Progress 09/01/19 to 02/28/22
Outputs
Target Audience:NY produces over 250 million pounds of onions from 7200 acres, on predominately muck soils, and accounts for 97% of the production in Northeast USA . This project involved growers from 16 farms in western NY, who participate in CCE educatorHoepting's onion scouting program which involves weekly meetings during the growing season to share scouting data, spray records, and management recommendations. These growers make up approximately 50% of the muck acreage in NY.. Further updates have been given to growers in the Orange County (eastern NY), through participation in the CCE Orange County Onion Schools in 2021 and 2022. Changes/Problems:No problems in the final reporting period. Previous problems that were documented through the project included Covid 19 restrictions in 2020 which prevented regular travel and field visits, leading to a restricted set of data. Malfunctions with dataloggers in the field also prevented collection of data for disease forecasting. The unusually large number of mutations in the SDH genes associated with fungicide resistance prevented the development of a practical molecular test to detect resistance to FRAC 7 fungicides. However, PCR primers to amplify the region and protocols to sequence the region were developed to allow molecular confirmation of fungicide resistance. What opportunities for training and professional development has the project provided?The project afforded the opportunity for a posdoctorate to gain experience in epidemiology and molecular techniques. How have the results been disseminated to communities of interest?Presentations were made to Empire State Producer's Expo in 2020 and 2021, the 69th Annual Muck Vegetable Growers Conference, 2021 and the Orange County Onion School 2021. A growers guide to recommended fungicide programs for control of SLB was provided on-line. Two articles on foliar disease were published in the grower magazine VegEdge. A guided tourof the 2021 SLB fungicide trialwas given to growers, crop consultants, and chemical industry representatives. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
Objective 1. To determine the utility of current disease forecasting models for optimizing timing of fungicide applications in onion. 2020 trial. Onion crops in Wayne Co. (1), Oswego Co. (2), and Elba (3) were sampled periodically in 2020 (Table 1). At each time 20 leaves were collected, sealed in plastic bags at high humidity for 7-10 days, then examined under the microscope for the presence of S. vesicarium. Due to Covid-19 restrictions spore traps were not deployed . Restrictions also prevented regular visits to the field, so issues with dataloggers prevented the collection of sufficient weather data to assess the BSPcast forecaster. However, information was collected on epidemic progression. All but two crops had a low level of infection on June 6-10. Incidence remained low until July 13-15 when it increased, or began to increase in most crops (Table 1). By July 27-29, all crops had over 40% incidence, and all but one crop had incidence over 80% by Aug 3-5 (Table 1). The data illustrated the potential for SLB to rapidly build up in onion crops in mid-season. Table 1. Incidence of Stemphylium vesicarium in onion leaves in five commercial crops in New York during the 2020 season. Farm and location: Sample time: Dis (Oswego) Duns (Oswego) Torr Nth (Elba) Mort (Elba) Tutt (Wayne) June 6-10 8 2 16 0 22 June 15-17 0 4.2 0 2 4.4 June 22-24 0 0 0 0 0 June 29-Jul 1 12.2 0 6 2 0 July 6-8 14 2 0 0 0 July 13-15 58 22.4 18.4 8 14 July20- 22 13 6 34.0 18.4 30 July 27-29 66 54 48 46 100 Aug 3-5 86 89.6 93.8 87.8 44 Aug 10-12 100 98 96 94 88 In 2021, four onion crops in Elba NY were monitored from June to September (Table 2). Data loggers measuring air temperature and leaf wetness at 15 minute intervals were deployed in each field. At each sample time, leaves (30) were collected, and assessed as for 2020, to determine fungal incidence. Spore traps were deployed (rotorod and Burkard) in each field. S. vesicarium was detected at between 5-20% incidence in all 4 crops at the earliest sample time, Jun 9 (Table 2). Stemphylium vesicarium built up more rapidly in transplant crop M8, with