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InvisiblePestile

Registered: 05/02/13
Posts: 875
Loc: Northern Europe
MYA agar dishes acting strange, 2 contams and 2 not wanting to jump-off
    #19274915 - 12/14/13 02:11 PM (10 years, 2 months ago)

Hi guys!
I poured some plates and made a couple transfers to them 10 days ago and I don't know what to make of this;

I made it with these ingredients:
10g. ME
10g. agar
1g. yeast
500ml water

which I PC'd for 40 minutes at 15psi.


I inoculated 4 dishes with two different cultures which I have in some vials, these are PE and P. stipticus. The PE's are showing contaminations and the stipticus won't even jump off.

So now I have these plates, 2 with a good amount of the wrong mycelium and 2 plates with barely any mycelium and I can't figure out why that is.

What did I do wrong?
Will P. stipticus not grow on MYA?
Did I overcook the agar?

The two small stipticus wedges I put on agar looks really good and I don't understand why it won't jump off when the PE/contam is thriving?!

Can I take these small wedges and transfer to a wood-based agar or should I just forget about them? They look healthy enough. Slow, but healthy.


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OfflineCamtaro420
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Registered: 11/25/13
Posts: 707
Loc: New Mexico
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Re: MYA agar dishes acting strange, 2 contams and 2 not wanting to jump-off [Re: Pestile]
    #19274933 - 12/14/13 02:15 PM (10 years, 2 months ago)

Pics? your cooking procedure? Your sterilization procedures? More info in general?


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OfflineStromriderM
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Registered: 06/02/13
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Re: MYA agar dishes acting strange, 2 contams and 2 not wanting to jump-off [Re: Camtaro420]
    #19274940 - 12/14/13 02:17 PM (10 years, 2 months ago)

Can you over cook agar? :confused:
I always pc for 30 min. I probably don't even need to go that long


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OfflineCamtaro420
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Registered: 11/25/13
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Re: MYA agar dishes acting strange, 2 contams and 2 not wanting to jump-off [Re: Stromrider]
    #19274960 - 12/14/13 02:24 PM (10 years, 2 months ago)

Quote:

Stromrider said:
Can you over cook agar? :confused:
I always pc for 30 min. I probably don't even need to go that long




Are you pressure cooking the agar in the pan? or the agar in a vessel like a jar or something if it's in a jar then you could go well beyond 30 mins most do 90-120.

Also your recipe doesn't look as good as it could have been.

The important thing to know is, after you PC'd your agar, how did you pour it into your dishes?

Were the dishes sterilized?

Did you use a SAB?

Pics would help to see what exactly is going on.


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InvisiblePestile

Registered: 05/02/13
Posts: 875
Loc: Northern Europe
Re: MYA agar dishes acting strange, 2 contams and 2 not wanting to jump-off [Re: Stromrider]
    #19274982 - 12/14/13 02:30 PM (10 years, 2 months ago)

I think you can, but I don't know for sure.

I put all ingredients into a pot and heated it a little while stirring, poured it into my whiskey bottle and PC'd 40 minutes, afterwards I let it cool about 4 hours before pouring the plates.
I poured them in my SAB and wrapped them in some plastic wrap when they had set.
Next day I inoculated and put the dishes in zip lock bags where they have been since.


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OfflineCamtaro420
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Registered: 11/25/13
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Re: MYA agar dishes acting strange, 2 contams and 2 not wanting to jump-off [Re: Pestile]
    #19274988 - 12/14/13 02:33 PM (10 years, 2 months ago)

Quote:

Pestilence said:
I think you can, but I don't know for sure.

I put all ingredients into a pot and heated it a little while stirring, poured it into my whiskey bottle and PC'd 40 minutes, afterwards I let it cool about 4 hours before pouring the plates.
I poured them in my SAB and wrapped them in some plastic wrap when they had set.
Next day I inoculated and put the dishes in zip lock bags where they have been since.





what kind of yeast did you use, and you innoced in the SAB as well right?

Also did you use a syringe off the internet or one you made yourself?


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InvisiblePestile

Registered: 05/02/13
Posts: 875
Loc: Northern Europe
Re: MYA agar dishes acting strange, 2 contams and 2 not wanting to jump-off [Re: Pestile]
    #19275029 - 12/14/13 02:43 PM (10 years, 2 months ago)

The dishes are glass. Washed in warm water and soap, wrapped in several layers tinfoil, PC'd 30 minutes and then baked in the oven for about 1-2 hours at 170°C to remove water from them. So they should be clean and sterile.

