High-resolution in situ genotyping of Legionella pneumophila populations in drinking water by multiple-locus variable-number tandem-repeat analysis using environmental DNA.
Cast your vote
You can rate an item by clicking the amount of stars they wish to award to this item.
When enough users have cast their vote on this item, the average rating will also be shown.
Your vote was cast
Thank you for your feedback
Thank you for your feedback
MetadataShow full item record
AbstractCentral to the understanding of infections by the waterborne pathogen Legionella pneumophila is its detection at the clonal level. Currently, multiple-locus variable-number tandem-repeat (VNTR) analysis (MLVA) of L. pneumophila isolates can be used as a tool for high-resolution genotyping. Since L. pneumophila is difficult to isolate, the isolation of outbreak strains often fails due to a viable but nonculturable (VBNC) state of the respective environmental population. Therefore, we developed a cultivation-independent approach to detect single clones in drinking water. This approach is based on the extraction of DNA from drinking water followed by PCR using a set of eight VNTR primer pairs necessary for MLVA genotyping of L. pneumophila. The PCR amplicons were analyzed by single-strand conformation polymorphism (SSCP) and capillary electrophoresis to obtain the respective MLVA profiles. Parallel to the high-resolution analysis, we used the same environmental DNA to quantify the number of L. pneumophila cells in drinking water using real-time PCR with 16S rRNA gene-targeted primers. We used a set of drinking water samples from a small-scale drinking water network to test our approach. With these samples we demonstrated that the developed approach was directly applicable to DNA obtained from drinking water. We were able to detect more L. pneumophila MLVA genotypes in drinking water than we could detect by isolation. Our approach could be a valuable tool to identify outbreak strains even after the outbreak has occurred and has the potential to be applied directly to clinical material.
CitationHigh-resolution in situ genotyping of Legionella pneumophila populations in drinking water by multiple-locus variable-number tandem-repeat analysis using environmental DNA. 2010, 76 (18):6186-95 Appl. Environ. Microbiol.
AffiliationDepartment of Vaccinology and Applied Microbiology, Helmholtz Centre for Infection Research (HZI), 38124 Braunschweig, Germany.
The following license files are associated with this item:
- Temperature-Dependent Growth Modeling of Environmental and Clinical Legionella pneumophila Multilocus Variable-Number Tandem-Repeat Analysis (MLVA) Genotypes.
- Authors: Sharaby Y, Rodríguez-Martínez S, Oks O, Pecellin M, Mizrahi H, Peretz A, Brettar I, Höfle MG, Halpern M
- Issue date: 2017 Apr 15
- Spatial distribution of Legionella pneumophila MLVA-genotypes in a drinking water system.
- Authors: Rodríguez-Martínez S, Sharaby Y, Pecellín M, Brettar I, Höfle M, Halpern M
- Issue date: 2015 Jun 15
- Virulence Traits of Environmental and Clinical Legionella pneumophila Multilocus Variable-Number Tandem-Repeat Analysis (MLVA) Genotypes.
- Authors: Sharaby Y, Rodríguez-Martínez S, Pecellin M, Sela R, Peretz A, Höfle MG, Halpern M, Brettar I
- Issue date: 2018 May 15
- Development of a genus-specific next generation sequencing approach for sensitive and quantitative determination of the Legionella microbiome in freshwater systems.
- Authors: Pereira RP, Peplies J, Brettar I, Höfle MG
- Issue date: 2017 Mar 31
- Identification of variable-number tandem-repeat (VNTR) sequences in Legionella pneumophila and development of an optimized multiple-locus VNTR analysis typing scheme.
- Authors: Pourcel C, Visca P, Afshar B, D'Arezzo S, Vergnaud G, Fry NK
- Issue date: 2007 Apr