There is a stark difference in plant vigor between an ASD-treated plot (left) and a standard untreated plot in an organic field infected with charcoal rot. Photo by Joji Muramoto

Anaerobic soil disinfestation helps suppress weeds, disease without fumigants

Troubled by puny plants, low yields and persistent mite problems, third-generation Southern California strawberry grower Glen Hasegawa was ready to give up on his transition from conventional to organic 12 years ago.

“I've always liked a challenge – but it turned out to be more of a challenge than I thought it would be!” he said.

But then, with the help of scientists including Oleg Daugovish, UC Cooperative Extension strawberry and vegetable crop advisor in Ventura County, Hasegawa tried a technique called anaerobic soil disinfestation (ASD). When applied correctly, the multi-step ASD process creates a soil environment that suppresses pathogens and weeds and makes for healthier, more robust crop growth.

“Back in the day, it was really hard to get the plant growing vigorously in organic,” said Hasegawa, owner of Faria Farms in Oxnard. “So we started using the ASD and then you could definitely see that the plant had more vigor and you could grow a bigger, better plant using it.”

Seeing that he could produce yields “in the neighborhood” of those grown in conventional strawberry fields fumigated with synthetic fumigants, Hasegawa was able to expand his original 10 acres of organic strawberries to 50 acres.

“I guess you could say I'm kind of a convert,” he said, noting that he now applies ASD to all his acreage each year in late spring.

Joji Muramoto, UC Cooperative Extension specialist in organic production based at UC Santa Cruz, has been experimenting with ASD since it was first brought to the U.S. from the Netherlands and Japan in the early 2000s. Carol Shennan, a professor in the Department of Environmental Studies at UCSC, and Muramoto were among the first to try the technique in California. They found that ASD successfully controlled an outbreak of Verticillium wilt – caused by the pathogen Verticillium dahliae – at UCSC's small organic farm in 2002.

Since then, Shennan, Muramoto, Daugovish and their colleagues have seen encouraging results at 10 trial sites across the state.

“We demonstrated that ASD can provide comparable yields with fumigants, in side-by-side replicated trials,” Muramoto said.

The strawberry plants in the ASD-treated plot (left) are more robust than those growing in untreated soil. A plastic mulch (typically opaque is used by growers, but clear was used here for research purposes) is crucial as part of the ASD process to improve plant productivity and control weeds. Photo by Oleg Daugovish

ASD promotes host of beneficial changes to soil ecosystem

ASD comprises three basic steps: incorporating a carbon source that is easily digestible by microbes in the soil (traditionally, rice bran has been used), further encouraging fermentation by covering the soil with plastic to limit oxygen supply, and finally adding water through drip irrigation to initiate the “anaerobic” decomposition of the carbon source and maintain the three-week “cooking” process.

The resulting cascade of chemical, microbiological and physical changes to the soil creates an ecosystem that is both conducive to strawberry growth – and inhospitable to pathogens and weeds.

“It's not like a pesticide where you have a mode of action, and thus resulting in ‘A' and ‘B' for you,” Daugovish explained. “There's a sort of cocktail of events that happens in the soil; they all happen interconnectedly.”

Compared to similar fields that did not undergo the process, ASD-applied organic strawberry fields across California have seen yields increase by 60% to 70% – and even doubling in some cases, according to Daugovish.

The UCCE advisor also shared the story of a longtime grower in Ventura County, who came to him with fields in “miserable” condition; they were plagued by one of the world's worst weeds, yellow nutsedge, and infected with charcoal rot, a disease caused by the fungus Macrophomina phaseolina. But after applying rice bran and following the ASD recipe, the grower saw phenomenal results.

“The only complaint he said to me was, ‘Now I have too many berries – we have to have more pickers to pick the berries!'” Daugovish recalled.

Via researchers' meetings, online resources, on-farm demonstration trials and word of mouth from peers, use of ASD by California strawberry growers has grown significantly during the past two decades. Tracking the purchase of rice bran, Muramoto estimated that about 2,500 acres were treated by the ASD-related practices in 2023 – covering roughly half of the 5,200 total acres of organic strawberries in California.

