Integrated Biocontrol Strategies for Disease Control on Greenhouse Ornamentalsusing Poinsettia as a Model System Progress Report– June 1999
Project Progress Report Cover Page & Requirements
11 Glen-Ed Professional Park, Glen Carbon, Illinois 62034 Telephone: 618.692.0045 Fax: 618.692.4045 Email: bstoeckl@endowment.org
Annual Report Deadline:
Reports must be received no later than June 1, 1999
Reporting Media:
An original hard copy and computer copy must be submitted.
- Submit 30 copies of the report by mail and
- An MSWord computer copy — two media options exist:
(1) Submit on 3.5″ diskette or (2) attach file to email.
Report Content:
Progress reports must include this cover sheet (fully completed). Do not exceed two typed pages (minimum font size = 11). Reports must (1) review industry needs addressed and project objectives, (2) summarize work conducted since inception outlining results and specific benefits to the industry especially new information identified, (3) outline next steps and future for project and anticipated industry benefit(s).
Purpose of Progress Report:
Progress reports enable the board and industry to keep abreast of the work accomplished in each project receiving AFE funding. They provide a tool to evaluate whether the work is meeting industry needs and original objectives. They are also used as a resource to develop publicity for AFE funded research.
IMPORTANT NOTE:
Information provided will be used as needed for publicity purposes unless the investigator indicates otherwise. If the material is confidential, an appropriate summary must be included that can be publicized, so the status of all projects can be released to industry.
Date: 5/12/99
Title of Project: Integrated biocontrol strategies for disease control on greenhouse ornamentals using poinsettia as a model system
Institution(s): North Carolina State University
AFE Grant Amount $19,536 Grant Period: 07-01-98 to 06-30-99
Project Completion Date: June 30, 1999
Project Leader: D. Michael Benson
Title: Professor
Address: Department of Plant Pathology, North Carolina State University, Campus Box 7616, Raleigh, NC 27695-7616
Telephone: 919-515-3966 Fax: 919-515-5657 Email: mike_benson@ncsu.edu
Additional Researchers (Names): Jae-soon Hwang, Graduate Student
Integrated biocontrol strategies for disease control on greenhouse ornamentals using poinsettia as a model system
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Industry needs and project objectives
The competitive nature of the floral industry within the USA and the stiff competition from foreign firms dictates that growers become more efficient in production. This means that adequate controls must be found for disease losses due to recurring pathogens like Botrytis, Pythium, and Rhizoctonia. Development of new technologies for the use of biological control is under way in the USA and elsewhere. Biological control takes advantage of environmentally safe microorganisms that are introduced to the production crop for disease control. Several biocontrol agents like Burkholderia cepacia, and binucleate Rhizoctonia fungi have been identified previously. The technology must now be developed to extent their range of effectiveness against multiple pathogens as well as to develop formulations that provide suitable shelf life, efficacy, and user-friendliness. Firms that adopt the use of biocontrol agents will enjoy an advantage of short re-entry periods under the Worker Protection Standard as well as a very low worker exposure hazard.
The objective of this research is to integrate the use of a nonpathogenic, binucleate Rhizoctonia fungus (BNR) and Burkholderia (Pseudomonas) cepacia, strain 5.5B, as biocontrol agents in sequential application approach to manage diseases caused by R. solani, P. aphanidermatum, and B. cinerea on poinsettia with potential applicability on a wide range of additional potted flowers and bedding plants. Industry benefits could potentially be wide-ranging and available in a reasonable period of time. Both biocontrol agents have controlled, in repeated trials, R. solani and have shown some activity against Pythium spp. Thus, the need to screen a wide diversity and number of potential biocontrol organisms is eliminated. In addition, each of the biocontrol agents exhibits a different mechanism of action. This would limit the ability of pathogens to develop resistance and enhance the target pathogen range. These pathogens are the principle fungal pathogens, worldwide, on greenhouse ornamentals and effectiveness in the poinsettia production system could allow applicability to other production systems and greenhouse crops.
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Summary of work conducted
A. Pyrrolnitrin production and biocontrol activity of B. cepacia
To investigate the relationship between pyrrolnitrin production by strain 5.5B of B. cepacia and biocontrol activity of stem rot of poinsettia, spontaneous mutation of 5.5B has been conducted and optimal cultural conditions for 5.5B have been studied to enhance pyrrolnitrin production by 5.5B.
