How to sterilize mushroom substrate without a pressure cooker

Sterilization of the substrate is one of the critical steps when it comes to mushroom cultivation. Without a good sterilized substrate, the level of contamination within the substrate is too high for the mycelium to grow. This, therefore, leads to low yield or even worse to a total loss.

While some mushroom farmers using a pressure cooker or autoclave, some don’t or even have one, especially in developed countries. This raises the question, “How to sterilize mushroom substrate without a pressure cooker?

If you want to sterilize your substrate, but you don’t have a pressure cooker at hand, then there are several methods from which you can choose to sterilize your substrate properly. I will describe each one of them in the next sections.

  • Composting
  • Chemical
  • Coldwater pasteurization
  • Hot water immersion (Scalding)
  • Pasteurization
  • Tyndallization

How to use COMPOSTING to sterilize your Substrate

Composting is typically used for growing button mushrooms (Agaricus). But it also found its way into growing oyster mushrooms (Pleurotus spp.).

Composting is a two-step process:

  • Phase 1, which is typically done outside (Figure 1), is a biological and chemical process and the first step to decompose the mixed raw materials. During this time this phase, the substrate itself heats up to 80°C.
Figure 1: View over piles of compost

Figure 1: View over piles of compost[1].

  • Phase 2, which is done inside (Figure 2 and Figure 3), is a biological process to finalize the decomposing. While in phase 1, the temperature is uncontrolled, in phase 2, the temperature is strictly controlled for a certain amount of time. In the first part of this phase – the pasteurization phase – the temperature is set to 56-60°C for 8 hours. After that, the temperature is dropped to 45°C for up to 7 days – the so-called conditioning period. The conditioning period ends after the volatile ammonium has been cleared form the process air.
Figure 2: Sketch of a tunnel

Figure 2: Sketch of a tunnel[2]

Figure 3: View into a tunnel

Figure 3: View into a tunnel[3].

COMPOSTING PROCESS

How does the process look like[4]?

Step 1 – Substrate Preparation

Pile your substrate up and create a heap. 

Step 2 – Moisturizing 

Water your substrate and mix thoroughly. The moisture content should be around 75%.

Step 3 – Composting

During the next 7 days, water and mix regularly your substrate. This prevents from creating hot spots inside the pile. The pile will heat up during the composting process to 60 to 70°C.

Step 4 – Filling the tunnels

After 7 days, fill the substrate in a tunnel. 

Step 5 – Pasteurization

Close the tunnel and keep the temperature at 65°C for 18 hours. For this process, you need aerated steam inside the tunnels.

Step 6 – Conditioning 

Reduce the heat to about 48°C and keep the temperature for 48 hours.

Step 7 – Cooldown

Stop the heating process and let the substrate cool down to 25°C.

Step 8 – Inoculation

Inoculate with a spawn rate of about 5%. 

Step 9 – Bagging

Fill your bags with the inoculated substrate and put them into your grow chamber.

RESULTS with Composting

Figure 4 shows several tests with the above-described process.

Figure 4: Mushroom yield of the investigated production series.

Figure 4: Mushroom yield of the investigated production series. The yield was determined as a percentage of wet weights of kilogram fresh mushrooms per 100-kilogram substrate block. Bars indicate standard deviation for individual production series, based on yield values of different mushroom production houses. The thick lines show the mean ± standard deviation range for the entire dataset[5],[6].

To put these findings into context, we come back to the definition of biological efficiency (BE). The BE is defined as 1 kg of fresh mushrooms for 1 kg dry substrate or 1 kg of fresh mushroom for 4 kg wet substrate (75% water content). As the author watered to a moisture content of 75%, we will use the second definition.

Example: 7-118, ~ 15% yield

1 kg of fresh mushrooms per 4 kg of substrate equals BE 100%

15 kg of fresh mushrooms per 100 kg of substrate equals BE 60%

If we repeat this calculation for all other values, we were able to better compare the findings of the author with other papers (Figure 5). The mean BE is around 95%, with a mean ± standard variation of 75% for the lower band and 114% for the upper band. As we now see, 5 out of 11 production series or 45% are below 100% BE.

