Alternaria Control Using Biocontrol Yeast in Organic Pistachio Production Systems

Project Overview

Project Type: Research and Education
Funds awarded in 2006: $110,286.00
Projected End Date: 12/31/2008
Region: Western
State: California
Principal Investigator:

Annual Reports


  • Nuts: general nuts


  • Education and Training: on-farm/ranch research
  • Pest Management: biological control

    Proposal abstract:

    This proposal addresses the need for an effective alternative to chemical control of Alternaria alternata, an economically important fungal pathogen of pistachio, in organic pistachio production systems. Specific objectives are 1) test of a selected biocontrol yeast for efficacy in controlling Alternaria alternata, and 2) development and testing of additional biocontrol agents (yeast strains) to replace chemical fungicides in organic and non-organic pistachio orchards with an environmentally benign alternative. A selected Pichia anomala yeast strain has been evaluated for three years with promising results for the control of Aspergillus flavus (which produces aflatoxin contamination), phytotoxicity, and field durability. A 2005 small-scale test in an organic production orchard showed statistically significant levels of control of Alternaria alternata. Proposed research includes a) expansion of field trials to evaluate Alternaria alternata control at a commercial scale, combined with evaluation prior to and at crop harvest, and b) in-vitro testing and small-scale field testing of additional promising yeast strains for A. alternaria control. Expected near-term outcomes include proof of concept for biocontrol of A. alternata with yeast, development of information needed for registration and commercialization of Pichia anomala yeast strain WRL-076 for control of A. alternata in pistachio, identification and testing of additional yeast strains to provide alternatives to chemical control of A. alternata in pistachio. Longer-term outcomes are reduced use of chemical pesticides in commercial pistachio orchards with reduced risk to the public and improved yield and profitability for producers of organically grown pistachios.

    Project objectives from proposal:

    1. Evaluate biocontrol yeast that has been released by USDA-ARS (presently going through registration as a control measure for Aspergillus flavus).
    2. Test additional biocontrol yeast strains that have shown particular efficacy against multiple A. alternaria strains found on pistachio.
    3. Test promising strains from objective 2 in the greenhouse and field.

    Our initial objective is to test the efficacy of a specific strain (WRL-076) of Pichia anomala that was isolated from the natural environment (a pistachio orchard) for control of A. alternata at a field scale in an organic production environment where possible interaction effects from prior pesticide sprays are absent.
    We will continue to screen additional strains of naturally occurring yeast for efficacy against multiple A. alternata biotypes. Initial tests will be done as in-vitro tests, followed by greenhouse tests in 2007. If successful control is shown in greenhouse tests, small-scale field tests will be conducted in 2008.

    Outreach Plan

    Our outreach plan has three components. Dr. Holtz, Madera county farm advisor, will be the primary agent for the outreach effort, since this is a major part of his position description. Initial outreach will be focused on on-farm demonstrations to growers of treatment efficacy and an explanation of the application procedures used and results obtained in cooperation with Mr. Braga on whose ranch the initial experiments are being conducted. As qualitative observations and quantitative evaluations are obtained, dissemination of results will expand to Dr. Holtz’s newsletter, which is available both in hardcopy and as a web page ( It is also widely accessible to growers throughout the state through the fruit and nut information center at UC Davis ( }. The University of California sponsors a ‘Pistachio Day’ for growers where applied research is demonstrated and current recommendations on production practices are highlighted as well as presentation of new and emerging technologies and problems.
    Popular dissemination of results and recommendations for use will occur through California Agriculture and the UC IPM Online, Statewide Integrated Pest Management Program website. The research is also expected to produce results suitable for refereed publications such as Plant Disease, Hort. Technology, and/or The Journal of the American Society for Horticultural Science. These conventional venues for information dissemination will be of use primarily for academic scientists, nationally and internationally, and other county farm advisors in California. California Agriculture is a technical/popular publication with wide distribution to the public by the University of California and will be used to disseminate results to the general public in a semi-technical format. In addition to the aforementioned avenues for information dissemination, the Western SARE program requires annual reports, which are posted on their website and which are available to the interested public.


