title: The fate of indigenous microbiota, starter cultures, Escherichia coli, Listeria innocua and Staphylococcus aureus in Danish raw milk and cheeses determined by pyrosequencing and quantitative real time (qRT)-PCR.
abstract: The purpose of this work was to study the bacterial communities in raw milk and in Danish raw milk cheeses using pyrosequencing of tagged amplicons of the V3 and V4 regions of the 16S rDNA and cDNA. Furthermore, the effects of acidification and ripening starter cultures, cooking temperatures and rate of acidification on survival of added Escherichia coli, Listeria innocua and Staphylococcus aureus in cheeses at different stages of ripening were studied by pyrosequencing and quantitative real time (qRT)-PCR. A high diversity of bacterial species was detected in raw milk. Lactococcus lactis, Streptococcus thermophilus, Lactobacillus casei and Lactobacillus rhamnosus were the main bacteria detected in raw milk and cheeses. Bacteria belonging to the genera Brevibacterium, Staphylococcus, Escherichia, Weissella, Leuconostoc, Pediococcus were also detected in both 16S rDNA and cDNA obtained from raw milk and cheeses. E. coli, which was added to milk used for production of some cheeses, was detected in both DNA and RNA extracted from cheeses at different stages of ripening showing the highest percentage of the total sequence reads at 7 days of ripening and decreased again in the later ripening stages. Growth of E. coli in cheeses appeared to be affected by the cooking temperature and the rate of acidification but not by the ripening starter cultures applied or the indigenous microbiota of raw milk. Growth of L. innocua and S. aureus added to milks was inhibited in all cheeses at different stages of ripening. The use of 16S rRNA gene pyrosequencing and qRT-PCR allows a deeper understanding of the behavior of indigenous microbiota, starter cultures and pathogenic bacteria in raw milk and cheeses.
authors.affiliation: Department of Food Science, Food Microbiology, University of Copenhagen, Rolighedsvej 30, DK-1958 Frederiksberg C, Denmark. wm@life.ku.dk
authors.forename: Wafa
authors.initials: W
authors.lastname: Masoud
authors.forename: Finn K
authors.initials: FK
authors.lastname: Vogensen
authors.forename: Søren
authors.initials: S
authors.lastname: Lillevang
authors.forename: Waleed
authors.initials: W
authors.lastname: Abu Al-Soud
authors.forename: Søren J
authors.initials: SJ
authors.lastname: Sørensen
authors.forename: Mogens
authors.initials: M
authors.lastname: Jakobsen
mesh_terms: D000818:Animals; D001419:Bacteria / Q000254:growth & development* / Q000302:isolation & purification; D002611:Cheese / Q000382:microbiology*; D004269:DNA, Bacterial / Q000302:isolation & purification; D003718:Denmark; D004926:Escherichia coli / Q000254:growth & development / Q000302:isolation & purification; D005516:Food Microbiology; D013294:Lactococcus lactis; D007934:Leuconostoc; D008087:Listeria / Q000254:growth & development / Q000302:isolation & purification; D054892:Metagenome; D008892:Milk / Q000382:microbiology*; D012329:RNA, Bacterial / Q000302:isolation & purification; D060888:Real-Time Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D013211:Staphylococcus aureus / Q000254:growth & development / Q000302:isolation & purification; D013696:Temperature
chemical_list: D004269:DNA, Bacterial; D012329:RNA, Bacterial