Assessing Anaerobic Soil Disinfestation for Improving Weed and Soilborne Disease Management in High-tunnel and Open-field Salad Green Production

Final report for GS20-221

Project Type: Graduate Student
Funds awarded in 2020: $16,499.00
Projected End Date: 08/31/2023
Grant Recipient: University of Florida
Region: Southern
State: Florida
Graduate Student:
Major Professor:
Dr. Xin Zhao
University of Florida
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Project Information


In response to national and local consumer demand, organic crop production in Florida has expanded during the last decade. In recent years, high-tunnels have emerged as a protected production system increasingly used by Florida organic growers because of their utility for season extension of high-value vegetables and improvement of produce quality. As organic production in high-tunnel systems continues to expand, research is warranted to guide producers in terms of pest and nutrient management. One method which has proved effective against several soilborne diseases across a wide range of horticultural crops and environments and shows promise as a weed management strategy is anaerobic soil disinfestation (ASD). ASD is a preplant method based on creating anaerobic soil conditions by incorporating labile carbon sources, irrigating to fill the soil pores with water, and covering the soil with a gas impermeable barrier. ASD promotes soil microbial shifts toward facultative and obligate anaerobes which produce short-chain organic acids and volatile compounds that are likely toxic to and inhibit weed seed germination. While previous studies have focused on disease management and the underlying mechanisms, limited field studies have investigated the effects of ASD treatment on soil and crop nutrient dynamics as well as ASD for managing weeds in high-tunnel systems.   

To our knowledge, limited field studies have been conducted to determine the impacts of ASD soil treatment on baby leafy green production and soil nutrient dynamics in organic high-tunnel systems. Organic fertilizers are commonly applied preplant in organic baby leafy green production. While the nitrogen (N) application rate for growing organic baby leafy greens is yet to be determined in Florida sandy soil conditions, the influence of preplant N rates through organic fertilization on the ASD soil treatment with respect to anaerobic condition development and crop and soil nutrient dynamics as well as crop performance also needs to be evaluated. This study examined different preplant N application rates involved in the ASD treatment in organic high-tunnel and open-field systems for developing recommendations on ASD application in production of direct seeded baby leafy greens. Based on linear and nonlinear models applied in each season, the optimum N application rate associated with ASD soil technique and production of baby leafy lettuce in HT and OF systems ranged from 136 kg N ha-1 to 233 kg N ha-1. Within this range, yield was not reduced compared with a reference N rate based on conventional commercial production, while limiting negative effects on lettuce color and phytochemical quality attributes at harvest. Project findings contribute to improving ASD as an environmentally-friendly method for soilborne pathogen, weed and nutrient management in organic production systems. 

Based on responses provided by participants in an online webinar, 100% or respondents indicated they are 'somewhat or extremely likely' to implement ASD as an integrated pest management plan. Similarly, 75% or respondents indicated they are 'somewhat or extremely likely' to transfer the knowledge and resources gained during the webinar. Moreover, 93% of farmers who participated in our field day indicated that they are 'somewhat or extremely likely' to transfer the knowledge and resources gained, while 86% of participants indicated they are 'somewhat or extremely likely' to consider using eco-based practices including ASD for managing pests. 

Project Objectives:

With the long-term goal of developing cost-effective and environmental-friendly ASD application for improving organic baby leafy green production systems, the specific objectives of this study included:


Objective 1: Assess the impact of N application rate on soil cumulative redox potential in ASD-treated soils.


Objective 2: Assess the ASD soil treatment impact on soil nutrient dynamics in high-tunnel and open-field systems. 


Objective 3: Compare different N application rates for baby leafy green production under ASD treatment in high-tunnel and open-field systems.


