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Molecular Profile of Phage Infection: A Novel Approach for the Characterization of Erwinia Phages through qPCR

Due to the emergence of antibiotic resistance, phage-mediated biocontrol has become an attractive alternative for pathogen management in agriculture. While the infection characteristics of many phages can be adequately described using plaque assays and optical density, the results from phages of the...

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Autores principales: Parcey, Michael, Gayder, Steven, Castle, Alan J., Svircev, Antonet M.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7014438/
https://www.ncbi.nlm.nih.gov/pubmed/31952282
http://dx.doi.org/10.3390/ijms21020553
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author Parcey, Michael
Gayder, Steven
Castle, Alan J.
Svircev, Antonet M.
author_facet Parcey, Michael
Gayder, Steven
Castle, Alan J.
Svircev, Antonet M.
author_sort Parcey, Michael
collection PubMed
description Due to the emergence of antibiotic resistance, phage-mediated biocontrol has become an attractive alternative for pathogen management in agriculture. While the infection characteristics of many phages can be adequately described using plaque assays and optical density, the results from phages of the apple pathogen Erwinia amylovora have low reproducibility with these techniques. Using quantitative real-time PCR (qPCR), the stage of the lytic cycle was determined through a combination of chloroform-based sampling, centrifugation, and DNase treatment. Monitoring the transition of phage genomes through the lytic cycle generates a molecular profile from which phage infection characteristics such as adsorption rate and burst size can be determined. To our knowledge, this is the first report of qPCR being used to determine these infection parameters. The characteristics of four different genera of Erwinia phages were determined. The phage ΦEa46-1-A1 was able to adsorb at a rate up to 6.6 times faster than ΦEa35-70 and ΦEa9-2. The low enrichment titer of ΦEa9-2 was shown to be due to the absence of lysis. The ΦEa46-1-A1 and ΦEa21-4 phages had the highest productivity, with burst sizes of 57 virions in 38 min and 185 virions in 98 min, respectively, suggesting these genera would make stronger candidates for the phage-mediated biocontrol of E. amylovora.
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spelling pubmed-70144382020-03-09 Molecular Profile of Phage Infection: A Novel Approach for the Characterization of Erwinia Phages through qPCR Parcey, Michael Gayder, Steven Castle, Alan J. Svircev, Antonet M. Int J Mol Sci Article Due to the emergence of antibiotic resistance, phage-mediated biocontrol has become an attractive alternative for pathogen management in agriculture. While the infection characteristics of many phages can be adequately described using plaque assays and optical density, the results from phages of the apple pathogen Erwinia amylovora have low reproducibility with these techniques. Using quantitative real-time PCR (qPCR), the stage of the lytic cycle was determined through a combination of chloroform-based sampling, centrifugation, and DNase treatment. Monitoring the transition of phage genomes through the lytic cycle generates a molecular profile from which phage infection characteristics such as adsorption rate and burst size can be determined. To our knowledge, this is the first report of qPCR being used to determine these infection parameters. The characteristics of four different genera of Erwinia phages were determined. The phage ΦEa46-1-A1 was able to adsorb at a rate up to 6.6 times faster than ΦEa35-70 and ΦEa9-2. The low enrichment titer of ΦEa9-2 was shown to be due to the absence of lysis. The ΦEa46-1-A1 and ΦEa21-4 phages had the highest productivity, with burst sizes of 57 virions in 38 min and 185 virions in 98 min, respectively, suggesting these genera would make stronger candidates for the phage-mediated biocontrol of E. amylovora. MDPI 2020-01-15 /pmc/articles/PMC7014438/ /pubmed/31952282 http://dx.doi.org/10.3390/ijms21020553 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Parcey, Michael
Gayder, Steven
Castle, Alan J.
Svircev, Antonet M.
Molecular Profile of Phage Infection: A Novel Approach for the Characterization of Erwinia Phages through qPCR
title Molecular Profile of Phage Infection: A Novel Approach for the Characterization of Erwinia Phages through qPCR
title_full Molecular Profile of Phage Infection: A Novel Approach for the Characterization of Erwinia Phages through qPCR
title_fullStr Molecular Profile of Phage Infection: A Novel Approach for the Characterization of Erwinia Phages through qPCR
title_full_unstemmed Molecular Profile of Phage Infection: A Novel Approach for the Characterization of Erwinia Phages through qPCR
title_short Molecular Profile of Phage Infection: A Novel Approach for the Characterization of Erwinia Phages through qPCR
title_sort molecular profile of phage infection: a novel approach for the characterization of erwinia phages through qpcr
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7014438/
https://www.ncbi.nlm.nih.gov/pubmed/31952282
http://dx.doi.org/10.3390/ijms21020553
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