I use brewers yeast, something called nutri-yeast.
Yes, I always use my SAB with wet walls and wet towel in the bottom.
I didn't use syringe, I have the cultures in vials in the fridge. :smile:


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OfflineCamtaro420
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Registered: 11/25/13
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Re: MYA agar dishes acting strange, 2 contams and 2 not wanting to jump-off [Re: Pestile]
    #19275046 - 12/14/13 02:47 PM (10 years, 2 months ago)

Seems like you did everything pretty much to the letter. Only thing I could think of at this point is somewhere between adding the culture to the dish is where contaims happened. Maybe in the cultures, or how you were adding them to the petris.

How did you add the cultures to the petris?


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Edited by Camtaro420 (12/14/13 02:52 PM)


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OfflinecronicrFacebook
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Re: MYA agar dishes acting strange, 2 contams and 2 not wanting to jump-off [Re: Camtaro420]
    #19275091 - 12/14/13 02:55 PM (10 years, 2 months ago)

for trouble shooting my technique i like to take a plate to my sab and do what i would do to inoculate but don't inoculate, open the dish/dishes up for a few secs and close them back up. if they contam it's me if not it's my prints


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InvisiblePestile

Registered: 05/02/13
Posts: 875
Loc: Northern Europe
Re: MYA agar dishes acting strange, 2 contams and 2 not wanting to jump-off [Re: Camtaro420]
    #19275102 - 12/14/13 02:59 PM (10 years, 2 months ago)

Using a scalpel. Flamed it red hot and let it cool down inside the SAB before transferring the pieces.

I'm not really worried about my sterile procedure/technique, it's more why the stipticus won't jump off when the contam obviously will.
I have 10 plates with this MYA waiting to be inoculated and I'll be slightly annoyed if all I can do with them is dump them in the trashcan.

I know contams can thrive in places normal mycelium would perish. So that's kinda what I'm afraid is going on here, "I screwed up the agar and now the only thing that will (and can) grow on it is contaminations." :crazy:


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Invisible36fuckin5
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Registered: 08/11/03
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Re: MYA agar dishes acting strange, 2 contams and 2 not wanting to jump-off [Re: Pestile]
    #19275209 - 12/14/13 03:28 PM (10 years, 2 months ago)

You're supposed to use yeast nutrient, not actual yeast. Trash those plates. You don't even need the yeast nutrient, although it helps.


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If you're offended I don't give a shit and don't come see me no more.

Pat The Bunny said:
A punk rock song won't ever change the world, but I can tell you about a couple that changed me.

bodhisatta said:
i recommend common sense and figuring it out.

These are the TEKs I use. They're all as cheap and easy as possible, just like your mom.


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OfflineCamtaro420
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Registered: 11/25/13
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Re: MYA agar dishes acting strange, 2 contams and 2 not wanting to jump-off [Re: 36fuckin5]
    #19275368 - 12/14/13 04:08 PM (10 years, 2 months ago)

Quote:

36fuckin5 said:
You're supposed to use yeast nutrient, not actual yeast. Trash those plates. You don't even need the yeast nutrient, although it helps.




Brew yeast is fine as long as it is not active. Nutritional Yeast is just better in most cases. the important thing is inactive yeast.


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Offlineloki44
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Registered: 08/01/13
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Re: MYA agar dishes acting strange, 2 contams and 2 not wanting to jump-off [Re: Camtaro420]
    #19275386 - 12/14/13 04:10 PM (10 years, 2 months ago)

in, this thread is relevant to my interests


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InvisiblePestile

Registered: 05/02/13
Posts: 875
Loc: Northern Europe
Re: MYA agar dishes acting strange, 2 contams and 2 not wanting to jump-off [Re: 36fuckin5]
    #19275515 - 12/14/13 04:40 PM (10 years, 2 months ago)

I am using nutritional yeast. At first I had purchased bakers yeast, but FH informed me that it had to be inactive and bakers yeast is active, so that's why I bought this product from a brewer.

Did I make the agar too nutritious when I added the yeast?
I know it's not required at all, I'm just trying things out. But I must admit, I did not expect it would mess so much with me. :smile:

Next batch will be a pure MEA recipe, no doubt about that. I should probably just stick to the basics until I get a better understanding of this agar culturing.


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