Figure 5: Estimated biological efficiency (BE)

Figure 5: Estimated biological efficiency (BE) based on figure 20[7]

The use of CHEMICAL sterilization

Chemical sterilization is typically used because they are inexpensive. During my research, I found many scientific studies in which different chemicals are used to sterilize the substrate. Table 1 gives an overview of these chemicals and their classification accordingly to their risks.

A big disclaimer: The shown information represents the view, and the opinion of mine and are just for education and information purpose only and should not be construed as legal advice or as an offer to perform legal services. The provided information contains general information and may not reflect current legal developments or information. I do not make any representation or warranties concerning the accuracy, applicability, fitness, or completeness of the information. The information is not intended to substitute for professional advice. Please inform yourself by reading the instruction manual, material safety data sheet carefully, and ask your local specialist before using any of these mentioned chemicals. I, therefore, disclaim any and all liability to any party for any direct, indirect, implied, punitive, special, incidental or other consequential damages arising directly or indirectly from this information, which is provided as is, and without warranties.

Table 1: Overview of chemicals used in science papers for the sterilization of mushroom substrate

Table 1: Overview of chemicals used in science papers for the sterilization of mushroom substrate[8]

With that said, I do not recommend any of them, because the risks of using them are for me too high. They are also poison, toxic, or hazardous to handle.

How to use COLD WATER LIME PASTEURIZATION to sterilize your Substrate

This one is somehow an exception. Attention: It is corrosive and harmful, which means you should carefully read the material safety data sheet, contact an expert and going back to what I wrote in the disclaimer section, before using it, but it has low toxicity and is widely used in the food industry (E526)[9].

The lime you want to use should be low in magnesium (< 2%) and high in calcium. Therefore, not all lime is created equally. If your lime contains more than 2%, the mycelium growth will be inhibited[10].

If you are adding lime to water, it increases the pH level. As many contaminations prefer an acidity regime, they will get therefore killed off. 

How much do you need? Add 355 g hydrated lime to 200 liters of water.

COLD WATER LIME PROCESS

How does the process look like?

Step 1 – Substrate preparation 

Chop your substrate (typically straw) into 1-6 cm long pieces. Chopping the substrate helps the breakdown and leads to higher biological efficiency (Figure 6).

Figure 6: Unshredded straw vs Shredded straw biological efficiency comparison

Figure 6: Unshredded straw vs Shredded straw biological efficiency comparison[11].

Step 2 – Prepare the water

Add about 7 grams of hydrated lime for every 4 liters of water.

Step 3 – Add your substrate and soak

After adding your substrate mix thoroughly.

Soak for 12-24 hours.

Step 4 – Drain

Remove the substrate from the water by placing the substrate onto a metal rack and let it drain for a maximum of 2 hours. The longer you dried the substrate, the more likely it is to get contaminated again. 

Step 5 – Inoculate

Inoculate the dried substrate with roughly 17% or 1 pound of spawn per 6 pound bag[12].

Step 6 – Bagging

Fill your bags with the inoculated substrate and put them into your grow chamber.

RESULTS with COLD WATER LIME

Figure 7 compares the different treatments. As indicated, lime produced in this test, the highest biological efficiency.

Figure 7: Regular straw vs. Shredded straw treatment averages

Figure 7: Regular straw vs. Shredded straw treatment averages[13]

Again, please take care and wear gloves, safety glasses, and a mask (read the disclaimer). If you are getting it on your skin, it can cause rashes and burns. Breathing it into your lungs can cause respiratory issues, and getting it in your eyes can be especially dangerous.

How to use HOT WATER IMMERSION (SCALDING) to sterilize your Substrate

For scalding the substrate is typically immersed for 1 to 1,5 hours in hot water (Figure 8).

Figure 8: Example of hot water immersion

Figure 8: Example of hot water immersion[14]

SCALDING PROCESS

How does the process look like[15]?

Step 1 – Substrate Preparation

Chop your substrate (typically straw) into 1-6 cm long pieces. Chopping the substrate helps the breakdown.

Step 2 – Prepare the water

Heat the water up to about 80°C.

Step 3 – Add your substrate

After adding your substrate mix thoroughly.

Let the process run for 1 hour.

