# Description and identification of the new phytopathogenic bacterium causing bacterial black spots on Corn salad (Valerianella locusta), as Acidovorax valerianellae

C. Grondeau, Régine Samson, Louis Gardan

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## To cite this version:

C. Grondeau, Régine Samson, Louis Gardan. Description and identification of the new phytopathogenic bacterium causing bacterial black spots on Corn salad (Valerianella locusta), as Acidovorax valerianellae. 10. International Conference of Plant Pathogenic Bacteria, Jul 2000, Charlottetown, Canada. pp.445. hal-01600999

HAL Id: hal-01600999

https://hal.science/hal-01600999

Submitted on 4 Jun 2020

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# Description and Identification of the new Phytopathogenic Bacterium causing Bacterial Black Spots on Corn Salad (*Valerianella locusta*), as *Acidovorax valerianellae*

Grondeau C., Samson R., Gardan L., Comité Départemental de Développement Maraîcher / Institut National de la Recherche Agronomique, 42 rue Georges Morel, BP 57, 49071 Beaucouzé cedex, FRANCE. Email: grondeau@angers.inra.fr

## Introduction

In France since 1985, corn salad (*Valerianella locusta*) production has been increasing by 820 t a year and now reaches nearly 15 000 t. This increase was followed by the outbreak of a new disease in the main production area. In 1991, a pathogenic bacterium was isolated from black and greasy leaf spots. Studies based on phenotypic and genetic characteristics have been carried out to describe this new bacterium first identified as a non-fluorescent *Pseudomonas* (1). As *Phoma valerianellae* is a fungus also responsible for black leaf spots on corn salad, a reliable method of diagnosis was needed to identify the disease in routine. Immunofluorescence performed on symptoms and double-diffusion on pure cultures were tested.

## Description of *Acidovorax valerianellae*

**Biochemical tests.** Numerical analysis of phenotypic characteristics indicated that corn salad strains were clustered in one discrete phenon related to *Acidovorax* reference strain and distantly related to *Pseudomonas*, *Ralstonia* and *Burkholderia* reference strains.

**DNA relatedness.** DNA hybridizations showed that corn salad strains were 84 to 100% similar to the type strain CFBP4730 and constituted a discrete genomospecies.

**16s rRNA gene sequence analysis.** Corn salad strains, branched in the vicinity of *Acidovorax facilis* and *A. anthurii*, a new pathogenic bacterium of *Anthurium*, belonged to the genus *Acidovorax*.

## Identification by Ouchterlony double-diffusion on pure cultures and path-test

Three hundred and ten strains isolated from corn salad which were non fluorescent, cytochrome c oxydase and HR on tobacco positive, levan sucrase and arginine dihydrolase negative, were inoculated by injection of a bacterial suspension in the oldest leaves of corn salad (Coquille de Louviers). Two hundred and fifty strains gave a brown and greasy necrosis of the infiltrated area within fifteen days, and were positive by double-diffusion with polyclonal antibodies. Sixty strains remained negative for the two tests.

Three strains isolated from water collected in pools near crops, that cross-reacted by double-diffusion, were negative to the pathogenicity test, and used
mannose, L+arabinose or glucose whereas the reference strain of *A.valerianellae* was negative for those three carbon sources.

**Direct diagnosis in spots by immunofluorescence**

From 1994 to 1998, immunofluorescence staining with polyclonal antibodies was performed directly on lesions cut in a drop of sterile water and taken on a large number of plants received at different times from different locations. The same drop was simultaneously streaked on King B medium and typical colonies of *Acidovorax* were identified by fluorescence, cytochrome c oxidase, levan sucrase, arginine dihydrolase, hypersensitivity on tobacco and double-diffusion. The two techniques gave the same result for more than 90% of the spots analysed.

**Conclusion**

Corn salad strains should be considered as a new phytopathogenic bacterium belonging to the β-subclass of Proteobacteria and the genus *Acidovorax*. The name *A. valerianellae* is proposed. In routine, *A. valerianellae* can be characterised by no fluorescent pigment, no levan sucrase and arginine dihydrolase, by the presence of a cytochrome c oxidase, by hypersensitivity on tobacco and double-diffusion positive. Double-diffusion can be replaced by pathogenicity test. Then, *A. valerianellae* was identified in Belgium (1998) and in Germany (2000).

**Reference**

1. Rat and Gardan, 1993. Fruits et Légumes 111, 42 - 43.