Muramoto directly links the growth of California organic strawberry production – which now comprises about 13% of total strawberry acreage in the state – with the increasing adoption of ASD.

“If you remove the acreage with the applied rice bran over the last 10 years or so, organic strawberry acreage is just flat,” he said.

Applying rice bran provides the easily digestible carbon source that soil microbes feed on. Once covered by plastic mulch, anaerobic conditions develop and microorganisms that prefer such an environment thrive, while pathogenic organisms and germinating weeds are suppressed. Before planting, holes are cut in the plastic to let oxygen back into the soil and encourage the growth of the strawberry plants. Photo by Oleg Daugovish

Within the last decade, acreage of organic strawberries with ASD-related practices increased by 1,640 acres, which is a boon for air quality, human health and long-term soil vitality. According to Muramoto's calculations, that increase in organic acreage translates to a reduction of about 465,000 pounds of fumigant active ingredients that would have been used in growing conventional strawberries.

“There are hundreds of reports of acute illnesses related to fumigation in the record, so it's very important to find alternatives to fumigants,” said Muramoto, citing California Department of Pesticide Regulation documents.

Research continues to make ASD more economical, effective

The popularity of ASD has come at a price, however, for organic strawberry growers.

“There's more organic out there, and I think most of the organic guys are using it, so there's more demand on the rice bran; the price has been steadily going up every year, like everything else,” said Hasegawa, adding that he has been trying to decrease the amount of carbon while maintaining ASD's efficacy.

On top of greater demand from other growers and from beef cattle and dairy producers (who use rice bran as feed), the price also has increased due to higher costs in transporting the material across the state from the Sacramento Valley. So Daugovish and his colleagues – including Peter Henry, a U.S. Department of Agriculture plant pathologist – have been searching for a cheaper alternative.

“We all want an inexpensive, locally available, reliable, easy to use and functional carbon source, which sounds like a big wish list,” Daugovish said.

Carbon sources such as bark, wood chips, or compost are ineffective, as the crucial ASD microorganisms are choosy about their food.

In a field of 'Victor' strawberries, researchers saw a 41% increase in yield using ASD with midds, in comparison to untreated soil. Photo by Oleg Daugovish

“Microbes are just like cows; you can't feed them straight wood; they get pretty angry,” Daugovish explained. “And if you feed them something with too much nitrogen, they can't digest it – they get the runs. Microbes are the same way – you have to have the right proportion of stuff so they feel comfortable doing what they're doing.”

In search of an ideal replacement, researchers tried and ruled out grass clippings, onion waste, glycerin and coffee grounds. Finally, they pivoted to a material with properties very similar to rice bran: wheat bran, in the form of wheat middlings (also called midds, a byproduct of flour milling) and dried distillers' grain (DDG, a byproduct of ethanol extraction).

After field experiments in Santa Paula, the UC and USDA researchers found that midds and DDG were just as effective at controlling soilborne pathogens and weeds as rice brain – but at 25% to 30% less cost. Their results were published last year in the journal Agronomy.

“Not surprisingly, the wheat bran has worked almost exactly the same as rice bran,” Daugovish said.

He and Muramoto are now conducting trials with wheat bran at commercial fields, and the initial results are promising. Daugovish said the grower at one site in Ventura County has seen a 90% reduction in Macrophomina phaseolina, the causal pathogen of charcoal rot, in the soil – and an 80% to 90% drop in yellow nutsedge germination. They are waiting for final yield numbers after the coming summer.

While ASD has been beneficial to organic productivity and soil health, both Daugovish and Muramoto acknowledged specific limitations in suppressing the “big three” strawberry diseases: Verticillium wilt, Fusarium wilt and charcoal rot. In coastal areas with cooler soil temperatures, for example, ASD can actually exacerbate the latter two diseases, as the fungal pathogens feed on the rice bran.

“We know it works at warmer temperatures, but, practically, it's hard to do in coastal California,” Muramoto said. “It would be nice if we can find a way to suppress Fusarium wilt at a lower temperature, but we don't have it right now.”

