The potential of a polyphasic PCR-dGGE approach in evaluating microbial diversity of natural whey cultures for water-buffalo Mozzarella cheese production: bias of culture-dependent and culture-independent analyses.
A polyphasic PCR-DGGE approach was used to describe the microbial population occurring in natural whey cultures (NWCs) for water-buffalo Mozzarella cheese production. Total microbial community was assessed without cultivation by analyzing DNA directly extracted from the original samples of NWC. In addition, DNA extracted from bulks of cells formed by harvesting colonies from the serial dilution agar plates of a variety of culture media was used to profile the "cultivable" community. The 16S rDNA V3 region was amplified using DNA from NWC as well as DNA from bulks as templates and the amplicons were separated by DGGE. The microbial entities occurring in NWCs were identified by partial 16S rDNA sequencing of DGGE bands: four lactic acid bacteria (LAB) closest relative of Streptococcus thermophilus, Lactococcus lactis, Lactobacillus delbrueckii and Lactobacillus crispatus were revealed by the analysis of DNA directly extracted from NWC while two other LAB, Lactobacillus fermentum and Enterococcus faecalis, were identified by analyzing DNA from the cultivable community. The developed PCR-DGGE analysis of the "cultivable" community showed good potential in evaluating microbial diversity of a dairy environment: it usefully highlighted the bias introduced by selective amplification when compared to the analysis of the total community from NWC and allowed suitability of media and growth conditions to be evaluated. Moreover, it could be used to complete the culture independent study of microbial diversity to give information on concentration ratios among species occurring in a particular environment and can be proposed for rapid identification of dominant microorganisms in alternative to traditional tools.
Ercolini
D
D
Dipartimento di Scienza degli Alimenti, Alimentare e Ambientale e di Igiene, Stazione di Microbiologia Industriale, Università degli Studi di Napoli Federico II, Portici, Italy.
Moschetti
G
G
Blaiotta
G
G
Coppola
S
S
D000818:Animals; D001483:Base Sequence; D002020:Buffaloes; D002611:Cheese / Q000382:microbiology*; D015169:Colony Count, Microbial; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D013293:Enterococcus faecalis / Q000235:genetics / Q000302:isolation & purification; D005516:Food Microbiology; D007558:Italy; D007778:Lactobacillus / Q000235:genetics / Q000302:isolation & purification; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012336:RNA, Ribosomal, 16S / Q000737:chemistry / Q000235:genetics; D017422:Sequence Analysis, DNA; D013291:Streptococcus / Q000235:genetics / Q000302:isolation & purification
D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S