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The crown gall of grapevines is an economic importance bacteriosis caused by Rhizobium vitis strains. Five essential oils (EO’s) were tested against the autochthonous 2btm strain of R. vitis for their capacity to inhibit the phytopathogen growth, by in vitro and in planta tests. The measurements regarding in vitro inhibition zone were performed. Subsequently, the ability of patentable formula based on essential oils to prevent/reduce tumor formation in tomato and vine plants has been tested. After 30 days for tomato plants, the evaluation of disease incidence and effectiveness of treatments applied at two points in time (T0 and T30), were determined. In vine plants, tumorigenesis is initiated up to 9 months after inoculation, so the results are being processed. Efficacy values of 94.7% at both T0 and T30 and, a very low disease incidence of 5% compared to positive control were recorded. Also, the tumor of EO’s treated tomato plants were smaller in size and did not form a complete ring as positive control plants. The use of patentable formula as prevented treatment, reduced the number of plants developing crown gall symptoms and tumor size compared to positive control plants.

Nonpathogenic strain ARK-1 of Rhizobium vitis has antagonistic activity against a tumorigenic (Ti) strain of R. vitis and works as a biocontrol agent, but the temporal dynamics of the bacterial colonization in plants needs to be understood. Here we showed that the incidence of galls induced by Ti strain was reduced to about 60% when both upper and lower positions on the grapevine stem were inoculated with ARK-1 3 cm from the site that was inoculated with Ti 5 days later. A bacterial cell detection assay in the grapevine tissue revealed that ARK-1 moved at least 3 cm in 5 days. [ABSTRACT FROM AUTHOR]

Crown gall disease in grapevine is caused by pathogenic strains of Rhizobium vitis with a tumor-inducing (Ti) plasmids. A nonpathogenic strain, VAR03-1 of R. vitis, has been isolated from the grapevine root of nursery stock and it was shown to act as a biological control agent to crown gall disease. Its disease-suppressive effect was observed even when it was coinoculated with the pathogen in a 1:1 ratio. Here, we present the complete genome data of R. vitis VAR03-1, assembled by sequencing reads obtained by both PacBio and Illumina technologies with annotation. This genome sequence could contribute to investigations of the molecular basis underlying the biocontrol activity as well as the root-colonization ability of this bacterial strain.[Graphic: see text] Copyright © 2020 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Rhizobium vitis strain VAT03-9 (MAFF 211676) is a causal agent of crown gall disease in grapevine. It is one of the pathogenic strains of R. vitis isolated from graft unions of grapevine in Okayama Prefecture, Japan. Inoculation tests verified its virulence for gall formation on grapevine, tomato, and sunflower. It harbors tumor-inducing plasmid. Here, we present the complete genome sequence with annotation of R. vitis VAR03-9 obtained by assembling reads from PacBio and Illumina-sequencers. This genome sequence should be useful for the analyses of pathogenicity and evolutionary lineage of the pathogens of crown gall disease.[Graphic: see text] Copyright © 2020 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Rhizobium (Agrobacterium) is one genus in the family Rhizobiaceae. Most of the species are epi- or endophytic bacteria which include tumorigenic or rhizogenic pathogens, root nodule bacteria, and commensal endosymbionts. Rhizobium vitis strain VAR06-30 is a commensal bacterium without pathogenicity which was isolated from a rootstock of grapevine in Japan. It also does not have antagonistic activity to the pathogenic strain of R. vitis. Here, we show the complete genome sequence data with annotation of R. vitis VAR06-30 which was analyzed by sequence reads obtained from both PacBio and Illumina platforms. This genome sequence would contribute to the understanding of evolutionary lineage and characteristics of Rhizobium commensal bacteria.[Graphic: see text] Copyright © 2020 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Abstract Objective To gain insights into the virulence suppressive mechanism of a nonpathogenic strain of Rhizobium vitis ARK-1, we co-inoculated ARK-1 with a tumorigenic (Ti) strain of R. vitis to examine the expression of two essential virulence genes (virA and virG) and one non-essential gene (virD3) of the Ti strain at the wound site of grapevine. Results Co-inoculation of ARK-1 with a Ti strain VAT03-9 at a 1:1 cell ratio into grapevine shoots resulted in significantly lower expression of the virulence genes virA, virD3, and virG of VAT03-9 at 1 day after inoculation compared with those when shoots were inoculated only with VAT03-9. ARK-1 was not able to catabolize acetosyringone, which is the plant-derived metabolites inducing the entire vir regulon in Ti strains, suggesting the direct effect of ARK-1 on the induction of broad range of vir genes of R. vitis Ti strains.

