title: DNA-based methodologies for the quantification of live and dead cells in formulated biocontrol products based on Pantoea agglomerans CPA-2.
abstract: Pantoea agglomerans strain CPA-2 is an effective biocontrol agent (BCA) against the major postharvest pathogens present on pome and citrus fruits. Dehydration, such as freeze-drying, spray-drying and fluidized bed drying is one of the best ways to formulate BCAs. In this work, the survival of CPA-2 cells after formulation was determined by dilution plating and molecular methods as qPCR alone and combined with a sample pretreatment with a propidium monoazide dye (PMA-qPCR) and they were used to calculate treatment concentrations in efficacy trials on postharvest oranges. Furthermore, no significant differences in CPA-2 survival were observed as determined by dilution plating and PMA-qPCR after both the freeze drying and fluidized bed drying processes; however, an interesting significant difference was observed in the spray dried product comparing all quantitative methods. A difference of 0.48 and 2.17 log10 CFU or cells g/dw was observed among PMA-qPCR with qPCR and dilution plating, respectively. According to our study, dilution plating was shown to be an unreliable tool for monitoring the survival of CPA-2 after spray drying. In contrast, the combination of PMA and qPCR enabled a quick and unequivocal methodology to enumerate viable and VBNC CPA-2 cells under stress-dried conditions. Efficacy trials showed that, after 3 days, spray drying formulation rehydrated with 10% non-fat skimmed milk (NFSM) was as effective as fresh cells to control Penicillium digitatum in oranges.
authors.affiliation: Food Technology Department, Lleida University, XaRTA-Postharvest, Agrotecnio Center, Av. Rovira Roure, 191, 25198 Lleida, Catalonia, Spain.
authors.forename: Lourdes
authors.initials: L
authors.lastname: Soto-Muñoz
authors.affiliation: IRTA, XaRTA-Postharvest, Edifici Fruitcentre, Parc Científic i Tecnològic Agroalimentari de Lleida, Parc de Gardeny, 25003 Lleida, Catalonia, Spain.
authors.forename: Rosario
authors.initials: R
authors.lastname: Torres
authors.affiliation: IRTA, XaRTA-Postharvest, Edifici Fruitcentre, Parc Científic i Tecnològic Agroalimentari de Lleida, Parc de Gardeny, 25003 Lleida, Catalonia, Spain.
authors.forename: Josep
authors.initials: J
authors.lastname: Usall
authors.affiliation: Food Technology Department, Lleida University, XaRTA-Postharvest, Agrotecnio Center, Av. Rovira Roure, 191, 25198 Lleida, Catalonia, Spain.
authors.forename: Inmaculada
authors.initials: I
authors.lastname: Viñas
authors.affiliation: IRTA, XaRTA-Postharvest, Edifici Fruitcentre, Parc Científic i Tecnològic Agroalimentari de Lleida, Parc de Gardeny, 25003 Lleida, Catalonia, Spain.
authors.forename: Cristina
authors.initials: C
authors.lastname: Solsona
authors.affiliation: IRTA, XaRTA-Postharvest, Edifici Fruitcentre, Parc Científic i Tecnològic Agroalimentari de Lleida, Parc de Gardeny, 25003 Lleida, Catalonia, Spain. Electronic address: neus.teixido@irta.cat.
authors.forename: Neus
authors.initials: N
authors.lastname: Teixidó
mesh_terms: D001386:Azides / Q000494:pharmacology; D058491:Bacterial Load / Q000379:methods*; D002957:Citrus / Q000382:microbiology*; D005612:Freeze Drying; D056265:Microbial Interactions; D050296:Microbial Viability / Q000187:drug effects; D020636:Pantoea / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D010407:Penicillium / Q000502:physiology; D010572:Pest Control, Biological / Q000379:methods*; D011419:Propidium / Q000031:analogs & derivatives / Q000494:pharmacology; D060888:Real-Time Polymerase Chain Reaction
chemical_list: D001386:Azides; C533957:propidium monoazide; D011419:Propidium
keywords: Bed fluidized-drying; Dilution plating; Freeze-drying; PMA; Spray-drying; Survival; qPCR