That's why researchers emphasize that ASD is not a “silver bullet.” It's just one tool in the organic toolbox, which includes careful crop rotation, disease-resistant strawberry varieties and better diagnostic tests that help growers pinpoint outbreaks and make the application of various methods more targeted and more efficient.

And scientists will continue to optimize ASD to make it more effective and economical for growers in the different strawberry regions of California – from the Central Coast to the Oxnard Plain.

“We know it can work really well; it's just finding the most sustainable way to do this in our region,” Daugovish said. “We've got to just have an open mind and keep trying.”

Although the strawberry industry has lagged behind crops like tomato and wheat on genetic and technical innovation, enhanced plant breeding, gene editing and other techniques are seen as crucial to ensuring strawberry crops are sustainable. Photo by Tetiana Bykovets on Unsplash

The $6.2 million grant centers on protecting crops in the future

The federal government is awarding $6.2 million to University of California, Davis, to study how to use breeding and genetic information to protect strawberry crops from future diseases and pests.

The four-year grant from the National Institute of Food and Agriculture (NIFA) centers on addressing expanding and emerging threats to strawberries, a popular fruit packed with Vitamin C and key to the diets of many Americans.

Enhanced plant breeding, gene editing and other technologies will be key to ensuring strawberry crops are sustainable in the face of climate change and possible restrictions on chemical use, said Steve Knapp, director of the Strawberry Breeding Center and a distinguished professor in the Department of Plant Sciences.

“We need to have the technology so that we can deal with the challenges strawberries face around the world,” Knapp said. “Can we use genetic knowledge to change the DNA in a specific way to get the resistance we need?”

USDA funding

The grant award was one of 25 announced Oct. 5 by NIFA – an agency of the U.S. Department of Agriculture – as part of the Specialty Crop Research Initiative program, which addresses “key challenges of national, regional and multistate importance in sustaining all components of food and agriculture…,” the agency said.

The strawberry industry has lagged behind crops like tomato and wheat when it comes to genetic and technical innovation, Knapp said, and the grant signifies that “now they want the foot on the accelerator.”

A key priority is identifying whether changing DNA molecules can improve disease resistance and what technologies would be needed. Ensuring some genes are expressed while others are suppressed would be part of the analysis.

“We're trying to build in natural resistance to pathogens through the genes that already exist but could be modified with this knowledge,” Knapp said. “If we were able to edit a gene that improves disease resistance, people would want us to use that in breeding.”

The intent is to produce disease-resistant cultivars and identify better ways to diagnose, prevent and manage disease. The research project will also include an economic forecast evaluating the consequences of production changes and communicating with farmers about the laboratory advances, according to the grant proposal.

Gitta Coaker from plant pathology and Mitchell Feldmann, Marta Bjornson and Juan Debernardi from plant sciences are participating in the research, as are scientists from California Polytechnic State University, UC Agriculture and Natural Resources, UC Berkeley, University of Florida and USDA's Agricultural Research Service.

Botrytis cinerea infection appears as wilted flowers and a layer of spores on ripe fruit.  Photo by Surendra Dara

Botrytis fruit rot or gray mold caused by Botrytis cinerea is an important disease of strawberry and other crops damaging flowers and fruits.  Pathogen survives in the plant debris and soil and can be present in the plant tissues before flowers form.  Infection is common on developing or ripe fruits as brown lesions.  Lesions typically appear under the calyxes but can be seen on other areas of the fruit.  As the disease progresses, a layer of gray spores forms on the infected surface.  Severe infection in flowers results in the failure of fruit development.  Cool and moist conditions favor botrytis fruit rot development.  Sprinkler irrigation, rains, or certain agricultural practices can contribute to the dispersal of fungal spores.

Although removal of infected plant material and debris can reduce the source of inoculum in the field, regular fungicide applications are typically necessary for managing botrytis fruit rot.  Since fruiting occurs continuously for several months and fungicides are regularly applied, botrytis resistance to fungicides is not uncommon.  Applying fungicides only when necessary, avoiding continuous use of fungicides from the same mode of action group (check FRAC mode of action groups), exploring the potential of biological fungicides to reduce the risk of resistance development are some of the strategies for effective botrytis fruit rot management.  In addition to several synthetic fungicides, several biological fungicides continue to be introduced into the market offering various options for the growers.  Earlier field studies evaluated the potential of various biological fungicides and strategies for using them with synthetic fungicides against botrytis and other fruit rots in strawberry (Dara, 2019; Dara, 2020).  This study was conducted to evaluate some new and soon to be released fungicides in fall-planted strawberry to support the growers, ag input industry, and to promote sustainable disease management through biological and synthetic pesticides.