Crown gall is a globally distributed and economically important disease of grapevine and other important crop plants. The causal agent of grapevine crown gall is tumorigenic Allorhizobium vitis (Ti) strains that harbor a tumor-inducing plasmid (pTi). The epidemic of grapevine crown gall has not been widely elucidated. In this study, we investigated the genetic diversity of 89 strains of Ti and nonpathogenic A. vitis to clarify their molecular epidemiology. Multi-locus sequence analysis (MLSA) of the partial nucleotide sequences of pyrG, recA, and rpoD was performed for molecular typing of A. vitis strains isolated from grapevines with crown gall symptoms grown in 30 different vineyards, five different countries, mainly in Japan, and seven genomic groups A to F were obtained. The results of MLSA and logistic regression indicated that the population of genetic group A was significantly related to a range of prefectures and that the epidemic of group A strains originated mainly in Hokkaido in Japan through soil infection. Moreover, group E strains could have been transported by infected nursery stocks. In conclusion, this study indicates that both soil infection and transporting of infected nursery stocks are working as infection source in Hokkaido.

Plants protect themselves from pathogen attacks via several mechanisms, including hypersensitive cell death. Recognition of pathogen attack by the plant resistance gene triggers expression of carboxylesterase genes associated with hypersensitive response. We identified six transcripts of carboxylesterase genes, Vitis flexuosa carboxylesterase 5585 (VfCXE5585), VfCXE12827, VfCXE13132, VfCXE17159, VfCXE18231, and VfCXE47674, which showed different expression patterns upon transcriptome analysis of V. flexuosa inoculated with Elsinoe ampelina. The lengths of genes ranged from 1,098 to 1,629 bp, and their encoded proteins consisted of 309 to 335 amino acids. The predicted amino acid sequences showed hydrolase like domains in all six transcripts and contained two conserved motifs, GXSXG of serine hydrolase characteristics and HGGGF related to the carboxylesterase family. The deduced amino acid sequence also contained a potential catalytic triad consisted of serine, aspartic acid and histidine. Of the six transcripts, VfCXE12827 showed upregulated expression against E. ampelina at all time points. Three genes (VfCXE5585, VfCXE12827, and VfCXE13132) showed upregulation, while others (VfCXE17159, VfCXE18231, and VfCXE47674) were down regulated in grapevines infected with Botrytis cinerea. All transcripts showed upregulated expression against Rhizobium vitis at early and later time points except VfCXE12827, and were downregulated for up to 48 hours post inoculation (hpi) after upregulation at 1 hpi in response to R. vitis infection. All tested genes showed high and differential expression in response to pathogens, indicating that they all may play a role in defense pathways during pathogen infection in grapevines.

Rhizobium vitis: strain VAR03-1 is a biological control agent that suppresses grapevine crown gall disease caused by a tumorigenic strain of R. vitis (Ti). Both acetosyringone-induced expression of a virulence gene and the growth of Ti were suppressed in vitro when it was cultivated in the VAR03-1 culture filtrate. These inhibitory effects were reduced by high-temperature treatment or incubation for 72 h. Both activities were detected in the high molecular weight fraction (> 100 kDa) of the filtrate. Our results suggest that the antagonistic effects of VAR03-1 on Ti are mediated by large particle(s) released in the culture media. [ABSTRACT FROM AUTHOR]

