Development and validation of a real-time quantitative PCR assay to detect Xanthomonas axonopodis pv. allii from onion seed

Robène Isabelle, Perret Marion, Jouen Emmanuel, Escalon Aline, Maillot Marie-Véronique, Chabirand Aude, Moreau Aurélie, Laurent Annie, Chiroleu Frédéric, Pruvost Olivier. 2015. Development and validation of a real-time quantitative PCR assay to detect Xanthomonas axonopodis pv. allii from onion seed. Journal of Microbiological Methods, 114 : 78-86.

https://doi.org/10.1016/j.mimet.2015.04.017

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Quartile : Q3, Sujet : BIOCHEMICAL RESEARCH METHODS / Quartile : Q3, Sujet : MICROBIOLOGY

Résumé : Bacterial blight of onion is an emerging disease threatening world onion production. The causal agent Xanthomonas axonopodis pv. allii is seed transmitted and a reliable and sensitive tool is needed to monitor seed exchanges. A triplex quantitative real-time PCR assay was developed targeting two X. axonopodis pv. allii-specific markers and an internal control chosen in 5.8S rRNA gene from Alliaceae. Amplification of at least one marker indicates the presence of the bacterium in seed extracts. This real-time PCR assay detected all the 79 X. axonopodis pv. allii strains tested and excluded 85.2% of the 135 non-target strains and particularly all 39 saprophytic and pathogenic bacteria associated with onion. Cross-reactions were mainly obtained for strains assigned to nine phylogenetically related X. axonopodis pathovars. The cycle cut-off was estimated statistically at 36.3 considering a risk of false positive of 1%. The limit of detection obtained in at least 95% of the time (LOD 95%) was 5 × 103 CFU/g (colony forming unit/g). The sensitivity threshold was found to be 1 infected seed in 32,790 seeds. This real-time PCR assay should be useful for preventing the long-distance spread of X. axonopodis pv. allii via contaminated seed lots and determining the epidemiology of the bacterium.

Mots-clés Agrovoc : Allium cepa, Xanthomonas, semence, diagnostic, PCR, Xanthomonas axonopodis

Mots-clés géographiques Agrovoc : La Réunion, France

Classification Agris : H20 - Maladies des plantes
F03 - Production et traitement des semences

Champ stratégique Cirad : Axe 4 (2014-2018) - Santé des animaux et des plantes

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