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Offlineverytastycheese
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What happened?
    #27078974 - 12/08/20 05:51 PM (1 month, 18 days ago)

Alright so admittedly it was my first crack at Agar but far from my first grow. Oddly all my success so far has been with multispore G2G but this time I thought I had a healthy culture. It grew like mycelium, It smelled like mushrooms, it colonized quickly... and it turned to trich. 16 bins and not a single pin. Is it possible this was never actually mycelium in the first place? Did I get that rare non-pinning culture?



A Couple magnified images:




I admit this stuff looks a little fuzzier than what I'm growing now. While I don't have a pic of the original agar plate I do have pics of some of its babies which all look more like cotton balls than mycelium.




Do other molds have that same fingery growth and mushroom smell just to throw me off? Did I just have some bacteria I couldn't see or smell in any way?

All the wisdom here says go back to the spawn, so that's what I'm doing. I took 4 transfers off my original spore plate and abandoned the failed series and moved to one of its siblings. So far I like how it looks... but I'm no expert.

This is the plate I transferred off and let continue to grow. Disturbance is from the transfers.

Some zooms of what I'm fairly certain is real mycelium this time





How can I be sure this is clean and real mycelium? How can I know it will pin? How does it look to you?

Thanks in advance for any advice!


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Offlinekanemush
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Re: What happened? [Re: verytastycheese]
    #27078988 - 12/08/20 06:03 PM (1 month, 18 days ago)

jars look a little wet to me so possible contimnation there. you cant really tell with agar if it will pin. unless you decide to let it try and pin on the agar then clone it. let some others chime in and help you out.


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OfflineJustweed
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Re: What happened? [Re: verytastycheese]
    #27079000 - 12/08/20 06:18 PM (1 month, 18 days ago)

That definitely looks like healthy mycelium on your plates. You can't ever 'guarantee' a fruiting strain, but rhizomorphic growth typically fruits a lot better than the fluffy stuff. Make sure you've thoroughly cleaned your grow area after disposing the trich growth, and be extra clean during spawning.


--------------------


Keep 'em high and tight guys....


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OfflineCrackatoaS
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Re: What happened? [Re: Justweed]
    #27079130 - 12/08/20 07:38 PM (1 month, 17 days ago)

I would def take some more transfers from the edges, as little as possible. There's something riding along. I've been there before. Trich seems to have a way of hiding during jar colonization, I always do 2 shakes to make sure its got good recovery. I always like to do an extra transfer or two for safety sake unless its just beautiful. Could also have gotten something during the G2G process.


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Offlineverytastycheese
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Re: What happened? [Re: Crackatoa]
    #27079519 - 12/08/20 11:07 PM (1 month, 17 days ago)

I'm wondering about the fuzziness riding on top of my cultures. Otherwise they seem to look alright but is this just tomentose growth or a contam?



This little guy was transferred the size of a pinhead, hopefully that helped get an isolation but he just looks like a ball of fuzz so far. Is this how they usually begin?



Zoomed in



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Offlineverytastycheese
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Re: What happened? [Re: Crackatoa]
    #27080279 - 12/09/20 12:57 PM (1 month, 17 days ago)

Quote:

Crackatoa said:
I would def take some more transfers from the edges, as little as possible. There's something riding along. I've been there before. Trich seems to have a way of hiding during jar colonization, I always do 2 shakes to make sure its got good recovery. I always like to do an extra transfer or two for safety sake unless its just beautiful. Could also have gotten something during the G2G process.




Are you saying you can see something riding along (The fuzziness) on my latest plates, or just that a ridealong must have been the reason for my past failures? Thanks!


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OfflineQuirkmeister92
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Re: What happened? [Re: verytastycheese]
    #27080286 - 12/09/20 01:02 PM (1 month, 17 days ago)

Well I wouldn't use the growth you have without transfers so it's possible there is something riding along. Like crack said  take more transfers. You want uniform growth. Additionally your sterile procedures must be on par or better.


--------------------
Come over to the oat side...


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OfflineThe Mycologist
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Re: What happened? [Re: Quirkmeister92]
    #27080298 - 12/09/20 01:08 PM (1 month, 17 days ago)

The density of the plates doesnt look 100% healthy.

That mycelium looks stressed. Its too dense.


--------------------
"That you are here—that life exists, and identity;
That the powerful play goes on, and you will contribute a verse.”
― Walt Whitman, Leaves of Grass
:acidfire::tmckenna:


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Offlineverytastycheese
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Re: What happened? [Re: Quirkmeister92]
    #27080625 - 12/09/20 04:32 PM (1 month, 17 days ago)

Quote:

Quirkmeister92 said:
Well I wouldn't use the growth you have without transfers so it's possible there is something riding along. Like crack said  take more transfers. You want uniform growth. Additionally your sterile procedures must be on par or better.




I've taken more transfers and I believe my sterile procedures are pretty on point as I've never had anything grow on a clean plate or anywhere I didn't place it, nor have I had any sort of contam on a plate I could detect by sight or smell. If anything it's a hidden myc looking contam that's coming along with my transfers, or just tomentose growth on top of rhizo?

Here's the newer transfers just starting off.






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OfflineCrackatoaS
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Re: What happened? [Re: verytastycheese]
    #27080905 - 12/09/20 07:49 PM (1 month, 16 days ago)

How small of a piece are you taking? My transfers usually have more empty space than myc.