an incidence of 82% on 6/23. By comparison, the incidence in direct seeded crops M, G and B at this time was 45%, 5% and 13% respectively (Table 2). The incidence of S. vesicarium increased more rapidly in Crop M, than Crops G or B. The incidence in Crop M reached 53% on 7/7 and 96.7% on 7/28. However, Crops G and B remained below 30% incidence until Jul 28, at which time incidence increased in crops G and B to 70.0 and 93.3% respectively (Table 2). Table 2. Incidence of Stemphylium vesicarium in onion leaves in four commercial crops in Elba, New York during the 2021 season. SLB incidence (% of leaves infected) Field 6/9/2021 6/23/2021 7/7/2021 7/14/2021 7/21/2021 7/28/2021 8/4/21 M8 20.0 82.0 100.0 100.0 93.3 n/a n/a M 10.0 45.0 53.0 73.0 56.7 96.7 96.7 G 18.0 5.0 13.0 28.0 13.3 70 50.0 B 5.0 13.0 30.0 28.0 23.3 93.3 86.7 The forecasting program BSPcast is used in Europe to predict periods of infection of S. vesicarium, cause of Brown Spot in pear. A daily risk value R is calculated from combinations of temperature and periods of leaf wetness, with R≥0.2 equating to infection. However, BSPcast was a poor predictor of increases in incidence of S. vesicarium in onion crops in Elba, NY. For example, in Crop M, R≥0.2 events were associated with increases in incidence for sample times 1, 3, and 5. However, at time 2 there was little increase in incidence despite the previous 4 of 7 days having R≥0.2 events (Table 3). Further, at time 4 there was a reduction in incidence despite previous R≥0.2 events (Table 3). Similar results were obtained at other sites. Future work should re-examine the relationship between infection and leaf wetness/temperature, to modify BSPcast to the onion pathosystem. Table 3. Lack of relationship between BSPcast risk values (R) and increase in the incidence of Stemphylium vesicarium at intervals during the 2021 season for Crop M. Sample time: 6/9/2021 - 6/22/2021 6/23/2021 - 7/6/2021 7/7/2021 - 7/13/2021 7/14/2021 - 7/20/2021 7/21/2021 - 7/28/2021 Days during sampling interval 14 14 7 7 7 Days with R ≥ 0.2 2 9 4 3 1 Change in incidence of S. vesicarium 10-45% 45-53% 53-73% 73-57% 57-96% Stemphylium vesicarium reportedly has a sexual stage (Pleospora allii) which produces ascospores in pseudothecia, important for long range transmission. In haploid ascomycetes, fertility depends on the configuration of genes at the mating type locus (MAT). Self-sterile individuals have only one of two single-copy genes (MAT1-1 or MAT1-2) at this locus, whereas haploid self-fertile individuals contain both. A multiplex PCR was developed and tested on 131 isolates of S. vescarium. All individual isolates contained both MAT1-1 and MAT1-2, indicating the isolates to homothallic and potentially self-fertile. Objective 2 Develop and deploy tools to quickly monitor changes in the fungicide resistance status of S. vesicarium. FRAC 7 fungicides Several fungicide active ingredients in the FRAC 7 group are utilized in onion production in NY. In fungi, mutations in the Sdh B, Sdh C and Sdh D genes are associated with resistance to the FRAC 7 fungicides. PCR primers were designed to allow amplification and sequencing of these genes in S. vesicarium. Sensitivity of 234 isolates of S. vesicarium to fluxapyroxad, fluopyram, boscalid and penthiopyrad was assessed by growing fungi on petri plates containing media amended with different concentrations of fungicide and calculating the effective concentration necessary to inhibit fungal growth by 50% in comparison to growth on non-amended plates (EC50). Isolates were considered sensitive (EC50 < 1 µg/ml), moderately insensitive (EC50 1-10 µg/ml) or highly insensitive EC50 > 10 µg/ml) For isolates sensitive to fluopyram, fluxapyroxad, penthiopyrad and boscalid, only 3%, 8%, 9% and 0% of isolates possessed mutations in the Sdh genes respectively. Only two mutations (H134R and D126N) on the Sdh C and Sdh D genes respectively were detected in sensitive isolates. For isolates moderately insensitive to fluopyram, fluxapyroxad, penthiopyrad and boscalid, some 75%, 60%, 60% and 30% respectively had mutations in the Sdh genes. There was a wide variety of gene mutations detected in moderately insensitive isolates including P230H (Sdh B), H134N, H134R and