Selective Quantification of Erwinia amylovora Live Cells in Pome Fruit Tree Cankers by Viability Digital PCR
Abstract
The accurate assessment of Erwinia amylovora live cell populations in fire blight cankers by classic microbiology methods has major limitations. Some of them are the presence of competitive microbiota in samples that inhibit E. amylovora’s growth and the release of toxic compounds by plant material during sample processing, which may hamper the pathogen’s ability to form colonies on solid media. Digital PCR (dPCR) combined with the photo-reactive DNA-binding dye propidium monoazide (PMA) allows selective detection and quantification of live E. amylovora cells in woody samples while overcoming the constraints of culture-dependent methods. This work describes a reliable viability dPCR procedure to determine E. amylovora live cell concentrations in fire blight cankers from pome fruit trees. This protocol can be adapted for the analysis of other types of plant material and enables investigation of ecological, epidemiological, and management significance of cankers as a relatively underexpl...ored part of the fire blight disease cycle.
Keywords:
Fire blight / Cankers / Bacterial survival / Bacterial viability / Propidium monoazide / dPCR / Molecular detection methods / Apple / Pear / Asian pearSource:
Plant Pathology, Part of the Methods in Molecular Biology book series (MIMB,volume 2536), 2022, 231-249Publisher:
- Springer
Funding / projects:
- National Institute of Food and Agriculture, U.S. Department of Agriculture, Hatch/Multistate Research Program projects NYG-625835 and NYG-625857 under 1009897 and 1014444
- NY State Farm Viability Institute grant number 81927/A001-FVI 17 006
- NY State Specialty Crop Block Grant Program grant number SCG 82535/A001-SCG 17 005 to SGA
- Ministry of Education, Science and Technological Development, Republic of Serbia, Grant no. 200010 (Institute for Plant Protection and Environment, Belgrade) (RS-200010)
DOI: 10.1007/978-1-0716-2517-0_14
ISBN: 9781071625163
ISSN: 1064-3745
Scopus: 2-s2.0-85134427071
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Institution/Community
IZBISTY - CHAP AU - Delgado Santander, Ricardo AU - Gašić, Katarina AU - Aćimović, Srđan PY - 2022 UR - https://plantarum.izbis.bg.ac.rs/handle/123456789/765 AB - The accurate assessment of Erwinia amylovora live cell populations in fire blight cankers by classic microbiology methods has major limitations. Some of them are the presence of competitive microbiota in samples that inhibit E. amylovora’s growth and the release of toxic compounds by plant material during sample processing, which may hamper the pathogen’s ability to form colonies on solid media. Digital PCR (dPCR) combined with the photo-reactive DNA-binding dye propidium monoazide (PMA) allows selective detection and quantification of live E. amylovora cells in woody samples while overcoming the constraints of culture-dependent methods. This work describes a reliable viability dPCR procedure to determine E. amylovora live cell concentrations in fire blight cankers from pome fruit trees. This protocol can be adapted for the analysis of other types of plant material and enables investigation of ecological, epidemiological, and management significance of cankers as a relatively underexplored part of the fire blight disease cycle. PB - Springer T2 - Plant Pathology, Part of the Methods in Molecular Biology book series (MIMB,volume 2536) T1 - Selective Quantification of Erwinia amylovora Live Cells in Pome Fruit Tree Cankers by Viability Digital PCR EP - 249 SP - 231 DO - 10.1007/978-1-0716-2517-0_14 ER -
@inbook{ author = "Delgado Santander, Ricardo and Gašić, Katarina and Aćimović, Srđan", year = "2022", abstract = "The accurate assessment of Erwinia amylovora live cell populations in fire blight cankers by classic microbiology methods has major limitations. Some of them are the presence of competitive microbiota in samples that inhibit E. amylovora’s growth and the release of toxic compounds by plant material during sample processing, which may hamper the pathogen’s ability to form colonies on solid media. Digital PCR (dPCR) combined with the photo-reactive DNA-binding dye propidium monoazide (PMA) allows selective detection and quantification of live E. amylovora cells in woody samples while overcoming the constraints of culture-dependent methods. This work describes a reliable viability dPCR procedure to determine E. amylovora live cell concentrations in fire blight cankers from pome fruit trees. This protocol can be adapted for the analysis of other types of plant material and enables investigation of ecological, epidemiological, and management significance of cankers as a relatively underexplored part of the fire blight disease cycle.", publisher = "Springer", journal = "Plant Pathology, Part of the Methods in Molecular Biology book series (MIMB,volume 2536)", booktitle = "Selective Quantification of Erwinia amylovora Live Cells in Pome Fruit Tree Cankers by Viability Digital PCR", pages = "249-231", doi = "10.1007/978-1-0716-2517-0_14" }
Delgado Santander, R., Gašić, K.,& Aćimović, S.. (2022). Selective Quantification of Erwinia amylovora Live Cells in Pome Fruit Tree Cankers by Viability Digital PCR. in Plant Pathology, Part of the Methods in Molecular Biology book series (MIMB,volume 2536) Springer., 231-249. https://doi.org/10.1007/978-1-0716-2517-0_14
Delgado Santander R, Gašić K, Aćimović S. Selective Quantification of Erwinia amylovora Live Cells in Pome Fruit Tree Cankers by Viability Digital PCR. in Plant Pathology, Part of the Methods in Molecular Biology book series (MIMB,volume 2536). 2022;:231-249. doi:10.1007/978-1-0716-2517-0_14 .
Delgado Santander, Ricardo, Gašić, Katarina, Aćimović, Srđan, "Selective Quantification of Erwinia amylovora Live Cells in Pome Fruit Tree Cankers by Viability Digital PCR" in Plant Pathology, Part of the Methods in Molecular Biology book series (MIMB,volume 2536) (2022):231-249, https://doi.org/10.1007/978-1-0716-2517-0_14 . .