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ShroomDAS
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Mycelium or mold - new to shrooms and trying Agar
#27453540 - 09/02/21 12:02 PM (2 years, 8 months ago) |
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I’ve been lurking for a while and finally tried to do some science, so I got hold on a MSS syringe for A+ and made some MEA agar with activated charcoal:
- Water: 300ml
- Agar agar: 6g
- Malt: 6g
- Yeast: 0.6hg
- Activated Charcoal: 1.2g
They sent the syringe and a jar of inoculated grains and pasteurized substrate, time passed and everything seemed going well when I opened them to transferred them to substrate I did the sniff test an it was sweet, then when breaking it up I noticed some green spots so… I throw it away.. well just dig it in a lemon tree flowerpot but before that I took a sample of mycelium and put it on Agar.
It started growing and did some transfers inside the same plate just to practice and then to a new agar plate, but I’m somewhat worried that is not mycelium and instead I’m breeding a happy mold.
Original Plate with the Inter-transfer:

What I think might be some mycelium:

Additional transfers as practice:

Then on August 27, 2021 (6 days ago) I venture with some droplets from the syringe into the agar using a SAB as a way to prevent outside contamination and I would greatly appreciate help to identify if is something good or bad, should I keep it or kill it with fire?
Mold or mycelium?


Thanks in advance! and sorry for the English, it is not my first language.
Edited by ShroomDAS (09/02/21 05:51 PM)
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gone-pear-shaped
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: ShroomDAS]
#27457021 - 09/05/21 11:31 AM (2 years, 8 months ago) |
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This isn't advanced mycology. That may contribute to you not getting answers sooner.
The last plates are bacteria since they basically look wet. The ones that you think might be mycelia indeed might be mycelia. It's very hard to be totally sure. But since the mushroom spores outnumber mold spores a million fold in a spore syringe, mushroom mycelia is what we should expect, unless it changes color or grows really fast. As it grows, it may become more clear, because mushroom mycelia have some growth patterns that contaminant molds never have.
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ShroomDAS
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: gone-pear-shaped]
#27457166 - 09/05/21 01:35 PM (2 years, 8 months ago) |
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Thanks for the replied and clarification I think I should had post this in maybe Getting Started or Mushroom Cultivation.
The first picture which I though mycelia grew, has become like heavy, pressing down into the agar and seems dead.
From the 2 pictures it indeed grew mold and what looked some mycelia which I transferred to another Agar and now I'm hoping for the best.
So, if it is wet, there is really high probability that it's contam.
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gone-pear-shaped
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: ShroomDAS]
#27457177 - 09/05/21 01:46 PM (2 years, 8 months ago) |
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It's just that mycelia is fibers, sometimes looking like fuzz (cotton), and in the lower pictures I don't see either. But it's the most interesting looking bacteria I've seen. Looks like a picture of a coronavirus.
If you think some mycelia is dead or unhealthy, you can measure it or trace its outline on the top or bottom of the dish. If it's noticeably bigger in two days, it's fine.
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Smartattack
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: gone-pear-shaped]
#27457541 - 09/05/21 06:55 PM (2 years, 8 months ago) |
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Your first pics are a start. The last two are for the circular file. Nothing worth playing with there.
-------------------- * Smarts videos * Planet of the APES   I'm a fungal white supremacist.
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ShroomDAS
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: ShroomDAS]
#27458147 - 09/06/21 11:15 AM (2 years, 8 months ago) |
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Thanks for the feedback.
Just as follow-up:
Picture of what I though was promising but now seems dead as it hasn't grow as per latest measurement, also it seems dry:

The agar plate with the two additional transfers seems thriving (be it mycelia or mold):

The one with circular contam seems like a mix:
One side:

The other side:

I took some of both sides and transferred into different plates to see.
What is the general consensus about using antibiotics like gentamicin (0.05g per 1L) for agar plates.
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gone-pear-shaped
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: ShroomDAS]
#27458165 - 09/06/21 11:30 AM (2 years, 8 months ago) |
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Btw, the first plate looks like a mold I've seen a lot. If so, it's very fast, and starts light and nearly invisible. But I can't guess why it stopped growing. (I'm not positive it's mold, though.) The second plate looks great. Arguably ready for grain. Especially the top colony.
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Smartattack
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: gone-pear-shaped]
#27458251 - 09/06/21 12:53 PM (2 years, 8 months ago) |
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I dont care for those satellites whatever they are, but i would for sure take another transfer from the top colony on the second plate. Somewhere 7:00 away from those suspicious spots on the edge of the plate.
-------------------- * Smarts videos * Planet of the APES   I'm a fungal white supremacist.
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gone-pear-shaped
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: Smartattack]
#27458277 - 09/06/21 01:11 PM (2 years, 8 months ago) |
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Oh yeah, if those white spots aren't condensation on the plastic, then it should be transferred right away.
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ShroomDAS
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: gone-pear-shaped]
#27462993 - 09/10/21 11:43 AM (2 years, 8 months ago) |
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So as follow up,
The second plate state is and as recommendation took some of it @ 7'clock:

The circular contam dish is still growing, will defintely throw it out:

The isolation from the second plate is thriving, additional isolations are recommended?:

Thanks!
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gone-pear-shaped
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: ShroomDAS]
#27463100 - 09/10/21 01:11 PM (2 years, 8 months ago) |
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Yes. If you only want one culture sample (and that's usually best for beginners), go for the rhizomorphic parts (because that's pretty much never contaminated). A plate of mixed fuzz and rhizomorphic mycelium can be very clean (if the culture just likes to grow that way), but your plate probably has a lot going on. I'd wait a few days for it to grow out then transfer a strong looking rhizomorphic section. If you're impatient, now would be fine but try to take a small cut. And don't touch the part you don't intend to transfer.
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ShroomDAS
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: gone-pear-shaped]
#27463172 - 09/10/21 02:24 PM (2 years, 8 months ago) |
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Thanks, will wait and make at least 2 transfers from it and get back.
I've read that even rhizomorphic can have sections in it, the objective will be to take from a single section.
Today I also tried some cloning, though I wiped the mushroom in alcohol before opening it, I hope I didn’t ruined it, overall I consider it a successful failure: https://www.shroomery.org/forums/showflat.php/Number/27463156/vc/1#27463156
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ShroomDAS
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: ShroomDAS]
#27466967 - 09/13/21 03:20 PM (2 years, 8 months ago) |
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I did take 3 different sections and did one internal transfer.
Before sectioning:


After sectioning:

Shoud I take smaller sections or the sizes are ok?
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Krismn
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: ShroomDAS]
#27472317 - 09/17/21 10:32 PM (2 years, 7 months ago) |
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You can look up pictures of how species of mushroom mycelium typically looks. For example, psilocybes, lion's mane and king stropharia typically have rhizomorphic myceliums. Oysters cottony and so on. You can also get familiar with how they may change color or metabolite colors you should expect, such as shiitake browns as it ages and gets ready to fruit. You should also know if you have a contaminated petri dish u can transfer healthy mycelium until u get a clean culture, but it's best to do this in an environment that won't allow for mold spores to get all over your house and projects. If u can afford to start over this isn't necessary. But consider this, when professionals clone wild mushrooms properly, perfect technique and equipment including flowhood, the chances of contamination is high. They use the technique of transferring sections of clean culture early on, to obtain a clean petri dish culture. They don't wait for mold to grow out and get spores everywhere, if healthy mycelium grows out a bit, transfer. So if ur doing petri dish work, learn to recognize early growth of mold or bacteria, you can transfer before things get messy.
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ShroomDAS
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: Krismn]
#27472766 - 09/18/21 09:00 AM (2 years, 7 months ago) |
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Thanks, appreciate the feedback and heads up about contam and mold.
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Krismn
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: ShroomDAS]
#27473707 - 09/18/21 11:21 PM (2 years, 7 months ago) |
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If you do not have a flowhood, you can use a still air box. Still air box would be best for petri dish work however if you were to inoculate a sealed jar with a culture or spore syringe I would suggest a glove box you may load everything in, spray lysol, then close. I had a argument with a guy over this, he insisted glove boxes don't work. I had to explain to him that lysol is very toxic so it kills a wide variety of contams, therefore it would allow glove boxes to work by killing any disturbed particles in the box, but because it's toxic it could harm the mycelium during petri dish work.however, you may get away with it, if you let the lysol settle. Bring in lysol wipes for the gloves, spray seal, wipe gloves clean with lysol wipes which will also disturb air helping to kill anything in air, wait a half hour or so, as lysol settles to the box it'll further disinfect the box, which you already disinfected. Then the result is a very close to sterile glove box and air to work in. But less toxic particles to harm mycelium. The box may have so much chemical residue that any mold spores that land from ur project will die on contact, and it'll prevent mold spores going all over the house. Although still a good idea to disinfect once you are finished.
Edited by Krismn (09/18/21 11:29 PM)
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Krismn
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: Krismn]
#27473732 - 09/18/21 11:43 PM (2 years, 7 months ago) |
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Just FYI, I would be very cautious of any information you receive from ashtray. Reason is he claims to want to provide good info, however he is spreading misinformation ironically. For example I had asked him to provide a white mold that looks like mushroom example and he provided a cultivated aspergillus oryzae example. Most molds have a white mycelium, this spores green so that was misinformation he was providing all because he wanted to fabricate proof. I'm only warning you because it's important to not take my advice or anyone's on here without checking their info, to see if it's indeed true. Don't assume anyone on here knows what they know, do ur research to validate the information be told to you. Simply put, many on here may have experience. But few are professionals and all are inexperienced in many attributes to mycology simply because there are so many methods, and an astounding variety of cultivatable species. We all have a lot of room to learn and must be humble to that fact if we expect to learn any further. Like in the movie the happening, to be a good scientist we must understand there a forces at work beyond our understanding. If you are arrogant about your abilities, you won't get very far.
Edited by Krismn (09/18/21 11:49 PM)
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PBJ710
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: Krismn]
#27473861 - 09/19/21 02:59 AM (2 years, 7 months ago) |
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Quote:
Krismn said: I'm only warning you because it's important to not take my advice or anyone's on here without checking their info, to see if it's indeed true.
Yes OP, please check the sources giving your info. Just because someone acts like they know wtf they are talking about doesn't mean they do.
BTW, I've never seen ashtray give bad info - but I have seen them contradict your bad info
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gone-pear-shaped
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: Krismn]
#27474070 - 09/19/21 08:23 AM (2 years, 7 months ago) |
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Quote:
Krismn said: If you do not have a flowhood, you can use a still air box. Still air box would be best for petri dish work however if you were to inoculate a sealed jar with a culture or spore syringe I would suggest a glove box you may load everything in, spray lysol, then close. I had a argument with a guy over this, he insisted glove boxes don't work. I had to explain to him that lysol is very toxic so it kills a wide variety of contams, therefore it would allow glove boxes to work by killing any disturbed particles in the box, but because it's toxic it could harm the mycelium during petri dish work.however, you may get away with it, if you let the lysol settle. Bring in lysol wipes for the gloves, spray seal, wipe gloves clean with lysol wipes which will also disturb air helping to kill anything in air, wait a half hour or so, as lysol settles to the box it'll further disinfect the box, which you already disinfected. Then the result is a very close to sterile glove box and air to work in. But less toxic particles to harm mycelium. The box may have so much chemical residue that any mold spores that land from ur project will die on contact, and it'll prevent mold spores going all over the house. Although still a good idea to disinfect once you are finished.
This sounds like overkill, but also very speculative. That is, I read a lot about sanitizers (compared to a normal sane person), but no publications ever mention air sanitizing via lysol. I assume either it doesn't work, or using it in the quantities that would work would be prohibitively expensive (or toxic).
If you have a technique that you're making up, it would be helpful to say it's a technique you made up, and what your evidence for it is. For example, when I talk about ozone, I'm eager to say that "this is an interesting idea that I would try in that situation" rather than "you can use ozone to do X". Because it's not a commonly accepted idea, nor a technique I have evidence for.
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ShroomDAS
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Re: Mycelium or mold - new to shrooms and trying Agar [Re: gone-pear-shaped]
#27474715 - 09/19/21 04:39 PM (2 years, 7 months ago) |
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Okay, will certainly double check info after this and also take a quick look to users rating as PBJ710 recommended.
I don't have a flow hood I use an unmodified SAB, which means I put the box at the edge of the table then slip my hands underneath and from a week or two I did start using Oust (something similar to Lysol), mist the inside and wait 10-15 mins then start working and have noticed a decrease in agar contams by doing so, will post a picture later.
Also I read about the ozone UV lamp, keep it on then turn off and work but the harzards seems to downplay the benefits of Oust.
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