A PCR METHOD FOR THE DETECTION OF LISTERIA MONOCYTOGENES BASED ON A NOVEL TARGET SEQUENCE IDENTIFIED BY COMPARATIVE GENOMIC ANALYSIS

Authors


X. Shi, Department of Food Science, School of Agriculture and Biology, Shanghai Jiao Tong University, 800 Dong Chuan Road, Shanghai 200240, China. TEL: +86-21-3420-6616; FAX: +86-21-3420-6616; EMAIL: xmshi@sjtu.edu.cn

Abstract

ABSTRACT

Listeria monocytogenes is an opportunistic foodborne pathogen that is a serious health hazard worldwide. In this study, a polymerase chain reaction (PCR) assay was developed for the detection of L. monocytogenes using a novel species-specific target sequence (a region of lmo0754gene) identified by a comparative genomic approach. An internal amplification control was incorporated into this PCR system. The assay allowed amplification of a 331-bp fragment only from the genomic DNA of L. monocytogenes strains and not from other Listeria species, as well as some non-Listeria species. The detection limit of the PCR assay was 55 copies/PCR with L. monocytogenes genomic DNA. Applying this PCR assay to artificially contaminated milk samples, low levels of L. monocytogenes (1–10 cfu/mL of milk) were detected after 6–9 h incubation in selective culture enrichment (UVM1).

PRACTICAL APPLICATIONS

A novel species-specific target sequence was identified by a comparative genomic approach that offered a new molecular diagnostic marker for specific detection of Listeria monocytogenes. Using this target, a PCR assay with an internal amplification control was developed and shown to be specific, sensitive and applicable to the detection of L. monocytogenes from artificially contaminated foods. Moreover, this comparative genomic approach could also provide a new tool in mining unique targets for other bacteria with the increasing availability of public data of genome sequences.

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