C135 R (Sdh C), and D126E, Q12K, S120P and D126N (Sdh D). Similarly for isolates highly insensitive to fluopyram, fluxapyroxad, penthiopyrad and boscalid, some 95%, 100%, 100% and 57% of isolates respectively had mutations in the Sdh genes respectively. There were a wide variety of mutations in the highly insensitive category including P230H (Sdh B), G79R, H134R and C135R (Sdh C) and D126E, Q12K, S120P and D126N (Sdh D). The most common mutations associated with insensitivity to fluxapyroxad, fluopyram and penthiopyrad were G79R, H134R and C135R. Mutations have developed rapidly in S. vesicarium, with 0%, 65% and 100% of S. vesicarium isolates tested from 2016, 2018 and 2020 respectively, possessing mutation in the Sdh genes, associated with declining field performance of FRAC 7 fungicides. The unusually large number of SNP's associated with FRAC 7 resistance in S. vesicarium negated the development of a multiplex PCR to more rapidly detect resistance in isolates. FRAC 3 fungicides Mutations in the Cyp51 gene have been associated with resistance to FRAC 3 fungicides in some fungi. PCR primers and protocols were developed to amplify the Cyp51 gene in S. vesicarium. Sequencing of 5 isolates of S. vesicarium which were sensitive and 5 which were insensitive to FRAC 3 fungicides failed to detect any sequence differences in the Cyp51 gene, suggesting resistance to be associated with other mechanisms. This finding negated the development of a PCR method to more rapidly assess the fungicide resistance status to FRAC 3 fungicides.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2021
Citation:
Hay, F.S., Heck, D., Klein, A., Hoepting, C., and S.J. Pethybridge 2021. Spatio-temporal analysis of epidemics of Stemphylium leaf blight caused by Stemphylium vesicarium in onion crops in New York. Plant Disease. https://doi.org/10.1094/PDIS-07-21-1587-RE
- Type:
Journal Articles
Status:
Published
Year Published:
2022
Citation:
Hay, F. S., Stricker, S., Gossen, B. D., McDonald, M. R., Heck, D. W., Hoepting, C. A., Sharma, S., and Pethybridge, S. J. 2022. Stemphylium leaf blight of onion: A re-emerging threat to onion production in eastern North America. Feature Article. Plant Dis. 105:3780-3794. https://doi.org/10.1094/PDIS-05-21-0903-FE.
- Type:
Journal Articles
Status:
Published
Year Published:
2022
Citation:
Hay, F. S., Heck, D. W., Sharma, S., Klein, A., Hoepting, C., and Pethybridge, S. J. 2022. Stemphylium leaf blight of onion. Plant Disease Lesson. The Plant Health Instructor 22: 10.1094/PHI-P-2022-01-0001
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Progress 09/01/20 to 08/31/21
Outputs
Target Audience:Results of this project were communicated to growers as part of the following presentations made during 2021. Hay, F., Heck, D., Hoepting, C., and S. Pethybridge. 2021. Stemphylium leaf blight - current status of fungicide insensitivity. W3008 Multistate Project Annual Meeting. February 3, 2021. (20 attendees) Hay, F., Heck, D., and K. Ayer. 2021. Current status of Stemphylium leaf blight fungicide resistance in onion in New York. Empire State Producers Expo 2021. Fragile FRAC 7 Fungicides: Part I -Onion Critical Issues. Virtual meeting, Jan 14, 2021. (50 attendees). Heck, D., Hay, F., Hoepting, C.A. and S.J. Pethybridge (2021). 'Stemphylium leaf blight in onion - a continually evolving problem for growers in New York'. 69th Annual Muck Vegetable Growers Conference 2021. Virtual meeting. April 1, 2021. (50 attendees). Changes/Problems:The amount of field data collected as part of objective 1 was significantly reduced in 2020 due to Covid 19 restrictions and malfunctions of weather stations. Some further malfunctions of weather stations occurred during 2021, despite purchasing new equipment and trialing equipment prior to deployment in the field. The loss of data in 2021 was reduced in comparison to 2020 due to the lack of Covid 19 restrictions and hence the ability to visit fields more frequently. Unfortunately, while the project will present some valuable preliminary data, the amount of data collected during the project is likely to be insufficient to fully validate theuse of BSPcast as a tool for onion growers to schedule fungicides. A 6- month no-cost extension was granted to allow