Biofumigation for soil health in organic high tunnel and conventional field vegetable production systems

Final Report for LS06-185

Project Type: Research and Education
Funds awarded in 2006: $170,000.00
Projected End Date: 12/31/2009
Region: Southern
State: Kentucky
Principal Investigator:
Dr. Michael Bomford
Kentucky State University
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Project Information

Abstract:

Biofumigation and solarization were tested as possible organic controls of white mold (Sclerotinia sclerotiorum), a soil-borne pathogen of cool-season vegetable crops commonly found in high tunnels. Biofumigation was also tested as a possible control of the warm season vegetable pathogen Phytophthora capsici. Pacific Gold mustard was identified as a potential biofumigant crop with a combination of high biomass production and glucosinolate concentration. Laboratory studies showed both pathogens to be susceptible to glucosinolates extracted from the mustard, but soil incorporation of field-grown biomass did not introduce sufficient glucosinolate to reduce disease pressure. Summer solarization in high tunnels destroyed white mold sclerotia.

Project Objectives:
  1. Identify brassica varieties that inhibit survival and growth of S. sclerotiorum and P. capsici in lab-based bioassays. Determine the potential of brassica residue incorporation and solarization – alone and in combination – to reduce disease pressure from S. sclerotiorum and build biologically active soils in high tunnels used for year-round vegetable production. Determine the potential for brassica and non-brassica cover crops to reduce disease pressure from P. capsici and build biologically active soils in field vegetable production systems.
Introduction:

The purpose of this project was to develop sustainable soil-borne disease management tactics suitable for use in organic or conventional vegetable production systems. In particular, we examined the potential of soil-incorporation of brassicas to fight white mold (Sclerotinia sclerotiorum) in unheated high tunnels and Phytophthora blight in summer-grown melons. We aimed to develop tactics that build healthy soils and promote microbial ecosystems that challenge these potentially devastating, broad spectrum pathogens. Soil-borne pathogens attack all vegetable crops, causing a range of root rots, vascular wilts, crown rots, fruit rots, and foliar blights. They can be difficult to identify, and may survive in soil for many years. Management recommendations sometimes discourage growers from planting susceptible crops in infected soil for several years, which can be particularly difficult for those with land and equipment suited to a particular crop rotation. The fungicide and soil fumigant regimes often used by conventional vegetable growers to cope with soil-borne disease may be environmentally disruptive or incompatible with sustainable production philosophies: The most commonly used fumigant, methyl bromide, is being phased out because it is an ozone-depleting substance and broad-spectrum biocide. Vegetable diseases are of particular interest in areas where small farmers are turning to vegetable crops as a high-value alternative to tobacco production. Here, in Kentucky, the number of farms growing vegetables doubled from 1,086 to 2,123 between 1997 and 2007, while the number growing tobacco – the longtime backbone of the commonwealth’s small farm economy – fell by 83% from 46,850 to 8,113 (USDA 2007). A small, but rapidly growing, segment of our vegetable growers are taking advantage of the growing demand for organic food. The national market for organic food has increased by 5-22% annually for over a decade (OTA 2010). Fresh produce accounted for 38% of the $24.8 billion worth of organic food sold nationwide in 2009 (OTA, 2010). Although this market remains largely under-served in the south, the number of certified organic farms in Kentucky grew 10-fold, from 12 to 125, between 2006 and 2010 (Fitzgerald pers. comm.). Many more Kentucky farmers are seeking certification, or are incorporating organic methods into their operations without certifying. Our research is intended to serve both organic and conventional growers by exploring a disease-management strategy that is compatible with organic standards, but also fits within conventional production systems. Its adoption has the potential to enhance the sustainability of both. Organic vegetable growers do not have the option of using synthetic fungicides or fumigants, so they must rely on more ecologically benign tactics for disease management. Management options for soilborne diseases available to organic vegetable growers include crop rotation, destruction of infested crop residue, water management, the use of resistant varieties, and sanitation. Two additional tactics available to organic growers are biofumigation and solarization. Biofumigation is the incorporation of plant residues that release pesticidal compounds into the soil as they decompose. Cruciferous crops (e.g. mustard, cabbage, canola) are most commonly used as biofumigants because they contain compounds called glucosinolates, long recognized as powerful natural pesticides (Drobnica et al. 1967). When plant tissue is macerated plant enzymes convert glucosinolates into isothiocyanates, which are toxic to a wide array of microorganisms, weeds, and insects. The potential of isothiocyanates for disease control has been the subject of many past and current investigations (Brown and Morra 1997; Kirkegaard et al. 1998; Cohen et al. 2005). They are particularly interesting because they do not kill everything in the soil: Many beneficial bacteria, including Trichoderma and Pseudomonas species, are more abundant after biofumigation (Smith and Kierkegaard 2002). Some researchers have even suggested that particular brassica varieties or tissues could be selected for their production of isothiocyanates that target specific diseases (Bending and Lincoln 2000). SARE-funded studies in Tennessee and North Carolina have shown that biofumigation suppresses the fungus that causes Southern blight of tomato (Sclerotium rolfsii) and increases beneficial bacterial counts (Sams 2002a, 2002b, 2004). An ongoing SARE study in South Carolina is examining the potential for biofumigation to protect watermelon from Fusarium wilt (Keinath 2004). Solarization is the use of a clear plastic film on the surface of moistened soil to trap solar energy, raising soil temperatures high enough to kill pest organisms. The tactic was pioneered in Israel in the mid-1970s, and has since been the subject of several comprehensive reviews (Katan 1987; Chen et al. 1991; DeVay et al. 1991; Stapleton 1998, 2000) and SARE projects (Long 1988; Bandele 2000; Sams 2002a). It works best when air temperatures above the plastic remain high throughout the treatment period, as in greenhouses, cold frames, and areas rotated out of crop production during summer months due to excessive heat. Under optimal conditions, solarization can disinfest the upper layers of soils, creating a “biological vacuum” more readily recolonized by benign or beneficial microorganisms than plant pathogens (Stapleton 2000). Although solarization is widely, and successfully, used in the Mediterranean and Japan – particularly in greenhouses (Horiuchi 1991; Cartia 1998) – it remains an uncommon practice in the USA, where researchers report mixed results under field conditions. SARE-funded studies found no reduction in soilborne disease of field tomatoes in Tennessee (Sams 2002), or increase in strawberry yields in Texas (Long 1988) after solarization. No SARE projects have yet examined the potential for solarization in protected enclosures such as greenhouses, high tunnels, or cold frames. A cornerstone of the organic agriculture philosophy is the idea that healthy soils grow healthy plants. Soils high in organic matter are considered healthy because they support soil life, including beneficial microbial communities, capable of suppressing plant diseases (Hoitink and Boehm 1999; Van Bruggen and Termorshuizen 2003.). Biofumigation has the potential to enhance soil health by adding organic matter to the soil. Both biofumigation and solarization have the potential to shift the ecological balance in favor of beneficial microorganisms in the soil community. We tested the potential of these control strategies to manage two pathogens that frequently cause problems for southern vegetable producers: Sclerotinia sclerotiorum and Phytophthora capsici. S. sclerotiorum, the cause of white mold, infects at least 361 plant species, encompassing 64 families (Purdy 1979). Its vegetable hosts include beans, brassicas, cucumbers, lettuce, melons, tomatoes, and squash, but it does not infect corn or grasses. It produces long-lived, hard, black sclerotia that germinate under cool (50-70°F), wet conditions to produce mushroom-like structures (apothecia) that release millions of spores. Field observations in commercial high tunnel systems in Kentucky indicate that white mold can become a serious problem, due to sustained cultivation of host species under the cool conditions that favor the disease. Our examination of S. sclerotiorum was conducted in these commercial high tunnel systems. Limited tests of biofumigation against S. sclerotiorum suggest that the tactic may not kill the organism’s sclerotia by itself (Manici et al. 1999; Gilardi et al. 2000), but we saw potential for synergism with solarization: Heating the soil may enhance the release of isothiocyanates from brassica tissues incorporated as biofumigants. We sought to identify biofumigant crops that are particularly toxic to S. sclerotiorum in the lab before introducing them into our field studies. Solarization has considerable potential for S. sclerotiorum control in high tunnels, judging from previous reports of its success in combating the disease in Israel, Western Europe and Australia (Porter and Merriman 1985; Ben-Yephet 1988; Phillips 1990; Cartia and Asero 1994; Minuto et al. 2000). The high temperatures achieved by solarization can kill S. sclerotiorum or weaken its survival bodies (sclerotia), making them more susceptible to colonization by beneficial organisms. However, the efficacy of solarization against S. sclerotiorum has not previously been tested either in high tunnels or in the transition zone of the USA. Kentucky growers using high tunnels often grow their vegetable crops in flat, rather than raised beds, to improve soil heat retention. Solarization is known to be less effective at the edges of plastic covering raised beds outdoors (Grinstein et al. 1995), but we know of no previous tests for this “edge effect” in flat beds in high tunnels, where the tunnel should promote even heating by reducing air circulation. We identified a need for monitoring of this technique with respect to temperatures achievable, depth of effect, and time required to be effective. P. capsici infects cucumbers, eggplants, melons, peppers, squash and tomatoes. In recent years it has become one of the most destructive diseases of these crops (Ristaino and Johnston 1999; Babadoost 2004; Hausbeck and Lamour 2004). It spreads rapidly under warm (~80°F), wet conditions, releasing swimming spores that are attracted to host roots. From there spores can be splashed to infect all parts of the plant. Under favorable conditions a minor infection early in the season can grow to an epidemic before harvest, destroying entire fields. The fungus produces hard-walled oospores that can survive in soil for 5-10 years. No single control measure is effective against this pathogen; management approaches that integrate a variety of control tactics have had the best results, but are still be insufficient when weather conditions favor the disease (Ristaino and Johnston 1999; Babadoost 2004; Hausbeck and Lamour 2004). Given the biology of this pathogen and the difficulty of managing it, it is important to seek additional management practices that can reduce levels of primary inoculum (the pathogen propagules that initiate disease; in this case, oospores in soil). The biofumigation concept has been tested against Phytophthora species and other oomycetes (Manici et al. 1999; Coelho et al. 1999; Smith and Kirkegaard 2002). However, there are no published reports evaluating either the toxicity of naturally occurring isothiocyanates to oospores of P. capsici or the effectiveness of brassica incorporation on development of disease caused by this pathogen.

Cooperators

Click linked name(s) to expand
  • George Antonious
  • Kenneth Seebold
  • Paul Vincelli
  • Alison Wiediger
  • Paul Wiediger

Research

Materials and methods:
Objective 1

The following laboratory studies were conducted to accomplish the objective of identifying brassica varieties that inhibit survival and growth of S. sclerotiorum and P. capsici.

Brassica selection

Forty-seven brassica accessions were secured from the national brassica germplasm collection at Iowa State University (Table attached). These included 20 Indian mustard (Brassica juncea) accessions, 13 oilseed rape (B. napus), five Ethiopian mustard (B. carinata), five field mustard (B. rapa), three arugula (Eruca sativa), and one white mustard (Sinapis alba). Accessions were selected based on reports of high glucosinolate content, cold hardiness, and rapid flowering. Accessions were grown to flower initiation under greenhouse, high tunnel, and field conditions. Twelve accessions that demonstrated superior cold tolerance, biomass production, and early flower initiation were selected for further evaluation.

Brassica production

Greenhouse-grown plants were direct-seeded on 26 Jul., 2006 and 27 Jan., 2007 in a greenhouse with active heating and cooling systems set to maintain a temperature of 20 °C. Each accession was represented by 10 seeds grown in a 3.6 L (16.5 cm diam x 16.5 cm high) bench-top nursery container filled with a mixture of equal parts peat-based potting soil and aged compost. High tunnel and field-grown accessions were planted at 5 cm spacing in 1 m rows 15 cm apart in Elk silt loam amended with aged compost. Each row contained a different accession, with locations randomly assigned within two complete blocks. Seeds were planted in the unheated high tunnel on 7 Dec. 2006 and 2 Mar., 2009; and in the field on 11 Jun. 2007 and 2 Mar., 2009. All aboveground biomass from each accession was collected and weighed when >50% of plants had initiated flowering. Five plants from each accession harvested from the first planting date were randomly selected for extraction of glucosinolates using the procedure of Antonious et al. (2009). Shoots (stems and leaves) were cut into 1-3 cm and 100 g subsamples were dropped into boiling methanol (300 mL) on a steam bath in 1-L wide-mounted Erlenmeyer flasks (covered with a watch glass) for 15 min. After cooling, the material was blended and vacuum filtered. The contents were centrifuged (10,000 ×g for 10 min). Ten mL of the extract was filtered through an open glass chromatographic column of 1.5 × 20 cm containing 4 g of celite to give a purified homogeneous aqueous extract.

Glucosinolate analysis

Separation of total glucosinolates was accomplished by adsorption on DEAE-Sephadex A-25 (2-[diethylamino] ethyl ether) ion exchange resin of 40-125 µm bead size. We simplified the GSLs separation procedure by using 10 mL disposable pipette tips filled with DEAE, a weak base, that has a net positive charge when ionized and therefore bind and exchange anions (anion-exchange resin) (Figure 1). Five mL of the purified plant extracts were applied to disposable pipette tips (10 × 1.2 cm) containing a small glass wool plugs, ion-exchange Sephadex (preswelled overnight with 2M ammonioum acetate) to give a settled resin height of 5 cm, and the resin was washed with 10 mL of deionized water. The extract was passed through the pipette tip containing Sephadex and washed with 10 mL of deionized water. Two mL of myrosinase (thioglucosidase in 5 mM phosphate buffer of pH 7) were added to the contents of the pipette tip (column). When enzyme solution had completely entered the column, the column flow was stopped, capped, and incubated at 37OC for 18 hrs. After incubation, the columns were allowed to stand at room temperature and eluted with 10 mL of deionized water. A fraction of water extract that contains the hydrolysis products of the GSLs was subjected to a glucose determination procedure (VanEtten and Daxenbichler 1977; Antonious et al. 1996). Brassicae extracts without addition of purified horseradish myrosinase were used as controls. Quantification of GSLs was based on measurement of enzymatically released glucose. Moles of glucose released into the aqueous medium are equivalent to the moles of total GSLs. A calibration curve was carried out with each group of samples using 10 – 100 µg mL-1 glucose. The activity of myrosinase was optimized using sinigrin as substrate (Antonious et al. 1996). Standard materials of sinigrin monohydrate (allylglucosinolate), thioglucosidase, and pure glucose were obtained from Sigma Chemical Co. (St. Louis, MO). Representative samples of the shoots (20 g) were blended with 100 mL of 0.4% oxalic acid solution for ascorbic acid extraction. The homogenate was filtered through a Büchner funnel containing Whatman filter paper No.1. Ascorbic acid was determined by the potassium ferricyanide method (AOAC 1970). L-ascorbic acid of 100% purity (Sigma Chemical Company, St. Louis, MO, USA) was used to establish a calibration curve. Representative samples of the shoots (20 g) were blended with 150 mL of ethanol to extract phenols. Following filtration through Whatman No.1 filter paper, an aliquot was used for phenol determination. Total phenolic constituents were determined by Folin-Ciocalteu colorimetric method (McGrath et al. 1982). A standard calibration curve was obtained using pure tannic acid in the range of 1 to 16 ?g/mL. Aboveground biomass yield and concentrations of total glucose equivalent to GSL, ascorbic acid, and total phenol contents in Brassica shoots of the different accessions tested in relation to growing conditions (greenhouse, high tunnels, and field) were statistically analyzed using analysis of variance (ANOVA) procedure (SAS Institute, 2003). Tukey’s honestly significant difference test was used to compare means. Pearson correlation coefficients were calculated to test for correlations between yield and concentrations of GSLs phenols and absorbic acid.

Activity against S. sclerotiorum in vitro

Sclerotia of S. sclerotiorum were produced in the laboratory on a sterilized mixture of wheat and rye, then hand-separated from the grain in a laminar flow hood. Glucosinolate extracts from promising brassica accessions were added to scintillation vials containing 15 sclerotia, myrosinase (enzyme that catalyzes the breakdown of glucosinolates into volatile isothiocyanate) and sterile soil extract. Extract volume was calculated to be the amount of glucosinolate produced by 0, 0.25, 0.5, 1, or 2 g of aboveground fresh biomass for each accession. The volume of myrosinase solution (5 g/L thioglucosidase in phosphate buffer) added to each vial matched the volume of glucosinolate extract. Sufficient sterile soil extract was added to bring the total volume of each vial to 4 mL. A total of 80 vials were prepared, representing five concentrations of glucosinolate and 16 glucosinolate sources (pure sinigrin, 11 high tunnel-grown accessions and four field-grown accessions). All vials were sealed and stored at room temperature in a fume hood for 24 hours. Sclerotia were removed from vials, surface sterilized, and plated onto ~ 40 mL sterile soil in sealed Petri dishes. These were incubated at 16 °C and a 12:12 light:dark photoperiod for six weeks to germinate apothecia. Sclerotia that had not germinated after this time were considered dead. Sclerotial inhibition was calculated to be the difference between survival of sclerotia after exposure to glucosinolates and survival of sclerotia not exposed to glucosinolates, expressed as a percent of survival of unexposed sclerotia.

Activity against P. capsici in vitro

Eighty plastic culture flasks were prepared with 4 mL solutions of glucosinolate, myrosinase, and soil extract representing the same treatment combinations used to test for activity against S. sclerotiorum. Twenty laboratory-grown P. capsici oospores were added to each flask. Flasks were sealed and incubated in the dark at room temperature for two weeks, then the number of germinating oospores in each flask was counted under a dissecting microscope at 60-120x magnification.

Objective 2

The following studies were conducted to determine the potential of brassica residue incorporation and solarization to reduce disease pressure from S. sclerotiorum and build biologically active soils in high tunnels used for year-round vegetable production.

Activity against S. sclerotiorum in soil

Pacific Gold mustard tops were harvested at flower onset, ground in a household juicing machine (Champion, Lodi CA), and incorporated into Elk silt loam at 0, 15, 30, 60, and 120 g/L. Each concentration was added to three 2.5 L plant pots (15 cm diam. x 14 cm high), to simulate field treatment rates of 0, 2, 4, 8 and 16 kg of fresh mustard per square meter incorporated to a depth of 14 cm. A mesh bag containing 40 sclerotia of S. sclerotiorum was buried in the center of each pot. The procedure was repeated using fresh grass in place of Pacific Gold mustard. The 60 pots were watered and randomly arranged on a table in a greenhouse with active heating and cooling systems set to maintain a temperature of 20 °C. Bags of sclerotia were collected after three days and plated onto ~ 40 mL sterile soil in sealed Petri dishes. These were incubated at 16 °C and a 12:12 light:dark photoperiod for eight weeks to germinate apothecia. The entire study was conducted three times, beginning on 12 Jun 2008, 11 Nov 2008, and 15 May 2009.

Field tests of biofumigation and solarization in high tunnels

This study was conducted in two adjacent commercial high tunnels (6 x 29 m) with a history of Sclerotinia sclerotiorum near Bowling Green, Kentucky. The two tunnels have been used for continuous soil-based mixed vegetable production; one for 6 and the other for 8 years. The grower uses organic management practices, but has not renewed organic certification since 2002. Soil is a silt loam (pH 6.8-7.3, O.M. 1.6-2.0%). High tunnel beds were divided into sixteen 1.2 x 3 m plots to create four replicate blocks of four treatments. Treatments were: 1) an untreated control 2) biofumigation with 1 kg of Pacific Gold mustard sheet mulch per square meter; 3) solarization with transparent 6 mil UV-resistant polyethylene sheets, anchored by burying the edges with soil; and 4) combined biofumigation and solarization. All plots were roto-tilled to a depth of 15 cm following mustard application. Mesh bags containing 40 S. sclerotiorum sclerotia were buried 0, 5, 10, and 15 cm below the soil surface (three bags per depth) in the center and at the edge of each plot. Temperature probes connected to a CR-10X datalogger (Campbell Scientific, Logan UT) were placed next to the bags in one solarized plot and one unsolarized plot. Plots were hand-watered before plastic installation, then irrigated through three lines of drip tape that ran along each bed for the duration of the study. The study began on 23 July 2007 and was repeated beginning on 5 August 2008. Bags of sclerotia were collected from each location 2, 4, and 6 weeks after setup in 2007; and 48 hours, 2 weeks and 4 weeks after setup in 2008. The number of intact sclerotia in each bag was recorded after each collection. Intact sclerotia were washed, surface sterilized, and rinsed in distilled water before asceptic transfer into Petri dishes containing a sterilized 1:1 soil:sand mix. The number of sclerotia exhibiting carpogenic germination was recorded after 6 weeks of incubation at 16 °C and a 12:12 day:night photoperiod. Plastic was removed after the final collection, allowing the grower cooperators to resume their regular crop rotation in early September. Soil samples were collected from the top 15 cm of each plot immediately before and after each experiment was conducted. Samples were subjected to routine soil analysis (pH, P, K, Ca, Mg, Zn, organic matter, cation exchange capacity, soluble salts) by the University of Kentucky regulatory services laboratory, and to microbial activity analysis using the fluorescein diacetate hydrolysis assay, which measures the activity of several enzymes that are ubiquitous in the soil environment (Adam and Duncan 2000).

