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Blue EaLAMP—a specific and sensitive method for visual detection of genomic Erwinia amylovora DNA

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Abstract

Fire blight caused by the gram-negative bacterium Erwinia amylovora is a major disease of pome fruit like apple (Malus domestica) or pear (Pyrus communis). Detection of the pathogen is hampered by low titres usually being present during initial flower infection and the brief time frame for analysis to decide upon subsequent countermeasures. Here we describe a loop-mediated isothermal amplification of DNA (LAMP) assay for genomic DNA of Erwinia amylovora, which relies on a highly specific primer design that excludes amplification from typical DNA sources of the orchard biological system and from sample handling. The assay enables the fast detection of down to approximately 20 cfu of pure Erwinia amylovora or 100 fg genomic DNA (corresponding to approximately 25 E. amyolovora equivalents) per reaction within 45 min. Fast and reliable detection of E. amylovora in orchard samples by naked eye is achieved through a visual colour change indication with hydroxynaphthol blue. The assay avoids use of complex technical devices and is thus suitable for field testing.

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Acknowledgments

We acknowledge all institutions that provided microbial strains or plant material for this study (listed in Table 1). The study was supported by The Austrian Federal Ministry of Agriculture, Forestry, Environment and Water Management (DaFNE Project No. 100404). Elizabeth Schroeder-Reiter (LMU Munich) is gratefully acknowledged for correcting the English text.

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Authors and Affiliations

  1. Institute of Chemical Engineering, Vienna University of Technology, Getreidemarkt 9, 1060, Vienna, Austria

    Christian Gosch, Karl Stich & Thilo Christopher Fischer

  2. Institute for Sustainable Plant Production, Dept. for Molecular Diagnostics of Plant Diseases, Austrian Agency for Health and Food Safety (AGES), Spargelfeldstraße 191, 1220, Vienna, Austria

    Richard Alexander Gottsberger

  3. Plant Biochemistry and Physiology, Ludwig-Maximilians-University Munich, Grosshadernerstr. 2-4, D-82152 Planegg-Martinsried, Planegg-Martinsried, Germany

    Thilo Christopher Fischer

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  1. Christian Gosch
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  2. Richard Alexander Gottsberger
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  3. Karl Stich
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  4. Thilo Christopher Fischer
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Correspondence to Thilo Christopher Fischer.

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Gosch, C., Gottsberger, R.A., Stich, K. et al. Blue EaLAMP—a specific and sensitive method for visual detection of genomic Erwinia amylovora DNA. Eur J Plant Pathol 134, 835–845 (2012). https://doi.org/10.1007/s10658-012-0059-5

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