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OfflineBloo
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Help with bacterial contaminant
    #26930985 - 09/11/20 11:19 PM (3 years, 5 months ago)

I think it's bacterial, it grows on bacterial screening media.

Fully colonized quarts, grown on oats, are spawned into plastic tubs (dishwashered and wiped down with isopropyl, allowed to dry in a flow hood), and broken up with a sterile tool. I never touch anything with bare hands, never add just oats, only colonized grain. Sometimes there's a casing layer, sometimes not, there's no difference.

The cubensis grows just fine, but there's large blocks of oats that refuse to colonize, they turn dark brown, look kind of wet, and never fully colonize- again, this is from fully colonized quarts.

Once I grabbed a few of the suspect patches and put the grains on bacterial screening media and they grew bacteria, sure enough, I forget the odor but it wasn't remarkable.

The grains kind of colonize but they don't colonize completely, leaving large patches like this one without fungal growth. The containers never fruit, they never contract and pull away from the walls, they just sit there forever, doing nothing.

It doesn't matter if I add water or not after spawning them, they're pretty dry when the grain comes out of the quarts already.

What am I doing wrong?







Edited by Bloo (09/12/20 12:56 AM)


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OfflineLieutenant Pan
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Re: Help with bacterial contaminant [Re: Bloo]
    #26931029 - 09/12/20 12:26 AM (3 years, 5 months ago)

You see that yellow color that's bacteria contam from being too wet.  What did you mix the grains with in the tub?  Best thing you can do is bury the colonized portion in the yard and fruit.


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OfflineBloo
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Re: Help with bacterial contaminant [Re: Lieutenant Pan]
    #26931040 - 09/12/20 12:35 AM (3 years, 5 months ago)

I mixed them with nothing. Just took 100% colonized grains, straight to a tub. Sometimes I use a casing layer, sometimes not. It doesn't seem to matter.


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InvisibleGoatrider
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Re: Help with bacterial contaminant [Re: Lieutenant Pan]
    #26931043 - 09/12/20 12:36 AM (3 years, 5 months ago)

Perhaps try some rye next round.
Just change something and start over.
And please resize your pictures like that next time  :wink:




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OfflineBloo
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Re: Help with bacterial contaminant [Re: Goatrider]
    #26931058 - 09/12/20 12:55 AM (3 years, 5 months ago)

Sorry about the image, I'll try to edit my post.

I've tried rye, I can find only one place that sells it locally and it never works for me. I get great colonization with stone-formers, but they never form stones and then they dry out and nothing happens to them.


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OfflineLieutenant Pan
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Re: Help with bacterial contaminant [Re: Goatrider]
    #26931059 - 09/12/20 12:56 AM (3 years, 5 months ago)

Try pasteurizing some organic coco coir from the large gardening bags.  Basically mix 1 cup of colonized grain to 4 cups of coir.  With B+ a casing won't matter.


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OfflineBloo
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Re: Help with bacterial contaminant [Re: Lieutenant Pan]
    #26931078 - 09/12/20 01:15 AM (3 years, 5 months ago)

Pasteurized coir + grain for cubes? Or a casing layer?

I've never had luck with Pasteurized coir, only pressure cooked stuff; I invariably get stuff that looks like cobweb whenever I use Pasteurized coir. Even as a casing layer, I've had to switch to peat because coir just contaminates for me.


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OfflineLieutenant Pan
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Re: Help with bacterial contaminant [Re: Bloo]
    #26931082 - 09/12/20 01:17 AM (3 years, 5 months ago)

That's why you buy the organic coco that looks pretty out of the bag not those floor swept coir bricks on the cheap.  If you get cobweb its either your technique or some dirty coir suppliers.


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InvisibleGoatrider
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Re: Help with bacterial contaminant [Re: Lieutenant Pan]
    #26931088 - 09/12/20 01:26 AM (3 years, 5 months ago)

Not a single cubensis variety needs a casing layer.
If your tub fucks up before a first flush, and you do coir for bulk,
it`s almost always your spawn.
Coir doesn`t need a heat treatment, you also can do a cold hydration,
coir is very mold resistant.
Just keep it simple and don`t overdo it.
I suggest to try that tek:
https://www.shroomery.org/forums/showflat.php/Number/26009662


                    :cookiemonster:


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OfflineBloo
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Re: Help with bacterial contaminant [Re: Goatrider]
    #26931104 - 09/12/20 01:52 AM (3 years, 5 months ago)

I'll read over the SFF Shoebox Tek, thanks.

