title: A multicenter study of viable PCR using propidium monoazide to detect Legionella in water samples.
abstract: Legionella quantification in environmental samples is overestimated by qPCR. Combination with a viable dye, such as Propidium monoazide (PMA), could make qPCR (named then vPCR) very reliable. In this multicentre study 717 artificial water samples, spiked with fixed concentrations of Legionella and interfering bacterial flora, were analysed by qPCR, vPCR and culture and data were compared by statistical analysis. A heat-treatment at 55 °C for 10 minutes was also performed to obtain viable and not-viable bacteria. When data of vPCR were compared with those of culture and qPCR, statistical analysis showed significant differences (P < 0.001). However, although the heat-treatment caused an abatement of CFU/mL ≤1 to 1 log10 unit, the comparison between untreated and heat-treated samples analysed by vPCR highlighted non-significant differences (P > 0.05). Overall this study provided a good experimental reproducibility of vPCR but also highlighted limits of PMA in the discriminating capability of dead and live bacteria, making vPCR not completely reliable.
authors.affiliation: Istituto Superiore di Sanità, Rome, Italy. Electronic address: maria.scaturro@iss.it.
authors.forename: Maria
authors.initials: M
authors.lastname: Scaturro
authors.affiliation: Istituto Superiore di Sanità, Rome, Italy.
authors.forename: Stefano
authors.initials: S
authors.lastname: Fontana
authors.affiliation: Laboratorio di Sanità Pubblica, Ospedale S. Chiara, Trento, Italy.
authors.forename: Italo
authors.initials: I
authors.lastname: Dell'eva
authors.affiliation: Laboratorio di Sanità Pubblica, Ospedale S. Chiara, Trento, Italy.
authors.forename: Fabrizia
authors.initials: F
authors.lastname: Helfer
authors.affiliation: Laboratorio di Sanità Pubblica, Ospedale S. Chiara, Trento, Italy.
authors.forename: Michele
authors.initials: M
authors.lastname: Marchio
authors.affiliation: Agenzia Regionale Protezione e Ambiente, Novara, Italy.
authors.forename: Maria Vittoria
authors.initials: MV
authors.lastname: Stefanetti
authors.affiliation: Agenzia Regionale Protezione e Ambiente, Novara, Italy.
authors.forename: Mario
authors.initials: M
authors.lastname: Cavallaro
authors.affiliation: Agenzia Regionale Protezione e Ambiente, Novara, Italy.
authors.forename: Marilena
authors.initials: M
authors.lastname: Miglietta
authors.affiliation: Università degli Studi di Bari, Bari, Italy.
authors.forename: Maria Teresa
authors.initials: MT
authors.lastname: Montagna
authors.affiliation: Università degli Studi di Bari, Bari, Italy.
authors.forename: Osvalda
authors.initials: O
authors.lastname: De Giglio
authors.affiliation: Università degli Studi di Bari, Bari, Italy.
authors.forename: Teresa
authors.initials: T
authors.lastname: Cuna
authors.affiliation: Agenzia Regionale Protezione e Ambiente, Bologna, Italy.
authors.forename: Leonarda
authors.initials: L
authors.lastname: Chetti
authors.affiliation: Agenzia Regionale Protezione e Ambiente, Bologna, Italy.
authors.forename: Maria Antonietta Bucci
authors.initials: MAB
authors.lastname: Sabattini
authors.affiliation: Agenzia Regionale Protezione e Ambiente, Bologna, Italy.
authors.forename: Michela
authors.initials: M
authors.lastname: Carlotti
authors.affiliation: Agenzia Regionale Protezione e Ambiente, Bologna, Italy.
authors.forename: Mariagabriella
authors.initials: M
authors.lastname: Viggiani
authors.affiliation: Agenzia Provinciale per l'Ambiente di Bolzano, Laives, Italy.
authors.forename: Alberta
authors.initials: A
authors.lastname: Stenico
authors.affiliation: Agenzia Provinciale per l'Ambiente di Bolzano, Laives, Italy.
authors.forename: Elisa
authors.initials: E
authors.lastname: Romanin
authors.affiliation: Laboratori Spa-ACEA, Roma, Italy.
authors.forename: Emma
authors.initials: E
authors.lastname: Bonanni
authors.affiliation: Laboratori Spa-ACEA, Roma, Italy.
authors.forename: Claudio
authors.initials: C
authors.lastname: Ottaviano
authors.affiliation: Ospedale Amedeo di Savoia, Torino, Italy.
authors.forename: Laura
authors.initials: L
authors.lastname: Franzin
authors.affiliation: Ospedale Amedeo di Savoia, Torino, Italy.
authors.forename: Claudio
authors.initials: C
authors.lastname: Avanzini
authors.affiliation: Ospedale Amedeo di Savoia, Torino, Italy.
authors.forename: Valerio
authors.initials: V
authors.lastname: Demarie
authors.affiliation: Fondazione IRCCS Policlinico S. Matteo, Pavia, Italy.
authors.forename: Marta
authors.initials: M
authors.lastname: Corbella
authors.affiliation: Fondazione IRCCS Policlinico S. Matteo, Pavia, Italy.
authors.forename: Patrizia
authors.initials: P
authors.lastname: Cambieri
authors.affiliation: Fondazione IRCCS Policlinico S. Matteo, Pavia, Italy.
authors.forename: Piero
authors.initials: P
authors.lastname: Marone
authors.affiliation: Istituto Superiore di Sanità, Rome, Italy.
authors.forename: Maria Cristina
authors.initials: MC
authors.lastname: Rota
authors.affiliation: Istituto Superiore di Sanità, Rome, Italy.
authors.forename: Antonino
authors.initials: A
authors.lastname: Bella
authors.affiliation: Istituto Superiore di Sanità, Rome, Italy.
authors.forename: Maria Luisa
authors.initials: ML
authors.lastname: Ricci
mesh_terms: D001386:Azides / Q000378:metabolism*; D001431:Bacteriological Techniques / Q000379:methods; D004791:Enzyme Inhibitors / Q000378:metabolism*; D006801:Humans; D007875:Legionella / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology*; D050296:Microbial Viability; D011419:Propidium / Q000031:analogs & derivatives* / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D013696:Temperature; D014871:Water Microbiology
chemical_list: D001386:Azides; D004791:Enzyme Inhibitors; C533957:propidium monoazide; D011419:Propidium
keywords: Culture; Legionella; Propidium monoazide; Quantitative PCR; Viable PCR