further characterization of the molecular mechanisms associated with fungicide insensitivity and mating type, and to allow statistical analysis of weather data in relation to disease incidence and spore release. What opportunities for training and professional development has the project provided?The project has afforded opportunities for a postdoctorate to develop skills in developing PCR protocols and sequencing of genes related to fungicide resistance. How have the results been disseminated to communities of interest?Oral presentations to growers have been made at the following events: Hay, F., Heck, D., Hoepting, C., and S. Pethybridge. 2021. Stemphylium leaf blight - current status of fungicide insensitivity. W3008 Multistate Project Annual Meeting. February 3, 2021. (20 attendees) Hay, F., Heck, D., and K. Ayer. 2021. Current status of Stemphylium leaf blight fungicide resistance in onion in New York. Empire State Producers Expo 2021. Fragile FRAC 7 Fungicides: Part I -Onion Critical Issues. Virtual meeting, Jan 14, 2021. (50 attendees). Heck, D., Hay, F., Hoepting, C.A. and S.J. Pethybridge (2021). 'Stemphylium leaf blight in onion - a continually evolving problem for growers in New York'. 69th Annual Muck Vegetable Growers Conference 2021. Virtual meeting. April 1, 2021. (50 attendees). Results were incorporated into disease management recommendations to growers in the following articles in a grower's magazine. Hoepting, C.A., and Hay, F. 2021. Part I: Onion fungicide research updates and new recommendations for control of Botrytis and Stemphylium leaf blights, 2021 VegEdge 17(12):8-10. Hoepting, C.A., and Hay, F. 2021. Part II: Onion fungicide research updates and new recommendations for control of Botrytis and Stemphylium leaf blights, 2021. VegEdge 17(13), 8-9. What do you plan to do during the next reporting period to accomplish the goals?A selection of S. vesicarium isolates with particular gene mutations or combinations of mutations in the Sdh genes will be tested for fungicide sensitivity to FRAC 7 fungicides to determine actual EC50 values. This will assist with identifying the most important gene mutations conferring fungicide insensitivity and will assist in developing a molecular test for fungicide insensitivity targeting the most important mutations. A selection of isolates will be screened for mating type genes to confirm the homothallic nature of S. vesicarium which has been determined on two isolates. Weather data collected over the growing seasonfrom 4 onion fields in each of 2020 and 2021 will be input into the disease forecaster BSPcast to determine periods of critical risk of infection and associated with weekly changes in disease incidence to determine the ability of the forecaster to predict periods of disease increase. Rods and tapes from spore traps collected from 4 onion fields in Elbawill be assessed for the presence of S. vesicarium, and spore release statistically associated with weather conditions.
Impacts
What was accomplished under these goals?
Objective 1. To determine the utility of current disease forecasting models for optimizing timing of fungicide applications in onion as a means of reducing economically wasteful and environmentally undesirable fungicide applications. A similar trial to 2020 was conducted in 2021 to obtain further data points to enable correlation of periods of critical risk of infection by S. vesicarium calculated using the forecasting model BSPcast, with increases in the incidence of S. vesicarium within crops. . Four onion crops were monitored in Elba NY from early June to early September 2021. Spectrum Watchdog 1650 weather station, measuring air temperature and leaf wetness at 15 minute intervals were deployed in each field. Leaves (30) were collected from each field on 6/9/2021, 6/23/2021, 7/7/2021, 7/14/2021, 7/21/2021, 7/28/2021, 8/4/2021 and 8/11/2021. Leaves were incubated in bags under high humidity, and observed under the microscope for signs of S. vesicarium and the incidence of S. vesicarium determined. Due to Covid-19 restrictions spore traps were not deployed in the crop during 2020. In 2021, two spore traps were deployed (rotorod and Burkard) in each field. At weekly intervals, rods and tape were collected