Spontaneous mutants of B. cepacia and their biocontrol activity.
Two spontaneous mutants of 5.5B (21-2, 13-1) were selected by using rifampicin and tested in the greenhouse experiment. Strain 21-2 showed the same level of disease control as the wild type of 5.5B, while strain 13-1 showed no biocontrol activity on stem rot. Production of pyrrolnitrin by wild type strain 5.5B and strain 21-2 was detected at same concentration on TLC plate. However, only a faint signal was detected when strain 13-1 was analyzed. These results suggest that the production of pyrrolnitrin by B. cepacia is essential for biocontrol of Rhizoctonia stem rot of poinsettia during propagation.
The effect of cultural conditions on the production of pyrrolnitrin.
Nutrient broth (NB), Luria-Bertani (LB) and a minimum salts media for incubation of B. cepacia were used at two different pH. After 5 days of incubation, B. cepacia in NB and minimum salts media reached the same cell concentration, but the cell concentration in LB was lower. B. cepacia incubated in NB and minimum salts media showed the same amount of biocontrol ability in the greenhouse experiment, but the biocontrol activity of B. cepacia incubated in LB was slightly lower than other two media. No differences in biocontrol ability were observed when B. cepacia was cultured at different pH values and applied in the greenhouse experiment.
B. Sequential application of two biocontrol agents
To extend the disease protection period from propagation through transplanting and finishing, additional applications of a biocontrol agent, and integrated use of two or more biocontrol agents were investigated. Two biocontrol agents, BNR isolates and B. cepacia, with different possible mechanisms of biocontrol were applied sequentially to provide crop long protection. Disease severity of stem rot in B. cepacia-treated rooting cubes was 57% lower than in infested controls. No biocontrol activity was observed in rooting cubes treated with BNR. After transplanting, isolates of BNR were more effective in control of stem rot compared to B. cepacia. In the poinsettia-Rhizoctonia stem rot system, applying B. cepacia at the propagation stage and then using BNR at the time of transplanting was the most effective biocontrol strategy.
C. Induced resistance by BNR isolates
Non-pathogenic, binucleate Rhizoctonia are known to induce plant resistance to plant pathogens. Systemic resistance of poinsettia against Rhizoctonia stem rot induced by two BNR isolate (BNR621 and P9023) was investigated as a part of understanding mechanisms involved in biocontrol. Poinsettia cuttings acquired from BNR treated stock plants were challenge-inoculated with Rhizoctonia. A series of time gaps between BNR treatment and challenge inoculation (1 to 8 weeks) was provided to give BNR time to colonize in the root system and to induce systemic resistance. Induced resistance by BNR was observed in the treatments with two or more weeks between BNR treatment and challenge inoculation. Disease severity of stem rot on cuttings from these treatments was 32% lower than the infested control. No induced resistance was observed when only one week was given between BNR treatment and inoculation with Rhizoctonia, indicating that one week is not long enough for BNR to induce resistance.
D. Population dynamics of B. cepacia
Establishment of biocontrol agents in the plant root zone and maintenance of high population levels are critical for successful biocontrol. The relationship between rhizosphere (zone of root influence) and non-rhizosphere colonization by strain 5.5B and its biocontrol ability was studied. Strain 5.5B successfully colonized and maintained its population level (107/g root) up to 5 weeks after the application on the poinsettia root system. However, the population of 5.5B in non-rhizosphere declined rapidly and reached a non-detectable level at 7 weeks after application. Multiple application of 5.5B (1st application in rooting cubes and 2nd application at transplanting) showed the highest root colonization, but the biocontrol activity was not significantly improved compared to sequential application.
E. Biocontrol of Botrytis blight on poinsettia
Numerous experiments have been conducted with these biocontrol agents and other candidates for Botrytis control without success to date.
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Outline next steps.
Experiments will continue to focus on effectiveness of strain 5.5B during propagation combined with the sequential use of the BNR fungi at transplanting of poinsettia for biocontrol of Rhizoctonia to extend length of protection based on population dynamics and mode of action of these biocontrol agents. Given the success of the induced resistance phenomenon to protect against Rhizoctonia stem rot, longer intervals between application of BNR isolates and challenge with Botrytis will be tested.
