New Antagonistic Strains of Non‐Pathogenic Agrobacterium vitis to Control Grapevine Crown Gall

Abstract

Graft unions of nursery stock of grapevine (Vitis vinifera L.) collected in Japan yielded non‐pathogenic strains of Agrobacterium. On the basis of classic diagnostic tests, a sequence analysis and a previously reported multiplex PCR method, the non‐pathogenic strains ARK‐1, ARK‐2 and ARK‐3 were identified as Agrobacterium vitis. Stems of grapevine seedlings were inoculated with both a cell suspension of seven mixed strains of A. vitis (Ti) as a pathogen and one of a new strain or A. vitis strain VAR03‐1, one of the biological control agents against crown gall previously reported, as competitors to assay the suppression of tumour formation caused by the pathogen. In a test with a 1:1 cell ratio of pathogen/nonpathogen, strains ARK‐1, ARK‐2 and ARK‐3 reduced the tumour incidence.. In particular, strain ARK‐1 was strongest at inhibiting tumour formation in this study. Strain ARK‐1 established populations on roots of grapevine tree rootstock and persisted on roots for a year. ARK‐1, ARK‐2 and ARK‐3 did not produce a halo of inhibition against A. vitis (Ti) strain on YMA medium. Moreover, strain ARK‐1 did not reduce tumour incidence on the stems of grapevine when ARK‐1 was dead or only culture filtrate was used. This result indicates the possibility that these new strains inhibit grapevine crown gall in planta by a different mechanism other than VAR03‐1. In particular, one of the new strains, named ARK‐1, was most effective in inhibiting tumour formation on grapevine and appears to be a promising new agent to control grapevine crown gall.

Erratum to “Biological control for grapevine crown gall using nonpathogenic <i>Rhizobium vitis</i> strain ARK-1”

“…14)-16) Treatment using ARK-1 showed the highest control effect and reduced the incidence of galled plants by an average 85.2% of those on nontreated grapevine seedlings. 16) Thus, we verified that pre-inoculation of grapevine roots with ARK-1 was effective in reducing the percentage of plants with galls. These positive results encouraged us to conduct field trials on the assumption of the practical agricultural condition.…”

“…Each treatment consisted of a mixture of ARK-1 cells and an R. vitis (Ti) (tumorigenic) strain cells at 1 : 1 ratios, and these were inoculated onto stems of grapevine seedlings using the needleprick methods described above. 13),14), 16) Results showed that the live strain ARK-1 in a cell suspension significantly suppressed tumor incidence in stems compared to the control treatment using Ti strains (protective value F 89.1%). 16) On the other hand, the dead cells and CF suspension did not reduce tumor incidence (both protective values F 6.4%).…”

“…13),14), 16) Results showed that the live strain ARK-1 in a cell suspension significantly suppressed tumor incidence in stems compared to the control treatment using Ti strains (protective value F 89.1%). 16) On the other hand, the dead cells and CF suspension did not reduce tumor incidence (both protective values F 6.4%). 16) Basically, the mechanism of antagonistic microorganisms as biological control agents for plant disease involves bacteriocins or antibacterial materials.…”

“…13) Most of these strains were identified as nonpathogenic R. vitis using pathogenicity tests, bacteriological characteristics, specific multiplex polymerase chain reaction (PCR), and 16S rDNA sequencing analysis. 13), 16) Screening tests for biological control agents using nonpathogenic R. vitis by a needle-prick method were carried out, 13),14),16) with a total of 306 nonpathogenic strains to determine their activities against a pathogenic strain. Cell suspensions of the pathogenic and nonpathogenic strains were prepared and mixed in various combinations at cell ratios of 1 : 1.…”

“…Then, nonpathogenic R. vitis strains ARK-1, ARK-2, ARK-3, and VAR03-1 were selected as biological control agents against grapevine crown gall. 16) There are at least five genotypes in R. vitis (Ti) based on multilocus sequencing analysis of housekeeping genes pyrG, recA, and rpoD. 23) On grapevine seedlings, a 1 : 1 cell ratio of each strain ARK-1, ARK-2, ARK-3, or VAR03-1 to the mixture of seven different R. vitis (Ti) strains belonging to five different genotypes, which were isolated from Japan, Australia, and Greece, significantly suppressed tumor incidence in stems.…”