Pasteurization and Sterilization

Two main methods are used to prepare the substrate: pasteurization and sterilization.

Pasteurisation

This method involves heating the substrate to moderate temperatures (generally between 60°C and 80°C) for a specific period of time. The goal is to reduce populations of unwanted microorganisms without completely eliminating them, while preserving some beneficial bacteria that can help inhibit the growth of other contaminants. Pasteurization is often used in growing mushrooms like oyster mushrooms because it is less expensive and does not require complex equipment.
Sterilization

Unlike pasteurization, sterilization aims to eliminate all microorganisms present in the substrate. This is usually done by the application of heat at much higher temperatures (above 121°C) under pressure (autoclave) or by chemical methods. Sterilization is essential for growing mushrooms more sensitive to contaminants, such as shiitake mushrooms, and it ensures a completely aseptic environment.

Why should I pasteurize my substrate when it doesn’t happen like that in nature?

It's true that in nature, mushrooms grow without human intervention, but the conditions there are very different. Outdoors, fungi often have to compete with other organisms, and only the strongest survive. Furthermore, if mushrooms produce millions of spores, it is mainly because they actually have very little chance of survival. The chances of success in the wild are slim.

When growing mushrooms, the goal is to maximize the chance of success. By pasteurizing the substrate, you reduce the amount of potential contaminants, giving the mycelium an advantage to quickly colonize the substrate.

Additionally, by working cleanly, you avoid introducing new contaminants, which is essential for more sensitive varieties of mushrooms.

Cleanliness and pasteurization help create optimal conditions for the growth of the mushrooms you are growing, increasing your chances of success.

Pasteurize or sterilize?

Two main techniques are used for this purpose: pasteurization and sterilization. Although these terms may seem similar, they differ significantly in terms of process, purpose and application. To clarify these differences, here is a simple comparison table, followed by an explanation of the uses of each method.

Features	Pasteurisation	Sterilization
Temperature	60-80°C	>121°C (under pressure)
Duration	1 to 2 hours	15-30 minutes (at high temperature)
Objective	Reduce the amount of contaminants	Eliminate all microorganisms
Surviving Micro-organisms	Yes, some beneficial	None (sterile environment)
Typical Substrates	Straw, wood chips, compost	Mycelium on mycelium on grains , substrates rich in sugar
Risks of Post-Treatment Contamination	Low to moderate	High if the environment is not kept sterile
Applications	Oyster mushrooms on straw, Paris mushrooms	Shiitake, Eryngii, Lion's Mane

Note: each myciculturist develops his own techniques over time. The indications we give you here should not prevent you from carrying out your own experiments to enrich your skills.

Typical Usages

Straw-based substrates? -> Pasteurization

Straw is a common substrate for growing mushrooms like oyster mushrooms. It lends itself well to pasteurization because it requires treatment that eliminates most contaminants while retaining some beneficial microbial flora.
Mycelium on mycelium on grains ? -> Sterilization

The mycelium on grains used to grow the mycelium are rich in nutrients, making them very prone to contamination if unwanted microorganisms are present. Complete sterilization is necessary to eliminate any risk of contamination.
Compost substrates? -> Pasteurization

Composted substrates, often used for button mushrooms, are pasteurized to control microorganisms without eliminating the beneficial thermophiles that help compost decompose.
Wood chip substrates? -> Pasteurization

Wood chips, used for mushrooms such as stropharia or shiitake, benefit from pasteurization which reduces competitors while retaining a structure conducive to colonization by mycelium.
Substrates rich in sugars and nitrogen? -> Sterilization

These substrates, often used for more delicate or specific cultures, are sterilized to prevent any microbial competition, since sugars and nitrogen promote the rapid growth of contaminants.

Pasteurization Techniques for Mushroom Cultivation

1) Hot Water Pseudo Pasteurization

pseudo hot water pasteurization is a quick and simple method often used by amateur mushroom growers or for small-scale cultivation . This technique consists of pouring hot water, typically from a kettle, directly onto the substrate. Although this method is not as effective as traditional pasteurization due to the limited heat exposure time, it can still significantly reduce the microbial load of the substrate.