Step 4 – Drain

Remove the substrate from the water by placing the substrate onto a metal rack and let it drain for a maximum of 2 hours. The longer you dried the substrate, the more likely it is to get contaminated again. 

Step 5 – Inoculate

Inoculate the dried substrate with 5% on a w/w wet weight basis.

Step 6 – Bagging

Fill your bags with the inoculated substrate and put them into your grow chamber.

RESULTS with SCALDING

As the results in figure 9, indicating a higher temperature doesn’t always lead to a higher yield. The best outcome for this trail was achieved at 80°c and 1 hour

.

Figure 9: Influence of different disinfection methods on biological efficiency

Figure 9: Influence of different disinfection methods on biological efficiency[16]

How to use PASTEURIZATION to sterilize your Substrate

Pasteurization at 60-80°C, up to 5 days, and 0 Psi.

Super-Pasteurization at 80-100°C for around 15 hours and 0 Psi.

Pasteurization and especially Super-Pasteurization, which was introduced by Paul Stamets, is a widely used method when it comes to sterilizing the substrate.

PASTEURIZATION PROCESS

How does the process look like[17]?

Step 1 – Substrate Preparation

Chop your substrate (typically straw) into 1-6 cm long pieces. Chopping the substrate helps the breakdown.

Step 2 – Bagging

Fill your bags with the chopped substrate.

Step 3 – Prepare the water

Heat the water up to about 100°C. The higher the temperature, the shorter the processing time. 

Step 4 – Fill the barrel

Add your bags to a barrel with a pressure relief valve (0 Psi).

Introduce the hot steam from your water tank into the barrel (e.g., at the bottom).

Let the process run for 12 to 15 hours.

During the process, make sure you have enough water in your tank. 

Step 5 – Drain

Remove the bags and let them cool down for at least 24 hours. The time depends on the density of your substrate. The temperature inside the bags should be below 28°C. Otherwise, the heat will kill the mycelium.

Step 6 – Inoculate

Inoculate the dried substrate with 5% on a w/w dry weight basis[18] and put them into your grow chamber.

RESULTS with PASTEURIZATION

In figure 10, several different sterilization methods were compared. As you can see, the pasteurization method comes in second.

Figure 10: Influence of the sterilization methods on the biological efficiency for different substrates

Figure 10: Influence of the sterilization methods on the biological efficiency for different substrates[19]

Tyndallization – The forgotten sterilization method

Sterilization method developed by John Tyndall (1820-1873). 

With this method, the product is treated in cycles. Each cycle contains the following steps.

  1. 30 min, 100°C
  2. 12 hours, 37°C
  3. 30 min, 100°C
  4. Repeat Step 2 and 3 until 72 hours are reached

During the germination phase (step 2), the bacteria start growing. In the sterilization phase (step 1), these bacteria will then get killed.

During these three days, there will be a total of 5 to 6 cycles.

https://www.umsl.edu/microbes/files/pdfs/tyndallization.pdf

Alternative process

  1. Prepare your bags/bottles/jars
  2. 60 min, 100°C
  3. 24 hours, RT
  4. Repeat steps 2 and 3 3 times. 

SUMMARY

As you could see, you don’t need a pressure cooker to sterilize your substrate. If you don’t have one, don’t worry, the results are quite promising, while at the same time, the processes are not that complicated.

But be aware that these results are only for demonstration purposes. Your results may vary depending on the substrate you are using in comparison to the substrate used in the tests and many other factors.

With that said, you should still use them as a benchmark and see if you are getting better results or not.

The links to the used articles and books can be found in the literature section.

Now I want to hear from you:

Which sterilization method from this article are you most excited to try?
And which method are you using today?

Let me know by leaving a quick comment.

RECOMMENDED READINGS

How your Sterilization Method can Impact Your Mushroom Yield

This article goes even deeper into the aspect of sterilization. Some parts are overlapping with this article but it describes each sterilization method in more detail. In addition, I will show way more results of the different sterilization processes on the mushroom yield.

How Your Substrate can Influence Your Mushroom Yield

Starting with the basics, I then will talk especially about the composition of your substrate and how they correlate with the growth rate and the yield. I will address the influence of the particle size and how different types of substrate will affect your mushroom yield. And finally, I will go over the impact of supplementation on the yield.