I don't use casing layers all the time, I get no more success with casing layers than without, it doesn't change anything for me.

But help me understand my original problem of contamination. I have more than 20 years of experience in axenic culture, I know how to keep things sterile. The jars look beautiful when I spawn them into clean boxes, and then they turn to shit.

I thought the problem was moisture, that they were too dry, so I took one that had large uncolonized patches, hydrated it, and got this.



The top and middle sections flushed with new growth, but the uncolonized patches didn't do anything at all. There's something bacterial in them and it keeps the boxes from fruiting. This is with multiple strains, including one that is a super-strong colonizer. I just can't figure out where it's coming from.


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Offlinemeowjinx
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Re: Help with bacterial contaminant [Re: Bloo]
    #26931106 - 09/12/20 01:54 AM (3 years, 5 months ago)

Quote:

Bloo said:
I mixed them with nothing. Just took 100% colonized grains, straight to a tub. Sometimes I use a casing layer, sometimes not. It doesn't seem to matter.




So you're trying to fruit on pure spawn w/o bulk substrate? Where did you see this tek working that made you want to try it? I've seen this done w/ mason jars on a small scale, but if you spawn to a tub the surface area for evaporation will be much greater and thus would probably dry out your spawn much faster

The bulk sub serves a very important purpose, it's a water reservoir for your cubes. Mushrooms are almost entirely water (by volume), so it's not surprising that you're never able to produce any fruits

As for the bacteria, I'm a bit confused. How is it that your tubs are not 100% colonized if they are only made up of 100% colonized spawn?
And what do you mean when you say that the spawn is "broken up with a sterilized tool"? What kind of tool is it? You shouldn't need a tool to spawn to a tub, just gently spread it out with your hands. A tool could mush up your spawn and that would be a much greater vector for contams than just touching the colonized surface of those grains

Honestly, to troubleshoot your problems it would help if you described in more detail the TEKs you've been using, from inoculation to colonization to fruiting


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OnlineBrian Jones
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Re: Help with bacterial contaminant [Re: Bloo]
    #26931115 - 09/12/20 02:08 AM (3 years, 5 months ago)

I'm disagreeing with a lot of what I'm reading here. You don't have to switch grains, although you might need to dry them out more before putting them in jars. And you don't have to pasteurize coir. That would be pointless.

Using clean inoculated petri dishes of agar and good SAB technique is what you need to do.


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OfflineLieutenant Pan
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Re: Help with bacterial contaminant [Re: Brian Jones]
    #26931139 - 09/12/20 02:42 AM (3 years, 5 months ago)

Brian Jones the Mac Daddy himself RR said unpasteurized coir is hard for mycelium to digest. Forgot if he meant cube or pan mycelium but the whole point of pasteurizing everything is to kill off the bad bacteria and keep the good. Coir is no exception. 

The OP is fruiting grains that had a bacterial contam to boot. It's time to go back to step 1. At least you're not Norman at Oscorp hearing you have to go back to formula.


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OfflineBloo
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Re: Help with bacterial contaminant [Re: meowjinx]
    #26931141 - 09/12/20 02:44 AM (3 years, 5 months ago)

Quote:

meowjinx said:
So you're trying to fruit on pure spawn w/o bulk substrate? Where did you see this tek working that made you want to try it? I've seen this done w/ mason jars on a small scale, but if you spawn to a tub the surface area for evaporation will be much greater and thus would probably dry out your spawn much faster

The bulk sub serves a very important purpose, it's a water reservoir for your cubes. Mushrooms are almost entirely water (by volume), so it's not surprising that you're never able to produce any fruits

As for the bacteria, I'm a bit confused. How is it that your tubs are not 100% colonized if they are only made up of 100% colonized spawn?
And what do you mean when you say that the spawn is "broken up with a sterilized tool"? What kind of tool is it? You shouldn't need a tool to spawn to a tub, just gently spread it out with your hands. A tool could mush up your spawn and that would be a much greater vector for contams than just touching the colonized surface of those grains

Honestly, to troubleshoot your problems it would help if you described in more detail the TEKs you've been using, from inoculation to colonization to fruiting




It's not that I've not gotten fruits- I've gotten fruits in tubs that didn't get contaminated. That's about 1/3 of the time.

I, too, am confused as to why there's any contamination with 100% colonized jars. I open up quart jars after they are heavily colonized, no un-colonized grains visible, and then gouge them out with a sterile stainless steel spatula. I used to mix them 1:1 with pressure cooked grain, but those often developed the uncolonized patches and I thought- let's see what happens with 100% grain. So far, it's 1/3, the same as when I added pressure cooked grains.

I went from BRF cakes to colonized jars (described in Psilocybin Magic Mushroom Grower's Guide) to bulk. No one single tek has worked all the way through, and I've tried piecmealing teks in order to find out where the problem lies.

Step 1: Isolate fast-growing, rhizomorphic strains on agar.

Step 2: Pressure cook grains, using bagged oats from local supplier. I've tried plain oats, I've tried oats + hydrogen peroxide + limestone, + gypsum, + ground oyster shell, I've tried organic rye. I've tried cooking it first and PC'ing it hot, I've tried cooking and drying it, I've tried cooking and PC'ing the next day. Times range from 55 minutes to 120 minutes, I got one run with trichoderma, otherwise they're visually clean.

Step 3: Containers are vented with either hole + Tyvek (doesn't work) or widemouth synthetic filter disks (which tend to dry out the grain).

Step 4: Inoculate with wedges from agar. Shake after colonies reach 6-7 cm diameter.

Step 5: Allow containers to fully colonize: 100% of grains visibly immersed in a sea of white mycelium.

Step 6: Open plastic shoebox that has been dishwashered, wiped down with isopropanol, and allowed to air dry in HEPA filtered air. Open jars, break up mycelium with sterile spatula. Smell jar when finished for any signs of contamination. Sometimes add more PC'd grain, sometimes not.

Step 7: Level out mycelium. Some get casing layer, most do not; coir casing layers tend to contaminate in my experience.

Step 8: Air out 2x/day (open, fan 30 seconds, mist with clean water), or not. I have noticed no difference in contamination between those that are fanned and those that are not.

The dark-grain-contamination thing also happened in jars when I used the instructions in the O.T. Oss and O.N. Oeric book mentioned above. Jars would just halt growth, and never fruit, the grain would never pull away from the walls of the jars like they would in successful containers.

I know- follow one tek. And I have. And when that didn't work, I tried another. And then another. I've tried a lot of different stuff and there's no consistent success, just a lot of this nasty contamination that I can't sort out. Whatever the hell it is, it carves out a niche in an otherwise perfect box of spawn and keeps the thing from fruiting.

The last one I did, I practically bent my spatula to get chunks of the colonized grain out of it- it's some really aggressive mycelia, the strongest and fastest grower I've ever isolated. 9 days later, I get what you see in the first image I've posted, an ugly brown intrusive wedge of I-don't-know-what, with the rest of it very nicely colonized. If it goes like the other boxes, it'll probably stop growing and do nothing for two months until I eventually dispose of it.


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OfflineLieutenant Pan
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Re: Help with bacterial contaminant [Re: Bloo]
    #26931142 - 09/12/20 02:49 AM (3 years, 5 months ago)

maybe the middle of the jar didnt colonize due to bacteria and you couldn't see it from the outside?  Either way you don't fruit grains in a tub like that you add bulk substrate and let the mycelium thrive


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OfflineBloo
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Re: Help with bacterial contaminant [Re: Lieutenant Pan]
    #26931150 - 09/12/20 03:08 AM (3 years, 5 months ago)

Quote:

Lieutenant Pan said:
maybe the middle of the jar didnt colonize due to bacteria and you couldn't see it from the outside?  Either way you don't fruit grains in a tub like that you add bulk substrate and let the mycelium thrive




One reason why I root around with the spatula. :smile: They're well-colonized by the time I pop them open.


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Offlinemeowjinx
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Re: Help with bacterial contaminant [Re: Bloo]
    #26931158 - 09/12/20 03:27 AM (3 years, 5 months ago)

Quote:

Bloo said:
Quote:

meowjinx said:
So you're trying to fruit on pure spawn w/o bulk substrate? Where did you see this tek working that made you want to try it? I've seen this done w/ mason jars on a small scale, but if you spawn to a tub the surface area for evaporation will be much greater and thus would probably dry out your spawn much faster

The bulk sub serves a very important purpose, it's a water reservoir for your cubes. Mushrooms are almost entirely water (by volume), so it's not surprising that you're never able to produce any fruits

As for the bacteria, I'm a bit confused. How is it that your tubs are not 100% colonized if they are only made up of 100% colonized spawn?
And what do you mean when you say that the spawn is "broken up with a sterilized tool"? What kind of tool is it? You shouldn't need a tool to spawn to a tub, just gently spread it out with your hands. A tool could mush up your spawn and that would be a much greater vector for contams than just touching the colonized surface of those grains

Honestly, to troubleshoot your problems it would help if you described in more detail the TEKs you've been using, from inoculation to colonization to fruiting




It's not that I've not gotten fruits- I've gotten fruits in tubs that didn't get contaminated. That's about 1/3 of the time.

I, too, am confused as to why there's any contamination with 100% colonized jars. I open up quart jars after they are heavily colonized, no un-colonized grains visible, and then gouge them out with a sterile stainless steel spatula. I used to mix them 1:1 with pressure cooked grain, but those often developed the uncolonized patches and I thought- let's see what happens with 100% grain. So far, it's 1/3, the same as when I added pressure cooked grains.

I went from BRF cakes to colonized jars (described in Psilocybin Magic Mushroom Grower's Guide) to bulk. No one single tek has worked all the way through, and I've tried piecmealing teks in order to find out where the problem lies.

Step 1: Isolate fast-growing, rhizomorphic strains on agar.

Step 2: Pressure cook grains, using bagged oats from local supplier. I've tried plain oats, I've tried oats + hydrogen peroxide + limestone, + gypsum, + ground oyster shell, I've tried organic rye. I've tried cooking it first and PC'ing it hot, I've tried cooking and drying it, I've tried cooking and PC'ing the next day. Times range from 55 minutes to 120 minutes, I got one run with trichoderma, otherwise they're visually clean.

Step 3: Containers are vented with either hole + Tyvek (doesn't work) or widemouth synthetic filter disks (which tend to dry out the grain).

Step 4: Inoculate with wedges from agar. Shake after colonies reach 6-7 cm diameter.

Step 5: Allow containers to fully colonize: 100% of grains visibly immersed in a sea of white mycelium.

Step 6: Open plastic shoebox that has been dishwashered, wiped down with isopropanol, and allowed to air dry in HEPA filtered air. Open jars, break up mycelium with sterile spatula. Smell jar when finished for any signs of contamination. Sometimes add more PC'd grain, sometimes not.

Step 7: Level out mycelium. Some get casing layer, most do not; coir casing layers tend to contaminate in my experience.

Step 8: Air out 2x/day (open, fan 30 seconds, mist with clean water), or not. I have noticed no difference in contamination between those that are fanned and those that are not.

The dark-grain-contamination thing also happened in jars when I used the instructions in the O.T. Oss and O.N. Oeric book mentioned above. Jars would just halt growth, and never fruit, the grain would never pull away from the walls of the jars like they would in successful containers.

I know- follow one tek. And I have. And when that didn't work, I tried another. And then another. I've tried a lot of different stuff and there's no consistent success, just a lot of this nasty contamination that I can't sort out. Whatever the hell it is, it carves out a niche in an otherwise perfect box of spawn and keeps the thing from fruiting.

The last one I did, I practically bent my spatula to get chunks of the colonized grain out of it- it's some really aggressive mycelia, the strongest and fastest grower I've ever isolated. 9 days later, I get what you see in the first image I've posted, an ugly brown intrusive wedge of I-don't-know-what, with the rest of it very nicely colonized. If it goes like the other boxes, it'll probably stop growing and do nothing for two months until I eventually dispose of it.





