Pasteurization and Sterilization

Pasteurization or sterilization of the substrate

Pasteurize or sterilize your substrate?

  • Pasteurization

    This method involves heating the substrate to moderate temperatures (generally between 60°C and 80°C) for a set period. The goal is to reduce populations of undesirable microorganisms without completely eliminating them, while preserving some beneficial bacteria that can help inhibit the growth of other contaminants. Pasteurization is often used in mushroom cultivation like oyster mushrooms, as 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 applying heat at much higher temperatures (above 121°C) under pressure (autoclave) or by chemical methods. Sterilization is essential for cultivating 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?

Question and answer

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

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

Moreover, by working cleanly, you avoid introducing new contaminants, which is essential for more sensitive mushroom varieties.

Cleanliness and pasteurization create optimal conditions for the growth of the mushrooms you cultivate, 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, objective, and application. To clarify these differences, here is a simple comparison table, followed by an explanation of the uses of each method.

Characteristics Pasteurization Sterilization
Temperature 60-80°C >121°C (under pressure)
Duration 1 to 2 hours 15-30 minutes (at high temperature)
Goal Reduce the amount of contaminants Eliminate all microorganisms
Surviving Microorganisms Yes, some beneficial None (sterile environment)
Typical Substrates Straw, wood chips, compost Mycelium on grains, sugar-rich substrates
Post-Treatment Contamination Risks Low to moderate High if the environment is not kept sterile
Applications Pleurotes sur paille, Champignons de Paris Shiitaké, Eryngii, Lion's Mane
Note: each mushroom grower develops their own techniques over time. The indications we give you here should not prevent you from conducting your own experiments to enrich your skills.

Typical Uses

  • Straw-based substrates? -> Pasteurization
    Pasteurization icon

    Straw is a common substrate for cultivating mushrooms like oyster mushrooms. It is well suited to pasteurization because it requires a treatment that eliminates most contaminants while preserving some beneficial microbial flora.

  • Mycelium on grains? -> Sterilization
    Sterilization icon

    Grains, used to cultivate mycelium, are nutrient-rich, making them very prone to contamination if unwanted microorganisms are present. Complete sterilization is necessary to eliminate any risk of contamination.

  • Compost substrates? -> Pasteurization
    Pasteurization icon

    Composted substrates, often used for button mushrooms, are pasteurized to control microorganisms without eliminating beneficial thermophiles that help decompose the compost.

  • Wood chip-based substrates? -> Pasteurization
    Pasteurization icon

    Wood chips, used for mushrooms like stropharia or shiitake, benefit from pasteurization which reduces competitors while preserving a structure conducive to colonization by the mycelium.

  • Substrates rich in sugars and nitrogen? -> Sterilization
    Sterilization icon

    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

The hot water pseudo pasteurization is a simple and quick method often used by amateur mushroom growers or for small-scale cultivation. This technique involves pouring boiling 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 over the substrate, the heat kills some of the microorganisms present. However, the substrate does not remain at a sufficiently high temperature (between 60°C and 80°C) for long enough to completely eliminate the most resistant contaminants, as is possible with traditional pasteurization.
  • Limits : Due to the short exposure time, this method does not offer the same guarantee of eliminating 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 straw pellets. Compressed straw pellets are an ideal substrate for oyster mushroom cultivation. Using pseudo pasteurization, you can quickly and effectively prepare these pellets for successful cultivation.
  • Beech Pellets for Shiitakes and Lion’s Mane : Similarly, beech pellets, suitable for cultivating shiitakes and Lion’s Mane (Hericium erinaceus), can also benefit from this technique. Although beech is a denser wood than straw, boiling water can still reduce contamination risks, providing a relatively clean substrate for inoculation.
Advantages:
  • Speed : Pseudo pasteurization is quick to implement, making it a convenient method for small amounts of substrate.
  • Simplicity : It requires no specialized equipment, just a kettle and a container to hold the substrate.
Disadvantages:
  • Limited Effectiveness : This method does not eliminate all microorganisms, which can increase the risk of contamination during cultivation.
  • Not Suitable for All Substrates : For denser or nutrient-rich substrates, such as those used for grain mycelium, sterilization or traditional pasteurization would be necessary.
2) Steam Pasteurization

Heat pasteurization is one of the oldest and most commonly used methods to prepare the substrate for mushroom cultivation. This technique involves 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.

Heat Pasteurization Steps:
  • Substrate Preparation : The substrate, often straw, wood chips, 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, 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 mycelium. This cooling must be rapid to prevent contaminants from entering during this period.
Advantages of Heat Pasteurization:
  • Effectiveness : Effectively eliminates the majority of unwanted microorganisms.
  • Adaptability : Suitable for various types of substrates, especially straw and compost-based substrates.
  • Simplicity : Requires only basic equipment like a heating container or steam generator.

 

3) Lime Pasteurization (Calcium Hydroxide)

Lime pasteurization is an alternative method to heat pasteurization for preparing mushroom growing substrate. This method uses slaked lime (calcium hydroxide) to raise the substrate's pH, which inhibits the growth of many bacteria and molds, while allowing mushroom mycelium such as pleurotus to develop.

