How to Cultivate Bioregional Mycorrhizal Fungi for Plants
Instructions adapted from Danielle Stevenson, D.I.Y. Fungi, This content was taught at workshop with Danielle for Healthy Soils, Healthy Trees program and part of tree drought research with TreeFolks. Thanks to Ecology Action and City of Austin Urban Forestry Grant for supporting this program.
There are several types of mycorrhizal fungi, but ectomycorrhizal (EMF) and arbuscular mycorrhizal fungi (AMF) are the most common. It is a good idea to look up what kind of mycorrhizal relationships the plant(s) you’re trying to cultivate form and cultivate those.
Watch part one of our workshop series with Danielle to learn more about mycorrhizal fungi.
Do you need to restore mycorrhizal populations? Yes in:
intensively managed agricultural lands (industrial ag- frequently tilled, heavily
fertilized, fungicide use)
degraded soils (remediation of mine sites and brownfields, ecological restoration and reforestation efforts)
nurseries (and hydroponic or container gardens)
You can restore mycorrhizal fungi through agricultural and land management practices or through introduction of mycorrhizal inoculum. Management practices that encourage mycorrhizal fungi:
Crop rotation including deep rooting of mycorrhizal plants
Cover cropping - do not leave soil bare
Reduced/no till
Deep plowing
Reduced fertilization (esp Phosphorus)
Where to source your MYCORRIZAL SPORES?
You can purchase commercial mycorrhizal spores to inoculate plants and soil but recent meta analysis has shown they are less effective than using methods that introduces native, cultivated mycorrhizae. Mycorrhizal spores, pieces of mycorrhizal roots and viable mycorrhizal hyphae are active propagules that can be used as inoculum.
Methods OF CULTIVATING AND AMPLIFYING Native mycorrhiza
Whole Soil Method
Trap Culture Method
Nurse Plant Method
Spore Slurry Method (EMF)
Species-specific AMF Method
Whole Soil METHOD
Takes soil from a healthy reference ecosystem as inoculum
It inoculates with intact rhizosphere soil containing the complete array of the soil community including AM fungi, beneficial bacteria such as nitrogen fixing Rhizobia, pathogens, soil-dwelling insects, nematodes, plant roots, and even seeds.
Destructive of healthy ecosystems and requires huge amounts of soil (150-10,000 gallons per acre).
Workshop series with Danielle at Circle Acres on how to cultivating bioregional mycorrhizal fungi using trap culture methods.
Trap Culture Methods
The following method is what we are using for our Healthy Soils, Healthy Trees research.
Works to amplify the volume of whole soil microbes so that a few liters of whole soil inoculum can be used to inoculate hundreds of plants.
Trap cultures are a way of ‘bulking up’ the AM fungal community present in whole soil before it is used as inoculum in a restoration. The AM fungi + plant dependent soil microbes are amplified by growing with plant hosts in a pot and this allows their propagation for future use.
Step-by-Step: Trap Method for Cultivating Native Mycorrhizal Fungi
Materials needed:
Clean Nursery pots or fabric grow bags. (The size of the pot should consider the root depth of the host plant at maturity.)
Native soil (from under healthy, mature trees)
Sterile potting mix or compost (See DIY mix below)
Host Plant Seeds: Bluestem, Corn, Sorghum
Shade cloth or partial shade location
Access to Water
Optional: mesh or hardware cloth to deter critters
Collecting soil from mother trees in the winter.
1. Gather Native Soil: Collect small amounts of soil from mature, healthy native plants you plan to propagate. This soil contains mycorrhizal fungi, root fragments, and microbes — the soil microbiome.
2. Mix Native Soil with Coarse Sand: Combine at a 1:1 ratio by volume (not weight).
3. Mix Your Own Sterile Potting Soil (Optional): It is becoming more common for AMF or EMF fungi to be added commercial mixes of potting soil. To ensure you have a sterile potting soil, we recommend a blend of coco coir and vermiculite at a 1:1 ratio by volume.
4. Combine Native Mix with Sterile Soil: Mix the native soil and sand with the sterile mix at a 1:1 ratio. Fill into clean nursery pot/s. Exact ratios aren’t critical — what matters is seed contact with native soil.
Winter rye grass planted with native soil after planting at Circle Acres in February 2025.
5. Seed Host Plant into Pots: Use grasses like bluestem, corn, or sudangrass, which tolerate heat and rely on AMF. Overseed the pot.
6. Water Regularly: Keep soil moist but not soggy. Mimic natural rainfall. Use rainwater if possible.
7. Provide Shade: Place pots in partial shade to reduce seedling stress and encourage fungal growth.
8. Grow Host Plant for 3–6 Months: Over time, fungi from the native soil colonize the roots. You’re “farming” fungal partnerships.
9. Stop Watering and Let Plants Dry Out: Leave pots undisturbed for 2–3 weeks. This triggers the host plant to invest carbon into AMF. As it dies back, AMF sporulate. Then, cut the shoots at soil level.
Rye grass in May 2025 before we stopped watering.
10. Harvest, Store, and Use Inoculum: Harvest the soil or chop the roots and mix with clean sand. This root-sand mix is your concentrated AMF inoculum. Store in plastic containers in a cool, dry place — it remains viable for at least a year. Use it for new pot cultures, in potting mixes, or to inoculate “nurse” plants in the greenhouse or field. Apply it in gardens, restoration sites, or urban forests.
