title: Use of propidium monoazide for the enumeration of viable Oenococcus oeni in must and wine by quantitative PCR.
abstract: Malolactic fermentation is an important step in winemaking, but it has to be avoided in some cases. It's carried out by lactic acid bacteria belonging mainly to the genus Oenococcus, which is known to be a slow growing bacterium. Classical microbiological methods to enumerate viable cells of Oenococcus oeni in must and wine take 7-9 days to give results. Moreover, RT-qPCR technique gives accurate quantitative results, but it requires time consuming steps of RNA extraction and reverse transcription. In the present work we developed a fast and reliable quantitative PCR (qPCR) method to enumerate cells of Oenococcus oeni, directly, in must and wine. For the first time we used a propidium monoazide treatment of samples to enumerate only Oenococcus oeni viable cells. The detection limit of the developed method is 0.33 log CFU/mL (2.14 CFU/mL) in must, and 0.69 log CFU/mL (4.90 CFU/mL) in wine, lower than that of the previously developed qPCR protocols.
authors.affiliation: Dipartimento di Scienze degli Alimenti, Università degli Studi di Udine, via Sondrio 2/A, 33100 Udine, Italy.
authors.forename: Marco
authors.initials: M
authors.lastname: Vendrame
authors.forename: Lucilla
authors.initials: L
authors.lastname: Iacumin
authors.forename: Marisa
authors.initials: M
authors.lastname: Manzano
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Comi
mesh_terms: D001386:Azides / Q000737:chemistry*; D015169:Colony Count, Microbial / Q000379:methods; D017931:DNA Primers; D004269:DNA, Bacterial / Q000302:isolation & purification; D005285:Fermentation; D005506:Food Contamination / Q000032:analysis; D005516:Food Microbiology; D056585:Oenococcus / Q000302:isolation & purification*; D011419:Propidium / Q000031:analogs & derivatives* / Q000737:chemistry; D012329:RNA, Bacterial / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014920:Wine / Q000382:microbiology*
chemical_list: D001386:Azides; D017931:DNA Primers; D004269:DNA, Bacterial; D012329:RNA, Bacterial; C533957:propidium monoazide; D011419:Propidium