Materials and methods:

Field trials were conducted in two production seasons on certified organic land at the University of Florida Plant Science Research and Education Unit in Citra, Florida. A spring trial was conducted between April – May 2021 and repeated in fall 20202 between November – December. In both trials, a split-plot design was used, with production system [high-tunnel (HT) vs. open-field (OF)] as the whole-plot factor and N application rate as the subplot factor. Soil treatments consisted of a seven N rates (Nrate) (0, 68, 136, 204, 271, 339, and 407 kg N ha-1) using Everlizer® (EV), heat-treated chicken litter (3%N-3%P2O5-3%K2O). All ASD treated soils were amended with 6.9 m3 ha−1 of molasses (M) (Agricultural Carbon Source, TerraFeed, LLC, Plant City, FL). A non-amended control treatment that did not receive EV or M was also included. In both years, lettuce (Lactuca sativa cv. Red Salad Bowl, Johnny’s Selected Seeds, Winslow, ME, USA) was direct seeded following ASD treatment period. After application of EV and M, soil was tilled with a rotary cultivator, and ASD beds were covered with a 0.025 mm black VaporSafe® TIF (Raven Industries Inc., Sioux falls, SD, USA) polyethylene mulch with an ethylene vinyl alcohol (EVOH) barrier layer. The beds undergoing ASD treatment were the only treatments irrigated one time using a sprinkler irrigation sprinkler system with an output of 0.66 L min-1 at 241.3 kPa water pressure. Irrigation was placed 1.5 m above crop height. The total run time required to saturate the top 5 cm of soil was approximately 4.7 hr. Control treatments were covered with a 6-mil silage tarp to prevent plots from receiving irrigation during initiation of ASD soil treatment.

During spring 2021 trial, ASD treatment period was 21 days. Based on results from subsequent trials the ASD treatment period was shortened to 8 days during the fall 2022 season.

Following the protocol outlined by Paudel et al. (2018), oxidation–reduction potential sensors (PT combination electrodes, Ag/AgCl reference, Sensorex, Garden Grove, CA) were installed to a depth of 15 cm in each plot to measure the redox potential, and thus, to evaluate the level of anaerobiosis achieved in the soil during the first 3 weeks after treatment application. Soil cores were taken randomly in each subplot at a depth of 15 cm, using a 1.9-cm diameter soil probe (Oakfield Apparatus Company, Oakfield, WI). Soil cores were pooled into one composite sample per plot. In each trial, baseline samples were taken prior to ASD treatment. During spring 2021 trial, subsequent soil samples were taken at 1, 7, 14, and 21 days after treatment application (DAT). During fall 2022 trial, subsequent soil samples were taken at 1 and 8 DAT. Soil samples taken during ASD treatment period were analyzed in a commercial lab for soil NH4-N and NO3-N content and soil pH. Soil temperature and volumetric water content were continuously monitored during the ASD treatment period. Soil temperature/moisture sensors (CS655, Campbell Scientific, Logan, UT) were installed to a depth of 15 cm in each plot before initial irrigation.

At harvest, whole plots were cut approximately 2.5 cm above the soil using a hand-operated Quick-cut Greens Harvester (Farmers Friend, Centerville, TN) to determine fresh weight biomass. Yield response models were used to describe the expected yield [E(Y)] as a predetermined function of N (X) using five linear and nonlinear models commonly cited in the literature, including: (1) linear-plateau, (2) quadratic-plateau, (3) exponential, (4) quadratic, and (5) square root models (Djidonou et al., 2015). Determination of lettuce quality attributes included leaf color, soluble solids content, total titratable acidity, total phenolic content, total antioxidant capacity and ascorbic acid. Lettuce leaf color was measured immediately after harvest in both spring 2021 and fall 2022 trials using a chromameter (CR-400, Konica Minolta Sensing Americas, Inc., Ramsey, NJ).


For the analysis of linear and non-linear yield response models, the errors are assumed to be independent and identically distributed from a normal distribution with a mean equal to 0 and variance σ2. For each trial, all models were estimated using nonlinear least squares estimation using Proc NLIN in SAS (Version 9.4; SAS Institute, Cary, NC). The models were compared based on five measures commonly used in model selection including the adjusted coefficient of determination (Adj-R2), the Akaike information criterion (AIC), the root mean squared error (RMSE), P value, and F value.

Research results and discussion:

Objective 1: Assess the impact of N application rate associated with ASD soil treatment on yield of direct seeded baby lettuce.

HT systems have shown promise for extending crop production seasons relative to OF systems. Previous studies conducted in the Southeastern U.S. region have reported improved marketable yields for a variety of high-value horticultural commodities grown in HT systems (Zhao et al., 2009; Laur et al., 2021). However, information highlighting the effect of varying N application rates in HT compared with OF systems remains to be explored. Furthermore, in the context of ASD technique, no N rates have been previously established on Florida sandy soils for direct-seeded baby leafy greens.

