Development and on-farm training of biologically based methods for integrated crop management of stone fruits in New England

Development and on-farm training of biologically based methods for integrated crop management of stone fruits in New England

Summary

In order to obtain isolates of bacteria (Xanthomonas arboricola pv. pruni =Xap) and bacteriophage (=”phage”), nineteen orchards in Connecticut, New York, and Massachusetts, were sampled for plant material obtained from peach, nectarine, and plum trees. A total of 121 plant samples were collected and tested for the presence of Xap as well as phage. From these, 23 Xap strains were obtained, purified, and stored. Additionally, 43 phage strains were obtained, identified as Xap-specific bacteriophage based on their ability to form phage plaques on Xap cultures. Laboratory studies were conducted to determine phage diversity, resulting in the conclusion that all 43 phage strains are functionally equivalent, based on cross-reactivity experiments using Xap strains lysogenic for a subset of 15 phage strains from the phage collection. A presentation on the goals, experimental plan, and research results to date was made in December 2010 to Connecticut growers at the annual meeting of the Connecticut Pomological Society. In January 2010, 60 O’Henry peach seedlings were potted and forced in the greenhouse. Bioassays were attempted in the greenhouse in March-April of 2010, but failed due to inability to infect trees under greenhouse conditions that exceeded the range of those acceptable for bacterial infection. All trees are now undergoing a 750-hour chill period in preparation for forcing in February 2011, after which branches will be tested in detached branch bioassays in the growth chamber, where conditions can be more accurately controlled. Assuming we are able to effect infections, we will then test the bacteriophage/bacterium interaction in vivo.

In order to obtain isolates of bacteria (Xanthomonas arboricola pv. pruni =Xap) and bacteriophage (=”phage”), nineteen orchards in Connecticut, New York, and Massachusetts, were sampled for plant material obtained from peach, nectarine, and plum trees. A total of 121 plant samples were collected and tested for the presence of Xap as well as phage. From these, 23 Xap strains were obtained, purified, and stored. Additionally, 43 phage strains were obtained, identified as Xap-specific bacteriophage based on their ability to form phage plaques on Xap cultures. Laboratory studies were conducted to determine phage diversity, resulting in the conclusion that all 43 phage strains are functionally equivalent, based on cross-reactivity experiments using Xap strains lysogenic for a subset of 15 phage strains from the phage collection. A presentation on the goals and experimental plan was made in December 2009 to an audience of approximately 60 Connecticut growers at the annual meeting of the Connecticut Pomological Society. In January 2010, 60 O’Henry peach seedlings were potted in the greenhouse, and are being maintained in preparation for greenhouse bioassays, which will test the bacteria/phage interaction in vivo.

Objectives/Performance Targets

Develop effective bacteriophage-based biological control strategy for controlling peach bacterial spot, which would provide an environmentally friendly and feasible alternative to the current chemical-based strategies. Additional target is a publication of the results of this project in a peer-reviewed scientific journal.

Develop effective bacteriophage-based biological control strategy for controlling peach bacterial spot, which would provide an environmentally friendly and feasible alternative to the current chemical-based strategies. Additional target is a publication of the results of this project in a peer-reviewed scientific journal.

Accomplishments/Milestones

MILESTONE 1 (Isolate 40 bacteriophages from New England stone-fruit orchards). Progress: 43 phages were isolated in the spring and summer of 2009 from a total of 19 orchards: two from New York, one from Massachusetts, and the remainder in Connecticut. Each phage was tested against the 23 Xap strains. Additionally, we have demonstrated that all 43 phages behave identically, and that phage immunity conferred through lysogenicity with one phage strain constitutes immunity to all 42 others. This experiment was repeated. An important conclusion from this result is that we will not have a mixture of genetically variable phage strains to use in the bioassays, greenhouse, and orchard trials.
MILESTONE 2 (Ten phages that show potential as biological control agents based on preliminary laboratory tests will be evaluated in bioassays).
Progress: This experiment is about to be underway. However, as noted above, we could identify no functionally genetic differences among the phage strains; therefore, we will test five phages representing the maximum geographic distribution.
MILESTONE 3 (Five phages that show potential based on bioassays will be mixed together and used in subsequent greenhouse trials). Progress: No progress to date; to be performed winter of 2010-2011.
MILESTONE 4 (Three phage-based treatments (phage mixture alone or together with other biological control agents) that provide equal or superior disease suppression compared to the currently used copper/antibiotic-based control methods in greenhouse trials will be chosen for further trials). Progress: 160 Sweet Dream peach trees on Lovell rootstock will be arriving in April for planting in three Connecticut orchards. Bioassay trials will be performed spring 2012.
MILESTONE 5 (Two phage-based treatments that provide equal or superior disease suppression compared to the currently used copper/antibiotic-based control methods in orchard trials will be recognized as environmentally friendly alternatives). Progress: No progress to date; to be performed spring-summer 2012.

Impacts and Contributions/Outcomes

Forty-three phage strains have been obtained from stone fruit orchards in Connecticut, Massachusetts, and eastern New York. These are equally virulent on all 23 of the Xap isolates that were collected from the same distribution of orchards. Therefore, we are making progress towards our goal of developing a phage-based biological control strategy for controlling bacterial spot of stone fruit. We will attain this outcome through greenhouse bioassays to be performed this spring, followed in winter/spring 2010/2011 by greenhouse trials comparing the use of phage against and in combination with currently used chemical and antibiotic based methods of bacterial spot control. Orchards of peach trees planted this spring will then be used in orchard trials in spring/summer 2012, comparing phage against and in combination with two other methods, which will be chosen based on results of greenhouse trials. Once results are obtained from these experiments, we will publish the results and conclusions in a peer-reviewed journal, such as Phytopathology.

Collaborators: