Biorational approaches for management of bacterial wilt and bacterial spot on tomato

Biorational approaches for management of bacterial wilt and bacterial spot on tomato

Summary

We are involved in a research project to develop strategies in integrated management of bacterial wilt. Tin the first aspect of the study we looked at the efficacy of several compounds for control of bacterial wilt of tomato caused by Ralstonia solanacearum under greenhouse conditions when applied as foliar spray or as a soil drench before transplanting tomato seedlings and once every 7 days thereafter until four weeks after transplanting. Soils used for transplanting were infested with Ralstonia solanacearum strain Rs 5 (race 1, biovar 1) at an initial density of 5.5 x 107 CFU/ml of soil. The plants were incubated in a greenhouse and arranged as a randomized complete block design with 5 replications. The results indicated that application of STBX-16 (Phyton 27), in conjunction with QRD 141, consistently provided significant protection of tomato plants against bacterial wilt. Disease incidence of treated plants was 15% four weeks after transplanting while 100% of untreated plants wilted in two weeks. QRD 600 (Muscodor) reduced disease incidence significantly compared to the untreated control, however, it was less effective compared with STBX-16 plus QRD 141(Serenade Max). We conclude that these compounds exhibited the capacity to significantly reduce bacterial wilt on the susceptible tomato cultivar used in this study. In other greenhouse tests we looked at using combinations of QRD600 with Actigard (active ingredient acibenzolar-S-methyl, ASM) for control of bacterial wilt in the greenhouse. Application of QRD 600 in conjunction with Actigard showed a tendency to enhance disease suppression. We concluded that QRD 600 exhibited the capacity to significantly reduce bacterial wilt on the susceptible tomato cultivar used in this study. The efficacy of several biorational products was evaluated for control of bacterial wilt on tomato in 2006 under field conditions in Quincy, FL. A susceptible tomato cultivar Phoenix was used in the study. Field soil was uniformly infested with the bacterial pathogen Ralstonia solanacearum strain Rs5 (race 1 biovar 1). Disease incidence reached 81.7% in the untreated control plots at the end of the experiment while only 20% in the plots treated with thymol. K-Phite, either alone or in combination with Serenade or Phyton 27, provided significant disease reduction. ProMax also reduced disease significantly compared with the untreated control but was less effective than other products or their combinations. All the treatments increased tomato yield significantly compared to the untreated control. In the second area our objective was to determine if lower rates of Actigard can be applied to enhance disease control without affecting tomato yield and identify resistant lines to determine if they respond to PGPRs. Greenhouse experiments to evaluate the efficacy of reduced rate of Actigard for managing bacterial spot on tomato were conducted in 2006 at the University of Florida North Florida Research and Education Center located in Quincy, FL. In an initial experiment, tomato varieties Solar Fire, FL 8000, FL 8314 and 684490 were used to select resistant lines for other experiments. In the untreated control, bacterial spot AUDPC (Area Under Disease Progress Curve) was 354.3 per plant for the four tomato varieties, which was significantly higher than any of the Actigard combinations, of which bacterial spot incidence ranged from 18.87 to 77.37 per plant. Actigard treatments consistently provided significant protection against bacterial spot on the four different varieties. The reduction of bacterial spot disease of Actigard treatment was not significantly different from the farmer standard copper and mancozeb treatment. More greenhouse experiments were conducted using susceptible tomato cultivar Solar Fire and resistant cultivar 684490 for fine-tuning the Actigard rate and optimizing the application rate combination in bacterial spot management. All rates of Actigard significantly reduced disease severity compared with untreated control on both susceptible and resistant genotypes. On resistant genotype, 684490, efficacy of different rates of Actigard was not significantly different; however, on the susceptible genotype, Solar Fire, the higher Actigard doses provided better disease control. These data indicate that the use of resistant cultivars allows application of reduced rates of Actigard. The efficacy of reduced Actigard rates (1X, 1/2X and 1/10X) on tomato bacterial spot disease development was compared with the standard copper-mancozeb treatment and evaluated concurrently in spring field trials in two locations, at the UF-NFREC in Quincy, FL and at the UF Plant Science Unit in Citra, FL in 2007. In Quincy, the standard copper-mancozeb treatment was most effective in controlling bacterial spot, whereas the Actigard treatments provided only marginal control. In Citra none of the treatments provided any significant control. Finally, we conducted experiments to determine if a resistant genotype responds to PGPR in greenhouse experiments. Two PGPR strains Bacillus pumilus B122 and Pseudomonas fluorescens B130 were used to test their efficacy on reduction of bacterial spot disease using tomato cultivars, Solar Fire and 684490 under greenhouse conditions. Experiments were conducted twice independently in Quincy. Both tomato lines responded to PGPRs similarly: Bacillus pumilus B122 significantly reduced disease severity, however Pseudomonas fluorescens B130 did not. Combination of the two PGPR strains worked similarly to Bacillus pumilus B122 alone.

