title: Novel PCR-RFLP system based on rpoB gene for differentiation of Cronobacter species.
abstract: Bacteria from the genus Cronobacter are opportunistic foodborne pathogens that can cause severe infections. More rapid, cost-effective and reliable methods are still required for the species identification of Cronobacter spp. In this study, we present a novel PCR-RFLP-based method that uses a newly designed pair of primers for the PCR-amplification of a partial rpoB gene sequence (1635 bp). The amplified products of DNA from 80 Cronobacter strains were separately digested with three restriction endonucleases (Csp6I, HinP1I, MboI). Using the obtained restriction patterns, a PCR-RFLP identification system was created to enable differentiation between all seven currently-known Cronobacter species. The functionality of our method was successfully verified on real food samples. Moreover, the relationships between the Cronobacter species were determined via a phylogenetic tree created from the RFLP patterns.
authors.affiliation: Department of Biochemistry and Microbiology, Faculty of Food and Biochemical Technology, University of Chemistry and Technology Prague, Technická 3, 166 28 Prague, Czech Republic.
authors.forename: Jiří
authors.initials: J
authors.lastname: Vlach
authors.affiliation: Department of Biochemistry and Microbiology, Faculty of Food and Biochemical Technology, University of Chemistry and Technology Prague, Technická 3, 166 28 Prague, Czech Republic.
authors.forename: Barbora
authors.initials: B
authors.lastname: Javůrková
authors.affiliation: Department of Biochemistry and Microbiology, Faculty of Food and Biochemical Technology, University of Chemistry and Technology Prague, Technická 3, 166 28 Prague, Czech Republic.
authors.forename: Ludmila
authors.initials: L
authors.lastname: Karamonová
authors.affiliation: Department of Biochemistry and Microbiology, Faculty of Food and Biochemical Technology, University of Chemistry and Technology Prague, Technická 3, 166 28 Prague, Czech Republic. Electronic address: martina.blazkova@vscht.cz.
authors.forename: Martina
authors.initials: M
authors.lastname: Blažková
authors.affiliation: Department of Biochemistry and Microbiology, Faculty of Food and Biochemical Technology, University of Chemistry and Technology Prague, Technická 3, 166 28 Prague, Czech Republic.
authors.forename: Ladislav
authors.initials: L
authors.lastname: Fukal
mesh_terms: D059126:Cronobacter / Q000145:classification* / Q000235:genetics; D017931:DNA Primers; D004269:DNA, Bacterial; D012321:DNA-Directed RNA Polymerases / Q000235:genetics*; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D005506:Food Contamination / Q000032:analysis; D005516:Food Microbiology; D005798:Genes, Bacterial; D010802:Phylogeny; D016133:Polymerase Chain Reaction / Q000191:economics / Q000379:methods*; D012150:Polymorphism, Restriction Fragment Length; D017422:Sequence Analysis, DNA
chemical_list: D017931:DNA Primers; D004269:DNA, Bacterial; D012321:DNA-Directed RNA Polymerases; D015252:Deoxyribonucleases, Type II Site-Specific; C024405:GATC-specific type II deoxyribonucleases; C037130:GCGC-specific type II deoxyribonucleases; C040158:GTAC-specific type II deoxyribonucleases
keywords: Cronobacter spp.; PCR-RFLP; Species identification; rpoB gene