    Evaluations consists of 3 parts, a scientific evaluation of spray efficacy, an evaluation of the effectiveness of the outreach effort, and commercialization. Scientific evaluation of the treatment efficacy will be the primary responsibility of Drs. Parfitt and Holtz. Evaluations will consist of a visual score of isolated leaves for lesions (year 1), and presence or absence of Alternaria damage on nut clusters in the field as well as visual evaluations of leaf damage in the field (generally quite obvious when there is a significant problem). Lesions from nuts and leaves will be sampled and cultured to confirm the presence of A. alternata. Yield evaluations will be done in years 2 and 3 to confirm the increased value to be derived from application of biocontrol yeast vs. the controls.

    Evaluation of outreach efficacy will be done by Dr. Holtz. This will consist of grower feedback through attendance at field day demonstrations, with question and answer sessions, and a questionnaire to be made available to growers at the annual UC Pistachio Day event, which is typically attended by several hundred growers. Technical evaluation of the research results will occur through the mechanism of publication submission, review by external journal referees, and publication in refereed journals.

    Commercialization of biocontrol yeast is the ultimate measure of the program’s success. This evaluation criteria can be measured by registration of specific biocontrol yeasts identified as effective by the program. As registration is being pursued (responsibility of Dr. Hua), commercial vendors will be solicited for marketing of the product.


    Part 1:
    May 2006: Prepare WRL-076 yeast inoculum
    June 2006: Apply first spray of WRL-076 inoculum. Collect nut subsamples to monitor residual yeast levels.
    July 2006: Do second spray of WRL-076 inoculum. Collect nut subsamples to monitor residual yeast levels.
    August 2006: Do third spray of WRL-076 inoculum. Take cluster count field observations of percent damage from A. alternata for all treatments. Collect nut subsamples to monitor residual yeast levels.
    September 2006: Do final spray of WRL-076 inoculum. Take cluster count field observations of percent damage from A. alternata for all treatments. Collect nut subsamples to monitor residual yeast levels. Commercial nut harvest of the treatments will be done by block. Yield and grade out data to be collected from the harvest.
    October 2006: Continue to process previously collected nut subsamples. Perform data analysis on cluster counts for A. alternata cluster data.
    November 2006: Perform data analysis on yield data from harvested nuts.
    December - January 2006: Finish data analysis of yield data and yeast survival evaluations. Presentation of results at grower meetings.
    Part 2:
    June 2006 - August 2006: Evaluate additional yeast strains for their ability to control A. alternata in-vitro.
    August-September 2006: Collect new inoculum of A. alternata from northern, central, and southern growing areas. Test yeast strains against new cultures. Obtain or graft potted trees for 2007 greenhouse tests.

    Part 1:
    Continue spray schedule as shown for 2006, with all treatments to be applied to the same trees used in the 2006 protocol.
    Part 2:
    October 2006 - Feb. 200: Test yeast strains against new inoculum.
    April - Sept. 2007. Conduct greenhouse test using strains selected in 2006

    Part 1:
    If results from 2006 and 2007 show significant differences between control and treatment (WRL-076 yeast), the trial will be repeated as for 2006 and 2007. 2008 will be an ‘on’ year as will 2006. Pistachio demonstrates a marked alternate bearing habit, with high nut production years followed by low nut production years. In ‘on’ years, Alternaria alternata is much more prevalent on pistachio than in ‘off’ years.
    October 2008 - May 2009: Complete data analysis, write up manuscripts, present talks at grower meetings.
    Part 2-3:
    Sept. 2007 - March 2008: Obtain permission for small-scale field test from CDFA and appropriate county officials. Produce yeast inoculum for field test.
    June 2008 - September 2008: Conduct small-scale field test using the format described in the current stage of knowledge section for the 2005 field test (e.g. 4 paired sets of trees, evaluated by cluster count and single tree yield.
    October 2008 - May 2009: Complete data analysis, write up manuscripts, present talks at grower meetings.

    Data collection and analysis: Data to be collected for the Alternaria alternata control evaluation: Cluster counts (dead vs. viable clusters) will be taken on each tree. Subsample and block error terms will be computed and used for ANOVAs. Each block x treatment will be harvested separately. Yield and quality data will be taken and ANOVAs performed. Yeast concentrations will also be tested on these nuts at the same time using differential media treatments that have been developed and refined by Dr. Hua. Yeast levels will be monitored on normal 100 nut samples (per treatment x replicate) across the season at 3 week intervals.
    Yeast counts will be transformed using the square root transformation and subjected to ANOVA.

    Any opinions, findings, conclusions, or recommendations expressed in this publication are those of the author(s) and do not necessarily reflect the view of the U.S. Department of Agriculture or SARE.