Objectives/Performance Targets

Objectives
1. To develop strategies in integrated management of bacterial wilt:

a. Evaluate the efficacy and application methods of new biofumigants and reduced risk compounds in control of R. solanacearum on tomato under greenhouse and field conditions.

b. Determine efficacy of the SAR inducer, Actigard, in reducing bacterial wilt on susceptible tomato cultivars under field conditions at different inoculum levels, and evaluate integrated effectiveness and economics of field application of Actigard, biofumigant (thymol), and commercial tolerant or resistant tomato genotype (FL 7514, BHN 669) in the management of bacterial wilt.

c. Using the data obtained in objectives 1a and 1b to develop and implement best management strategies for bacterial wilt on tomato in naturally infested commercial tomato fields. On-farm research and demonstrations will be conducted in collaboration with growers and extension agents in north Florida and southern Georgia.

2. To optimize integrated management of bacterial spot with the SAR inducer Actigard, PGPRs and bacteriophages.

a. Determine if lower rates of Actigard can be applied to enhance disease control without affecting tomato yield and identify resistant lines to determine if they respond to PGPRs.

b. Determine the effects of modified application strategies of the SAR inducer (Actigard) and PGPRs in combination with bacteriophages.

c. Combine the best strategies in 2a and 2b for management of bacterial spot in field experiments to achieve maximum reduction of the disease and copper bactericide application. On-farm research and demonstration will be conducted in north Florida and southern Georgia and economic benefits will be analyzed.

3. To determine, through Cost Benefit Analysis of each field trial, the management strategies yielding the greatest financial returns to the grower.

Accomplishments/Milestones

1. To develop strategies in integrated management of bacterial wilt.

The efficacy of several compounds was evaluated for control of bacterial wilt of tomato caused by Ralstonia solanacearum under greenhouse conditions. STBX-016 (Phyton 27) was applied as foliar spray and soil drench at a concentration of 1ml/liter starting 15 days before transplanting tomato seedlings and once every seven days thereafter until four weeks after transplanting. QRD 141 (Serenade Max) was applied to the same treatment as STBX-016 at a rate of 4 g/liter by foliar spray and soil drench 12 and 5 days before transplanting and by foliar spray once every week after transplanting. For soil drench, 5 ml of STBX-16 or QRD 141 was applied to each transplant cell. Soils used for transplanting were infested with Ralstonia solanacearum strain Rs 5 (race 1, biovar 1) at an initial density of 5.5 x 107 CFU/ml of soil. The soils were then treated with STBX-016 at a rate of 2 ml/liter of soil two hours after soil infestation with the pathogen. In a separate treatment, QRD 600 (Muscodor) was used to treat the infested soil as STBX-16 but at a rate of 7.5 g per liter of soil. Soils were treated with these products for four days in closed plastic bags and six-week old tomato seedlings were transplanted into treated soils three days later. The plants were incubated in a greenhouse and arranged as a randomized complete block design with 5 replications. Disease incidence was recorded weekly after transplanting and quantified as the percentage of plants wilted. The data were analyzed using the ANOVA or GLM procedures of the Statistical Analysis System (SAS). The results indicated that application of STBX-16, in conjunction with QRD 141, consistently provided significant protection of tomato plants against bacterial wilt. Disease incidence of treated plants was 15% four weeks after transplanting while 100% of untreated plants wilted in two weeks. QRD 600 reduced disease incidence significantly compared to the untreated control, however, it was less effective compared with STBX-16 plus QRD 141. These compounds exhibited the capacity to significantly reduce bacterial wilt on the susceptible tomato cultivar used in this study.