Test of solarization and biofumigation with high tunnel-grown biomass

The high tunnel biofumigation and solarization study was repeated in the Kentucky State University high tunnel (9 x 12 m, Elk silt loam soil) in 2009 using a modification of the study design previously used in the commercial high tunnels. Instead of importing field-grown sheet mulch, plots designated for biofumigation treatment were seeded to Pacific Gold mustard on 30 June, 2009. Remaining plots were seeded to buckwheat on the same day. Aboveground biomass was flail-mowed and incorporated by roto-tilling at flower initiation on 5 Aug, 2009, 36 days after seeding. Experimental setup was conducted as in the commercial high tunnels immediately after biomass incorporation. Bags of sclerotia were collected 48 hours and four weeks after biomass incorporation and initiation of solarization, and germinated using the methods already described for the commercial high tunnel studies.

Objective 3

The following studies were conducted to determine the potential for brassica and non-brassica cover crops to reduce disease pressure from P. capsici and build biologically active soils in field vegetable production systems.

Mustard planting date effect on field biomass production

Weather permitting, Pacific Gold mustard was direct-seeded weekly between 24 Apr and 3 Jul, 2008, in three replicated field plots measuring 10 square meters each. Individual plots were roto-tilled on the day of planting. Three rows in each plot were planted 33 cm apart using a walk behind seeder. Weed control was done by hand as needed throughout the season. Plots were not irrigated. Aboveground biomass was collected for measurements of fresh weight, dry weight, and glucosinolate content by randomly harvesting a square meter of each replicate at the onset of flowering.

Activity against P. capsici in soil

Sixteen liters of organic planting mix (Sungro Horticulture Inc., Bellevue WA) was thoroughly mixed with 80 ml of Phytophthora capsici inoculant (Vincelli, Lexington KY) and divided into 12 equal portions. Aboveground portions of a spring cover crop of Pacific Gold mustard or a winter cover crop mixture of rye and vetch, were harvested at first flower, finely chopped, and mixed into planting media at 0, 13.5, 27, 54, 108, or 216 g/L. Media mixtures containing each combination of cover crop and rate were randomly assigned to fill two of 24 evenly-spaced 65 ml cells in each of 10 seedling plug trays (128 cells/tray, Hummert International, Earth city MO). A freshly-germinated yellow squash (Cucurbita pepo) seedling was buried just below the media surface in each cell. Trays were randomly assigned to flood irrigation with untreated tap water or tap water treated with 4.2 g/L of a commercial fertilizer containing beneficial microbials (Organica Plant Growth Activator). Plug trays were partially submerged in the irrigation liquid to ensure thorough drenching, and incubated for 5d at 26 °C. Seedling survival and shoot length were recorded for each treatment combination in each of 5 replicates.

Effect of biofumigation on field vegetable production under P. capsici pressure

This study was originally proposed for a one-acre field with a recent history of severe P. capsici infestation, jointly owned by members of the West Kentucky Grower Co-op. The proposed site could not be used due to the bankruptcy of the West Kentucky Grower Co-op and subsequent sale of the land. A second commercial field with a recent history of P. capsici infection also proved unsuitable for the study because no P. capsici occurred at the site in the drought years of 2007 and 2008 and the grower cooperator withdrew from the project. In 2008 the project advisory committee recommended inoculation of an isolated field on University of Kentucky research land in order to ensure sufficient disease pressure to conduct a field study in 2009. The site was inoculated in 2008 by growing a crop of pumpkin, infecting fruit with laboratory-raised P. capsici, and allowing fruit to decompose in the field over winter. The success of the 2008 inoculation was tested with an early crop of yellow squash in 2009. Disease was allowed to develop on the yellow squash crop between first observation of P. capsici infection, on 18 May, and 20 Jun, when the crop was incorporated into the soil by disking. Sixteen research plots (5 x 8 m) were laid out in four replicate blocks, each with four randomly assigned cover crop treatments: 1) no cover crop; 2) Pacific Gold mustard; 3) Buckwheat; and 4) Pacific Gold mustard and buckwheat. Cover crops were direct-seeded on 20 Jul, then mowed and incorporated into the soil by roto-tilling on 24 Aug, following flower set. Beds were shaped and covered with black plastic immediately after cover crop incorporation on 24 Aug. Twenty-four yellow squash seedlings were transplanted into each plot on 3 Sep. Plant mortality due to P. capsici infection was recorded on 28 Sep, 5 Oct and 15 Oct.

Research results and discussion:
Glucosinolate analysis

Aboveground biomass production was negatively correlated with concentration of glucosinolates: Brassica yield was lower in the field than in the other environments, but glucosinolate concentration in the shoots was higher. These effects largely cancelled each other out, so that growing environment had little effect on total glucosinolate yield. The highest tissue concentration of glucosinolates was consistently found among the B. juncea accessions, but only PI 649112/Ames 8887 had a significantly higher glucosinolate concentration than the Eruca sativa accession PI 633215 across the range of environments tested. This accession had a lower biomass yield than the B. napus accession. The B. juncea accession ‘Pacific gold’, recently released as cover crop for biofumigation, did not differ significantly from other accessions in its biomass production or glucosinolate concentration, but tended to have a higher glucosinolate yield overall. Our results suggest that B. juncea accession ‘Pacific gold’ is the most promising biofumigation cover crop among those tested.