For the Healthy Soils, Healthy Trees research we will store all of the soil in 5 gallon pots indoors until it is time to plant trees in the late winter. Watch recording of workshop where we harvest the roots and validated that we cultivated AMF.
Variations:
INVAM method: International Culture Collection of (Vesicular) Arbuscular Mycorrhizal Collection at West Virginia University.
Rodale-USDA method: On Farm Arbuscular Mycorrhizal Inoculum Production. Rodale Institute.
Example of in-ground trap method in Africa.
Low-Tech, In-Ground Trap Cultures: In-Ground Trap Cultures: Missouri Botanical Garden and the Restoration EcologyLab at Virginia Tech
Fill a pit (150 cm long × 50 cm wide × 30 cm deep) lined with sacks with topsoil collected from around the roots of three native tree species
Then grow maize and beans in this soil for three months before cutting these plants down
Letting the substrate dry out for two weeks. The substrate remaining in the pit is the inoculum
Use by adding one tablespoon inoculum to each seedling container.
The nurse plant method
Inoculating potted seedlings with AM fungi before planting into a site allows them to act as nurse plants
Enhances the likelihood that the inoculum will remain viable in the field
AM fungi can spread to nearby un-inoculated plants, over distances of up to two meters within a single growing season
SPORE SLURRY METHOD FOR Cultivating native Ecomycorrhizal fungi (EMF)
REQUIREMENT: Mushrooms (that you know are ecto) must be gathered from near known ecto-mycorrhizal fungal trees.
Spores or macerated fruiting bodies of some ectomycorrhizal mushrooms, puffballs, or truffles (and false truffles) provide good inoculum. Truffles (Ascomycotina) and false truffles (Basidiomycotina), from now on together referred to as truffles, are uniquely suited for this because their fruiting body tissue consists mostly of spore-bearing tissue and the fruiting bodies can be quite large. We recommend using fresh spores whenever possible, but have stored spore suspensions of various Rhizopogon species up to 3 years without a significant reduction in inoculum effectiveness (Castellano 1987).
Rinse freshly collected fruiting bodies with tap water to remove adhering soil or organic matter
Cut into pieces (1 - 3 cm) and blend with tap water at high speed for 2 to 3 minutes, until all pieces are thoroughly blended. Note: The final consistency is similar to thick chocolate milk.
It is unnecessary to purify spore suspensions. Li and Castellano (1987) and Li (1987) have found beneficial microorganisms within and on the surface of mature fruiting bodies of various ectomycorrhizal fungi; these organisms should be encouraged, not excluded.
Spore concentrations within the resulting suspension are determined with a hemacytometer (blood cell counter).
Store slurry in the dark in the refrigerator until used (up to 3 years) at up to 5 °C or 41 °F).
To then inoculate plants:
In a nursery or for nurse plants, apply spore slurry 6 to 12 weeks after sowing, either with a standard watering can or through the existing irrigation system. Most truffle spores are less than 50 μm in diameter and will pass freely through most filters and nozzle tips.
The desired amount of spores is mixed into a watering can containing sufficient water to cover a certain number or area of seedlings (Styroblocks® or racks of plastic tubes).
Applying spores twice, 2 to 3 weeks apart, works best to assure even distribution (fig. 5.2.53), especially when using the irrigation system instead of watering cans. Alternatively, spores can be applied to the seed before sowing. Although we have not tried this method, it may prove more effective than the watering can method in inoculating each seedling. Seed treatment would also allow finer control in matching ecotypes of fungi to specific seed sources.
Species-specific AMF method
Arbuscular mycorrhizal (AM) cultures contain only living propagules of single or multiple mycorrhizal fungal species). Cultures of a single species usually must be started from carefully selected spores. These spores can originate from a field soil if they are healthy. Step by step process for Establishment of Monospecific Cultures from INVAM
RECORDING: Cultivating mycorrhizal Fungi Workshop with Danielle Stevenson (THREE Parts)
PHOTOS: Cultivating mycorrhizal Fungi Workshop with Danielle Stevenson
Resources
The Container Tree Nursery Manual, Volume 5. Agric. Handbk. 674. Castellano, M.A.; Molina, R. 1989. Mycorrhizae. In: Landis, T.D.; Tinus, R.W.; McDonald, S.E.; Barnett, J.P. Washington, DC: U.S. Department of Agriculture, Forest Service: 101-167.
Use of Mycorrhizae in Restoration of Hawaiian Habitats. Gemma, J.L., R.E. Koske. Hawaii Conservation Alliance.
The Use of Mycorrhizae in Native Plant Production. Lorraine Brooks, Deborah Brown, Sierra Smith, and Samantha Sprenger. June, 2006: University of Washington.
A Practical Guide to Inoculation with Arbuscular Mycorrhizal Fungi in Ecological Restoration. Liz Koziol, Peggy A. Schultz, James D. Bever. University of Kansas, Lawrence Geoffrey House, Jonathan Bauer Indiana University, Bloomington. Elizabeth Middleton, Missouri Department of Conservation. June 2017.
Trap Culture Methods. International Culture Collection of (Vesicular) Arbuscular Mycorrhizal Collection at West Virginia University.
On Farm Arbuscular Mycorrhizal Inoculum Production. Rodale Institute.