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InvisibleJosex
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Re: What happened? [Re: Crackatoa] * 1
    #27080934 - 12/09/20 08:11 PM (1 month, 16 days ago)




I can't believe nobody has seen the mold in these pics. Gimme a sec and I'll circle it for you.


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InvisibleJosex
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Re: What happened? [Re: Josex]
    #27080945 - 12/09/20 08:18 PM (1 month, 16 days ago)



Here you go.


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OfflineJustweed
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Re: What happened? [Re: Josex]
    #27080946 - 12/09/20 08:19 PM (1 month, 16 days ago)

Quote:

Josex said:
I can't believe nobody has seen the mold in these pics. Gimme a sec and I'll circle it for you.




OP already pointed out that they have trich. We're talking about his agar now.


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Keep 'em high and tight guys....


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InvisibleJosex
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Re: What happened? [Re: Justweed]
    #27080948 - 12/09/20 08:21 PM (1 month, 16 days ago)

That he had trich in his tubs....


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InvisibleJosex
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Re: What happened? [Re: Josex]
    #27080981 - 12/09/20 08:37 PM (1 month, 16 days ago)

His plates may need some more work but I don't think they're contaminated, at any rate not with mold.

It looks like that mold got in the jars upon inoculation.

Those jars in the pic would have needed a shake if they had had no mold, it would have sped things up a lot.


Edited by Josex (12/09/20 08:48 PM)


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Offlinealland
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Re: What happened? [Re: verytastycheese]
    #27081076 - 12/09/20 09:30 PM (1 month, 16 days ago)

Quote:

verytastycheese said:
... If anything it's a hidden myc looking contam that's coming along with my transfers, or just tomentose growth on top of rhizo?





I think i've got this exact scenario going on with a few of my plates.  Thinking of trying Josex's "clean yo shit" tek to isolate.  Will be following your progress for sure.  Please keep us posted and thanks.


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Always learning.. always growing.


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OfflineQuirkmeister92
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Re: What happened? [Re: Josex]
    #27081357 - 12/10/20 01:33 AM (1 month, 16 days ago)

Quote:

Josex said:


Here you go.



I think we all saw it although OP may not have I guess...

Moving beyond the painfully obvious point of; don't use moldy spawn...

Those transfers are looking decent. Smaller samples are definitely helpful.


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Offlineverytastycheese
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Re: What happened? [Re: Quirkmeister92]
    #27081906 - 12/10/20 12:16 PM (1 month, 16 days ago)

Thanks all. Yeah I caught the mold in those jars but what I'm wondering is if the myc I had was actually real, was that trich in its infancy, myc that never mated and won't pin, really thick cobweb mould, or something else? In my closeups of those jars I don't see much for rhizo or organized growth. Same for the plates I still have of that culture, just looking like a thick cotton ball.

Not that it matters anyway, let's see how these new plates look today:

Here's the transfer plate these are from, looking pretty uniform except where I disturbed it IMO, though The Mycologist says it looks dense/stressed.


The transfer to this plate shifted while I was shaking it trying to get rid of some of the condensation. I'm struggling to photograph the fuzziness on top. Wondering if I should try taking a transfer of just that without cutting into the agar to see if I get more of the same or just a puffball.


Some zooms of my little agar sandwiches from Dec 7. These just look like puffballs right now too but maybe that's just them in their infancy before sending out the rhizo feelers?


And here's my little puffball from Dec 3. Man it takes so much longer to get started when you take a poppyseed sized transfer. Looks like I can still see more than one culture there though.


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InvisibleJosex
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Re: What happened? [Re: verytastycheese]
    #27081919 - 12/10/20 12:28 PM (1 month, 16 days ago)

Honestly I can't see anything wrong with those plates. I think the reason myc is looking dense like that is because of too much nutrition. Reminds me of some plates I had when I started with the hobby where I usually went a bit too heavy on the honey for the potato recipe.

Correlation doesn't mean causation in some cases, but you got mold in those jars via technique it looks like, which makes think maybe you need to tighten up your game there. Mold can definitely hide very well in your spawn sometimes.

Also, my looking puffy in the center happens sometimes too. It's just growing from the wedge itself trying to find food above only to find air. Your wedges are pretty big too, so that makes even more sense.


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Offlineverytastycheese
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Re: What happened? [Re: Josex]
    #27092282 - 12/16/20 04:25 PM (1 month, 10 days ago)

Thanks. I like the theory that they're just overfed. I have a few plates that I poured thick and a few that I poured sideways and I can see that the thicker the pour the more it tends towards tomentose growth. IE:

This one was a lot more Rhizo when it started, but it also started on the thin side (I shook it after dropping my wedge... oops) It has been slowing down and thickening out lately.


I'm making more peroxide agar today and dropping from 10gLME to 7g per 500ml water and 10g agar. Hopefully that helps, although these T4 plates are looking pretty good now. So another question, does a plate have to be fully colonized before dropping it to master jars? At what point would you drop these in?





I did an experiment with the T3 mother plate to hopefully confirm that this genetics will actually pin for me. Just cut it in half and dropped both halves into a jar, first transfer done under my LFH! Looking like good growth so far :smile:


I'm anxious that I'm not spotting any contams on any of these though. What's the best color for agar to spot contams on?


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