The efficacy of QRD600 (Muscodor) was evaluated for control of bacterial wilt of tomato caused by Ralstonia solanacearum under greenhouse conditions. Soils used for transplanting were infested with Ralstonia solanacearum strain Rs 5 (race 1, biovar 1) at an initial density of 4.2 x 107 CFU/ml of soil. QRD 600 (Muscodor) was used to treat the infested soil at a rate of 7.5 g per liter of soil in closed plastic bags and six-week old tomato seedlings were transplanted into treated soils three days later. Actigard was applied as foliar spray and soil drench. The initial foliar application of Actigard was performed two weeks after seedling emergence at a concentration of 50 ug/ml, followed by a foliar and soil drench application 5 days prior to plant inoculation with the pathogen. For soil drench, 5 ml of Actigard solution at a concentration of 25 ug/ml was applied to each transplant cell. Actigard was sprayed to tomato foliage once every week after transplanting at a concentration of 50 g/ml. After transplanting the plants were incubated in the greenhouse and arranged as a randomized complete block design with 4 replications. Disease incidence was recorded weekly after transplanting and quantified as the percentage of plants wilted. The data were analyzed using the ANOVA or GLM procedures of the Statistical Analysis System (SAS). The results indicated that QRD 600 reduced disease incidence significantly compared to the untreated control. Disease incidence of QRD 600-treated plants was 45% four weeks after transplanting while 100% of untreated plants wilted in three weeks. Application of QRD 600 in conjunction with Actigard showed a tendency to enhance disease suppression. QRD 600 significantly reduced bacterial wilt on the susceptible tomato cultivar used in this study.

The efficacy of several biorational products was evaluated for control of bacterial wilt on tomato in 2006 under field conditions in Quincy, FL. A susceptible tomato cultivar Phoenix was used in the study. Field soil was uniformly infested with the bacterial pathogen Ralstonia solanacearum strain Rs5 (race 1 biovar 1). Solutions of these products were applied in the soil under plastic mulch by drip irrigation for 3 h. Three days after application, holes were pierced through the plastic mulch for transplanting and to allow aeration of excess volatiles. The period of aeration was 4 days. Tomato seedlings were transplanted into the field one week after application of these compounds. The experimental design was a randomized complete block with four replications. Disease incidence reached 81.7% in the untreated control plots at the end of the experiment while only 20% in the plots treated with thymol. K-Phite, either alone or in combination with Serenade or Phyton 27, provided significant disease reduction. ProMax also reduced disease significantly compared with the untreated control but was less effective than other products or their combinations. All the treatments increased tomato yield significantly compared to the untreated control.

Field experiments are being conducted this spring and again next fall. Fall field experiments in 2007 were unsuccessful in terms of several technical problems.

2. To optimize integrated management of bacterial spot with the SAR inducer Actigard, PGPRs and bacteriophages.

a. Determine if lower rates of Actigard can be applied to enhance disease control without affecting tomato yield and identify resistant lines to determine if they respond to PGPRs.

Evaluation of the efficacy of reduced rate of Actigard in greenhouse experiments and identification of resistant lines.

Greenhouse experiments to evaluate the efficacy of reduced rate of Actigard (active ingredient acibenzolar-S-methyl, ASM) for managing bacterial spot on tomato were conducted in 2006 at the University of Florida North Florida Research and Education Center located in Quincy, FL. In an initial experiment, tomato varieties Solar Fire, FL 8000, FL 8314 and 684490 were used to select resistant lines for other experiments. Tomato plants were grown in greenhouse in Soil-less medium Sungro Metro Mix 200 series (Sungro Horticulture Canada Ltd.) in expanded polystyrene flats 16 x 8 cells with cell size of 3.5 x 3.5 cm2. Tomato seedlings were transplanted to 10-cm pot 10 days after seedling emergence. Three weekly applications of Actigard were made in three independent experiments. Ten combinations of different doses of ASM, including 1 x (ASM 27.5 ug/mL), ½ x and 1/10 x of ASM, were evaluated. The initial foliar application of ASM was done two weeks after seedling emergence. Plants were sprayed till run-off with ASM solution using a hand held garden sprayer.

