A Comprehensive Guide to Agar in Mushroom Cultivation | Mycology Start (2024)

What is Agar?

Agar, a gelatinous substance derived from red seaweed, plays a crucial role in mushroom cultivation. It's primarily used in petri dishes or jars to germinate spores and culture mycelium. The clear, nutrient-rich medium allows cultivators to observe the growth and health of the mycelium, isolate pure cultures, and identify any contamination.

Make Agar at Home

While the above agar media are commercially available, homemade alternatives are also possible for mushroom cultivation. The most common is Light Malt Extract Agar (LMEA), made from light malt extract, agar-agar, and water. It is suitable for a wide range of mushroom species.

Remember, no matter the type of agar you choose, the most important factor in mushroom cultivation is maintaining a clean, contamination-free environment. Ensure that you sterilize all your tools, substrates, and the agar itself before use.

How to Pour Agar

After pressure cooking the pre-mixed agar, allow the mixture to cool to around 115-125°F (46-52°C) before pouring. This temperature range ensures that the agar remains liquid for easy pouring but is not so hot as to compromise sterility or burn you. As you prepare to pour the agar, ensure that your workspace is sterilized. Working in a still-air box or in front of a laminar flow hood is highly recommended for maintaining a sterile environment.

Before you begin, sterilize your gloves and work surface one more time using isopropyl alcohol to minimize the risk of contamination. Once everything is set, carefully pour the cooled agar into your sterile Petri dishes or ketchup-cups. Aim to fill each container just enough to cover the bottom, taking care to pour smoothly to avoid the formation of air bubbles. If air bubbles do occur, you can remove them by gently passing a sterilized loop or needle over the surface of the agar.

After pouring, allow the agar to solidify at room temperature, which usually takes around 20 to 30 minutes. Once solidified, seal the Petri dishes or ketchup-cups with their respective lids or with plastic wrap. Make sure to label each container with the date and any other relevant information for future reference. Your agar is now ready for use or can be stored in a cool, dark place until needed.

Agar Shelf Life & Storage

Storing blank agar plates properly is crucial for maintaining their sterility and effectiveness for future use. Once the agar has solidified, the plates should be wrapped in plastic wrap or sealed in a plastic bag to prevent contamination. Store them in a refrigerator at a temperature range of 35-40°F (2-4°C), away from raw food items to minimize the risk of bacterial or fungal spores contaminating the surface. Some people prefer to store them in a dedicated lab refrigerator. Always label your stored plates with the date of preparation to keep track of their shelf life.

The longevity of blank agar plates can vary, but generally, they can last up to a month when stored under these ideal conditions. As time passes, the risk of contamination or drying increases, making the plates less effective for mycological work. Therefore, it's advisable to use them within 2-4 weeks for optimal results. Always check for signs of contamination like mold spots, discoloration, or an off-odor before using a stored agar plate for inoculation.

Ways to Use Agar (Transfers)

• Spore to Agar: Transferring spores to agar initiates their germination into mycelium.
• Agar to Agar: This involves transferring a healthy mycelium sample from one agar plate to another.
• Grain to Agar: A sample of colonized grain can be transferred to agar for isolation or testing.
• Liquid Culture to Agar: You can inoculate an agar plate with liquid culture for mycelial growth.
• Agar to Spawn: Transferring a colonized agar wedge to grain or sawdust to begin the spawn run.
• Agar to Liquid Culture: Agar samples can be used to inoculate liquid cultures.
• Agar to Culture Slant: For long-term storage, transfer a sample of mycelium on agar to a culture slant.

How to Incubate Agar

Once you've inoculated your agar plates with mushroom spores or a mycelium sample, the next critical step is incubation. Proper incubation provides the ideal environment for the mycelium to grow and colonize the agar surface. Start by placing your inoculated agar plates in a clean, sealable container like a plastic tote or a specialized incubator. Ensure that the container is clean and has been wiped down with isopropyl alcohol to minimize the risk of contamination.

The optimal temperature for incubating most types of mushroom mycelium ranges from 75-80°F (24-27°C). You can achieve this temperature using a heating mat, temperature controller, or even a warm room that consistently stays within this range. Keep in mind that too high of a temperature can encourage the growth of contaminants, while too low of a temperature can slow down mycelium growth.

Position the plates in your chosen container so that they are not touching each other, which allows for good air circulation. Stack them upside-down (agar side up) to prevent condensation from dripping onto the agar surface. Close the container securely to maintain a humid and sterile environment, but make sure there's some level of gas exchange; this can be achieved by poking small holes in the container or leaving the lid slightly ajar.

Monitor the plates regularly to check for signs of mycelial growth and potential contamination. If you spot any contaminants, it's advisable to remove the affected plate immediately to prevent the spread to other plates. The mycelium will usually take several days to a couple of weeks to fully colonize the agar, depending on the mushroom species and the incubation conditions.

Once the agar is fully colonized, you're ready to move on to the next stage of mushroom cultivation: transferring the mycelium to a substrate or directly to a fruiting chamber. Proper incubation is key to a successful mushroom cultivation journey, and mastering this stage sets the foundation for robust, healthy mycelial growth.

Types of Agar Media and Their Uses

Agar media come in various types, each formulated to serve different purposes in mushroom cultivation. Understanding the different types of agar media and their uses in mushroom cultivation can significantly increase your success rate, whether you're a hobbyist or a commercial cultivator. It's all about providing your mushrooms with the ideal conditions for growth.

1. Malt Extract Agar (MEA)

Malt Extract Agar is another widely used agar medium in mycology. It contains malt extract, which provides the necessary nutrients for the growth of the mycelium. MEA is suitable for the germination of spores and the culture and maintenance of mushroom mycelium.

2. Potato Dextrose Agar (PDA)

Potato Dextrose Agar is a common type of agar medium used in mushroom cultivation. As the name suggests, it's made from potato infusion and dextrose. PDA is an excellent medium for culturing a wide range of mushroom species and is especially useful in spore germination and maintaining mother cultures.

3. Sabouraud Dextrose Agar (SDA)

Named after the French dermatologist Raymond Sabouraud, this type of agar is primarily used in medical mycology but can also be used in mushroom cultivation. SDA is useful for the selective isolation of dermatophytes, yeasts, molds, and other acid-tolerant fungi.

4. Yeast Extract Peptone Dextrose (YPD) Agar

YPD agar is rich in nutrients, providing a conducive environment for the growth of a variety of yeasts and molds. It is often used in research and molecular genetics and can be used to culture certain mushroom species.

5. Czapek Dox Agar

Czapek Dox Agar is a type of agar medium used for the cultivation of fungi where the nutritive sources are strictly defined. It's often used in physiological studies and for determining the fungal flora in various substrates.

6. Oatmeal Agar

Oatmeal Agar is a simple and economical type of agar used for growing mycelium. It consists of common oatmeal and agar-agar. Oatmeal agar is perfect for hobbyists looking to cultivate mushrooms at home.