Allelopathic cover crops, roller crimping, and soil steaming as an integrated non-chemical weed management strategy in tomato

1. Greenhouse trial

• 1.1. Screening and identification of allelopathic cover crops (CC) against problematic weeds in tomato (MSU-Tseng & UA-Burgos): We will assess the weed-suppressive capabilities of six cover crops (Crimson clover, cereal rye, hairy vetch, wheat, sorghum-sudangrass, and buckwheat) on three problematic weed species—yellow nutsedge, Palmer amaranth, and large crabgrass. This evaluation will take place in a modified stair-step system (Figure 1) within a controlled greenhouse environment (maintained at 27/18°C day/night temperatures, 60-70% humidity, and 400-500 μmol/m2s light intensity) through separate experiments. A video of the construction and working of our stair-step system can be viewed using this link https://www.jove.com/video/60764?status=a62770k. The stair-step system is designed with one column dedicated to a weed species as a control, and six treatments (CC species) along with their respective controls arranged in columns of four plant samples descending the steps (Figure 1). Figure 1 illustrates the treatment layout in the stair-step apparatus, representing a single replication of a cover crop (donor) against a weed species (target). The experiment will be conducted with four replications and repeated in time. Each pot, measuring 6 inches in diameter, will be planted with either the cover crop (10 plants per pot) or the weed species (10 plants per pot). Leachate from each column will be cycled back to the top using a pump set to activate every 6 hours for 1 minute. Allelochemicals, if present, will be concentrated after successive cycles. The impact of allelochemicals from the donor on the weedy (target) plants will be observed through the reduction in plant height. Pots will be arranged in the stair-step system within designated trays (Figure 1), and plant heights will be measured at 1, 7, 14, and 21 days after treatment (DAT). Comparisons among cover crop (CC) species will be based on percent height inhibition data. The mean inhibition of weed species by each CC species will determine the highest and lowest-ranking accessions. Root and shoot biomass will be measured at 21 DAT. Data analysis will involve using ANOVA to assess overall differences among cover crop species for percent height inhibition, root biomass, and shoot biomass. If significant differences are detected, Tukey's HSD will be employed to identify specific pairwise differences.
• 1.2. Identify the most effective soil steaming depth x steaming duration combination for killing yellow nutsedge tubers (MSU-Tseng & UA-Burgos): Briefly, 8 inch plant containers will be planted with ten yellow nutsedge tubers planted in clay-loam field soil, followed by irrigation. Two days after weed planting, the soil surface in each pot will be exposed to steaming until soil temperature reaches 65 ± 5ºC at three different depths (4, 6, 8, and 12 inches), after which the steaming will continue for 0, 5, 20, and 45 mins, for container assigned to each depth. The temperature will be monitored using wireless soil temperature sensors connected to the phone. Yellow nutsedge is found at various depths below the soil surface, with tubers present at multiple levels. Studies have shown that yellow nutsedge tubers can remain viable and persist in the soil up to a depth of 8 inches (Stoller and Wax, 1973). An unsteamed control will be included. The steamer to be used will be a 120 V power supply electric steamer (SG15 Steam Generator, Gothic Arch Greenhouses, Mobile, AL; Figure 2; See Equipment Justification and Quote), with a 42-gallon fuel tank, 60 psi ultra-dry steam pressure, and 500 lb per hour steam outflow. To prioritize researcher safety, protective gear such as heat-resistant clothing and gloves will be provided. Researchers will receive training on the safe operation of the 120 V power supply electric steamer (SG15 Steam Generator), including clear protocols for handling steam-related equipment and emergency procedures. Regular safety checks will be conducted to maintain a secure working environment. Treatments will be arranged in a randomized complete block design with four replications and repeated twice. Plant height, injury, and density of yellow nutsedge will be counted at 7 DAT and every 7 d after until 21 DAT. At 21 DAT, three random plants in each pot will be removed, and their root length and root and shoot dry biomass will be recorded, in addition to plant height, injury, and weed density. The steam treatments will be scored according to the average level of weed growth inhibition it caused. Upon completion of the experiment, data collected on plant height, injury, density of yellow nutsedge, and various plant metrics at different time points will undergo statistical analysis using JMP software (v.17.2, SAS Institute, Cary, NC). We will use Analysis of Variance (ANOVA) to assess overall differences among soil steaming depth x steaming duration combinations. Tukey's Honestly Significant Difference (HSD) will be employed for pairwise comparisons if significant differences are detected. Repeated measures ANOVA may also be utilized to account for the experimental design with four replications and repeated measures.

2. Field trial

• 2.1. To test the integration of allelopathic cover crops (identified from Objective #1), roller crimping, and soil steaming (depth and duration identified from Objective #1) to suppress weeds and improve tomato yield/quality (MSU-Tseng/Broderick, UA-Burgos, Rodale-Stallworth): The sequence of events (timeline) is presented in Figure 3. The experiment will use a completely randomized block design of 12 treatments (Table 1), each replicated four times across MS and AR. Two best-performing cover crop species from Objective #1 (based on weed suppression efficacy) will be planted in October-November. Treatment numbers 3, 4, 7, 8, 11, and 12 will undergo roller crimping utilizing the ZRX 30” integrated rollers (Dawn Equipment, Sycamore, IL), coupled with double-disk row-cleaners. Two most effective soil steaming treatment combinations identified from Objective #1 (steaming depth and duration combinations on weed suppression), will be applied. Conventional land preparation will be conducted according to commercial practices. Weed counts (measured by the number of plants/plot, by weed species) and tomato height will be assessed at intervals of 0, 14, 28, and 42 days. Additionally, the tomato yield and fruit quality parameters will be gathered at the time of harvest. We will utilize descriptive statistics to summarize collected data and employ analyses of variance (ANOVA) to assess treatment effects on weed counts, tomato height, yield, and fruit quality parameters. Tukey's HSD will facilitate pairwise comparisons. Temporal changes will be analyzed using repeated measures ANOVA. Correlation and regression analyses will explore relationships among variables. The JMP software (v.17.2, SAS Institute, Cary, NC) will be the platform for all statistical analyses, with a significance level set at α = 0.05.
• 2.2. To determine the effect of cover crop x roller crimping x soil steaming on soil health and diversity (MSU-Tseng/Shanmugam, UA-Burgos, Rodale-Stallworth): Soils will be sampled (3-in depth) at tomato transplanting and once during the growing season. Samples will be collected from all four replications of the 12 treatments (from Objective #2.1) at both field locations, MS and AR (96 soil samples/location), and analyzed for:
  • Physical: Soil texture, record in situ penetrometer resistance (0 to 12 in.), record in situ soil moisture (0 to 12 in) by TDR (Dane et al., 2002).
  • Chemical: pH, electrical conductivity, cation exchange capacity, extractable macro- and micro-nutrients, total C and N (C:N) (Sparks, 1996).
  • Soil-microbial indicators: C availability index, N availability index, bacterial and fungal diversity, microbial biomass, and respiration.
  • Microbial diversity and weed suppression: Mantel tests will delineate the precise relationships between microbial matrix and weed indices (density, biomass, and diversity). Soil functional prediction analysis will be performed to understand the relationship between soil microbial diversity and weed suppression (as a function). A partial mantel test will also be performed to examine the correlation between the degree of similarity in microbial community composition with environmental, weed, and soil variables (Winter et al., 2013).

Sampling points within each treatment will follow a systematic spatial distribution pattern, covering the entire experimental area to ensure representative data. Grid-based or transect sampling methods will be employed, with strategic placement to capture variations across the field. The spatial distribution will be designed to account for potential heterogeneity in soil conditions and weed populations.

3. Testing the most effective cover crop x roller crimping x soil steaming combinations at grower’s field (Certified Naturally Grown and conventional) (MSU-Tseng/Broderick, Growers): To determine the most effective cover crop x roller crimping x soil steaming combination, field screenings will be conducted, and the selected combination will be implemented at four growers' fields in Mississippi. The evaluation criteria encompass superior weed suppression, improved soil health indicators, enhanced crop yield, and seamless integration into growers' practices. Economic feasibility and practical implementation will also be considered to ensure the selected combination is ecologically and agriculturally beneficial. The experimental design will utilize a split-plot approach, with cover crop treatments (cover crop vs. fallow) as the main plot and soil steaming treatment as the subplot (with and without soil steaming). This design will be replicated four times in each farm for robust statistical analysis. In Certified Naturally Grown farms, sustainable and natural methods will be employed, avoiding synthetic chemicals and GMOs. Conventional farms will undergo comparative testing to evaluate the efficacy of non-chemical methods against a herbicide weed management treatment. The data collection and analysis during these trials will mirror the parameters outlined in Objective #2.1, ensuring consistency and allowing a direct comparison of results. This includes an in-depth analysis of weed suppression, soil health indicators, and crop yield. The sampling strategy, monitoring, and adaptation mechanisms outlined in the methodology will be applied to capture a comprehensive understanding of the on-farm performance. Continuous monitoring during on-farm trials will assess the performance of the cover crop x roller crimping x soil steaming combination. We will conduct regular site visits and communication with growers to gain valuable insights and real-world feedback. Regular communication with farmers allows for swift adjustments to the experimental plan, ensuring it works effectively on different farms. Embracing flexibility acknowledges the diversity in agricultural practices. This collaborative methodology seeks to provide valuable insights into applying sustainable agricultural practices in various farming contexts.
4. Economic Analysis (MSU-Tseng, UA-Burgos, Rodale-Stallworth): To evaluate the long-term economic sustainability of our non-chemical weed management approach, we will conduct a comprehensive economic analysis, which will include a comparison with current organic production practices. Key components of this economic analysis comprise:
   • Labor Cost Assessment: We will track and record labor costs associated with weed control for our proposed non-chemical approach, as well as existing organic production methods. This data will provide insights into the labor efficiency and cost-effectiveness of our approach over the long term.
   • Material Cost Comparison: We will conduct a comprehensive cost comparison that extends beyond our non-chemical approach to encompass the costs of materials used in current organic production practices, such as cover crops and specialized equipment, along with conventional chemical weed control methods.
   • Data Collection and Duration: The economic analysis will span an extended period to capture the long-term financial implications. Data will be collected with precision, and the assessment period will extend well beyond the immediate project timeframe.
   • Cost-Benefit Analysis: We will perform a comprehensive cost-benefit analysis that compares the long-term costs of our non-chemical approach with both current organic production practices and traditional chemical-based weed control methods. This analysis will provide a comprehensive understanding of the economic advantages, potential savings, and sustainability of our approach for organic tomato growers.