; qPCR
Elenco autori:
P. Russo, G. Botticella, M.L. Amodio, G. Colelli, M. Cavaiuolo, A. Ferrante, S. Massa, G. Spano, L. Beneduce
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