In the current two-year study, harvest fresh weight (FW) was significantly impacted by Nrate in both seasons. Moreover, in spring 2021 trial the interaction of PS  Nrate significantly impacted FW. Compared with open field (OF), high tunnel (HT) resulted in greater FW accumulation in ASD136, ASD270, ASD339, and ASD­407­. Although the difference was not significant, ASD204increased FW by 93% in HT compared with OF system. During the fall 2022 trial, ASD136significantly increased FW vs. unamended control, but did not differ from ASD0 or ASD68. Interestingly, increasing N­­rate above 136 kg N ha-1 did not result in significant FW increases. Based on multiple goodness-of-fit statistics including F value, P value, Adj-R2, RMSE, and AICC the linear plateau model was identified as the best fitting model in both spring 2021 and fall 2022 trials. Based on the linear-plateau junction point which occurs at the intersection of the linear and plateau lines, the critical rate of fertilization was determined to be 136 kg N ha-1 and 233 kg N ha-1 during spring 2021 and fall 2022 trials, respectively. Similarly, when data was pooled across both production seasons, and across both OF and HT systems, the linear-plateau model was identified as the best-fitting model, and the junction point occurred at 147 kg N ha-1. However, FW exhibited variation among production seasons and production systems. Therefore, in order to provide growers with robust N rate recommendations for direct seeded baby leafy greens associated with ASD soil treatment, further research is warranted to refine the N application ranges identified in this study.

Objective 2: Assess the impact of N application rate associated with ASD soil treatment on quality attributes of direct seeded baby lettuce in high-tunnel and open-field systems. 

ASD impacts have been investigated in several high-value fruiting vegetables and tree fruit including strawberry (Fragaria × ananassa), eggplant (Solanum melongena), tomato (Solanum lycopersicum), pepper (Capsicum annuum), cucumber (Cucumis sativus), and apple (Malus domestica). Yet, the effects of ASD on leaf quality attributes of direct-seeded baby leafy greens, particularly in HT organic production systems remain to be understood. The lack of relevant information pertaining to the effects of ASD on lettuce leaf quality attributes provided justification for the thorough examination of ASD soil treatments amended with varying N rates.

In both spring 2021 and fall 2022 trials, leaf quality attributes including SSC, AA, TPC, and FRAP were impacted by N rate. Similarly, leaf color attributes including a* and b* values were significantly impacted by PS, Nrate or their interaction. Moreover, a clear distinction in H* values was evident between treatments that did not receive organic granular N, compared with those treatments that did. Treatments that received organic granular N had significantly higher H* values indicating leaves appeared more green/yellow compared with treatments that did not receive organic granular fertilizer and therefore appeared less green/yellow and more red/blue.

Increasing N application rates resulted in decreased TPC in both seasons compared with the unamended Control treatment that did not receive organic granular N application. In general, Control treatment resulted in increased TPC compared with ASD-treated soils receiving at least 136 kg N ha-1 in both years. This stratification among soil treatments was most evident during fall 2022 trial, where ASD-treated soils receiving at least 136 kg N ha-1 were significantly reduced compared with Control and ASD0 treatments. However, TPC was not found to differ significantly in both years among ASD treated soils receiving at least 204 kg N ha-1.

Ascorbic acid (AA) content significantly responded to in spring 2021 and fall 2022 trials but was not significantly affected by PS or the interaction of PS × Nrate. While the effect of PS was not significant, AA content trended higher in OF compared with HT during both trials. In general, AA content in baby lettuce leaves was negatively associated with Nrate in both seasons. During spring 2021, AA content was well differentiated between treatments receiving up to 136 kg N ha-1, and those that received at least 204 kg N ha-1. A similar trend was observed during fall 2022, but the difference in AA content between Control and ASD339 was not significant. During both trials, the highest AA content was obtained in treatments without application of EV (Control, ASD0), and ASD68. The lowest AA content in both trials was obtained in ASD407.

Production system and N­rate significantly impacted total ferric reducing antioxidant capacity (FRAP) during spring 2021 trial, while Nrate significantly impacted FRAP during fall 2022 trial. In both seasons, a significant interaction effect of PS × Nrate was not observed. In general, increasing N­rate resulted in decreased FRAP content in both trials, whereas the impact of PS was inconsistent during spring and fall. Interestingly, the trends observed in FRAP activity were similar to those of AA and TPC. In both seasons AA content, TPC content, and FRAP activity tended to decrease as N rate increased. Among the higher N application rates (≥ 204 kg N ha-1), these quality attributes were not significantly different, yet compared with an unamended control and ASD-treated soils receiving up to 136 kg N ha-1 these attributes were lower.