How your Inoculation Method can Impact Your Mushroom Yield

While inoculation seams straight forward it isn’t. While going over several aspects of inoculation, I then will introduce a six-step process to reduce contamination. At the same time, this six-step process improves the process of inoculation itself.

LITERATURE

Stamets 1983 The Mushroom Cultivator

https://amzn.to/33RtruM

Holliday 2012 https://www.researchgate.net/publication/224947271_Simplified_and_Lower_Cost_Methods_for_Culinary-Medicinal_Mushrooms_Cultivation/link/5b29251d4585150c63dcf0d9/download

Atila 2016

https://www.researchgate.net/publication/309596170_Effect_of_Different_substrate_Disinfection_Methods_on_the_Production_of_Pleurotus_ostreatus

Oseni 2012

https://www.researchgate.net/publication/279189762_Effect_of_Substrate_Pre-treatment_Methods_on_Oyster_Mushroom_Pleurotus_ostreatus_Production

Vajna 2010

https://www.researchgate.net/publication/40454271_Microbial_community_structure_changes_during_oyster_mushroom_substrate_preparation

Alexander 1994 The Best Growing Edge

https://amzn.to/33RNxE3

Kashangura 2008 http://citeseerx.ist.psu.edu/viewdoc/download?doi=10.1.1.1021.4118&rep=rep1&type=pdf

Jones 1964

https://nph.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1469-8137.1965.tb05378.x

Kumar 2018

https://scialert.net/fulltextmobile/?doi=ppj.2018.19.24

Shahid 2006

http://www.fspublishers.org/published_papers/62199_..pdf


[1] Source

[2] Paul Stamets (1983)

[3] Source

[4] Vajna 2010

[5] Baláz Vajan (2010)

[6] The first number indicates the year of production, while the second number indicates the production series

[7] Own calculation

[8] Own table based on different sources

[9] Source

[10] Alexander 1994

[11] Holliday 2012

[12] Holliday 2012

[13] Holliday 2012

[14] Source

[15] Atila 2016

[16] Own figure based on Atila 2016

[17] Stamets 1983

[18] Oseni 2012

[19] Own figure based on Oseni (2012)

4 thoughts on “How to sterilize mushroom substrate without a pressure cooker

  1. Hi Rouven, I am interested in the steam pasteurization method for button mushroom production. However, I would like to know- can one substitute steam with hot air? I have engaged a steam boiler maker (not in mushroom business) to make my pasteurization unit and he suggested the use of elements for hot air blowing into unit as an easier alternative to steam boiler, is there a difference if we use hot air from steam or does it have to be steam for button mushroom production? Why steam?

    1. Hi Mwaka, you can use hot air for sterilization, but if it is a good strategy for sterilization of substrate, that is a totally different story. If only hot air is used, the temperature is usually higher (e.g., 170°C), and takes longer, which means your cost of running the process is getting higher as well. The problem I see with using only hot air for sterilization of your substrate is that you will lose moisture during the process. As your substrate should contain between 60-70% of moisture (depending on the mushroom species), this process is counterproductive. There is maybe a workaround for this problem. You test how much moisture content you will lose during the sterilization process, and then you will increase the moisture content before the sterilization process. Therefore, the moisture content after the sterilization is in the mentioned range. But I don’t know someone who used only hot air for the sterilization of substrate, and I haven’t read an article about that.

  2. Hi, nice site for people starting on mushroom cultivation like me. Thank you. Have you heard of pasteurization/sterilization methods other than using a pressure cooker or autoclave, ie safer and cheaper methods? How about putting sawdust in an oven or using some kind of ultrasound device to kills the contamination?

    1. Hi, thanks for reaching out and your kind words. Good to hear that you found my articles helpful. As you are referring to sawdust, I guess that is what you are planning to use. The problem with sawdust is that you want to supplement it to increase the nutrient density and, therefore, get a higher yield. In doing so, you are increasing the likelihood of contamination. Here autoclaving or steam pasteurization (no pressure) makes sure to reduce the number of contaminations. If you put your sawdust or any kind of substrate in an oven, you start drying it. But to use it for cultivating mushrooms, you need a moisture content of around 60-70%). If you know how much you will lose during this process, you could adjust.

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