title: Study of the bacterial diversity of foods: PCR-DGGE versus LH-PCR.
abstract: The present study compared two culture-independent methods, polymerase chain reaction denaturing gradient gel electrophoresis (PCR-DGGE) and length-heterogeneity polymerase chain reaction (LH-PCR), for their ability to reveal food bacterial microbiota. Total microbial DNA and RNA were extracted directly from fourteen fermented and unfermented foods, and domain A of the variable regions V1 and V2 of the 16S rRNA gene was analyzed through LH-PCR and PCR-DGGE. Finally, the outline of these analyses was compared with bacterial viable counts obtained after bacterial growth on suitable selective media. For the majority of the samples, RNA-based PCR-DGGE revealed species that the DNA-based PCR-DGGE was not able to highlight. When analyzing either DNA or RNA, LH-PCR identified several lactic acid bacteria (LAB) and coagulase negative cocci (CCN) species that were not identified by PCR-DGGE. This phenomenon was particularly evident in food samples with viable loads<5.0 Logcfug-1. Furthermore, LH-PCR was able to detect a higher number of peaks in the analyzed food matrices relative to species identified by PCR-DGGE. In light of these findings, it may be suggested that LH-PCR shows greater sensitivity than PCR-DGGE. However, PCR-DGGE detected some other species (LAB included) that were not detected by LH-PCR. Therefore, certain LH-PCR peaks not attributed to known species within the LH-PCR database could be solved by comparing them with species identified by PCR-DGGE. Overall, this study also showed that LH-PCR is a promising method for use in the food microbiology field, indicating the necessity to expand the LH-PCR database, which is based, up to now, mainly on LAB isolates from dairy products.
authors.affiliation: Dipartimento di Scienze Agrarie, Alimentari ed Ambientali, Università Politecnica delle Marche, Via Brecce Bianche, 60131 Ancona, Italy.
authors.forename: Cristiana
authors.initials: C
authors.lastname: Garofalo
authors.affiliation: Dipartimento di Scienze degli Alimenti, Università degli Studi di Parma, 43124 Parma, Italy.
authors.forename: Elena
authors.initials: E
authors.lastname: Bancalari
authors.affiliation: Dipartimento di Scienze Agrarie, Alimentari ed Ambientali, Università Politecnica delle Marche, Via Brecce Bianche, 60131 Ancona, Italy. Electronic address: v.milanovic@univpm.it.
authors.forename: Vesna
authors.initials: V
authors.lastname: Milanović
authors.affiliation: Dipartimento di Scienze Agrarie, Alimentari ed Ambientali, Università Politecnica delle Marche, Via Brecce Bianche, 60131 Ancona, Italy.
authors.forename: Federica
authors.initials: F
authors.lastname: Cardinali
authors.affiliation: Dipartimento di Scienze Agrarie, Alimentari ed Ambientali, Università Politecnica delle Marche, Via Brecce Bianche, 60131 Ancona, Italy.
authors.forename: Andrea
authors.initials: A
authors.lastname: Osimani
authors.affiliation: Dipartimento di Scienze degli Alimenti, Università degli Studi di Parma, 43124 Parma, Italy.
authors.forename: Maria Luisa Savo
authors.initials: MLS
authors.lastname: Sardaro
authors.affiliation: Dipartimento di Scienze degli Alimenti, Università degli Studi di Parma, 43124 Parma, Italy.
authors.forename: Benedetta
authors.initials: B
authors.lastname: Bottari
authors.affiliation: Dipartimento di Scienze degli Alimenti, Università degli Studi di Parma, 43124 Parma, Italy.
authors.forename: Valentina
authors.initials: V
authors.lastname: Bernini
authors.affiliation: Dipartimento di Scienze Agrarie, Alimentari ed Ambientali, Università Politecnica delle Marche, Via Brecce Bianche, 60131 Ancona, Italy.
authors.forename: Lucia
authors.initials: L
authors.lastname: Aquilanti
authors.affiliation: Dipartimento di Scienze Agrarie, Alimentari ed Ambientali, Università Politecnica delle Marche, Via Brecce Bianche, 60131 Ancona, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Clementi
authors.affiliation: Dipartimento di Scienze degli Alimenti, Università degli Studi di Parma, 43124 Parma, Italy.
authors.forename: Erasmo
authors.initials: E
authors.lastname: Neviani
authors.affiliation: Dipartimento di Scienze degli Alimenti, Università degli Studi di Parma, 43124 Parma, Italy.
authors.forename: Monica
authors.initials: M
authors.lastname: Gatti
mesh_terms: D001419:Bacteria / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D044822:Biodiversity; D004269:DNA, Bacterial / Q000235:genetics; D058645:Denaturing Gradient Gel Electrophoresis / Q000379:methods*; D005516:Food Microbiology; D016133:Polymerase Chain Reaction / Q000379:methods*; D012336:RNA, Ribosomal, 16S / Q000235:genetics
chemical_list: D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S
keywords: Bacterial microbiota; DNA; Fermented and unfermented foods; LH-PCR; PCR-DGGE; RNA