Activity against S. sclerotiorum in vitro

Sclerotial germination was reduced by glucosinolate extracts from all of the mustard accessions tested at the range of rates tested. The B. juncea accession ‘Pacific Gold’ was the most effective, completely inhibiting germination of sclerotia after exposure to the amount of glucosinolate extracted from 0.5 g of plant tissue in a vial containing 4 mL of liquid medium. PI 649112/Ames 8887 (Accession #12) was the only other accession to completely inhibit germination with glucosinolate extract from 1 g of fresh tissue. Most accessions completely inhibited germination after addition of extracts from 2 g of fresh tissue. The calculated dose response curve showed 50% inhibition of sclerotial germination after exposure to extracts from 0.65 g of tissue, on average. This effective concentration is equivalent to 0.16 g/mL, or 16 kg of fresh tissue per square meter of soil, incorporated to a depth of 10 cm. This rate is more than twice the highest rate of aboveground biomass production observed in our mustard test plantings.

Activity against P. capsici in vitro

All of the tested brassica accessions completely inhibited germination of P. capsici oospores after exposure to the amount of glucosinolate extracted from 0.5 g of plant tissue in a flask containing 4 mL of liquid medium. Inhibition averaged 96% at half this rate, and no lower rates were tested. A dose response curve could not be calculated because oospore germination was only observed at the lowest rate tested. Our in vitro tests suggest that P. capsici oospores are more susceptible to biofumigation than sclerotia of S. sclerotiorum.

Activity against S. sclerotiorum in soil

Slight inhibition of sclerotial germination was associated with higher rates of biomass incorporation into soil. Incorporation of mustard biomass offered no greater inhibition than incorporation of biomass from grass. No biofumigation effect was observed.

Field tests of biofumigation and solarization in high tunnels

Four weeks of summer solarization in a high tunnel completely inhibited germination of S. sclerotiorum sclerotia buried in the top 15 cm of soil in 2007. Sclerotial germination was also reduced by solarization in 2008, but complete inhibition of germination was only observed in sclerotia buried in the top 5 cm. August 2007 was much warmer than August 2008 at the experiment site (average 29 and 24 °C, in Aug 2007 and 2008, respectively). Solarization was as effective at the edge of solarized plots as in the middle. Treatments had no significant effect on soil enzyme activity, as measured by the soil’s ability to hydrolyze fluorescein diacetate before and after treatment. Biofumigation by incorporating fresh Pacific Gold mustard green manure did not reduce sclerotial germination in either year. Grower cooperators offered anecdotal reports of reduced S. sclerotiorum disease pressure after incorporating a high tunnel-grown cover crop of Pacific Gold mustard, instead of incorporating field-grown sheet mulch. There is potential for more mustard biomass to be added to the soil by a cover crop because it contributes root biomass in addition to aboveground biomass. A cover crop also offers the potential advantage of increasing the time in which soil-borne sclerotia are exposed to root exudates from the mustard. Our attempt to reproduce this reported effect under controlled conditions in 2009 was not successful. A high tunnel-grown cover crop of Pacific Gold mustard produced an average 0.6 kg of aboveground biomass per square meter at flowering, which was chopped by flail mowing and immediately incorporated into the soil. This offered no greater inhibition of S. sclerotiorum than a control crop of buckwheat, which produced a similar amount of biomass but none of the glucosinolates thought to be necessary for biofumigation. Four weeks of solarization in August 2009 was sufficient to completely inhibit germination of all sclerotia buried up to 15 cm below the soil surface.

Mustard planting date effect on field biomass production

All fall-seeded brassica accessions were winter killed in 2006 and 2007, leading to the decision to evaluate the potential for spring or summer plantings of biofumigant cover crops. Aboveground biomass produced at flowering by direct-seeded, rain-fed field plots of Pacific Gold mustard declined with each successive planting date between 24 Apr and 12 Jun, 2008. The average decline was 271 g per square meter per week, beginning at 1882 g per square meter for mustard seeded on 24 Apr, and falling to 6 g per square meter for that seeded on 12 Jun. Two plantings in late June and another in early July failed completely. The time required for flower initiation also declined with successive planting dates, from 56 days for the 24 Apr planting to 35 days for the 12 Jun planting. The observed planting date effect was likely due to warmer weather and drier soils as spring progressed. We conclude that Pacific Gold mustard is well suited to early spring production under field conditions in Kentucky. In other studies we succeeded in growing Pacific Gold mustard in June and July using irrigation.

Activity against P. capsici in soil

Incorporation of Pacific Gold mustard biomass into potting soil inhibited growth of yellow squash seedlings but did not protect the seedlings from P. capsici infection. Observed disease incidence was positively correlated with mustard incorporation rate.

Effect of biofumigation on field vegetable production under P. capsici pressure

Eighty-four percent of yellow squash plants died within six weeks of transplanting due to P. capsici infection. The cover crop incorporated before transplanting had no significant effect on mortality.

