Legionella spp. survival after different disinfection procedures: Comparison between conventional culture, qPCR and EMA–qPCR

Mansi, Antonella; Amori, I; Marchesi, Isabella; Marcelloni, Anna Maria; Proietto, Anna Rita; Ferranti, Greta; Magini, V.; Valeriani, Federica; Borella, Paola

doi:10.1016/j.microc.2013.09.017

Cited by 28 publications

(24 citation statements)

References 29 publications

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“…These intercalating dyes are intended to enter damaged cell membranes and covalently bind to DNA after photo activation, preventing downstream PCR amplification of DNA from membrane-compromised cells. Several studies have indicated promise of an EMA/PMA-based qPCR method for Legionella (Adela Yanez et al, 2011; Chen and Chang, 2010; Delgado-Viscogliosi et al, 2009; Inoue et al, 2015; Mansi et al, 2014; Slimani et al, 2012), mycobacteria (Lee et al, 2015; Nocker et al, 2007), P. aeruginosa (Gensberger et al, 2013, 2014), and Acanthamoeba (Chang et al, 2013) cells by testing against heat-treated, chlorine-treated cells and/or environmental water samples (e.g. drinking water, spa water, swimming pool water) as controls.…”

Section: Detection Methodsmentioning

confidence: 99%

Methodological approaches for monitoring opportunistic pathogens in premise plumbing: A review

et al. 2017

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Opportunistic pathogens inhabiting premise (i.e., building) plumbing (OPPPs, e.g., L. pneumophila, M. avium complex, P. aeruginosa, Acanthamoeba, and N. fowleri) are a significant and growing source of disease. Because OPPPs establish and grow as part of the native drinking water microbiota, they do not correspond to fecal indicators, presenting a significant challenge to common and effective monitoring strategies. Further, different OPPPs present distinct requirements for sampling, preservation, and analysis, creating a significant impediment to their parallel detection. The aim of this critical review is to synthesize the state of the science of monitoring OPPPs and to identify a path forward for their simultaneous detection and quantification in a manner commensurate with the need for reliable data to inform risk assessment and mitigation. Water and biofilm sampling procedures, as well as factors influencing sample representativeness and detection sensitivity, are critically evaluated with respect to the five representative bacterial and amoebal OPPPs noted above. Available culturing and molecular approaches are discussed in terms of their advantages, limitations, and applicability. Knowledge gaps and research needs are identified.

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Section: Detection Methodsmentioning

confidence: 99%

Methodological approaches for monitoring opportunistic pathogens in premise plumbing: A review

et al. 2017

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“…The results were expressed as numbers of CFU per liter, and the limit of detection (LOD) of the procedure was 25 CFU liter Ϫ1 . DNA was extracted by use of a QIAamp DNA minikit (Qiagen, Hilden, Germany) according to the manufacturer's instructions, as reported elsewhere (19). The extracted DNA was stored at Ϫ20°C until use.…”

Section: Methodsmentioning

confidence: 99%

“…For each treatment, the first 22 negative samples with culture (total of 88 samples) were analyzed by qPCR, with and without the EMA pretreatment. The water samples were treated with EMA (Sigma Chemical Co., St. Louis, MO) at a final concentration of 6 M (2.53 g ml Ϫ1 ) prior to DNA extraction, as reported by Mansi et al (19).…”

Section: Methodsmentioning

confidence: 99%

“…In order to overcome this problem, new PCR-based strategies, collectively called molecular viability analyses, have been developed (18). Among these, methods based on DNA-intercalating dyes, such as ethidium monoazide (EMA), have been proposed in combination with qPCR (19)(20)(21). EMA selectively binds the DNA of cells with compromised membranes, while intact cell membranes represent a barrier for the dye.…”

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confidence: 99%

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Control of Legionella Contamination and Risk of Corrosion in Hospital Water Networks following Various Disinfection Procedures

