Occurrence of crown gall of the grapevine in Tunisia and characterization of Tunisian Agrobacterium vitis and A. tumefaciens strains.

摘要

The occurrence of crown gall disease in Tunisian vineyards was investigated. Crown gall symptoms were detected in several vineyards in the northern and central regions of the country but with low incidence (9.72%). Different Agrobacterium species were isolated and characterized. Morphological characterization, molecular assays and pathogenicity tests were conducted to identify Agrobacterium isolates. A set of 380 isolates with morphologically Agrobacterium-like colonies on MG medium was obtained from different parts of diseased vines and rhizospheric soils. Eighty out of them were chosen for detailed study in comparison with Agrobacterium reference strains obtained from the French CFBP collection. Biovars 1, 2 and 3 were first distinguished on the three semi-selective media 1A, 2E and 3AN respectively. Further, the isolates were differentiated by BOX-PCR analysis. It revealed that 11 Tunisian isolates could be identified as A. tumefaciens and four as A. vitis. The assignment of six A. tumefaciens strains to genomic species was achieved using the housekeeping gene recA. Results confirmed those of the BOX-PCR analysis and showed 99% similarity of three strains with the A. tumefaciens G4 strain (CFBP7128) and a comparable similarity (99%) of two strains was recorded with the A. tumefaciens G4 strain (CFBP7126). A non pathogenic strain was identified as A. tumefaciens G7 with 98% similarity to CFBP 7129. Two strains were confirmed to belong to the A. vitis by analysis of their 16S genes. The virulence and tumourigenicity of these Tunisian isolates were analyzed by vir-PCR and pathogenicity tests on carrot disks and indicator plants. The study revealed a relatively low percentage of highly tumourigenic isolates in Tunisia. Only 16% of the 300 isolates tested on carrot disks proved to be highly tumourigenic (++) and to form typical tumours around the cambium of carrot slices, whereas 36%, induced not clearly discernible tumours (+). The rest of the strains (48%) were negative (-). Of the 80 isolates tested on indicator plants, only two elicited tumours on tomato and tobacco plants. The vir-PCR test of 10 strains revealed the pathogenicity of five of them.