Crown gall-affected grapevine samples were collected during 2009–2010 from major vineyards, located in different Turkish provinces. One hundred and three bacterial strains were obtained from 88 vineyards and 18 grapevine varieties; they were tumorigenic when inoculated in tobacco, sunflower and Datura stramonium plants and were identified as Rhizobium vitis using biochemical and physiological tests as well as PCR and specific primers. Nineteen R. vitis strains presented a number of anomalous biochemical and physiological characters. PCR and opine-specific primers revealed the presence of octopine/cucumopine-type plasmid in 82 R. vitis strains, nopaline-type plasmids in 18 strains and vitopine-type plasmids in three strains. Clonal relationship of strains was determined using Pulsed Field Gel Electrophoresis following digestion of genomic DNA with the restriction endonuclease PmeI. The greatest genetic diversity was found for the strains from Denizli, Ankara and Nevşehir provinces. Nopaline and vitopine-types of Rhizobium vitis were detected for the first time in Turkey.

Pathogenic Rhizobium vitis and R. radiobacter strains in symptomless plants and grapevine tumors were revealed by PCR technique. The comparative tests with five primer pairs to different siquences of bacterial genome were carried out. It was shown that in plants of Cabernet Sauvignon cultivar 18.5% from total Rhizobium population were represented by pathogenic strains.

Bacteriocins of 25% of investigated Rhizobium vitis strains isolated from grapevines in southern Ukraine caused in vitro lysis of crown gall agents of Rhizobium radiobacter Ñ58 strain. Treatment with bacteriocins from three R. vitis strains in vivo on test-plants kalanchoe and grapevine green cuttings lead to significant decrease in tumour formation.

Isolated Rhizobium vitis and Pantoeae agglomerans strains showed bacteriocinogenic activity against closely related strains. Investigated cultures had different susceptibility to bacteriocines. Comparing to mitomycin induction, spontaneous induction resulted in less quantity of lysis spots on test-culture lawns, but some strains still showed marked bacteriocinogenic activity.

Mildew resistance locus o (Mlo) is a specific seven-transmembrane gene family that confers resistance against several abiotic and biotic stresses in plants. In this study, we identified nine Mlo-like loci, VfMlo- like3243 ( Vitis flexuosa powdery- mildew resistance locus o-like3243), VfMlo- like4098, VfMlo- like5057, VfMlo- like6833, VfMlo- like7881, VfMlo- like8120, VfMlo- like14439, VfMlo- like14557, and VfMlo- like18484, which showed differential expression upon transcriptome analysis conducted using next generation sequencing (NGS) of V. flexuosa infected with Elsinoe ampelina. These genes are 1773-2548 bp long and encode deduced full-length proteins of 482-587 amino acids. The predicted amino acid sequences of all genes show typical Mlo domains containing seven highly conserved transmembrane domains, a calmodulin-binding domain and 30 important amino acid residues for Mlo function. While 3 VfMlo-like genes were downregulated, 5 VfMlo-like genes were up-regulated in grapevines infected with Erysiphe necator, among which four ( VfMlo- like6833, VfMlo- like8120, VfMlo- like14439 and VfMlo- like18484) are orthologous to the AtMlo2, AtMlo6 and AtMlo12 genes involved in powdery mildew susceptibility. All genes except VfMlo- like17881 showed upregulated expression at 48 hpi after infection of Rhizobium vitis. The results revealed that the tested genes are related in responses to pathogens of grapevines and can be utilized as useful resources for investigating their roles in disease resistant responses in grapevines. [ABSTRACT FROM AUTHOR]