Marchesi

Mansi

Marcelloni

et al. 2016

Appl Environ Microbiol

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bPhysical and chemical disinfection methods have been proposed with the aim of controlling Legionella water contamination. To date, the most effective procedures for reducing bacterial contamination have not yet been defined. The aim of this study was to assess the long-term effectiveness of various disinfection procedures in order to reduce both culturable and nonculturable (NC) legionellae in different hospital water networks treated with heat, chlorine dioxide, monochloramine, and hydrogen peroxide. The temperature levels and biocide concentrations that proved to give reliable results were analyzed. In order to study the possible effects on the water pipes, we verified the extent of corrosion on experimental coupons after applying each method for 6 months. The percentage of positive points was at its lowest after treatment with monochloramine, followed by chlorine dioxide, hydrogen peroxide, and hyperthermia. Different selections of Legionella spp. were observed, as networks treated with chlorinebased disinfectants were contaminated mainly by Legionella pneumophila serogroup 1, hyperthermia was associated with serogroups 2 to 14, and hydrogen peroxide treatment was associated mainly with non-pneumophila species. NC cells were detected only in heat-treated waters, and also when the temperature was approximately 60°C. The corrosion rates of the coupons were within a satisfactory limit for water networks, but the morphologies differed. We confirm here that chemical disinfection controls Legionella colonization more effectively than hyperthermia does. Monochloramine was the most effective treatment, while hydrogen peroxide may be a promising alternative to chlorine-based disinfectants due to its ability to select for other, less virulent or nonpathogenic species.

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“…Despite these shortcomings, culture remains the method of choice. Other methods, such as qRT-PCR, can yield faster results and are highly sensitive and specific (16,17). However, qRT-PCR has the disadvantage that it cannot differentiate between live and dead bacteria, therefore it may yield false high counts (15).…”

Section: Discussionmentioning

confidence: 99%

First study in Qatar to reveal high Legionella counts in cooling towers

AbuOdeh

Aziz

Moussa

et al. 2017

East Mediterr Health J

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Legionella spp. is transmitted from water to humans by aerosol-generating devices, including cooling towers (CTs). There have not been published reports about Legionella in these systems in Qatar. Ten CTs in Qatar University were sampled on a monthly basis. Bacteria were recovered from 90 water samples by filtration and concentration. Legionella DNA copy number (CN) was assessed by quantitative RT-PCR. Legionella DNA was detected in 100% of the samples. The bacterial counts ranged from 0.006 to 199.56 CFU/mL, and critical counts were found in 51 (56.7 %) samples. Moreover, 7 (7.8%) samples showed a count of more than 100 CFU/mL. The highest counts were found in the months of May and June. These results suggest that this organism is found in high number in tested CTs, presenting a potential health risk to the local population.https://doi.org/10.26719/2017.23.10.703Première étude au Qatar pour mettre en évidence la forte présence de légionelles dans les tours de refroidissement RÉSUMÉ Les Legionella spp se transmettent de l'eau à l'homme par les dispositifs générateurs d'aérosols, notamment les tours de refroidissement. Aucun rapport n'a été publié sur la présence de légionelles dans ces systèmes au Qatar. Des prélèvements mensuels ont été effectués dans dix tours de refroidissement de l'Université du Qatar. Des bactéries ont été retrouvées dans 90 échantillons d'eau par filtration et concentration. Le nombre de copies de l'ADN des Legionella a été évalué par PCR quantitative en temps réel. L'ADN des Legionella a été détecté dans 100 % des échantillons. La numération bactérienne était comprise entre 0,006 et 199,56 CFU/mL et des numérations critiques ont été constatées dans 51 échantillons (56,7 %). En outre, 7 échantillons (7,8 %) présentaient une numération supérieure à 100 CFU/mL. Les numérations les plus élevées ont été relevées aux mois de mai et de juin. Ces résultats semblent indiquer que cet organisme est présent en grand nombre dans les tours de refroidissement ayant fait l'objet de prélèvements, ce qui constitue un risque sanitaire potentiel pour la population locale.

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Legionella spp. survival after different disinfection procedures: Comparison between conventional culture, qPCR and EMA–qPCR

Cited by 28 publications

References 29 publications

Methodological approaches for monitoring opportunistic pathogens in premise plumbing: A review

Methodological approaches for monitoring opportunistic pathogens in premise plumbing: A review

Control of Legionella Contamination and Risk of Corrosion in Hospital Water Networks following Various Disinfection Procedures

First study in Qatar to reveal high Legionella counts in cooling towers

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