Principle and Limits :

Principle : When boiling water is poured onto the substrate, the heat kills some of the microorganisms present. However, the substrate does not remain at a high enough temperature (between 60°C and 80°C) for a long enough time to completely eliminate the most resistant contaminants, as is possible with traditional pasteurization.
Limitations : Due to the short exposure time, this method does not offer the same guarantee of elimination of unwanted microorganisms. Some more resistant contaminants may survive and potentially compete with the mycelium once the substrate is inoculated.

Applications and Examples :

Straw Pellets for Oyster Mushrooms : This method works well with the Straw pellets . Straw compressed into pellets is an ideal substrate for growing oyster mushrooms. Using pseudo pasteurization, you can quickly and efficiently prepare these pellets for successful cultivation.
Beech Pellets for Shiitakes and Lion's Mane : Likewise, beech pellets, suitable for cultivation shiitakes and Lion's Mane (Hericium erinaceus), can also benefit from this technique. Although beech is a denser wood than straw, boiling water can further reduce the risk of contamination, providing a relatively clean substrate for inoculation.

Benefits :

Speed : Pseudo pasteurization is quick to implement, making it a practical method for small quantities of substrate.
Simplicity : It requires no specialist equipment, just a kettle and a container to hold the substrate.

Disadvantages:

Limited Effectiveness : This method does not eliminate all microorganisms, which may increase the risk of contamination during cultivation.
Not Suitable for All Substrates : For denser or nutrient-rich substrates, such as those used for mycelium on grains based mycelia, traditional sterilization or pasteurization would be necessary.

2) Steam Pasteurization

Heat pasteurization is one of the oldest and most commonly used methods of preparing substrate for growing mushrooms. This technique consists of heating the substrate to a temperature of 60 to 80°C for a period of 1 to 2 hours. This process kills the majority of unwanted microorganisms, while leaving some beneficial microbes intact that can help inhibit more aggressive contaminants.

Steps of Heat Pasteurization:

Preparation of the Substrate : The substrate, often straw, wood shavings or sawdust, is first moistened to ensure even heat distribution during the process. It is recommended to soak your substrate in water for 12 to 24 hours to "wake up" the spores and bacteria present.
Heating : The substrate is then placed in a heat-resistant container or bag and then immersed in hot water or exposed to steam. The temperature must be maintained between 60 and 80°C for at least one hour to ensure effective pasteurization.
Cooling : After heat treatment, the substrate must be cooled to room temperature before being inoculated with the mycelium. This cooling must be rapid to prevent contaminants from being introduced during this period.

Advantages of Heat Pasteurization:

Efficacy : Effectively eliminates the majority of unwanted microorganisms.
Adaptability : Suitable for various types of substrates, especially straw and compost-based substrates.
Simplicity : Only requires basic equipment such as a heating container or a steam generator.

3) Pasteurization with Lime (Calcium Hydroxide)

Lime pasteurization is an alternative method to heat pasteurization for preparing mushroom growing media. This method uses slaked lime (chalky white lime) to increase the pH of the substrate, which inhibits the growth of many bacteria and molds, while allowing the mycelium of mushrooms like oyster mushrooms to grow.

Steps of Lime Pasteurization:

Use slaked lime (white calcareous lime). Avoid limes high in magnesium.
Mix about 1 cup of slaked lime in about 50 liters of water. This proportion may vary slightly depending on the quantity of substrate and the concentration of lime.
Immerse the substrate in the lime solution. Make sure the entire substrate is well submerged. Let the substrate soak in the lime solution for about 12 hours.
After soaking, drain the substrate well to remove excess lime solution. The substrate should be moist but not soggy.
Mix the treated substrate with 5 to 10% mushroom mycelium relative to the wet weight of the substrate.

Advantages of Lime Pasteurization:

Simplicity : Less equipment required compared to heat pasteurization.
Efficacy : Inhibition of bacteria and mold thanks to the increase in pH.
Adaptability : Can be used for various types of lignocellulosic substrates.