Methodology

This study was conducted at the Manzanita Berry Farms, Santa Maria in strawberry variety 3024 planted in October, 2020.  While Captan and Switch were used as synthetic standards, a variety of biological fungicides of microbial, botanical, and animal sources were included at various rates and different combinations and rotations.  Products and active ingredients evaluated in this study included Captan Gold 4L (captan) from Adama, Switch 62.5 WG (cyprodinil 37.5% + fludioxinil 25%) from Syngenta, NSTKI-14 (potassium carbonate 58.04% + thyme oil 1.75%) from NovoSource, A22613 [A] (botanical extract) from Syngenta, Regalia (giant knotweed extract 5%) from Marrone Bio Innovations, EXP14 (protein 15-20%) from Biotalys, Gargoil (cinnamon oil 15% + garlic oil 20%) and Dart (caprylic acid 41.7% + capric acid 28.3%) from Westbridge, Howler (Pseudomonas chlororaphis strain AFS009), Theia (Bacillus subtilis strain AFS032321), and Esendo (P. chlororaphis strain AFS009 44.5% + azoxystrobin 5.75%) from AgBiome, ProBlad Verde (Banda de Lupinus albus doce – BLAD, a polypeptide from sweet lupine) from Sym-Agro with Kiplant VS-04 (chitosan 2.3%) or Nu-Film-P spreader/sticker, AS-EXP Thyme (thyme oil) from AgroSpheres, and AgriCell FunThyme (thyme oil) provided by AgroSpheres.

Table 1. List of treatments color coded according to the kind of fungicide (light blue=synthetic fungicide; dark blue=synthetic+biological fungicide active ingredient; peach=synthetic and biological fungicides alternated; green=biological fungicides)

Excluding the untreated control, rest of the 24 treatments can be divided into synthetic fungicides, a fungicide with synthetic + biological active ingredients (a formulation with two application rates), synthetic fungicides alternated with biological fungicides, and various kinds of biological fungicides  (Table 1).  Treatments were applied at a 7-10 day interval between 22 April and 17 May, 2021.  Berries for pre-treatment disease evaluation were harvested on 19 April, 2021.  Each treatment had a 5.67'X15' plot replicated four times in a randomized complete block design.  Strawberries were harvested 3 days before the first treatment and 3-4 days after each treatment for disease evaluation.  On each sampling date, marketable-quality berries were harvested from random plants within each plot during a 30-sec period and incubated in paper bags at outdoor temperatures under shade.  Number of berries with botrytis infection were counted on 3 and 5 days after harvest (DAH) and percent infection was calculated.  This is a different protocol than previous years' studies where disease rating was made on a 0 to 4 scale.  Treatments were applied with a backpack sprayer equipped with Teejet Conejet TXVK-6 nozzle using 90 gpa spray volume at 45 PSI.  Water was sprayed in the untreated control plots.  Dyne-Amic surfactant at 0.125% was used for treatments that contained Howler, Theia, Esendo, AgriCell FunThyme, AS-EXP Thyme, and EXP 14.  Research authorization was obtained for some products and crop destruction was implemented for products that did not have California registration.

Percent infection data were arcsine-transformed before subjecting to the analysis of variance using Statistix software.  Significant means were separated using the least significant difference test.

Results

Pre-treatment infection was very low and occurred only in some treatments with no statistical difference (P > 0.05).  Infection levels increased for the rest of the study period.  There was no statistically significant difference (P > 0.05) among treatments for disease levels 3 or 5 days after the first spray application.  Differences were significant (P = 0.0131) in disease 5 DAH after the second spray application where 13 treatments from all categories had significantly lower infection than the untreated control.  After the third spray application, infection levels were significantly lower in eight treatments in 3 DAH observations (P = 0.0395) and 10 treatments in 5 DAH observations (P = 0.0005) compared to untreated control.  There were no statistical differences (P > 0.05) among treatments for observations after fourth spray application or for the average of four applications.  However, there were numerical differences where infection levels were lower in several treatments than the untreated control plots.

