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Testing for LC contamination on agar vs Grain?


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#1 ShortcutSlim

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Posted 13 May 2022 - 03:46 PM

I'm wondering if contamination would show faster on agar or on grains in a half pint jar? How long would contamination take to show on the average? 10 days to 2 weeks,or is it faster with LC? (generally speaking and allowing for different genetics).


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#2 FunnyFarmer

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Posted 13 May 2022 - 11:26 PM

It took about 5 days to show up on agar and you can really see what is growing on the agar compared to grain. I add some activated charcoal to the agar too to add contrast to what is growing on it. The bad news was my PB was nothing but trich even though it looked like mycelia in the jar. Four other LC jars tested fine as well as a jar each  of PESAmazon and Argentina, those grows were the test detailed in another thread.


Edited by FunnyFarmer, 13 May 2022 - 11:33 PM.

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#3 ShortcutSlim

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Posted 14 May 2022 - 01:06 AM

That answers my question then @FunnyFarmer. I'm just nervous about Agar and looking for an alternative more than anything else. I guess I'll just need to get up the nerve and do it. I have 2 jars of LC that I saw a small white dot that looked about the size of a small cannabis seed.It looked soft to me like it was fuzzy. Both dots are now gone,but I suspect that there is something unwanted in both jars. Guess I'll be finding out here presently.



#4 FunnyFarmer

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Posted 14 May 2022 - 10:25 AM

Once you dip your toes into agar you'll find its not all that hard. I use the so called no pour TEK. Pouring in a SAB didn't work for me, I got more than 60% contam rate before I even used the plate. It dropped to less than 10% when I went with the no pour TEK. The other thing I did was use the large mouth 4 oz glass containers instead of Petri dishes. I can get a mycelia culture going in one and use a syringe to squirt some prepard LC juice in it through the injection port, shake it up good, then transfer it to the LC jar with the syringe. As long as you flame sterilize the needle you don't need a SAB for that operation.
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#5 ElPirana

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Posted 14 May 2022 - 02:05 PM

Once you dip your toes into agar you'll find its not all that hard. I use the so called no pour TEK. Pouring in a SAB didn't work for me, I got more than 60% contam rate before I even used the plate. It dropped to less than 10% when I went with the no pour TEK. The other thing I did was use the large mouth 4 oz glass containers instead of Petri dishes. I can get a mycelia culture going in one and use a syringe to squirt some prepard LC juice in it through the injection port, shake it up good, then transfer it to the LC jar with the syringe. As long as you flame sterilize the needle you don't need a SAB for that operation.

I agree. I tried pouring agar the traditional way in my SAB once several years ago, my first attempt at agar. What a huge PITA!! I went to a no pour tek and never looked back. Probably 5 years into doing agar this way and have never had an agar jar contaminate….at least not before I open the lid to inoculate lol.

ShortcutSlim, making agar is not much more difficult than making LC. I’d say go for it, it can be a lot of fun.

Edited by ElPirana, 14 May 2022 - 02:05 PM.

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#6 ShortcutSlim

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Posted 17 May 2022 - 04:03 PM

I sure appreciate the help with this. @FunnyFarmer mentioned injection ports,and suddenly a light went on the deep dark recesses of my mind.Genius says I ! :biggrin: Decision is now made,no pour is the tek I'll use. Thanks to @ElPirana  for the encouragement too.

 

Folks at Mycotopia are tops.

 

 

On edit: I've seen no mention of GE or FE with regards to Agar. Do you folks suppose grain jar lids could be used in a pinch even though there are FE holes in the lids as well as injection ports?


Edited by ShortcutSlim, 17 May 2022 - 04:14 PM.

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#7 ElPirana

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Posted 17 May 2022 - 06:41 PM

On edit: I've seen no mention of GE or FE with regards to Agar. Do you folks suppose grain jar lids could be used in a pinch even though there are FE holes in the lids as well as injection ports?


I’ve found that it helps to have some gas exchange. I use the exact same lids for grain jars as I do with agar. I use tyvek for the filter. Synthetic filter discs work well too, I used to use those.
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#8 jkdeth

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Posted 17 May 2022 - 08:53 PM

I'm on the fence about GE there. I use petris, wrapped with glad wrap, half pints with no mod, and pp5 containers with a small hole and micropore, because I read they may collapse without it. All three seem to about the same. I do some no mod grain jars, and they've dine fine, but I feel they colonize slower. But using GE on half pint agar cant hurt.
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#9 ShortcutSlim

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Posted 18 May 2022 - 12:29 AM

By golly, it actually worked! My question now is...how to eliminate the condensation? I watched a video and it mentioned wiping up the condensation. How might that be done without introducing contaminants?