Objective 3: Assess the ASD soil treatment impact on soil nutrient dynamics in high-tunnel and open-field systems.

The impact of sampling date (DAT) and PS was evident on soil pH during spring 2021 and fall 2022 trials. Moreover, Nrate significantly affected soil pH during fall 2022 trial. A slight increase was observed between 0 – 1 DAT during spring 2021 trial, whereas pH remained relatively stable between 7 – 21 DAT. Similarly, pH reached its peak at 1 DAT before returning to pre-ASD levels during fall 2022 trial. Compared with OF, HT increase soil pH in both trials. During fall trial, ASD339 and ASD407 increased pH compared with unamended control treatment. Compared with baseline measurements and measurements taken at 1 DAT during spring 2021 trial, soil NO3-N content significantly increased at 7 DAT and remained relatively stable until the end of the ASD treatment period for ASD-treated soils receiving at least 136 kg N ha-1. A similar trend was observed in fall 2022 trial, where NO3-N content increased in all ASD treatments at 8 DAT compared with either baseline or samples taken at 1 DAT.

Soil NH4-N was impacted by the DAT × Nrate two-way interaction during spring 2021 trial, with ASD-treated soils receiving at least 136 kg N ha-1 reaching their peak at 1 DAT compared with ASD68 which did not peak until 7 DAT. Interestingly, NH­4-N content decreased in all treatments at day 14 compared with 7 and 21 DAT during spring 2021. In fall 2022 trial, Nratesignificantly impacted NH4-N content. Both ASD339 and ASD407 increased NH4-N content compared with all other treatments.

Participation Summary

Educational & Outreach Activities

1 Curricula, factsheets or educational tools
1 Online trainings
1 Tours
5 Webinars / talks / presentations
1 Workshop field days

Participation Summary:

15 Farmers participated
12 Ag professionals participated
Education/outreach description:

Our research findings were presented at the American Society for Horticultural Science 2022 and 2023 Annual Conferences. In addition, we hosted a field day during Fall 2022 for project dissemination to demonstrate the application of ASD in high-tunnel production systems for organic baby lettuce production. We also hosted an online webinar focused on pest management through ASD in organic agriculture. Research manuscript for publication in a peer-reviewed scientific journal is on-going. As well, farmer-oriented fact sheets for implementing ASD in urban gardens is on-going. 

Project Outcomes

25 Farmers reporting change in knowledge, attitudes, skills and/or awareness
Project outcomes:

The phase-out of methyl bromide as a broad spectrum fumigant necessitates the examination of integrated methods for managing soil borne pathogens, while considering the economic, environmental, and social sustainability. This project implemented ASD as a biologically-based method for managing such pathogens. Our study investigated impacts of ASD on soil  and crop nutrient dynamics, demonstrating the positive environmental benefits of combing ASD with high tunnel production systems for minimizing N loss through leaching compared with open field systems. Our results will serve as a groundwork for establishing N recommendations for direct seeded baby leafy greens associated with ASD in open field and high tunnel organic systems. Our project therefore contributes to the future of sustainable agriculture by providing farmers with environmentally-friendly alternatives to chemical fumigation. Although economic analyses were not conducted in this research, we have demonstrated in pilot trials that re-useable 6 mil silage tarp can serve as an alternative to single season plastic mulch during the ASD implementation. In this way, farmers can reduce their input cost associated with ASD.  The combination of reduced costs associated with ASD, and the reduced reliance of chemical fumigants will enable farmers to continue to meet the growing demand for organic produce at the state and national levels. 


Knowledge Gained:

During the course of this multi-year  project, we conducted extensive research, attended workshops, participated in academic conferences, hosted an online webinar, and engaged with experts in the field of sustainable agriculture. These experiences broadened our understanding and provided insight into emerging technologies and innovative approaches in sustainable agriculture. Moreover, we learned to apply emerging techniques and methods for managing soilborne pest and pathogens. In addition, our abilities to conceptualize, design, and implement research-oriented sustainable agricultural projects improved. 

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.