Table of contents:
1. What should a spore syringe look like?
2. Visual inspection
3. Microscopic examination
4. Sterility testing
5. What about a mass that’s floating in my syringe?
6. Prevention techniques for avoiding contamination when using a spore syringe
7. Suggestions on what to do if contamination is detected
What should a spore syringe look like?
A helpful starting place when assessing whether a spore syringe is contaminated is to know how your spore syringe should look. A healthy spore syringe typically appears clear or translucent, containing suspended spores without any debris, or discoloration. The solution should have a consistent appearance throughout and may exhibit a slight coloration depending on the spore species. The spores should be evenly dispersed when gently agitated by shaking. Clumps of spores can form showing very visible chunks of spores and is sometimes unavoidable. It is also possible that no spores are visible with the naked eye as the individual spores are invisible unless viewed under a microscope. No need to panic if you can’t see anything! It’s important to note that the exact appearance can vary based on the spore species and the preparation method. We hom*ogenise our spore syringes which helps break up the spores and can sometimes result in a translucent looking syringe. A clean and uncontaminated spore syringe should generally maintain transparency or slight coloration without any visible signs of contamination or irregularities. In this guide we will look at different ways to assess whether your spore syringe is contaminated as well as how to prevent contamination when using your spore syringe.
Visual inspection
Visually inspecting your spore syringe is great as a first port of call for picking up obvious contaminants. You will be able to see obvious contamination but visual inspection will not always be conclusive. You should check for the following:
1. Examine the Solution: Hold the syringe up against a well-lit background (sunlight is best) and observe the solution for any unusual changes in color, which might appear as cloudiness, discoloration, or the presence of floating blobs.
2. Check for Sediment: Gently swirl or shake the syringe. If the spores are visible check for an even distribution of spores. Look for any settling or sediment at the bottom, as this might indicate contamination or impurities.
3. Note Unusual Features: Take note of any unexpected or irregular characteristics in the spore solution, including changes in transparency or consistency. Any unusual blobs especially if coloured is a clear indication of contamination.
Microscopic examination
Microscopic examination will allow you to see much more about what is going on in your spore solution. Using a microscope to assess mushroom spores for contamination involves several steps:
1. Preparation: Place a small drop of the spore solution on a clean glass slide. Cover it with a coverslip to prevent drying and maintain clarity.
2. Microscope Setup: Place the prepared slide on the microscope stage and adjust the magnification to view the spores clearly. Start with a lower magnification (e.g., 100x) to locate the spores. Spores are usually best viewed at 400x.
3. Observation: Focus the microscope to view the spores. Healthy spores should appear uniform in shape, size, and color. Look for any irregularities, such as unusual shapes, sizes, colors, or the presence of foreign spores or debris.
4. Identify Contaminants: Examine the entire field of view systematically. Look for signs of contamination, which may include bacterial or mold growth, irregular spore shapes or sizes, clumps, or debris that shouldn’t be present in healthy spores.
By following these steps and using proper microscopy techniques, you can effectively assess mushroom spores for potential contamination, ensuring the quality and purity of the spore sample for cultivation or research purposes.
Sterility testing
The truth is very few spore syringes will be 100% free from all contaminants. When dealing with biological products a small amount of contamination will often find it’s way in to your syringe. This is not usually a problem as it does not interfere with most mycology projects. one exception is agar.
Spore syringes aren’t ideal for agar use. Occasionally, they may reveal bacterial presence on agar, but when applied directly to grain, no bacterial signs are apparent. On agar, the mushroom mycelium struggles against trace bacteria amounts, unable to compete effectively. However, when directly introduced to grain, the mycelium easily outperforms trace bacteria and thrives in a healthy environment. This insight comes from Paul Stamets, a prominent mycologist. Due to this, we advise against using spore syringes on agar, unless gentamicin sulfate antibiotic powder is employed. In our testing, we utilize gentamicin in agar to ensure no bacterial presence. Without gentamicin, your agar batches might show bacterial growth. Gentamicin can be challenging to procure but is available at ShroomSupply.com. For strain isolation or creating liquid cultures on agar, we recommend using spore prints. Additionally, excessive spore solution on agar can lead to bacterial growth; only a few drops are necessary.
What about a mass that’s floating in my syringe?
Often times when you have a jelly like mass surrounding a clump of spores floating in your syringe, but the liquid surrounding it is still clear, the syringe is often uncontaminated. What can happen is the spores metabolise their outer layer and germinate. This is essentially the start of a liquid culture forming. Spore syringes are made with no nutrient present (unlike liquid cultures) to avoid this from happening. There is no real way of controlling this when making spore syringes and the longer the spore syringe is left before use the more likely germination. The best advice is to try and use your spore syringe as quickly as possible after purchasing/ making it if you want to avoid this.
Prevention techniques for avoiding contamination when using a spore syringe
Aside from receiving a spore syringe that is contaminated the other consideration is avoding introducing contaminants when using said spore syringe. Here are some prevention techniques to employ when using a spore syringe:
1. Sterile Environment: Work in a clean, controlled environment using a laminar flow hood or a still air box to minimize airborne contaminants.
2. Proper Sterilization: Sterilize all equipment, including syringes, needles, containers, and work surfaces using methods like autoclaving and flame sterilization.
3. Use of Personal Protective Equipment: Wear gloves, masks, and other protective gear to prevent personal contamination of the spore solution. You can use fresh sterile gloves every time (best option) or clean your gloves with 70% iso alcohol solution.
4. Aseptic Technique: Maintain strict aseptic technique throughout the process, avoiding unnecessary exposure of the syringe to non-sterile surfaces or environments.
Suggestions on what to do if contamination is detected
The short answer is bin it!
If contamination is detected in a spore syringe, here are some suggested steps to manage the situation:
1. Isolate and Discard: Immediately isolate the contaminated syringe to prevent further spread. Safely discard the contaminated spore syringe and its contents.
2. Identify Source: Determine the source of contamination, if possible. If you made the spore syringe review your sterilization procedures and environment to identify potential points of failure. In the event you purchased the spore syringe you should email the vendor.
3. Start Anew: Obtain a fresh batch of spores from a reputable source.
4. Consult Experts: If contamination persists or if unsure about handling the situation, seek advice from experienced mycologists or forums dedicated to mycological practices. The shroomery is a fantastic resource and the folks are very kind with sharing there knowlege.
Addressing contamination promptly, revising procedures, and maintaining strict sterile practices is imperitive to minimizing the risk of recurrent issues and ensure successful mycological endeavors.
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