Crown gall-affected grapevine samples were collected during 2009-2010 from major vineyards, located in different Turkish provinces. One hundred and three bacterial strains were obtained from 88 vineyards and 18 grapevine varieties; they were tumorigenic when inoculated in tobacco, sunflower and Datura stramonium plants and were identified as liliizobium vitis using biochemical and physiological tests as well as PCR and specific primers. Nineteen R. vitis strains presented a number of anomalous biochemical and physiological characters. PCR and opine-specific primers revealed the presence of octopine/cucumopine-type plasmid in 82 R. vitis strains, nopaline-type plasmids in IS strains and vitopine-type plasmids in three strains. Clonal relationship of strains was determined using Pulsed Field Gel Electrophoresis following digestion of genomic DNA with the restriction endonuclease Pmel. The greatest genetic diversity was fouuid for the strains from Denizli, Ankara and Nevşehir provinces. Nopaline and vitopine-types of Rliizobium vitis were detected for the first time in Turkey. [ABSTRACT FROM AUTHOR]

Nucleotide binding site-leucine-rich repeat (NBS-LRR) genes are the largest gene resistance family in plants. Resistance to Pseudomonas syringae 5 (RPS5) is a member of the NBS-LRR family. RPS5 and its homologs are important disease resistance genes in plants including Arabidopsis. This study identified nine loci of RPS5-like genes in Vitis flexuosa that showed differential expression from transcriptome analysis by next generation sequencing (NGS) of V. flexuosa infected with Elsinoe ampelina based on structural analyses: VfRPS5-like1059 (Vitis flexuosa resistance to Pseudomonas syringae 5-like 1059), VfRPS5-like1833, VfRPS5-like4135, VfRPS5-like4833, VfRPS5-like6172, VfRPS5-like13564, VfRPS5-like20585, VfRPS5-like55532, and VfRPS5-like62178. These genes, 2236-4762 bp in length, exhibited the structure of a coiled coil-nucleotide binding site-leucine-rich repeat (CC-NBS-LRR) resistance gene. The predicted amino acid sequences of all nine genes contained typical NBS domains. Chromosomal localization revealed that the nine genes were located in six chromosomes of the V. flexuosa pseudo-genome. Of the nine loci, only VfRPS5-like55532 showed upregulated expression against all tested pathogens at all tested time points, except 24 h postinoculation (hpi) for E. ampelina. All tested genes also showed differential expression against major grapevine fungal and bacterial pathogens. The results presented herein will also serve as a useful reference dataset for functional analysis and utilization in molecular breeding of resistant grapevines. [ABSTRACT FROM AUTHOR]

This study was conducted to investigate the efficacy of antibacterial compounds isolated from hairy vetch ( Vicia villosa Roth) shoots against grapevine crown gall pathogen. The hairy vetch shoot powder was sequentially extracted and screened for antibacterial activity by the disc diffusion method. Ethyl acetate extract from the hairy vetch shoots was subjected to column chromatography on silica gel. The structures of the compounds were then elucidated by spectroscopic analysis and chromatographic assay. Three compounds were isolated and identified as bis (2-ethylhexyl) phthalate (compound 1), diethyl phthalate (compound 2), and p-hydroxybenzoic acid (compound 3). The compounds showed potential antibacterial effect as diameters of inhibition zones (7.3 ± 0.3 to 11.2 ± 0.4 mm) against Rhizobium vitis and Bacillus subtilis. The minimum inhibitory and minimum bactericidal concentrations of these compounds against the target bacteria were found to range from 62.5 to 125 and 125 to 250 μg·mL, respectively. The compounds also showed bactericidal activity based on the viable counts of the tested strain. These findings indicate that the isolated compounds could be used as possible candidates for control of crown gall disease in grapevines caused by R. vitis. [ABSTRACT FROM AUTHOR]

