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A loop-mediated isothermal amplification-based method for confirmation of Guignardia citricarpa in citrus black spot lesions

Lookup NU author(s): Professor Neil Boonham

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Abstract

Guignardia citricarpa Kiely (anamorph Phyllosticta citricarpa Van der Aa), the causal agent of citrus black spot disease, is subject to phytosanitary restrictions in the EU and USA, such that consignments of citrus are rejected at import if citrus black spot is identified on inspection. Due to the variability of black spot symptoms, positive identification solely on the basis of visual inspection is difficult, especially when lesions lack pycnidia (fruiting bodies of the anamorph Phyllosticta citricarpa). As an aid to visual inspection of symptoms, we have developed a method for detection of G. citricarpa using loop-mediated isothermal amplification (LAMP) which can be used to confirm the presence of G. citricarpa in black spot lesions, including those lacking pycnidia. The LAMP assay can be used to test crude extracts prepared directly from lesions on fruit, and the entire test can be completed in less than 40 min, making it faster than previously described PCR-based methods for detection of G. citricarpa. The method is sufficiently simple to allow deployment of the test in the field, for example in the course of import inspections. Recent years have seen the description of a number of newly recognised species in the genus Phyllosticta that are associated with citrus. As new species emerge, and the taxonomy of the genus is resolved, it will be important to periodically re-evaluate the performance of DNA-based methods for detection of G. citricarpa, including the LAMP assay described here, such that the accuracy of diagnosis can be assured. © 2013 UK Crown Copyright.

Publication metadata

Author(s): Tomlinson JA, Ostoja-Starzewska S, Webb K, Cole J, Barnes A, Dickinson M, Boonham N

Publication type: Article

Publication status: Published

Journal: European Journal of Plant Pathology

Year: 2013

Volume: 136

Issue: 2

Pages: 217-224

Print publication date: 01/06/2013

Online publication date: 25/01/2013

ISSN (print): 0929-1873

ISSN (electronic): 1573-8469

Publisher: Springer

URL: https://doi.org/10.1007/s10658-013-0168-9

DOI: 10.1007/s10658-013-0168-9

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