Discussion

Glucosinolates extracted from fresh mustard biomass inhibited survival of both S. sclerotiorum and P. capsici in the laboratory. The Indian mustard (B. juncea) variety ‘Pacific Gold’ demonstrated particular potential as a biofumigant crop in laboratory tests. Soil incorporation of fresh ‘Pacific Gold’ mustard biomass did not reduce disease incidence under greenhouse, high tunnel or field conditions. Possible reasons that success in the laboratory did not translate into success in the field include: 1. Low incorporation rate. Mustard biomass production under field conditions did not approach the range of application rates observed to reduce S. sclerotiorum survival in laboratory tests. 2. Timing. The highest rates of mustard biomass production were observed with early spring plantings. Early spring plantings of biofumigant cover crops may be compatible with field vegetable production systems, but are less compatible with high tunnel production systems, which rely on early spring cash crop production for much of their income. Mustards planted outdoors in the fall as a winter cover crop were winter killed before they contributed substantial biomass or initiated flowering. 3. Phytotoxicity. Growth of yellow squash seedlings was inhibited by mustard biomass incorporated into the soil. Sufficient time is needed between soil incorporation and planting to allow breakdown of mustards. We observed instances in which fresh biomass incorporation into soil reduced germination of S. sclerotiorum. This appeared to be an effect of biomass in general, as opposed to a biofumigation effect associated with release of glucosinolates from brassica biomass in particular. Incorporation of Pacific Gold mustard offered no greater inhibition of S. sclerotiorum than incorporation of equivalent amounts of non-brassica biomass from sources such as grass or buckwheat. Based on these results, we cannot recommend soil incorporation of fresh mustard biomass as a means of reducing disease pressure due to S. sclerotiorum or P. capsici in Kentucky. Summer solarization of high tunnel soil proved to be a very effective means of interrupting the S. sclerotiorum disease cycle. High tunnels are well adapted to solarization because they offer a protected environment in which soil temperature can be elevated substantially. We observed no reduction in the efficacy of solarization near the edge of solarized plots in high tunnels, in contrast to previous reports of “edge effects” in field studies. Solarization is best conducted at the hottest time of the year, which is not a key time for cash crop production in high tunnels. In three consecutive years we found that a month of solarization in August killed all sclerotia in the top 5 cm of high tunnel soil. In two of these years we saw complete inhibition of sclerotia down to 15 cm depth. High tunnel managers may enhance the efficacy of solarization by extending the solarization period beyond four weeks in unusually cool summers, and using shallow cultivation after solarization to avoid bringing viable sclerotia to the surface after solarization. Although solarization effectively interrupts the life cycle of S. sclerotiorum, it does not build soil organic matter. We found no significant effect of solarization on the ability of soil to hydrolyze fluorescein diacetate, which indicates soil enzyme activity. Previous work has demonstrated that solarization is more detrimental to mesophyllic organisms (most plant pathogens and pests) than to beneficial fungi and plant growth promoting bacteria (Stapleton and DeVay 1982, 1984; Gamliel and Katan 1991). The population of beneficial soil-dwelling organisms often expands rapidly after solarization, to fill the ecological vacuum left by the inactivation of pathogenic organisms. Some research suggests that soil inoculation with beneficial organisms is particularly effective after solarization (Ristaino et al. 1991; Gnanavel and Jayaraj 2003; Otieno et al. 2003; Minuto et al. 2006). Solarization has a place in an integrated, sustainable high tunnel system if organic matter can be built through other elements of the system.

