Inhibition of crown gall induction by Agrobacterium vitis strain F2/5 in grapevine and Ricinus
Biological control measures to prevent or reduce Agrobacterium vitis-caused losses in grapevine cultures are a worldwide increasing challenge. In the present study, tumour development in grapevine (Vitis vinifera L.) was induced in the sensitive cv. Kerner by infection with Agrobacterium vitis strain K306, carrying the p35Sgus-int plasmid with the gus gene as marker for transformation by the wild-type T-DNA. Pre-inoculation with the non-tumorigenic A. vitis strain F2/5 prevented tumour induction by K306(p35gus-int). Strain M1154, a Tn5 mutant of F2/5 in the luxR-like aviR gene, partially reduced the biocontrol efficiency compared to the wild-type F2/5. GUS-labelling by K306gus was poor in grapevine in contrast to A. tumefaciens 28l(p35gusint)- induced tumours in Arabidopsis, indicating plant species-dependent variable gus expression. To use the more reliable direct mRNA expression assay by RTPCR, a new experimental plant/A. vitis system was established with Ricinus communis as model plant. Ricinus! A. vitis galls were available within one week after K306gus inoculation, reached diameters up to 5 cm, and contained more abundant GUS staining. An additional transformation marker, mRNA expression of the T-DNA-located iaaM oncogene, coding auxin synthesis, was apparent only in tumours induced by the wild-type A. vitis strain K306 in the absence of the gus construct, which is under the control of the strong 35S CaMV promoter. F2/5 pre-inoculation suppressed GUS staining and gus mRNA expression. DAPI staining revealed the loss of vital fluorescent cell nuclei in F2/5-inoculated grapevine tissue and thus inhibition of any successful T-DNA transfer into host cell nuclei. Differentiation of typical circular vessels in globular vascular bundles in M1154-pretreated galls suggests interference with plant auxin metabolism. In conclusion, together with successfully establishing a new experimental model system, Ricinus!A. vitis, pre-treatment of host tissue with the non-pathogenic strain F2/5 resulted in preventing the integration and expression of the oncogenic T-DNA of A. vitis strains by locally necrotizing host cell nuclei.