Xanthomonas perforans strain 91-118 (race 3, T-3) isolated from tomato was used in the studies. Bacterial inoculation was performed 5 days after the 3rd Actigard application. Inoculation of the pathogen in the greenhouse was as follows, bacterial suspension containing 1 x 107 cells/mL was sprayed on tomato plants thoroughly (on both upper and back of the leaves) till run-off. Inoculated plants were then covered with plastic bags, and removed after 48 hr. Bacterial spot incidence was rated as bacterial spot number per plant. Among the four tomato varieties, Solar Fire was the most susceptible to Xanthomonas perforans strain 91-118 with bacterial spot severity of 100.28 per plant, whereas 684490 was the most resistant to Xanthomonas perforans strain 91-118 with bacterial spot severity of 38.25 per plant. The resistant level of the four varieties was significantly different based on LSD-Duncan test.
In the untreated control, bacterial spot AUDPC (Area Under Disease Progress Curve) was 354.3 per plant for the four tomato varieties, which was significantly higher than the any of the Actigard combination treated, of which bacterial spot incidence were ranging from 18.87 to 77.37 per plant. Actigard treatments consistently provided significant protection against bacterial spot on the four different varieties. The reduction of bacterial spot disease of Actigard treatment was not significantly different from the farmer standard copper and mancozeb treatment.

Integration of reduced Actigard rate and tomato resistant line on bacterial spot management in greenhouse experiments.

More greenhouse experiments were conducted using susceptible tomato cultivar Solar Fire and resistant cultivar 684490 for fine-tuning the Actigard rate and optimizing the application rate combination in bacterial spot management.
The production of tomato seedlings was as previously described. Two applications of Actigard were performed in two independent experiments. Three combinations of different dosage of ASM, including 1 x (ASM 27.5 g/mL) with 1 x, ½ x with ½ x, and 1/10 x with 1/10 x of ASM, were evaluated. The initial foliar application of ASM was done two weeks after seedling emergence. Plants were sprayed till run-off with ASM solution using a hand held garden sprayer. A second application of ASM was made one week after, 5 days prior to bacterial inoculation.
Xanthomonas perforans strain 91-118 (race 3, T-3) was used in the studies. Inoculation of the pathogen was different from previous experiments due to cold weather. Tomato seedlings were transferred to growth chamber (28 C, RH 90% and 16-hr photoperiod) before inoculation. Bacterial suspension containing 1 x 108 cells/mL was sprayed on tomato plants thoroughly (on both upper and back of the leaves) till run-off. Inoculated plants were then covered with plastic bags, and removed after 48 hr. Inoculated plants remained in the growth chamber until disease severity assessment. Bacterial spot incidence was rated as bacterial spot number per plant. All rates of Actigard significantly reduced disease severity compared with untreated control on both susceptible and resistant genotypes. On resistant genotype, 684490, efficacy of different rates of Actigard was not significantly different; however, on the susceptible genotype, Solar Fire, the higher Actigard doses provided better disease control. These data indicate that the use of resistant cultivars allows application of reduced rates of Actigard.

Reduced Actigard rate on bacterial spot management in field experiments.

The efficacy of reduced Actigard rates (1X, 1/2X and 1/10X) on tomato bacterial spot disease development was compared with the standard copper-mancozeb treatment and evaluated concurrently in spring field trials in two locations, at the UF-NFREC in Quincy, FL and at the UF Plant Science Unit in Citra, FL in 2007.
Bella Rosa, commercial variety resistant to tomato spotted wilt virus and heat, was used in both locations. In Quincy, 15 plants, spaced 20” apart constituted an experimental unit, whereas in Citra 10 plants, spaced 18” apart made up a plot.
Tomato seedlings were transplanted on April 26 in Citra, and April 13 in Quincy. The middle tomato plants of each plot were inoculated with X. perforans. Weekly foliar spray started 3-days after transplanting in Quincy (April 16 – July 3) and 5-days in Citra (May 1 – July ) for 10 weeks. Guidelines established by the University of Florida/IFAS were followed for land preparation, fertility, irrigation, weed management and insect control. White mulch was used to avoid heat during Summer time. Plants were treated as scheduled with the selected compounds as shown in Table 1 throughout the growing season using a CO2 pressurized hand-held 5-nozzle tomato boom in Quincy and Solo backpack sprayers in Citra. Untreated control (UTC) received no treatment of the selected compounds. Treatments were arranged in randomized complete block design with 4 replications. Disease assessments were carried out biweekly throughout the growing season. Disease severity was evaluated by visual assessments of percent defoliation using the Horsfall-Barratt scale. The area under the disease process curve (AUDPC) was calculated using the entire season’s ratings. The disease severity ratings were subjected to analyses of variance (ANOVA), and the treatment means were tested for significance by the Duncan-Waller K-ration test using SAS program package version 8.0. In Quincy, the standard copper-mancozeb treatment was most effective in controlling bacterial spot whereas
the Actigard treatments provided only marginal control. In Citra none of the treatments provided any significant control.

