title: Evaluation of propidium monoazide real-time PCR for enumeration of probiotic lactobacilli microencapsulated in calcium alginate beads.
abstract: The aim of the study was to evaluate real-time PCR coupled with propidium monoazide (PMA) treatment for enumeration of microencapsulated probiotic lactobacilli microencapsulated in calcium alginate beads. Lactobacillus gasseri K7 (CCM 7710) and Lactobacillus delbrueckii subsp. bulgaricus (CCM 7712) were analysed by plate counting and PMA real-time PCR during storage at 4 °C for 90 days. PMA was effective in preventing PCR amplification of the target sequences of DNA released from heat-compromised bacteria. The values obtained by real-time PCR of non-treated samples were in general higher than those obtained by real-time PCR of PMA-treated samples or by plate counting, indicating the presence of sub-lethally injured cells. This study shows that plate count could not be completely replaced by culture independent method PMA real-time PCR for enumeration of probiotics, but may rather complement the well-established plate counting, providing useful information about the ratio of compromised bacteria in the samples.
authors: 1 University of Ljubljana, Biotechnical Faculty, Department of Animal Science, Institute of Dairy Science and Probiotics, Groblje 3, 1230 Domžale, Slovenia.
authors: K
authors: K
authors: Oketič
authors: B Bogovič
authors: BB
authors: Matijašić
authors: T
authors: T
authors: Obermajer
authors: Z
authors: Z
authors: Radulović
authors: S
authors: S
authors: Lević
authors: N
authors: N
authors: Mirković
authors: V
authors: V
authors: Nedović
mesh_terms: D000464:Alginates; D001386:Azides / Q000378:metabolism*; D058491:Bacterial Load / Q000379:methods*; D018914:Cells, Immobilized; D004337:Drug Carriers; D004356:Drug Storage; D020723:Glucuronic Acid; D006603:Hexuronic Acids; D007778:Lactobacillus / Q000302:isolation & purification* / Q000378:metabolism; D019936:Probiotics; D011419:Propidium / Q000031:analogs & derivatives* / Q000378:metabolism; D011786:Quality Control; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D013194:Staining and Labeling / Q000379:methods*; D013678:Technology, Pharmaceutical / Q000379:methods; D013696:Temperature; D013997:Time Factors
chemical_list: D000464:Alginates; D001386:Azides; D004337:Drug Carriers; D006603:Hexuronic Acids; C533957:propidium monoazide; D011419:Propidium; D020723:Glucuronic Acid
keywords: Lactobacillus; calcium alginate beads; electrostatic droplet generation; propidium monoazide; viability real-time PCR