 

I assume a person should also wait a few days to make sure there was no contam in the fresh agar?

 

Thanks for all the help.  :smile: 

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#10 jkdeth

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Posted 18 May 2022 - 09:08 AM

Well, too late now of course, next round when pour, leave the lid off and let them cool dry any noticeable condensation before you pc, helps reduce it. Upping the agar in your recipe a bit can help. Pour thinner plates. I most often get some condensation in no pours. I dont worry about it. When its a ridiculous amount. I have given them a quick wipe with a sterile gauze pad. Any extra steps are more steps that can go wrong. You were testing LC right? And you went with grain grain lids with injection ports? I wouldn't do anything extra. Just go with a drop it or so of lc through the port and never have to open the jar.
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#11 ShortcutSlim

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Posted 18 May 2022 - 01:47 PM

Well, too late now of course, next round when pour, leave the lid off and let them cool dry any noticeable condensation before you pc, helps reduce it. Upping the agar in your recipe a bit can help. Pour thinner plates. I most often get some condensation in no pours. I dont worry about it. When its a ridiculous amount. I have given them a quick wipe with a sterile gauze pad. Any extra steps are more steps that can go wrong. You were testing LC right? And you went with grain grain lids with injection ports? I wouldn't do anything extra. Just go with a drop it or so of lc through the port and never have to open the jar.

It's never too late to get some education. :biggrin:  I followed the tek in the video,so thought I was covered. The suggestion of letting the agar cool before running through the PC was something I never considered,but makes a ton of sense. Thanks for suggesting that. I'll cut back on the depth I am pouring too. I thought that it would be a good idea seeing as how the bottom of the jars are slightly concave. So,any reason I couldn't just open the jars and wipe up the condensation and then run them through the PC again? I did see some separation in the agar after it had cooled,so will PC'ing the agar again result in even more separation? Maybe I'll just try it to see. It's not so troublesome to make,so I can always start over if it looks suspect.

 

Or, I could toss the first trial and start fresh. I use my Instant Pot to pc,so it isn't terribly inconvenient to do.



#12 jkdeth

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Posted 18 May 2022 - 03:00 PM

Never ran any a second time. Not sure how that would turn out. I dint think it should hurt.
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#13 Oldpunk

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Posted 18 May 2022 - 03:17 PM

I always just dump the excess out when I inoculate the jars. But I've never used injection ports so I have to open the lid anyway. And I usually squirt too much LC or spore solution in and it's wet anyway.

You could try 1 or 2. And dump/wipe out a couple to re sterilize and see what works best.
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#14 ShortcutSlim

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Posted 18 May 2022 - 03:35 PM

Never ran any a second time. Not sure how that would turn out. I dint think it should hurt.

I think I'll try it. It might separate further,or it might not have a negative effect at all.

 

I always just dump the excess out when I inoculate the jars. But I've never used injection ports so I have to open the lid anyway. And I usually squirt too much LC or spore solution in and it's wet anyway.

You could try 1 or 2. And dump/wipe out a couple to re sterilize and see what works best.

I will do as @jkdeth suggested and let the agar cool and dry a bit before running through the PC on the next round.

 

You bring up a good question tho @Oldpunk,and that is,how much LC to inject for contamination detection? Just a drop or 2 I would guess? Do you scratch it into the agar,or is a drop on the surface adequate?

 

If I wanted to inoculate the agar for a transfer of some sort,would I use a few drops , and should it be scratched into the agar,or would just dropping on the surface be sufficient?
 



#15 ElPirana

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Posted 18 May 2022 - 03:44 PM

Never ran any a second time. Not sure how that would turn out. I dint think it should hurt.

I think I'll try it. It might separate further,or it might not have a negative effect at all.

I always just dump the excess out when I inoculate the jars. But I've never used injection ports so I have to open the lid anyway. And I usually squirt too much LC or spore solution in and it's wet anyway.
You could try 1 or 2. And dump/wipe out a couple to re sterilize and see what works best.

I will do as @jkdeth suggested and let the agar cool and dry a bit before running through the PC on the next round.

