The aim of this study was the development of a new molecular assay for Pseudomonas aeruginosa identification in recreational water. The method includes bacterial cell concentration through membrane filtration, a short (6 h) culture enrichment step, DNA extraction and its amplification through a Real-Time PCR assay. The performance of the molecular approach was evaluated on 44 samples of swimming pool water and compared with the reference method UNI EN ISO 16266:2008. Positivity rates of 6% and 74% in pool and inlet water, respectively, with the standard culture method, and of 23% and 74% with the molecular method were found. Statistical analysis indicated ‘‘substantial agreement’’ (Cohen’s Kappa index: 0.6831) between the two approaches. RAPD typing of P. aeruginosa isolates showed identical fingerprint profiles, indicating their epidemiological correlation. The developed protocol showed very high specificity and a detection limit of 10 genomic units. This technique has the potential to screen large numbers of environmental samples, and could be proposed as part of a self-monitoring plan for recreational facilities, improving surveillance and early warning systems.
Molecular detection of Pseudomonas aeruginosa in recreational water
AMAGLIANI, GIULIA;BRANDI, GIORGIO;STOCCHI, VILBERTO;SCHIAVANO, GIUDITTA FIORELLA
2012
Abstract
The aim of this study was the development of a new molecular assay for Pseudomonas aeruginosa identification in recreational water. The method includes bacterial cell concentration through membrane filtration, a short (6 h) culture enrichment step, DNA extraction and its amplification through a Real-Time PCR assay. The performance of the molecular approach was evaluated on 44 samples of swimming pool water and compared with the reference method UNI EN ISO 16266:2008. Positivity rates of 6% and 74% in pool and inlet water, respectively, with the standard culture method, and of 23% and 74% with the molecular method were found. Statistical analysis indicated ‘‘substantial agreement’’ (Cohen’s Kappa index: 0.6831) between the two approaches. RAPD typing of P. aeruginosa isolates showed identical fingerprint profiles, indicating their epidemiological correlation. The developed protocol showed very high specificity and a detection limit of 10 genomic units. This technique has the potential to screen large numbers of environmental samples, and could be proposed as part of a self-monitoring plan for recreational facilities, improving surveillance and early warning systems.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