Lime, when used appropriately, is generally not considered toxic to the environment. However, it is important to use it wisely to avoid potential negative impacts. Here are some points to consider:

Effects on Soil: Excessive application of lime can change soil pH, which can affect soil biodiversity and nutrient availability to plants.
Surrounding waters: If lime is used in large quantities near bodies of water, it may increase the pH of the water, which could affect sensitive aquatic organisms to changes in pH.
Biodegradability: Lime is a mineral compound that does not decompose in the same way as organic substances. It remains in the environment but can gradually be neutralized.

4) Anaerobic Pasteurization

Anaerobic pasteurization is an alternative method that does not require heat or chemicals. It relies on the creation of an oxygen-free environment, which prevents the survival of unwanted aerobic microorganisms.

Steps of Anaerobic Pasteurization:

Extended Immersion : The substrate is submerged in water and stored in an airtight container for an extended period of time, typically 10 to 15 days. Deprived of oxygen, aerobic microorganisms die, while the mycelium, which can tolerate slightly anaerobic conditions, finds a favorable environment for its growth.
Fermentation : During this immersion period, the substrate can begin to ferment. This fermentation process can transform certain organic compounds into acids, which can make the substrate even more conducive to colonization by the mycelium of certain mushroom species, such as oyster mushrooms.

Advantages of Anaerobic Pasteurization:

Economical : Requires neither thermal energy nor chemicals.
Subtlety : Preserves the characteristics of the substrate while making it conducive to colonization by mycelium.
Adapted to certain mushrooms : Particularly effective for varieties such as oyster mushrooms, which appreciate lightly fermented substrates.

5) Super-Pasteurization

Super-pasteurization is an advanced technique, particularly used when the substrate is supplemented or has a high risk of contamination. It is the recommended technique with our optimal substrate For example.

How it works :

Temperature and Duration : Super-pasteurization is carried out with steam, at a temperature of around 90°C, without added pressure. The duration of the process is extended, between 12 and 24 hours, much longer than traditional pasteurization.
Advantages : This method is effective in eliminating not only unwanted microorganisms but also their spores, while preserving certain thermophilic microorganisms that can help maintain a biological balance in the substrate.
Usage : Recommended for supplemented substrates or those at high risk of contamination, super-pasteurization is a technique that helps reduce the risk of contamination while avoiding excessive alteration the composition of the substrate.

Typically, mushroom growers using this technique collect 200-liter metal barrels, in which they place an electrical resistor at the bottom. A raised floor in the form of a grid is installed at a height of 15-20 cm above the resistor, on which the substrates in bags micro-filtered are positioned. A temperature controller with a probe placed in the upper part of the barrel allows the temperature to be regulated: the electrical resistance is activated until the desired temperature (for example, 95°C) is reached, and stops once the temperature is reached. It is essential to check that the power of the resistance corresponds to that tolerated by the controller, otherwise it is recommended to use an electrical relay system. To improve energy efficiency, it is also advisable to insulate the outside of the barrel to minimize heat loss.

6) Pasteurization by Composting

Pasteurization by composting is a method commonly used to prepare the substrate for growing button mushrooms (Agaricus bisporus). This technique harnesses the heat generated by natural microbial activity during the composting process to pasteurize the substrate, thereby eliminating unwanted microorganisms and pathogens.

Steps of Pasteurization by Composting:

Preparation of the Substrate : Mix approximately 75% manure (ideally horse manure) with 25% straw. This mixture forms the basis of the compost. Add water to moisten the mixture, making sure it is not soggy, to ensure proper microbial breakdown.
Composting : Form a compost pile of at least 1 cubic meter to ensure a good rise in temperature. Turn the pile regularly (every 2 to 3 days) to aerate it and promote uniform decomposition.
Natural Heating : The natural microbial activity generated by composting raises the temperature of the pile. You should aim for an internal temperature of 60 to 70°C. Maintain this temperature for approximately 7 to 14 days to pasteurize the substrate.
Mixing : Turn the compost pile every 2 to 3 days. This helps aerate the compost, redistribute moisture and promote a uniform rise in temperature throughout the pile.
Cooling : After the pasteurization period, allow the compost to cool to room temperature before using it for seeding.
Inoculation : Once the compost has cooled, mix it with button mushroom mycelium. Use approximately 1-2% mycelium relative to the weight of the substrate to ensure effective colonization.