I see. I have a few recommendations for you, my friend:

1. Spawn earlier. I know this rock-hard mycelium you're talking about. I've allowed jars to colonize for too long and they're impossible to break up by shaking. I ended up having to scoop them out with my hands. If you spawn earlier, instead of waiting several days after 100% colonization, you are able to shake the grains apart and then they will slide out of the jar without you having to pierce it with a spatula

Do not pierce your grains with a spatula ever again. If you break through grains then you are exposing the inner core of it to open air when you spawn the jars. The SURFACE of the grain is 100% colonized, but the inside is still uncolonized, so when you spawn into your open air tubs EVERY single burst grain is highly susceptible to contamination. This is probably how the bacteria are getting into all of your grows

2. Do NOT spawn your 100% colonized grain jars onto more PCed grain. You cannot use PCed grain as a bulk substrate. What you are essentially doing is a G2G transfer, but to grains in open air. If you sterilize grains in a PC and then open up the jars and throw them in a tub then they are trash at that point. Sterilized grains are super susceptible to contams because they're sterilized so it's first come first serve, and bacteria can reproduce much faster than cubes mycelium

3. Use coir. I guarantee you that if you had contam issues w/ coir it was not the coir's fault, it just happened to be there when you got mold. A casing is a layer on top of your tub, so if you've only used coir as a casing then you need to start using it differently. You should be mixing coir with your spawn in, at least, equal amounts by volume. That is, a 1:1 ratio. Or you could do more coir than spawn, like a 1:2 ratio or something, but the point is that you should mix it evenly throughout the tub w/ your spawn. It doesn't just go on top, it's mixed with gently-broken-up spawn all throughout


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OfflineBloo
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Re: Help with bacterial contaminant [Re: meowjinx]
    #26934416 - 09/14/20 01:07 AM (3 years, 5 months ago)

Thanks for the reply.

Quote:

meowjinx said:
1. Spawn earlier. I know this rock-hard mycelium you're talking about. I've allowed jars to colonize for too long and they're impossible to break up by shaking. I ended up having to scoop them out with my hands. If you spawn earlier, instead of waiting several days after 100% colonization, you are able to shake the grains apart and then they will slide out of the jar without you having to pierce it with a spatula




I have done that as well. There are no differences in results. The main strain I use is a fast colonizer, it forms well-colonized mycelium, but I have used other strains and some will tumble out with just a bit of shaking. There is no difference in results.

Quote:

meowjinx said:
Do not pierce your grains with a spatula ever again. If you break through grains then you are exposing the inner core of it to open air when you spawn the jars. The SURFACE of the grain is 100% colonized, but the inside is still uncolonized, so when you spawn into your open air tubs EVERY single burst grain is highly susceptible to contamination. This is probably how the bacteria are getting into all of your grows




Again, there is no difference as to whether I use a tool or not.

Quote:

meowjinx said:
2. Do NOT spawn your 100% colonized grain jars onto more PCed grain.




I have tried it both ways; thinking it was the mixed grains that was the problem, I've switched to 100% colonized grains. Again, no difference has been noticed.

Another more recent example:

This tub was populated 6 days ago with two quart jars of well-colonized grain, and cased with something like two pint jars of PC'd peat with a bit of agricultural lime added.



A side pic for anyone who thinks this tub might be "too wet".



All this was unpacked in a unused, freshly-washed, freshly-disinfected plastic tub, in laminar flow hood, never contacted with hands. And still there are at least 2-3 of these ugly yellow-brown sites of contamination which, in my experience, will prevent the tub from doing anything.



So my original question remains: how do I find out what the hell this contaminant is? I appreciate all the advice about not doing this or trying a different tek, but it just doesn't matter. This contaminant stops colonization and fruiting 2/3rds of the time, and not a single change in technique has altered this.


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OfflineForresterM
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Re: Help with bacterial contaminant [Re: Bloo]
    #26934430 - 09/14/20 01:46 AM (3 years, 5 months ago)

Quote:

Bloo said:
So my original question remains: how do I find out what the hell this contaminant is?




I'm pretty sure it's bacteria, just like you said in the title of your post.  If you want to identify the species of bacteria, you will have to send it into a lab, we can't do that for you.


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Offlinemescalinechemist
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Re: Help with bacterial contaminant [Re: Forrester]
    #26934449 - 09/14/20 02:20 AM (3 years, 5 months ago)

Is it a common practice to mix colonized spawn with more grains in order to fruit in bulk? I have never heard of this. Everyone here spawns grain to coir, straw, manure etc. The bacteria present in pasteurized substrate helps fill niches in the uncolonized bulk fighting off contaminants until the mycelium arrives. I remember taking pressure cooked grain, exposing it to air for 5 seconds (jar with the lid off for 5 seconds, closing it and the next day it stank, badly. Sterile substrate is extremely prone to contamination because it is just vacant space (and highly nutritious) for anything that lands on it.


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