Crude extracts from shoot and root tissues of Astragalus sinicus, Brassica napus, Dactylis glomerata, Lolium perenne, Lolium multiflorum and Vicia villosa were investigated for their antimicrobial activity against Rhizobium vitis, Bacillus subtilis, Aspergillus niger, Botrytis cinerea, Colletotrichum gloeosporioides, C. acutatum, and Elsinoe ampelina. Extracts were prepared from dried and powdered shoots and roots using common solvents (methanol, ethyl acetate, and water) with different degrees of polarity. The diameter of inhibition zones of the shoot (2,000 µg/disc) and root (2,500 µg/disc) extracts in water and ethyl acetate solvents against R. vitis and B. subtilis were found to be in the range of 9.8±1.3 to 18.0±2.0 mm and 9.0±0.7 to 16.4±0.5 mm, respectively. However, methanolic extract did not show any inhibitory activity. All the extracts showed a minute inhibition zone against A. niger which was not measurable, whereas the other tested fungi were not inhibited. The minimum inhibitory concentration (MIC) values of shoot and root extracts against the tested bacteria were in the range of 3.12 to 12.5 and 3.12 to 25 mg/ml, respectively. A preliminary phytochemical screening of all the tested extracts indicated the presence of alkaloids, flavanoids, saponins, cardiac glycosides, steroids and terpenoids. Total polyphenol contents of extracts positively correlated with their inhibitory activities, suggesting their potential antibacterial efficacy. Our findings suggest that crude extracts from the shoots and roots of these cover crops may be useful in preventing Rhizobium vitis-induced crown gall on grapes. [ABSTRACT FROM AUTHOR]

Forty-one strains of Rhizobium vitis, either tumorigenic (Ti) or nonpathogenic, were characterized using multilocus sequence analysis (MLSA) of the partial nucleotide sequences of pyrG, recA, and rpoD. The strains separated into seven clades. Rhizobium vitis (Ti) strains isolated from Japan were divided into five genetic groups (A to E), and nonpathogenic R. vitis strains were divided into two genetic groups (F and G). This result suggests that there are new genetic groups of R. vitis in Japan. Among these groups, members of A and B groups are widely distributed throughout Japan. [ABSTRACT FROM AUTHOR]

Crown gall is a globally distributed and economically important disease of grapevine and other important crop plants. The causal agent of grapevine crown gall is tumorigenic Allorhizobium vitis (Ti) strains that harbor a tumor-inducing plasmid (pTi). The epidemic of grapevine crown gall has not been widely elucidated. In this study, we investigated the genetic diversity of 89 strains of Ti and nonpathogenic A. vitis to clarify their molecular epidemiology. Multi-locus sequence analysis (MLSA) of the partial nucleotide sequences of pyrG, recA, and rpoD was performed for molecular typing of A. vitis strains isolated from grapevines with crown gall symptoms grown in 30 different vineyards, five different countries, mainly in Japan, and seven genomic groups A to F were obtained. The results of MLSA and logistic regression indicated that the population of genetic group A was significantly related to a range of prefectures and that the epidemic of group A strains originated mainly in Hokkaido in Japan through soil infection. Moreover, group E strains could have been transported by infected nursery stocks. In conclusion, this study indicates that both soil infection and transporting of infected nursery stocks are working as infection source in Hokkaido. [ABSTRACT FROM AUTHOR]

Non-tumorigenic Rhizobium (syn. Agrobacterium) vitis strain ARK-1 (ARK-1) has been shown to reduce crown gall in grapevine (Vitis vinifera) seedlings when co-inoculated with Japanese tumorigenic isolates of R. vitis. The objectives of this study were to test ARK-1's efficacy as a biological control agent against R. vitis isolated from grapevines in Virginia, and to examine genetic diversity of R. vitis found in Virginia, USA and Japan. ARK-1 was co-inoculated into wine grape trunks (V. vinifera cv. 'Cabernet Sauvignon') with a tumorigenic R. vitis isolate at a 1:1 cell ratio (~108 cell/ml). A total of four R. vitis isolates were tested individually. Compared to treatments of a tumorigenic isolate alone, ARK-1 co-inoculation significantly decreased (P ≤ 0.05) the mean probability of gall formation in all cases, and the mean gall diameter in all, but one case (isolate ACME15). The average reduction was 90% and 92% in the mean probability of gall formation and mean gall diameter, respectively. We conducted exploratory data visualization with distance-based clustering and correspondence analysis using data from ISSR- and rep-PCR, which showed separation of Virginia and Japanese isolates and placement of ARK-1 near to Virginia isolates. These results suggest that ARK-1 is a good candidate for biocontrol of grapevine crown gall in the mid-Atlantic region of the USA. [ABSTRACT FROM AUTHOR]