Advantages of Pasteurization by Composting:

Nutritional Richness : Composting improves the nutritional quality of the substrate, making it ideal for growing button mushrooms.
Ecological : Uses natural resources (manure and straw) and relies on biological processes without requiring external heat or chemicals.
Effectiveness Against Pathogens : The natural rise in temperature effectively eliminates most contaminants and pathogens present in the substrate.

The Sterilization

Sterilization is a crucial technique for mushroom cultivation, especially when working with nutrient-rich substrates such as mycelium on grains . It involves eliminating all microorganisms present in the substrate by heating it to very high temperatures (usually above 121°C) under pressure, typically using an autoclave. This method is essential because nutrient-rich substrates provide a breeding ground for the rapid proliferation of contaminants, such as bacteria and mold, if they are not completely sterilized.

Precautions:

Sterile Environment: A sterile substrate is extremely vulnerable to contamination whenever it is exposed to air or handled. After sterilization, the substrate must be handled in a completely sterile environment to avoid the introduction of unwanted microorganisms. This includes the use of a laminar flow hood, sterile gloves, and strict aseptic techniques, such as regular disinfection of work surfaces and use of face masks. Even brief exposure to non-sterile air can introduce contaminant spores, ruining the entire sterilization job.
Post-Processing Contamination Risk: Sterilization is only effective if the substrate remains in a sterile environment throughout the cultivation process. Once sterilized, if aseptic procedures are not rigorously followed, the substrate can be quickly recontaminated. It is essential to maintain a sterile environment throughout the incubation period, until the mycelium has completely colonized the substrate. This involves working in controlled conditions and minimizing interventions after inoculation.

Use of the Autoclave :

When using an autoclave, it is crucial to follow the specific guidelines provided by the manufacturer regarding recommended sterilization times and temperatures, called "tables." These parameters can vary depending on several factors, such as the type of container used (glass, plastic, etc.), the quantity of substrate, the humidity level, and the granulometry (particle size) of the substrate. It is often necessary to adjust these parameters after a few trials to optimize the process based on these variables.

Adaptation of the Scales: Autoclaves of different powers and designs can have varied heating performances. Therefore, the sterilization time may be longer or shorter. It is recommended to experiment to determine the optimal settings based on the specifics of your autoclave and the substrates you are using.
Air Vacuum Function: Many high-quality autoclaves offer an “air vacuum” or “preheat” function, essential for uniform sterilization. This function allows air to be removed from the sterilization chamber, either by evacuation or by a preheating sequence that fills the chamber with water vapor. Removing air is crucial because water vapor is much more efficient at transferring heat than dry air. By removing air from the chamber, steam can penetrate the substrate evenly, ensuring that all parts of the substrate reach the required sterilization temperature for a sufficient duration.
Experimentation and Adjustment: Depending on the power of your autoclave, the temperature rise and maintenance time may vary. More powerful autoclaves will reach the desired temperature more quickly and may require adjustments in sterilization time to avoid overheating the substrate. In addition, the correct use of the vacuum and air evacuation function not only improves sterilization uniformity, but also prevents cold air pockets that could compromise sterilization. efficiency of the process.

In summary, pasteurization and sterilization are essential but distinct tools for growing mushrooms. Pasteurization, which is gentler, creates a favorable environment while leaving certain beneficial micro-organisms in place, while sterilization offers a substrate free from any microbial competition, then requiring extreme precautions to avoid recontamination.

The choice between these methods will depend on the type of substrate used, the mushrooms grown, and the growing conditions available.