You bring up a good question tho @Oldpunk,and that is,how much LC to inject for contamination detection? Just a drop or 2 I would guess? Do you scratch it into the agar,or is a drop on the surface adequate?

If I wanted to inoculate the agar for a transfer of some sort,would I use a few drops , and should it be scratched into the agar,or would just dropping on the surface be sufficient?
I always do what jkdeth suggested, wiping off the condensation and letting it cool before sterilizing. I usually have a little condensation afterwards, but minimal.

I don’t pour too shallow. My jars are concave in the bottom, like you mentioned, and I found that if it’s too thin in the center it can dry out a bit faster in that area.

Edited by ElPirana, 18 May 2022 - 03:44 PM.


#16 FunnyFarmer

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Posted 18 May 2022 - 09:38 PM

Sorry for being MIA, a few days ago I got a bug that wasn't very kind to my intestinal tract, no Hershey squirts but the end result wasn't very fun either. It laid me out for a couple days. The following day I had an endless supply of farts, fortunately not the juicy kind.

Anyway I always pour agar in open air and let the agar set with the lid off before PC'ing. It's helped control the condensation quite a bit. I use the same kind of lid for the agar as I do for grain jars when I want to work with LC, injection port and micropore tape. If I'm just selecting for mycelia no injection port or tape. Once I get the growth I'm looking for then I switch lids for the next round. That way I don't risk contaminating the mycelia by random chance. For the LC jar I use a syringe filter and injection port.

I'm sure some of you are familiar with the video detailing the use of peptone in the LC recipe, I finally tried it and it is much better at growing mycelia. Philly Golden teacher goes through the process for those who haven't heard of it yet.


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#17 ShortcutSlim

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Posted 19 May 2022 - 04:10 AM

I understand full well the discomfort of intestinal issues, I suffer from them myself @FunnyFarmer.

 

It's late and I want to get this posted,so I may come back tomorrow and edit it some....so please forgive the brevity.

 

I have seen the video mentioning Peptone. I may have to try it based on other comments I have read. Could anyone advise me on what LME to buy? There are numerous choices on the Ebay or whatever,and I would prefer to know what to look for. Sadly there are no brew stores locally.



#18 sandman

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Posted 19 May 2022 - 08:06 AM

Briess Organic Maltoferm 10001 Dry Malt Extract 


Edited by sandman, 19 May 2022 - 08:07 AM.

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#19 ShortcutSlim

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Posted 19 May 2022 - 10:43 PM

I forgot aout the edit time limit. :blush:   @FunnyFamer  . I hunted down the video that PGT did that discussed the LME and Peptone. I watched PGT's video,but didn't think to pause to see what brand he was using. I looked at Amazon,but there were quite a few different Malt Extracts offered and I didn't know if the differences were important or not..

 

Thanks to @sandman for sharing the LME I was looking for.

 

I redid the agar, and as suggested,the condensation was greatly reduced so:

 

Do I need to set the jars right side up and allow some time for any potential contamination to show before testing my LC?

 

Will that small amount of condensation be a problem of any kind?

 

Thx.



#20 FunnyFarmer

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Posted 20 May 2022 - 12:45 AM

I forgot aout the edit time limit. :blush:   @FunnyFamer  . I hunted down the video that PGT did that discussed the LME and Peptone. I watched PGT's video,but didn't think to pause to see what brand he was using. I looked at Amazon,but there were quite a few different Malt Extracts offered and I didn't know if the differences were important or not..

 

Thanks to @sandman for sharing the LME I was looking for.

 

I redid the agar, and as suggested,the condensation was greatly reduced so:

 

Do I need to set the jars right side up and allow some time for any potential contamination to show before testing my LC?

 

Will that small amount of condensation be a problem of any kind?

 

Thx.

 

The contam rate has dropped so much that I pretty much use it after it's cooled down but that being said it's still not a bad idea to let it sit for about 5 days before working with them. I'm not very particular about the LME I use so anything used for beer making will be fine. Don't go overboard on your LME purchase as it is very hygroscopic, I've had powder turn into little tar balls in a matter of seconds, it's really challenging working with it during times of high humidity. A while back I got some empty freezer pop bags to store shroom powder in and they are a better container than the bag it comes in so I transferred the powder. It's still challenging but easier to store and break up the rocks that form over time. Those bags are designed to exclude air and humidity so I use what I need, roll up the end and use alittle binder clip to close it off.
 


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