Steps of Lime Pasteurization:
    •  Use slaked lime (calcium hydroxide). Avoid magnesium-rich limes. 
    • Dissolve about 1 cup of slaked lime in about 50 liters of water. This proportion may vary slightly depending on the amount of substrate and lime concentration.
    • Immerse the substrate in the lime solution. Ensure the entire substrate is fully 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 waterlogged.
    • 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 needed compared to heat pasteurization.
    • Effectiveness : Inhibition of bacteria and molds through pH increase.
    • 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 the soil : Excessive application of lime can alter the soil pH, which can affect soil biodiversity and nutrient availability for plants.
    • Surrounding Waters: If lime is used in large quantities near bodies of water, it can increase the water's pH, which could affect aquatic organisms sensitive to pH changes.
    • 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 neutralize.
    4) Anaerobic Pasteurization

    Anaerobic pasteurization is an alternative method that requires neither heat nor chemicals. It relies on creating an oxygen-free environment, which prevents the survival of unwanted aerobic microorganisms.

    Steps of Anaerobic Pasteurization:
    • Prolonged Immersion : The substrate is immersed in water and stored in an airtight container for an extended period, usually 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 favorable for 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 the mycelium.
    • Suitable for certain mushrooms : Particularly effective for varieties like oyster mushrooms, which appreciate slightly fermented substrates.
    5) Super-Pasteurization

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

    How it works:
    • Temperature and Duration : Super-pasteurization is done with steam, at a temperature of about 90°C, without added pressure. The process duration is extended, between 12 and 24 hours, much longer than traditional pasteurization.
    • Advantages : This method is effective for 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 contamination risks while avoiding excessive alteration of the substrate composition.

    In general, mushroom growers using this technique recover 200-liter metal barrels, in which they place an electric heating element at the bottom. A false floor in the form of a grid is installed at a height of 15 to 20 cm above the heating element, on which the micro-filtered substrates in bags are positioned. A temperature controller with a probe placed in the upper part of the barrel regulates the temperature: the electric heating element turns on as long as the desired temperature (for example, 95°C) is not reached, and stops once the temperature is reached. It is essential to check that the power of the heating element matches that tolerated by the controller; otherwise, it is recommended to use an electric relay system. To improve energy efficiency, it is also advisable to insulate the outside of the barrel to minimize heat loss.

    Super-Pasteurization
    6) Pasteurization by Composting

    Pasteurization by composting is a commonly used method to prepare the substrate intended for growing button mushrooms (Agaricus bisporus). This technique exploits 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:
    • Substrate Preparation : Mix about 75% manure (ideally horse manure) with 25% straw. This mixture forms the base of the compost. Add water to moisten the mixture, ensuring it is not soaked, to ensure good microbial decomposition.
    • Composting : Form a compost pile of at least 1 cubic meter to ensure a good temperature rise. Turn the pile regularly (every 2 to 3 days) to aerate it and promote even decomposition.
    • Natural Heating : The natural microbial activity generated by composting raises the pile's temperature. You should aim for an internal temperature of 60 to 70°C. Maintain this temperature for about 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 temperature rise 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 about 1 to 2% mycelium relative to the substrate weight to ensure effective colonization.
    Advantages of Compost Pasteurization:
    • Nutritional Richness : Composting improves the nutritional quality of the substrate, making it ideal for cultivating button mushrooms.
    • Ecological : Uses natural resources (manure and straw) and relies on biological processes without requiring external heat or chemicals.
    • Effectiveness Against Pathogens : Natural temperature rise effectively eliminates most contaminants and pathogens present in the substrate.

     

    Sterilization

    Sterilization is a crucial technique for mushroom cultivation, especially when working with nutrient-rich substrates such as 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 fertile ground for the rapid proliferation of contaminants like bacteria and molds if not completely sterilized.

    Precautions:

    • Sterile Environment: A sterile substrate is extremely vulnerable to contamination as soon as it is exposed to air or handled. After sterilization, the substrate must be handled in a completely sterile environment to avoid introducing unwanted microorganisms. This includes using a laminar flow hood, sterile gloves, and strict aseptic techniques, such as regular disinfection of work surfaces and wearing face masks. Even brief exposure to non-sterile air can introduce contaminant spores, thus ruining all sterilization efforts.

    • Risk of Post-Treatment Contamination: Sterilization is only effective if the substrate remains in a sterile environment throughout the cultivation process. Once sterilized, if aseptic procedures are not strictly followed, the substrate can be quickly recontaminated. It is essential to maintain a sterile environment during the entire incubation period until the mycelium has fully colonized the substrate. This involves working under 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 sterilization times and recommended temperatures, called "barèmes". These parameters can vary depending on several factors, such as the type of container used (glass, plastic, etc.), the amount of substrate, humidity level, and the granulometry (particle size) of the substrate. It is often necessary to adjust these parameters after some trials to optimize the process according to these variables.

    • Adjustment of Scales: Autoclaves of different powers and designs may have varied heating performances. Therefore, sterilization time can be longer or shorter. It is recommended to conduct tests to determine the optimal settings based on the specifics of your autoclave and the substrates you use.

    • Air Vacuum Function: Many high-quality autoclaves offer an "air vacuum" or "preheating" function, essential for uniform sterilization. This function removes air from the sterilization chamber, either by vacuuming or by a preheating sequence that fills the chamber with water vapor. Removing air is crucial because water vapor is much more effective at transferring heat than dry air. By eliminating air from the chamber, steam can penetrate the substrate evenly, ensuring 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 heating-up and holding time may vary. More powerful autoclaves will reach the desired temperature faster and may require adjustments in sterilization time to avoid overheating the substrate. Additionally, the correct use of the air vacuum and air evacuation function not only improves sterilization uniformity but also prevents cold air pockets that could compromise the process's effectiveness.

    In summary, pasteurization and sterilization are essential but distinct tools for mushroom cultivation. Pasteurization, being gentler, creates a favorable environment while leaving some beneficial microorganisms in place, whereas sterilization provides a substrate free of 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 cultivated, and the available growing conditions.

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