Grapevine crown gall causes significant economic damage in vineyards and nurseries worldwide. Infected vines are not curable due to irreversible genomic transformation by the tumorigenic bacterium Rhizobium vitis. Crown gall results in a gradual decline in yield and vine vigor, then the complete collapse of the vine. R. vitis ARK-1, an antagonistic and non-tumorigenic strain, has been shown to inhibit gall formation when equal cell concentrations of antagonist and tumorigenic strains are co-inoculated in planta. In this study, we tested the efficacy of ARK-1 against higher cell numbers of tumorigenic isolates and evaluated timing and methods of ARK-1 application in in-planta assays in grapevine (Vitis vinifera 'Chardonnay', 'Cabernet Sauvignon', 'Merlot') and tomato (Solanum lycopersicum 'Beefsteak'). ARK-1 significantly (P ≤ 0.01) reduced gall incidence and gall diameter against four times higher tumorigenic bacterial cell numbers. Inoculation of ARK-1 up to 48 hours before or within 6 hours after inoculation with tumorigenic isolates significantly (P ≤ 0.01) reduced gall incidence and gall diameter. Root dipping of grapevine for 24 hours and tomato for 1 hour in the formulated ARK-1 suspension (Kumiai Chemical Industry Co. Ltd., Japan) prior to transplanting in tumorigenic bacteria-infested soil completely inhibited gall formation in the roots. These results are promising and support the development of ARK-1 as a biological control agent to manage grapevine crown gall. / Master of Science in Life Sciences / Grapevine crown gall is an economically significant disease in vineyards and nurseries worldwide. The pathogen of grapevine crown gall alters plant genome and causes tumor-like gall formation. Infected vines lose yield and vigor and eventually get killed. In this study, we tested the efficacy of a biological control agent called ARK-1. We challenged ARK-1 against higher cell numbers of pathogenic bacteria, evaluated timings, and practical methods of ARK-1 application. We conducted our greenhouse studies in grapevine (Vitis vinifera) cultivars 'Chardonnay', 'Cabernet Sauvignon', 'Merlot' and tomato (Solanum lycopersicum) cultivar 'Beefsteak'. In efficacy test, ARK-1 was effective and reduced 68% gall incidence against a four times higher pathogenic bacteria mixture. In timing assay, the application of ARK-1 up to 48 hours before inoculation of pathogenic bacteria reduced gall formation by more than 95%. Also, ARK-1 was effective when it was applied within 6 hours after inoculation of pathogenic bacteria. Root soaking of grapevine and tomato in the formulated ARK-1 suspension (KUF-1511, Kumiai Chemical Industry Co. Ltd., Japan), prior to transplanting in pathogenic bacteria-infested soil, completely inhibited gall formation in the roots. The promising result of this study is a step forward towards use of ARK-1 as a biological control agent to manage grapevine crown gall disease.

Crown gall of grapevine (Vitis spp.) is a serious and economically important disease caused by the bacterial pathogen Rhizobium vitis, which transforms healthy plant cell genomes leading to hypertrophic and hyperplastic growth of affected plant cells. Recent studies have documented a strong inhibitory effect against Japanese tumorigenic R. vitis isolates by a newly identified non-tumorigenic strain of R. vitis, ARK-1. We conducted co-inoculation assays in tomato (Solanum lycopersicum) and wine grape cultivars (Vitis vinifera) with four tumorigenic isolates of R. vitis from Virginia. These tumorigenic isolates were co-inoculated with ARK-1 in various ratios and resulting gall incidence and gall size were measured. Analysis was conducted with the generalized linear mixed model (GLIMMIX) in SAS (ver. 9.4). ARK-1 significantly reduced both the mean probability of gall formation and the mean gall size (P < 0.05). ARK-1 efficacy against combinations of two or four tumorigenic isolates and up to twice as many cells of tumorigenic isolates was also significant. However, there was an indication of a loss of efficacy when ARK-1 was challenged with four isolates at four times the cell number of ARK-1. Also, the efficacy of ARK-1 was influenced by both the specific isolate and host plant used in the study. Our results suggest that ARK-1 has promising potential as an effective biological control agent for grapevine crown gall in the United States. / MSLFS / Crown gall of grapevine (Vitis species) is a serious and economically important disease to the grape production industry caused by the bacterium Rhizobium vitis. This pathogen alters the genetic material of plant cells leading to cancer-like growth of affected plant cells (tumors); which may lead to the death of a grapevine. Recently, a non-tumor inducing strain of R. vitis (ARK-1) was documented to inhibit gall induction by tumor-inducing R. vitis strains in Japan. To see if ARK-1 would be a good candidate for biological control of the tumor-causing strains of R. vitis in the US, we tested it against four tumor-inducing strains in both tomato (Solanum lycopersicum) and grapevine (Vitis vinifera). The tumor-inducing strains were mixed with ARK-1, injected into plant stems in various ratios, and resulting gall incidence and gall size were measured 42 to 90 days later. ARK-1 significantly reduced both the probability of gall formation and the gall size in both tomatoes and grapevines. ARK-1 was an effective agent against mixtures of one, two, or four tumor-inducing strains and provided control even when outnumbered two to one by cells of tumor-inducing strains. The efficacy of ARK-1 was influenced by the specific strain, relative cell number to tumor-inducing strains, and host plant used in the study. Our results suggest that ARK-1 has promising potential as an effective biological control agent for grapevine crown gall in the United States.