Determine if resistant line responds to PGPR in greenhouse experiments.

Two PGPR strains Bacillus pumilus B122 and Pseudomonas fluorescens B130 were used to test their efficacy on reduction of bacterial spot disease using tomato cultivars, Solar Fire and 684490 under greenhouse conditions. Experiments were conducted twice independently in Quincy.
Tomato seedling growth was same as previously described. Bacterial spot pathogen, X. perforans strain 91-118 (5 x 108 CFU/mL) was used for bacterial inoculation. Inoculation was made on 4-leaf stage seedlings.
PGPR applications were performed with seed treatement and drench application 7 days prior to bacterial inoculation. Soak seeds in PGPR bacterial suspension in 0.85% Saline (5 x 108 CFU/mL) on an orbital shaker 150 rpm at room temperature for 20 min. Drain suspension using cheese cloth, and blot the excessive liquid using a paper towel. 0.85% Saline used as control. Apply PGPR strains once during seedling growth by drenching 10 mL of PGPR bacterial suspension (5 x 108 CFU/mL) into cell, 7-days before inoculation.
Treatments included UTC, standard control with Kocide 3000 + Manzate DF 75 (1.5 lb/acre : 2 lb/acre); 0.85% Saline-treated seeds; B130 treatment; B122 treatment; B130 and B122 treatment.
Disease assessment and data analysis were same as previously described. Both tomato lines responded to PGPRs similarly: Bacillus pumilus B122 significantly reduced disease severity, however Pseudomonas fluorescens B130 did not. Combination of the two PGPR strains worked similarly to Bacillus pumilus B122 alone. Saline did not have effect on disease symptoms.

Impacts and Contributions/Outcomes

Publications
1. Momol, M.T., P. Ji, A. Wen, J. B. Jones, S.M. Olson, B. Balogh, L. Ritchie, P.D. Roberts, R. Systma, C. Meister, and D.J. Norman. 2007. Evaluation of phosphorus acid-containing products for managing bacterial wilt and spot of tomato. Program and Abstract Book, the 2nd ISHS International Symposium on Tomato Diseases, Kusadasi, Turkey. p 70.

2. Wen, A, B. Balogh, M. T. Momol, S. M. Olson, and J. B. Jones. 2007. Integration of reduced rates of Actigard and host resistance in management of bacterial spot of tomato. Phytopathology. S121

3. Wen, A, B. Balogh, M. T. Momol, J. B. Jones, S. M. Olson, L. S. Ritchie, P. D. Roberts, R. Systma, and C. W. Meister. 2007. Management of bacterial spot of tomato with phosphorous acid containing compounds. Phytopathology. S121

4. Wen, A, B. Balogh, M. T. Momol, S. M. Olson, and J. B. Jones. 2007. Integration of reduced rates of Actigard and host resistance in management of bacterial spot of tomato. APS Annual Meeting. July 28-August 1, 2007, San Diego, CA. (Poster)

5. Wen, A, B. Balogh, M. T. Momol, J. B. Jones, S. M. Olson, L. S. Ritchie, P. D. Roberts, R. Systma, and C. W. Meister. 2007. Management of bacterial spot of tomato with phosphorous acid containing compounds. APS Annual Meeting. July 28-August 1, 2007, San Diego, CA. (Poster)

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