Learn all about mushroom inoculation so you can practice the process of growing mushrooms at home or commercially

The process of mushroom inoculation involves bringing spawn into contact with a food source.

Today I am going over the process of mushroom inoculation, including what it is and how to do it. First, I will briefly discuss substrate preparation.

Substrate preparation

All organic material is broken down by fungi and bacteria. The job of a cultivator is to get the mycelium of the desired edible mushroom established before anything else can take hold. For the highest chance of success, the material or “substrate” needs to be prepared. The word substrate refers to any material that is a food source for mushroom mycelium. The way the substrate is prepared will be guided by the type of substrate and the equipment that is available.

Preparation includes ensuring moisture content is optimal and the substrate is clean of contaminants, mixing substrates if desired, and sometimes placing the substrate into a bag. Common substrates include logs, stumps, woodchips, straw, sawdust, coffee grounds, grain hulls, and other carbon-rich materials. Some species are very particular about the type of substrate used, while others are flexible.

The process of mushroom inoculation

Mushroom inoculation involves bringing the spawn into contact with the substrate to initiate its growth and development. Depending on the substrate, inoculation may entail drilling holes into a log, cutting wedges into a stump, or mixing spawn into individual bags. Outdoor methods can generally be done with little concern for introducing contaminants (mostly molds), while most of the indoor methods require the substrate to be inoculated in a sterile space to avoid contaminating the substrate. There are three factors in this step to consider:

  1. Where to inoculate
  2. Inoculation rates
  3. Spawn distribution

Where the process of inoculation happens depends on the nutrient level of the substrate. Low-nitrogen, carbon-rich materials like logs, woodchips, and straw are fine to inoculate outside in the open air. During this form of inoculation it is helpful to be clean, using a clean table and clean hands but sterility is not a concern. Sterility becomes much more critical when using high nitrogen materials like agar, grain, and supplemented sawdust. These are easier for ambient microorganisms to grow on, so require the use of a lab-like setting for inoculations. On a small scale, it is possible to create low budget equipment to mimic a lab setting, but the success rate is lower. A commercial lab would include a separate space for lab use, cleaning and maintaining that space, using HEPA (high efficiency particulate air) filters, and equipment for steam treatment. If inoculations are done in a lab, being mindful of the seven vectors of contamination and implementing proper lab technique is critical.

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Mushroom inoculation rates

Inoculation rates are a balance between economics and speed of spawn run. The cultivator is trying to get the desired edible mushroom mycelium established before anything else. The higher the inoculation rate or amount of spawn added to the substrate, the faster the mycelium will grow through the substrate. The substrate likely will not produce a larger amount of mushrooms, so a high inoculation rate means more money spent on spawn per pound of mushrooms. For a new cultivator, using a high inoculation rate can really boost the success rate. As the grower dials in the process of cultivation, they can start to lower the inoculation rate and observe if there is a difference in colonization time.

At Fungi Ally, this sort of trial was conducted on supplemented sawdust. Five pound bags of shiitake spawn were used to inoculate 20 five-pound bags of bulk substrate. These bags were colonized in about 8 weeks. Slowly the amount of bags inoculated by the five pound bag of spawn was increased. At 30 five-pound bags of bulk substrate, colonization time was also 8 weeks and contamination rates did not go up. At -40 five-pounds bags, time until colonization was still 8 weeks, and little to no contamination increase was observed. At 50 five-pound bags both colonization time and contamination increased. The sweet spot was using one five-pound bag of spawn for every 200 pounds (40 bags) of substrate. The amount of money spent on spawn was decreased by 50% per pound of mushrooms. Saving .25 cents per pound of mushrooms adds up to a lot when growing 300 pounds per week – almost $4,000 per year!

Spawn distribution and mushroom inoculation

Spawn distribution also impacts the speed of colonization. Two methods of distribution commonly used are “top spawning” and “through spawning”. Top-spawning is the process of adding the spawn on top of the substrate and letting it grow down. This process is good when inoculating full containers like mason jars that can not be shaken, or for low stakes cultivation. Through-spawning refers to shaking the substrate after inoculation to distribute the spawn throughout the substrate. This shortens the length of spawn run but adds a small amount of labor. In low tech methods like wood chip beds and straw tubes, through-spawning is achieved by layering the substrate and spawn like lasagna. Alternating between the spawn and substrate allows for faster colonization than putting all the spawn on top. In supplemented sawdust this typically means shaking the substrate after inoculation to distribute the spawn evenly.

The impact of top vs. through-spawning was inadvertently tested at Fungi Ally. One day, the crew was feeling like shaving some time off of the inoculation process. Instead of shaking after inoculating they left the spawn on top for 50% of the bags. Four shelves containing 40 bags each, were filled in the incubation room. The top two shelves were top-spawned, and the bottom two shelves were through-spawned. After 14 days the bottom two through-spawned shelves were fully colonized while the top spawned bags were closer to half colonized. By day 24, the top shelves were also fully colonized. In total it took ten extra days for the same amount of spawn to colonize a five pound bag of substrate. It saved a small amount of labor but was not worth the extra time in incubation. Not only did the top-spawned bags take up more space for a longer amount of time, but the extra time also left more opportunity for contaminants to take hold.