Journal
FOOD MICROBIOLOGY
Volume 80, Issue -, Pages 1-8Publisher
ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD
DOI: 10.1016/j.fm.2018.12.009
Keywords
Listeria monocytogenes; PNA-FISH; Blocker probe; Enrichment procedure
Funding
- Fundo Europeu de Desenvolvimento Regional (FEDER) [POCI-01-0145-FEDER-006939, UID/EQU/00511/2013, POCI-01-0145-FEDER-006684, UID/BIO/04469]
- Coded-FISH through COMPETE2020 - Programa Operacional Competitividade e Internacionalizacao (POCI) [(PTDC/DTP-PIC/4562/2014) POCI-01-0145-FEDER-016678]
- FCT - Fundacao para a Ciencia e a Tecnologia [ENMed/0003/2014]
- North Portugal Regional Operational Programme (NORTE 2020), under the European Regional Development Fund (ERDF) [NORTE-01-0145-FEDER-000005, NORTE-01-0145-FEDER-000004]
- FCT Fundacao para a Ciencia e a Tecnologia [SFRH/BDE/51910/2012, SFRH/BPD/74480/2010]
- Fundação para a Ciência e a Tecnologia [SFRH/BDE/51910/2012, ENMed/0003/2014] Funding Source: FCT
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Listeria monocytogenes is one of the most important foodborne pathogens due to the high hospitalization and mortality rates associated to an outbreak. Several new molecular methods that accelerate the identification of L. monocytogenes have been developed, however conventional culture-based methods still remain the gold standard. In this work we developed a novel Peptide Nucleic Acid Fluorescence in situ Hybridization (PNA-FISH) method for the specific detection of L. monocytogenes. The method was based on an already existing PNA probe, LmPNA1253, coupled with a novel blocker probe in a 1:2 ratio. The method was optimized for the detection of L. monocytogenes in food samples through an evaluation of several rich and selective enrichment broths. The best outcome was achieved using One Broth Listeria in a two-step enrichment of 24 h plus 18 h. For validation in food samples, ground beef, ground pork, milk, lettuce and cooked shrimp were artificially contaminated with two ranges of inoculum: a low level (0.2-2 CFU/25 g or mL) and a high level (2-10 CFU/25 g or mL). The PNA-FISH method performed well in all types of food matrices, presenting an overall accuracy of 99% and a detection limit of 0.5 CFU/25 g or mL of food sample.
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