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A method to detect and quantify Phaeomoniella chlamydospora and Phaeoacremonium aleophilum DNA in grapevine-wood samples

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Abstract

Grapevines are sensitive to a wide range of fungal pathogens, including agents such as Phaeomoniella chlamydospora and Phaeoacremonium aleophilum that cause tracheomycosis. In the present study, a procedure for DNA extraction from grapevine woody tissue is first evaluated and shown to be suitable for quantitative analysis. Next, a multiplex real-time PCR method targeting the β-tubulin gene of the pathogens and the actin gene of plant material is developed and its quantitative capability is verified. This protocol was evaluated in inoculated grapevine-wood samples and in young vines from a nursery and was found to be reliable and highly specific. Results obtained from inoculated cuttings show that the fungal colonization process must be considered regardless of the wood phenotype. An analysis of samples of young vines from the nursery shows that a high rate of contamination occurs at the basis of plants and that this contamination is associated with low quantitative values. This finding provides evidence that in vine nurseries, these fungi may be efficient soil-borne pathogens.

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Acknowledgments

We would like to thank Pascal Lecomte, Gwenaëlle Comont and Philippe Larignon for providing isolates, and for their helpful informations on INRA UMR-SV collection strains. We thank Dr. Philippe Rolshausen for his helpful assistance on the improvement of this manuscript. This research was supported by Loire Vini Viti Distribution (LVVD, Z.A du Landreau, 49610 Mozé sur Louet, France), and “Agence Nationale de la Recherche et de la Technologie”.

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Author notes

  1. Jérôme Pouzoulet

    Present address: Department of Botany and Plant Sciences, University of California, Riverside, CA, 92521, USA

Authors and Affiliations

  1. Loire Viti Vini Distribution (LVVD), Z.A du Landreau, 49610, Mozé sur Louet, France

    Jérôme Pouzoulet & Xavier Besson

  2. INP‐Ecole d’Ingénieurs de PURPAN, Laboratoire d’Agrophysiologie, Equipe Vins Viticulture et Œnologie, Université de Toulouse, 75 Voie du TOEC, BP 57611, 31076, Toulouse Cedex 3, France

    Jérôme Pouzoulet, Nathalie Mailhac, Christel Couderc, Jean Daydé, Marie Lummerzheim & Alban Jacques

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  1. Jérôme Pouzoulet
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  2. Nathalie Mailhac
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Correspondence to Alban Jacques.

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Marie Lummerzheim and Alban Jacques contributed equally to this work.

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Pouzoulet, J., Mailhac, N., Couderc, C. et al. A method to detect and quantify Phaeomoniella chlamydospora and Phaeoacremonium aleophilum DNA in grapevine-wood samples. Appl Microbiol Biotechnol 97, 10163–10175 (2013). https://doi.org/10.1007/s00253-013-5299-6

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