Recipient Organization
WASHINGTON STATE UNIVERSITY
240 FRENCH ADMINISTRATION BLDG
PULLMAN,WA 99164-0001
Performing Department
Plant Pathology
Non Technical Summary
This Research-led project addresses the CPPM Focus Area (1) of Plant Protection Tactics and Tools and aims to develop adequate management strategies against gay mold, caused by Botrytis cinerea, one of the most important diseases of apple and pear in the Pacific Northwest. Recent regional large surveys revealed gray mold as the most important storage disease beside blue mold. Infections by B. cinerea occur in the orchards but because of the perception that gray mold is a storage problem, preharvest management is often delayed to the last two weeks before harvest or after harvest. The dearth of knowledge about the epidemiology of Botrytis in apple and pear in the PNW and elsewhere rendered current management strategies ineffective. Moreover, because none of the current apple or pear cultivars is resistant to B. cinerea, management relies solely on fungicide applications which differ between the various growing regions in the PNW. Whether the recent surge in gray mold is due to the emergence of resistance to fungicides, in use for more than three decades, remains unknown. Our project goals are to provide the pome fruit industries with: (1) epidemiological knowledge to aid growers and packers in gray mold management decisions, (2) regional documentation of fungicide resistance levels to support mid- and long-term control, (3) epidemiology-based and improved management tactics pertinent in various production areas, (4) economic analysis tools to increase profitability of stakeholders, and (5) continue regional and national extension efforts to educate pome fruit growers about gray mold best management tactics.
Animal Health Component
60%
Research Effort Categories
Basic
20%
Applied
60%
Developmental
20%
Goals / Objectives
This project aims to develop adequate management strategies against gay mold, caused by Botrytis cinerea, one of the most important diseases of apple and pear in the Pacific Northwest. Recent regional large surveys revealed gray mold as the most important storage disease beside blue mold. Infections by B. cinerea occur in the orchards but because of the perception that gray mold is a storage problem, preharvest management is often delayed to the last two weeks before harvest or after harvest. The dearth of knowledge about the epidemiology of Botrytis in apple and pear in the PNW and elsewhere rendered current management strategies ineffective. Moreover, because none of the current apple or pear cultivars is resistant to B. cinerea, management relies solely on fungicide applications which differ between the various growing regions in the PNW. Whether the recent surge in gray mold is due to the emergence of resistance to fungicides, in use for more than three decades, remains unknown. Our project goals are to provide the pome fruit industries with: (1) epidemiological knowledge to aid growers and packers in gray mold management decisions, (2) regional documentation of fungicide resistance levels to support mid- and long-term control, (3) epidemiology-based and improved management tactics pertinent in various production areas, (4) economic analysis tools to increase profitability of stakeholders, and (5) continue regional and national extension efforts to educate pome fruit growers about gray mold best management tactics. Specific Objectives are:OBJECTIVE 1. Acquire novel epidemiological knowledge on Botrytis in different growing regions of the PNW. [Research]. Hypothesis: Botrytis can cause latent infections through flower organs, and that fruit infections occurring at early stages of fruit setting are more difficult to control. Approach: We will use qPCR and selective media to track and quantify the fungus in relation to weather conditions occurring in different regions of the PNW. Impacts: In the short- and intermediate terms, targeted sprays will sanitize flowers and fruits at early stages to lower latent infections. This could be measured by reduction in Botrytis inoculum before harvest and reduction in fruit losses in storage. In the long-term, we will use epidemiological and meteorological data to develop a predictive model (not in this project) valid for different regions.OBJECTIVE 2. Provide quick, location-specific profiles to support mid- and long-term regionally-based fungicide resistance mitigation for gray mold. [Research]. Hypothesis: Resistance to preharvest fungicides has emerged at low frequencies and resistance to postharvest fungicides is slightly higher because of frequent usage and extended contact between the fungus and the fungicide. Approach: We will use large Botrytis populations (5,700) to test for fungicide resistance and provide feedback and recommendations to growers and packers. Impacts: Knowing the key timing for Botrytis infections (Obj. 1) and using the most effective fungicides will enhance gray mold control, reduce fruit losses, and increase returns. This could be measured by reduction in: i) fruit losses, ii) resistance frequencies, and iii) spray cost (number of fungicide sprays).OBJECTIVE 3. Improve the use of conventional products to develop targeted- preharvest spray programs. [Research]. Hypothesis. Current management strategy consisting of a late single fungicide application near harvest is not effective. Reducing early-season infections of flowers and fruit will reduce gray mold postharvest. Approach: We will develop timely spray programs to reduce preharvest Botrytis infections using most effective materials based on knowledge from fungicide resistance monitoring. Impacts: Timely application of most effective materials will reduce losses and increase returns. Moreover, better Botrytis management during the season will reduce overwintering inoculum and enhance orchard sanitation. This could be measured by the reduction in fruit losses in storage, the number of sprays and packer/grower returns.OBJECTIVE 4. Estimate profitability of new management approaches and conduct outreach activities. [Extension]. We will assess if the new management tactics, from Obj. 3, are economically feasible. For this we will estimate the costs and benefits of each proposed strategy and provide growers and packers with decision making tools to maximize their profits. We will use traditional and online extension resources to ensure effective technology transfer. Impacts: We will provide better training and up-to-date information to stakeholders in terms of best management practices using the knowledge gained from this project to empower them towards making most effective and economical management decisions.
Project Methods
OBJECTIVE 1. Acquire novel epidemiological knowledge on Botrytis in different growing regions of the PNW. Preharvest sampling: In spring of 2021, sixty apple and pear blossoms will be collected from each orchard. Five flowers will be collected at petal fall from different sections (lower-, mid, and top canopy) and the sides of each tree for a total of 60 samples per orchard. Afterward, samples will be collected similarly from the same tagged trees at 30 (fruitlet), 90 (unripe fruit), and 150 days (commercial maturity) post-bloom. Samples will be transported in separate clean bags to the Tree Fruit Research and Extension Center in Wenatchee. At each sampling time, the 60 samples will be divided into two subsamples of 30 samples each. One subsample will be used for DNA analyses by qPCR, whereas the second subsample will be used for Botrytis enumeration on a semi-selective medium (BSM) as described below. Sampling will be conducted similarly in 2022.Postharvest (cold storage) sampling: At commercial maturity, 160 fruit will be harvested from each orchard as described above (field sampling) and will be stored in regular atmosphere at 1°C. After 6 months of storage, thirty fruit will be used for qPCR and 30 for BSM analyses as explained below. The remaining 100 fruit will be stored for up to 9 months to estimate gray mold incidence and correlate with preharvest Botrytis load, sprays, and weather conditions among regions.Botrytis detection and quantification: The 30-subsample fruit collected at different preharvest time-points and after 6 months of storage will be processed immediately after sampling or kept at -80°C until used. Fruit will be peeled to analyze separately the cuticle and the flesh of the fruit in order to distinguish infestations (spores present on the surface) from endophyte infections (latent infections). Peelers will be sterilized after each fruit to avoid cross contaminations. Samples will be blended, and 1 g of each sample will be used for DNA extraction and qPCR analysis (9). After DNA extraction, the 10 samples from each sub-block, will be pooled to obtain a total of 9 samples per orchard. qPCR assays will be run in triplicate and performed twice.Data analysis for Activities 1a-b: The incidence of Botrytis will be calculated as % of samples positive for Botrytis based on qPCR and BSM tests; the data will be analyzed separately (for each test) and conjointly. The biomass of B. cinerea DNA will be expressed as ng/g of apple or pear tissue and will be analyzed separately for the flesh and the cuticle. ANOVA and repeated measure regressions will be used to conduct multiple range comparisons (incidence, DNA quantity) to find differences between orchards and sampling times. Cumulative annual values of gray mold incidence