Participation Summary

Educational & Outreach Activities

Participation Summary

Education/outreach description:
Presentations
  1. Winter Gardens in Solar Greenhouses. Bluegrass Energy Expo, Lexington, KY. 10-14-06 (http://organic.kysu.edu/WinterGardens.pdf). Pest management: What can an organic grower do about soil-borne diseases? Organic Third Thursday Thing, Frankfort, KY. 02-15-07 (http://organic.kysu.edu/BrianTTT.pdf). Organic Growing. Master Gardeners’ Class, Bath County, KY. 05-14-07 (http://organic.kysu.edu/OrganicGrowing.pdf). Season extension with high tunnels in Kentucky. American Society for Horticultural Sciences Annual Meeting, Scottsdale, AZ. 07-18-07 (http://organic.kysu.edu/KYHighTunnel.pdf). Organic Pest Management (building healthy soil without succumbing to weeds, insects & diseases). A Look at Organic Vegetable Production Workshop, University of Kentucky Cooperative Extension, Princeton, KY. 12-06-07 (http://organic.kysu.edu/OrganicPestManagement.pdf). Solarization for organic control of white mold in high tunnels. Kentucky Fruit and Vegetable Conference and Trade Show, Lexington, KY. 01-08-08 (http://organic.kysu.edu/Solarization.pdf). Sustainable Soil-Borne Disease Management: A Study of the Effects of Soil Solarization and Biofumigation on Sclerotinia sclerotiorum. Southern Sustainable Agriculture Working Group Meeting – Au Naturel Farm Field Trip, Smiths Grove, KY. 01-16-08 (http://organic.kysu.edu/SSAWGPoster.pdf). Effect of Planting Date on Biomass Production by Brassica juncea var. ‘Pacific Gold’ Cover Crops. Kentucky Academy of Science Meeting, Lexington, KY. 11-01-08 (http://organic.kysu.edu/Evaluation%20of%20Various%20Planting%20Dates%20of%20Brassica%20Juncea.pdf). Effect of Glucosinolate Exposure on Sclerotinia sclerotiorum and Phytopthora capsici. Kentucky Academy of Science Meeting, Lexington, KY. 11-01-08 (http://organic.kysu.edu/BomfordKAS2008.pdf). A Simplified Procedure for Glucosinolate Quantification. Kentucky Academy of Science Meeting, Lexington, KY. 11-01-08 (http://organic.kysu.edu/GlucosinolateQuant.pdf). Vegetable Production in High Tunnels. Fairview Produce Auction, Christian County, KY. 12-10-08 (http://organic.kysu.edu/HighTunnelVeg.pdf). Fresh Tomatoes in January: Can B-ISA Make them Sustainable? Building-Integrated Sustainable Agriculture Summit, Berkeley, CA, 12-13-08 (http://organic.kysu.edu/B-ISA-HighTunnel.pdf). Sustainable Soil-Borne Disease Management presentation, Southern Sustainable Agriculture Working Group High Tunnel Short Course. Chatanooga, TN, 01-22-09. (http://organic.kysu.edu/SSAWG2009.pdf) Organic Agriculture at Kentucky State University. Third Thursday Thing, Organic Agriculture Day, Frankfort, KY, 02-19-09. (http://organic.kysu.edu/OrganicTTT2009.pdf) Effectiveness of a Blend of Beneficial Microorganisms and Brassica Green Manures in Reducing Damage by Phytophthora capsici to Yellow Squash (Curbita pepo) Seedlings. Bio 410 Seminar Series, Kentucky State University, Frankfort, KY, 03-06-09. (http://organic.kysu.edu/Abdul-Phytophthora.pdf) Managing Sclerotinia sclerotiorum in High Tunnels with Biofumigation and Solarization. Plant Pathology Seminar Series, University of Kentucky, Lexington, KY, 03-09-09. (http://organic.kysu.edu/UKPathologySeminar.pdf) Organic Growing. Henderson County Master Gardeners’ Class, Henderson, KY, 03-19-09. (http://organic.kysu.edu/HendersonOrganicGardeners.pdf) Solarization and biofumigation for organic control of white mold in high tunnels. American Society for Horticultural Science Meeting, St. Louis, MO, 07-27-09. (http://ashs.org/db/horttalks/detail.lasso?id=685) Effect of Biofumigation and Soil Solarization on Sclerotinia sclerotiorum in High Tunnel Vegetable Production Systems of Kentucky. Kentucky Academy of Science Meeting, Highland Heights, KY, 11-14-09. (http://organic.kysu.edu/Geier-KAS2009-Biofumigation.pdf) Organic Agriculture. Russell County Extension FarmStart series. Russell County, KY, 04-08-10 (http://organic.kysu.edu/RussellCountyOrganic.pdf) Organic Agriculture. Lawrence County Extension Gardeners’ Toolbox Series. Lawrence County, KY, 04-27-10 (http://organic.kysu.edu/LawrenceCountyOrganic.pdf) High Tunnel Solarization. Horticulture Third Thursday Thing. Frankfort, KY, 06-17-10 (http://organic.kysu.edu/HTSolarization.pdf). High Tunnel Solarization. Kentucky State University Sustainable Farms, Families, and Farm Energy Field Day. Frankfort, KY, 07-15-10.
Print
  1. G.F. Antonious, M.K. Bomford & P.C. Vincelli. 2009. Screening brassica species for glucosinolate content. Journal of Environmental Science and Health 44: 311-316. Michael Bomford, Paul Vincelli, George Antonious, Brian Geier and Ed Dixon. 2009 (Abstract). Solarization and biofumigation for organic control of white mold in high tunnels. HortScience 44: 1041. M.K. Bomford, P.C. Vincelli, E.W. Dixon, and B.A Geier. 2007. M.K. Bomford, A.F. Silvernail and B.A. Geier. 2007 (Abstract). Season extension with high tunnels in Kentucky. HortScience 42: 988. Sara Gividen. High Life Helps Plants Survive Low Temps. Frankfort State Journal, p. A1. 04/18/2007. Evaluation of solarization and Contans WG for control of Sclerotinia sclerotiorum in high tunnels, 2006. Plant Disease Management Reports 1: V163. Other print publications being prepared from this work include a factsheet on solarization in high tunnels and another journal article detailing our findings.
Web
  1. Biofumigation Studies. (http://organic.kysu.edu/Biofumigation.shtml) On-Farm Study Results: Biofumigation and Soil Solarization. (http://organic.kysu.edu/biofumigation3.shtml) Organic Disease Management in High Tunnels: A Little Piece of the Sustainable Agriculture Puzzle (Post to Energy Farms blog). (http://energyfarms.wordpress.com/2008/11/26/organic-disease-management-in-high-tunnels-a-little-piece-of-the-sustainable-agriculture-puzzle/) Streaming Video: Organic Control of White Mold in High Tunnels. (http://organic.kysu.edu/SclerotiniaVideo.shtml)

Project Outcomes

Project outcomes:
  • More than 355 people attended workshops, tours, and presentations discussing this project (see outreach section) Preliminary results and presentation materials have been archived on the Kentucky State University Organic Agriculture Working Group website, which attracted 92,297 hits from 57,996 unique viewers between April 2006 and March 2010. Grower cooperators Paul and Alison Wiediger and at least four other other high tunnel growers in Kentucky have altered their production systems to include a period of summer solarization for management of S. sclerotiorum. Undergraduate and graduate students John Rodgers, Abdul Kakar, Amy Bateman, and Mike Ward received hands-on training in research techniques through involvement with this project. Project participants have responded to more than 30 requests for information about winter vegetable production and soil-borne disease management in Kentucky. The number of certified organic farms in Kentucky increased from 12 to 125 over the course of this project. The support offered by this, and other, projects may be a factor in this rapid growth. Recommendations generated by this research are making high tunnel vegetable production in Kentucky more sustainable by helping to build soil health, reducing the need for external inputs, and allowing diverse crop rotations that include the many high tunnel crops susceptible to S. sclerotiorum. Evidence that mustards are not a suitable winter cover crop for Kentucky, and do not contribute to control of P. capsici, allows field vegetable growers to select cover crops that offer other advantages, such as fixing nitrogen, protecting winter soil from erosion, and building soil organic matter more rapidly than mustard. We anticipate that new disease management tactics generated by this project will encourage Kentucky farmers to consider vegetable production as a high value alternative to tobacco.

Farmer Adoption

  • At least five farmers who have attended our workshops have been inspired to erect new high tunnels. At least five Kentucky farmers, including the grower cooperators involved with this project, are incorporating solarization periods into high tunnel rotations.
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.