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Evaluation of a microfluidic chip system for preparation of bacterial DNA from swabs, air, and surface water samples

The detection of bacterial pathogens from complex sample matrices by PCR requires efficient DNA extraction. In this study, a protocol for extraction and purification of DNA from swabs, air, and water samples using a microfluidic chip system was established. The optimized protocol includes a combinat...

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Detalles Bibliográficos
Autores principales: Julich, Sandra, Hotzel, Helmut, Gärtner, Claudia, Trouchet, Daniel, Fawzy El Metwaly Ahmed, Marwa, Kemper, Nicole, Tomaso, Herbert
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Academic Press 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5119575/
https://www.ncbi.nlm.nih.gov/pubmed/27520284
http://dx.doi.org/10.1016/j.biologicals.2016.06.013
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author Julich, Sandra
Hotzel, Helmut
Gärtner, Claudia
Trouchet, Daniel
Fawzy El Metwaly Ahmed, Marwa
Kemper, Nicole
Tomaso, Herbert
author_facet Julich, Sandra
Hotzel, Helmut
Gärtner, Claudia
Trouchet, Daniel
Fawzy El Metwaly Ahmed, Marwa
Kemper, Nicole
Tomaso, Herbert
author_sort Julich, Sandra
collection PubMed
description The detection of bacterial pathogens from complex sample matrices by PCR requires efficient DNA extraction. In this study, a protocol for extraction and purification of DNA from swabs, air, and water samples using a microfluidic chip system was established. The optimized protocol includes a combination of thermal, chemical and enzymatic lysis followed by chip-based DNA purification using magnetic particles. The procedure was tested using Gram-positive Bacillus thuringiensis Berliner var. kurstaki as a model organism for Bacillus anthracis and the attenuated live vaccine strain of Francisella tularensis subsp. holarctica as Gram-negative bacterium. The detection limits corresponded to 10(3) genome equivalents per milliliter (GE/ml) for surface water samples spiked with F. tularensis and 10(2) GE/ml for B. thuringiensis. In air, 10 GE of F. tularensis per 10 L and 1 GE of B. thuringiensis per 10 L were detectable. For swab samples obtained from artificially contaminated surfaces the detection limits were 4 × 10(3) GE/cm(2) for F. tularensis and 4 × 10(2) GE/cm(2) for B. thuringiensis. Suitability of the chip-assisted procedure for DNA preparation of real samples was demonstrated using livestock samples. The presence of thermophilic Campylobacter spp. DNA could be confirmed in air samples collected on pig and broiler farms.
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spelling pubmed-51195752016-11-28 Evaluation of a microfluidic chip system for preparation of bacterial DNA from swabs, air, and surface water samples Julich, Sandra Hotzel, Helmut Gärtner, Claudia Trouchet, Daniel Fawzy El Metwaly Ahmed, Marwa Kemper, Nicole Tomaso, Herbert Biologicals Article The detection of bacterial pathogens from complex sample matrices by PCR requires efficient DNA extraction. In this study, a protocol for extraction and purification of DNA from swabs, air, and water samples using a microfluidic chip system was established. The optimized protocol includes a combination of thermal, chemical and enzymatic lysis followed by chip-based DNA purification using magnetic particles. The procedure was tested using Gram-positive Bacillus thuringiensis Berliner var. kurstaki as a model organism for Bacillus anthracis and the attenuated live vaccine strain of Francisella tularensis subsp. holarctica as Gram-negative bacterium. The detection limits corresponded to 10(3) genome equivalents per milliliter (GE/ml) for surface water samples spiked with F. tularensis and 10(2) GE/ml for B. thuringiensis. In air, 10 GE of F. tularensis per 10 L and 1 GE of B. thuringiensis per 10 L were detectable. For swab samples obtained from artificially contaminated surfaces the detection limits were 4 × 10(3) GE/cm(2) for F. tularensis and 4 × 10(2) GE/cm(2) for B. thuringiensis. Suitability of the chip-assisted procedure for DNA preparation of real samples was demonstrated using livestock samples. The presence of thermophilic Campylobacter spp. DNA could be confirmed in air samples collected on pig and broiler farms. Academic Press 2016-11 /pmc/articles/PMC5119575/ /pubmed/27520284 http://dx.doi.org/10.1016/j.biologicals.2016.06.013 Text en © 2016 The Author(s) http://creativecommons.org/licenses/by/4.0/ This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Julich, Sandra
Hotzel, Helmut
Gärtner, Claudia
Trouchet, Daniel
Fawzy El Metwaly Ahmed, Marwa
Kemper, Nicole
Tomaso, Herbert
Evaluation of a microfluidic chip system for preparation of bacterial DNA from swabs, air, and surface water samples
title Evaluation of a microfluidic chip system for preparation of bacterial DNA from swabs, air, and surface water samples
title_full Evaluation of a microfluidic chip system for preparation of bacterial DNA from swabs, air, and surface water samples
title_fullStr Evaluation of a microfluidic chip system for preparation of bacterial DNA from swabs, air, and surface water samples
title_full_unstemmed Evaluation of a microfluidic chip system for preparation of bacterial DNA from swabs, air, and surface water samples
title_short Evaluation of a microfluidic chip system for preparation of bacterial DNA from swabs, air, and surface water samples
title_sort evaluation of a microfluidic chip system for preparation of bacterial dna from swabs, air, and surface water samples
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5119575/
https://www.ncbi.nlm.nih.gov/pubmed/27520284
http://dx.doi.org/10.1016/j.biologicals.2016.06.013
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