Specific Detection of Viable Legionella Cells by Combined Use of Photoactivated Ethidium Monoazide and PCR/Real-Time PCR

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Specific detection of viable Legionella cells by combined use of photoactivated ethidium monoazide and PCR/real-time PCR.

Legionella organisms are prevalent in manmade water systems and cause legionellosis in humans. A rapid detection method for viable Legionella cells combining ethidium monoazide (EMA) and PCR/real-time PCR was assessed. EMA could specifically intercalate and cleave the genomic DNA of heat- and chlorine-treated dead Legionella cells. The EMA-PCR assay clearly showed an amplified fragment specific...

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Comparison of ethidium monoazide and propidium monoazide for the selective detection of viable Legionella cells.

Ethidium monoazide (EMA) and propidium monoazide (PMA) have been utilized for selective PCR amplification of DNA from viable bacterial cells. In this study, we compared the abilities of EMA and PMA, together with real-time PCR, to specifically distinguish dead Legionella cells from viable cells. Several experiments showed that PMA or EMA treatment could specifically prevent the PCR amplificatio...

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Molecular Characterization of Viable Legionella spp. in Cooling Tower Water Samples by Combined Use of Ethidium Monoazide and PCR

Viable Legionella spp. in environmental water samples were characterized phylogenetically by a clone library analysis combining the use of ethidium monoazide and quantitative PCR. To examine the diversity of Legionella spp., six cooling tower water samples and three bath water samples were collected and analyzed. A total of 617 clones were analyzed for their 16S rRNA gene sequences and classifi...

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Quantification of viable Legionella pneumophila cells using propidium monoazide combined with quantitative PCR.

One of the greatest challenges of implementing fast molecular detection methods as part of Legionella surveillance systems is to limit detection to live cells. In this work, a protocol for sample treatment with propidium monoazide (PMA) in combination with quantitative PCR (qPCR) has been optimized and validated for L. pneumophila as an alternative of the currently used time-consuming culture m...

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Enumeration of viable Listeria monocytogenes cells by real-time PCR with propidium monoazide and ethidium monoazide in the presence of dead cells.

Propidium monoazide (PMA) and ethidium monoazide were used for enumeration of viable Listeria monocytogenes cells in the presence of dead cells. PMA had no antimicrobial effect on L. monocytogenes. Viable cell counts were linearly related to real-time PCR threshold cycle values for PMA-treated cells from planktonic and biofilm sources over a 4-log range.

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ژورنال

عنوان ژورنال: Applied and Environmental Microbiology

سال: 2009

ISSN: 0099-2240,1098-5336

DOI: 10.1128/aem.00604-08