from the rotorod and Burkard traps respectively. Burkard tape was cut into 6 h sections and along with rods have been stored. Leaves from the final two sampling periods are currently being assessed. The presence of S. vesicarium conidia on rods and tape from the spore traps will be detected during the latter part of 2021, by visualization of spores under the microscope, and DNA extraction and PCR with specific primers. Weather data from each of 2020 and 2021 will be input into the forecasting model BSPcast to calculate weekly critical risk values and these will be correlated with observed increases in SLB incidence between weeks. In addition weather data will be analyzed to determine any relationships with spore release either daily (Burkard) or weekly (Rotorod) Objective 2) Develop and deploy tools to quickly monitor changes in the fungicide resistance status of S. vesicarium to commonly used fungicides. Mechanisms of resistance to FRAC 7 fungicides in Stemphylium vesicarium Several fungicide active ingredients in the FRAC 7 group are utilized in onion production in NY, including fluxapyroxad, fluopyram, boscalid and penthiopyrad. Mutations in the Sdh B, Sdh C and Sdh D genes of fungi are associated with differences in sensitivity to the FRAC 7 fungicides. PCR primers were designed to allow successful amplification and sequencing of these genes in S. vesicarium. Isolates of S. vesicarium were chosen with differing sensitivity to 4 different FRAC 7 fungicide active ingredients (fluxapyroxad, fluopyram and penthiopyrad). Sensitivity was assessed by growing fungi on petri plates containing media amended with different concentrations of fungicide and calculating the effective concentration necessary to inhibit fungal growth by 50% in comparison to growth on non-amended plates (EC50). Isolates were placed into categories of sensitive (EC50 < 1 µg/ml), moderately insensitive (EC50 1-10 µg/ml) and highly insensitive EC50 > 10 µg/ml) For isolates sensitive to fluopyram, fluxapyroxad, penthiopyrad and boscalid, only 3%, 8%, 9% and 0% of isolates possessed mutations in the Sdh genes respectively. Only two mutations (H134R and D126N) on the Sdh C and Sdh D genes respectively were detected in sensitive isolates. For isolates moderately insensitive to fluopyram, fluxapyroxad, penthiopyrad and boscalid, some 75%, 60%, 60% and 30% respectively had mutations in the Sdh genes. There was a wide variety of gene mutations detected in moderately insensitive isolates including P230H (Sdh B), H134N, H134R and C135 R (Sdh C), and D126E, Q12K, S120P and D126N (Sdh D). Similarly for isolates highly insensitive to fluopyram, fluxapyroxad, penthiopyrad and boscalid, some 95%, 100%, 100% and 57% of isolates had mutations in the Sdh genes respectively. There were a wide variety of mutations in the highly insensitive category including P230H (Sdh B), G79R, H134R and C135R (Sdh C) and D126E, Q12K, S120P and D126N (Sdh D). The most common mutations associated with insensitivity to fluxapyroxad, fluopyram and penthiopyrad were G79R, H134R and C135R. Mutations have developed rapidly in S. vesicarium, with 0%, 65% and 100% of isolates tested from 2016, 2018 and 2020 respectively, possessing some mutation in the Sdh genes, associated with declining field performance of FRAC 7 fungicides. By contrast to other FRAC 7 active ingredients, a lower percentage of isolates which were highly insensitive to boscalid (57%) possessed mutations in the Sdh genes. This may suggest some additional mechanism of resistance other than mutations in the Sdh genes is operating in S. vesicarium to confer insensitivity to this fungicide. Mechanisms of resistance to FRAC 3 fungicides in Stemphylium vesicarium Mutations in the Cyp51 gene have been associated with resistance to FRAC 3 fungicides in some fungi. Protocols were developed to allow amplification of the Cyp51 gene in S. vesicarium. Sequencing of 5 isolates of S. vesicarium which were sensitive and an additional 5 which were insensitive to FRAC 3 fungicides failed to detect any differences in the sequences of the Cyp51 gene which could be associated with fungicide insensitivity. It is proposed that FRAC 7 fungicide resistance in S. vesicarium is associated with other mechanisms.