In general, the efficacy of both synthetic and biological fungicides varied throughout the study period among the treatments.  When the average for post-treatment observations was considered, infection was numerically lower in all treatments regardless of the fungicide category.  Multiple biological fungicide treatments either alone or in rotation with synthetic fungicides appeared to be as effective as synthetic fungicides.

Conclusions

Botanical and microbial fungicides can be effective against either for using alone or in rotation with synthetic fungicides for suppressing botrytis fruit rot in strawberry.  Additional studies can help optimize the application rates and use strategies for those fungicides that were not as effective as others.  Sanitation practices and use of synthetic and biological fungicides help manage botrytis fruit rot.

Acknowledgements: Thanks to AgBiome, AgroSpheres, Biotalys, NovaSource, Sym-Agro, Syngenta, and Westbridge for funding and Chris Martinez for his technical assistance.

References

Dara, S. K.  2019.  Five shades of gray mold control in strawberry: evaluating chemical, organic oil, botanical, bacterial, and fungal active ingredients.  UCANR eJournal of Entomology and Biologicals.  https://ucanr.edu/blogs/blogcore/postdetail.cfm?postnum=30729

Dara, S. K. 2020.  Evaluating biological fungicides against botrytis and other fruit rots in strawberry.  UCANR eJournal of Entomology and Biologicals. https://ucanr.edu/blogs/blogcore/postdetail.cfm?postnum=43633

Strawberry, a high-value specialty crop in California, suffers from several soilborne, fruit, and foliar diseases.  Verticillium wilt caused by Verticillium dahliae, Fusarium wilt caused by Fusarium oxysporum f. sp. fragariae, and Macrophomina crown rot or charcoal rot caused by Macrophomina phaseolina are major soilborne diseases that cause significant losses without proper control.  Chemical fumigation, crop rotation with broccoli, nutrient and irrigation management to minimize plant stress, and non-chemical soil disinfestation are usual control strategies for these diseases.  Botrytis fruit rot or gray mold caused by Botrytis cineaea is a common fruit disease requiring frequent fungicidal applications.  Propagules of gray mold fungus survive in the soil and infect flowers and fruits.  A study was conducted to evaluate the impact of drip application of various fungicides on improving strawberry health and enhancing fruit yields.

Methodology

This study was conducted in an experimental strawberry field at the Shafter Research Station during 2019-2020.  Cultivar San Andreas was planted on 28 October 2019.  No pre-plant fertilizer application was made in this non-fumigated field which had Fusarium wilt, Macrophomina crown rot, and Botrytis fruit rot in the previous year's strawberry planting.  Each treatment was applied to a 300' long bed with single drip tape in the center and two rows of strawberry plants.  Sprinkler irrigation was provided immediately after planting along with drip irrigation, which was provided one or more times weekly as needed for the rest of the experimental period.  Each bed was divided into six 30' long plots, representing replications, with an 18' buffer in between.  Between 6 November 2019 and 9 May 2020, 1.88 qt of 20-10-0 (a combination of 32-0-0 urea ammonium nitrate and 10-34-0 ammonium phosphate) and 1.32 qt of potassium thiosulfate was applied 20 times at weekly intervals through fertigation.  Treatments were applied either as a transplant dip or through the drip system using a Dosatron fertilizer injector (model D14MZ2).  The following treatments were evaluated in this study:

i)  Untreated control: Neither transplants nor the planted crop was treated with any fungicides.

ii) Abound transplant dip: Transplants were dipped in 7 fl oz of Abound (azoxystrobin) fungicide in 100 gal of water for 4 min immediately prior to planting.  Transplant dip in a fungicide is practiced by several growers to protect the crop from fungal diseases.

iii) Rhyme: Applied Rhyme (flutriafol) at 7 fl oz/ac immediately after and 30, 60, and 90 days after planting through the drip system.

iv) Velum Prime with Switch: Applied Velum Prime (fluopyram) at 6.5 fl oz/ac 14 and 28 days after planting followed by Switch 62.5 WG (cyprodinil + fludioxinil) at 14 oz/ac 42 days after planting through the drip system.

v) Rhyme with Switch: Four applications of Rhyme at 7 fl oz/ac were made 14, 28, 56, and 70 days after planting with a single application of Switch 62.5 WG 42 days after planting through the drip system.