Crown gall disease in grapevine is caused by pathogenic strains of Rhizobium vitis with a tumor-inducing (Ti) plasmids. A nonpathogenic strain, VAR03-1 of R. vitis , has been isolated from the grapevine root of nursery stock and it was shown to act as a biological control agent to crown gall disease. Its disease-suppressive effect was observed even when it was coinoculated with the pathogen in a 1:1 ratio. Here, we present the complete genome data of R. vitis VAR03-1, assembled by sequencing reads obtained by both PacBio and Illumina technologies with annotation. This genome sequence could contribute to investigations of the molecular basis underlying the biocontrol activity as well as the root-colonization ability of this bacterial strain.[Formula: see text] Copyright © 2020 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Rhizobium vitis strain VAT03-9 (MAFF 211676) is a causal agent of crown gall disease in grapevine. It is one of the pathogenic strains of R. vitis isolated from graft unions of grapevine in Okayama Prefecture, Japan. Inoculation tests verified its virulence for gall formation on grapevine, tomato, and sunflower. It harbors tumor-inducing plasmid. Here, we present the complete genome sequence with annotation of R. vitis VAR03-9 obtained by assembling reads from PacBio and Illumina-sequencers. This genome sequence should be useful for the analyses of pathogenicity and evolutionary lineage of the pathogens of crown gall disease.[Formula: see text] Copyright © 2020 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Rhizobium ( Agrobacterium ) is one genus in the family Rhizobiaceae. Most of the species are epi- or endophytic bacteria which include tumorigenic or rhizogenic pathogens, root nodule bacteria, and commensal endosymbionts. Rhizobium vitis strain VAR06-30 is a commensal bacterium without pathogenicity which was isolated from a rootstock of grapevine in Japan. It also does not have antagonistic activity to the pathogenic strain of R. vitis . Here, we show the complete genome sequence data with annotation of R. vitis VAR06-30 which was analyzed by sequence reads obtained from both PacBio and Illumina platforms. This genome sequence would contribute to the understanding of evolutionary lineage and characteristics of Rhizobium commensal bacteria.[Formula: see text] Copyright © 2020 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Plants protect themselves from pathogen attacks via several mechanisms, including hypersensitive cell death. Recognition of pathogen attack by the plant resistance gene triggers expression of carboxylesterase genes associated with hypersensitive response. We identified six transcripts of carboxylesterase genes, Vitis flexuosa carboxylesterase 5585 (VfCXE5585), VfCXE12827, VfCXE13132, VfCXE17159, VfCXE18231, and VfCXE47674, which showed different expression patterns upon transcriptome analysis of V. flexuosa inoculated with Elsinoe ampelina. The lengths of genes ranged from 1,098 to 1,629 bp, and their encoded proteins consisted of 309 to 335 amino acids. The predicted amino acid sequences showed hydrolase like domains in all six transcripts and contained two conserved motifs, GXSXG of serine hydrolase characteristics and HGGGF related to the carboxylesterase family. The deduced amino acid sequence also contained a potential catalytic triad consisted of serine, aspartic acid and histidine. Of the six transcripts, VfCXE12827 showed upregulated expression against E. ampelina at all time points. Three genes (VfCXE5585, VfCXE12827, and VfCXE13132) showed upregulation, while others (VfCXE17159, VfCXE18231, and VfCXE47674) were down regulated in grapevines infected with Botrytis cinerea. All transcripts showed upregulated expression against Rhizobium vitis at early and later time points except VfCXE12827, and were downregulated for up to 48 hours post inoculation (hpi) after upregulation at 1 hpi in response to R. vitis infection. All tested genes showed high and differential expression in response to pathogens, indicating that they all may play a role in defense pathways during pathogen infection in grapevines.