per orchard will be computed and analyzed using a generalized linear mixed model.OBJECTIVE 2. Provide quick, location-specific profiles to support mid- and long-term regionally-based fungicide resistance mitigation for gray mold.We collected about 5,700 Botrytis isolates from previous surveys in 2017 and 2018 from 600 lots representing 30 packinghouses. These isolates have been purified and stored in 20% glycerol at -20°C. Pre- and postharvest spray records obtained from all surveyed lots will be analyzed and correlated with potential resistance frequencies to map fungicide resistance in the region. We propose to test the sensitivity of 4,000 isolates to 4 preharvest and 4 postharvest fungicides. Boscalid (BOSC) and pyraclostrobin (PYRA) have been used preharvest since 2004 in the PNW, whereas fluopyram (FLUOP) and fluxapyroxad (FLUX) were registered in 2014 and 2016, respectively. Among the 4 current postharvest fungicides, thiabendazole (TBZ) has been in use for more than 4 decades, whereas pyrimethanil (PYRI) and fludioxonil (FDL) were registered in 2004 and have been used frequently by packers. Sensitivity assays will be conducted using spore germination and germ tube length assays (4,20). The selected discriminatory doses of fungicides and media were determined previously to be appropriate for fungicide sensitivity tests in B. cinerea.Each fungicide will be mixed with the corresponding autoclaved medium, poured into 150-mm Petri dishes and allowed to solidify. Fungicide-free plates will be used as controls. The 150-mm plate-based assay allows to test for 60 isolates at a time using a grid of 60 squares (4 x 4 mm) on a transparent film placed under the plate. Resistance frequencies will be compared to pre and postharvest spray records by lots to generate site-specific resistance profiles (RP). Each RP will include resistance frequencies to all fungicides, history of sprays and recommendations. RPs will be sent to participating growers and packers before the beginning of the 2022 growing season to make appropriate spray decisions before the start of the new seasons.Determine fungicide sensitivity phenotypes in Botrytis "Orchard Populations". We will test if resistance to some fungicides is selected in the orchard. Isolates of Botrytis that will be collected from preharvest samples from Activity 1-a will be tested for fungicide sensitivity for pre- and postharvest fungicides (Activity 2-a). The orchard sites (Activity 1-a,) have already been surveyed in 2019 and a representative of "Cold Storage" isolates are available. However, to compare "Orchard" and "Cold Storage" isolates from the same season, we will collect additional "Cold Storage" isolates from these 4 lots in 2021.OBJECTIVE 3. Improve the use of conventional products to develop timely targeted- preharvest spray programs. Evaluate multiple spray approaches in an experimental orchard: We will conduct field trials to test multiple spray programs and timings at an experimental 1-acre block at Sunrise orchard in Wenatchee, WA (47.31304, -120.067987). We will use a 12-year-old Fuji block planted on Nick 29 rootstock. In addition to the untreated control, we will test three preharvest spray types outlined in Table 3. We aim to compare the industry current standard program with timely applications. Trials will be set in a randomized complete block design with four replicate trees per treatment and fungicides will be sprayed using backpack sprayers between April 2021 and October 2021. At commercial maturity, 200 fruit/treatment (50 fruit/replicate tree) will be harvested, drenched with Penbotec, and stored at 1°C. Fruit will be inspected for gray mold after 3 months of storage and every two months thereafter for up to 9 months to determine incidence.Assess efficacy of management approaches in commercial orchards. We will conduct trials at one apple and one pear commercial orchards in Year 2 and 3.At each site, a one-acre block will be divided by 2 sub-blocks. One sub-block will be sprayed following the grower spray calendar (one 7 day preharvest spray) and the second sub-block will be sprayed using our proposed program of 3 to 4 sprays throughout the season based on preliminary data and results from Activity 3-a. At commercial maturity, fruit from each sub-block will be harvested and stored at respective warehouses until packed. Decayed fruit will be collected to determine incidence and decay type by Amiri team.OBJECTIVE 4: Estimate profitability of new management approaches and develop and conduct outreach regional activities.Profitability of new management approaches: PD Gallardo will work alongside PD Amiri and with participant orchard/warehouse managers to gather data on cost centers for apple and pear production.