Parameters observed during the study included leaf chlorophyll and leaf nitrogen (with chlorophyll meter) in February and May; fruit sugar (with refractometer) in May; fruit firmness (with penetrometer) in April and May; severity of gray mold (caused by Botrytis cinereae) twice in March and once in May, and other fruit diseases (mucor fruit rot caused by Mucor spp. and Rhizopus fruit rot caused by Rhizopus spp.) once in May 3 and 5 days after harvest (on a scale of 0 to 4 where 0=no infection; 1=1-25%, 2=26-50%, 3=51-75% and 4=76-100% fungal growth); and fruit yield per plant from 11 weekly harvests between 11 March and 14 May 2020.  Leaf chlorophyll and nitrogen data for the Abound dip treatment were not collected in February. Data were analyzed using analysis of variance in Statistix software and significant means were separated using the Least Significant Difference test.

Results and Discussion

Leaf chlorophyll content was significantly higher in plants that received drip application of fungicides compared to untreated plants in February while leaf nitrogen content was significantly higher in the same treatments during the May observation.  There were no differences in fruit sugar or average fruit firmness among the treatments.

The average gray mold severity from three harvest dates was low and did not statistically differ among the treatments.  However, the severity of other diseases was significantly different among various treatments with the lowest rating in Abound transplant dip on both 3 and 5 days after harvest and only 3 days after harvest in plants that received four applications of Rhyme.  Unlike the previous year, visible symptoms of the soilborne diseases were not seen during the study period to evaluate the impact of the treatments.  However, there were significant differences among treatments for the marketable fruit yield.  The highest marketable yield was observed in the treatment that received Rhyme and Switch followed by Velum Prime and Switch and Rhyme alone.  The lowest fruit yield was observed in Abound dip treatment.  Unmarketable fruit (deformed or diseased) yield was similar among the treatments.  Compared to the untreated control, Abound dip resulted in 16% less marketable yield and such a negative impact from transplant dip in fungicides has been seen in other studies (Dara and Peck, 2017 and 2018; Dara, 2020).  Marketable fruit yield was 4-28% higher where fungicides were applied to the soil. 

Although visible symptoms of soilborne diseases were absent during the study, periodic drip application of the fungicides probably suppressed the fungal inocula and associated stress and might have contributed to increased yields.  The direct impact of fungicide treatments on soilborne pathogens was, however, not clear in this study due to the lack of disease symptoms.  Considering the cost of chemical fumigation or soil disinfestation and the environmental impact of chemical fumigation, treating the soil with fungicides can be an economical option if they are effective.  While this study presents some preliminary data, additional studies in non-fumigated fields in the presence of pathogens are necessary to consider soil fungicide treatment as a control option.

Field assistants Tamas Zold and Marjan Heidarian Dehkordi

Acknowledgments: Thanks to FMC for funding this study and Marjan Heidarian Dehkordi and Tamas Zold for their technical assistance.

References

Dara, S. K. 2020.  Improving strawberry yields with biostimulants and nutrient supplements: a 2019-2020 study.  UCANR eJournal of Entomology and Biologicals. https://ucanr.edu/blogs/blogcore/postdetail.cfm?postnum=43631

Dara, S. K. and D. Peck.  2017.  Evaluating beneficial microbe-based products for their impact on strawberry plant growth, health, and fruit yield.  UCANR eJournal of Entomology and Biologicals.  https://ucanr.edu/blogs/blogcore/postdetail.cfm?postnum=25122

Dara, S. K. and D. Peck.  2018.  Evaluation of additive, soil amendment, and biostimulant products in Santa Maria strawberry.  CAPCA Adviser, 21 (5): 44-50.

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