Publications
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Progress 09/01/19 to 08/31/20
Outputs
Target Audience:
Nothing Reported
Changes/Problems:The Covid-19 shutdown and Cornell University restrictions on travel necessitated a reduction in laboratory and field work for 2020. This resulted in a decision not to deploy spore traps in growers fields, due to the need to collect spore samples at weekly intervals, and for samples to be processed and stored in the laboratory on receipt. Collection of leaves at weekly intervals was able to proceed as intended with leaves collected by members of Cornell Co-Operative Extension who were permitted access to growers fields.Leaf sampleswere processedin a laboratory in the PI's home. In addition, periodic malfunctions occurred in newly acquired dataloggers deployed in the field. This prevented sufficient local weather data being collected for analysis. Some analyses were able to be undertaken with data obtained from centralized weather stations. Plans are being made to acquire more reliable equipment for the 2021 season. What opportunities for training and professional development has the project provided?
Nothing Reported
How have the results been disseminated to communities of interest?Reports on increases in Stemphylium incidence in the six monitored fields were made available to Cornell Co-operative Extension educators during the season, and used in discussion with individual growers. However, as these results are of a preliminary nature, they have not been disseminated to a wider audience as yet. What do you plan to do during the next reporting period to accomplish the goals?1. Sequencing of the sdhC and sdhD genes of representative isolates will continue, to determine the range in mutations associated with FRAC 7 resistance in S. vesicariumisolates fromNY onion and to identify the major mutations. This will be used to inform Cornell Co-operative Extension educators andgrowers of the most suitable fungicide resistance management strategies. Once the major mutations associated with FRAC 7 resistanceare identified, a qPCR technique will be developed to enable the rapid identification of mutations within isolates, to improve the efficiency of fungicide resistance monitoring. 2.Field trials will be established in 2021 as outlined in the project proposal to (i) investigate improving the timing of initial fungicide applications by modelling spore release in spring and (ii) optimizingthe timing of in-season fungicides by validating models whih predict critical risk periods for infection. Covid restrictions and malfunctions in equipment prevented sufficient data to be collected in 2020. More reliable dataloggers will be sourced and spore traps deployed in 2021, to enable the collection of a more robust dataset. 3. Plans are being made for virtual presentationsat an onion workshop as part ofthe Empire Producers State Expo (January 2021). This will include anunderstanding of what mutations are associated with FRAC 7 fungicide resistance and strategies to contain the development of resistance.
Impacts
What was accomplished under these goals?
(i) Improving the timing of initial fungicide applications by modelling spore release in spring and (ii) optimize the timing of in-season fungicides by validating models whih predict critical risk periods for infection. Progress on theseaspect of the project has been limited due to the Covid-19 shutdown and travel restrictions at times during the season. The restrictions prevented the intended deployment of spore traps in the 2020 season. In addition, newly purchased dataloggers deployed in three of four commercial fields during the season had periodic malfunctions which adversely affected the amount of in-field weather data available for analysis. However, some aspects of the work were successfully achieved. Leaf samples (n = 20 to 50) were collected from six commercial onion fields at weekly intervals between late May and late August 2020. Leaves were incubated in plastic bags under high humidity and examined for signs of S. vesicarium (conidia and pseudothecia) under the microscope to identify periods of disease incidence increase. Preliminary analysis has been conducted to associate weather data from centralized weather stations with disease in field. (iii) Develop tools to better monitor fungicide resistance and better understand pathogen biology. Resistance to FRAC 7 fungicides in fungi is associated with mutations in the succinate dehydrogenase which is made up of four subunits (SDHA, SDHB, SDHC and SDHD). Mutations in genes (sdhB, sdhCand sdhD) lead to amino acid substitutions in the respective subunits, which are associated with resistance to various FRAC 7 fungicides. PCR primers and techniques were optimized to allow the amplification of the sdhB region of the S. vesicarium genome, a region in fungi in which mutations associated with resistance to FRAC 7 fungicides commonly occur. The finding of only 5 isolates with a mutation in the sdhBgene occurreddespite laboratory tests indicating substantially more of theisolates selected to be insensitive to FRAC 7 fungicides. This suggests that FRAC 7 resistance in isolates of S. vesicarium may be associated with mutations in other genes (sdhCand sdhD), or that other mechanisms of fungicide resistance are present in S. vesicarium. Primers are currently being designed and optimized to allow amplification and identification of any mutations in the sdhCand sdhDregions.
Publications
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