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OfflinePs.NoName
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Registered: 08/03/18
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Bacteria inside the spore syringe ?
    #26817711 - 07/11/20 04:03 PM (3 years, 7 months ago)

Hey Shroomery.  I am having some bacteria troubles and trying to figure out where it is coming from.

Every few years I do a small batch of PF cakes.  Apparently I waited too long between grows and print I had left seems to be not viable.  It grew nothing.

I orders some spore syringes from a vendor that has been around a long time.  Made cakes likes I have done 10+ times before but every single one got bacteria and it seemed to start at the inoculation point.

I almost never have contamination problems.  Maybe 1-2 jars slow down and bacteria out, but this time lost 100% cakes.  Figured I did something wrong with sterilization or something.... No big deal.  Made new cakes, used two different syringes from the same vendor.  Again 100% of the cakes got bacteria again.  I did not check on them for 1 week so hard to say if it started at inoculation point, but looks that way.  The bacteria in the 2nd round cakes has the same grossly sweet smell the first syringe ones did.

I have one untouched spore syringe from that same vendor left but now I am wondering if I should not use it.  I know the vendor means well and don't want to throw them under the bus, but do vendors ever have trouble with bacteria?  It is a very well known vendor I think I used 10+ years ago.

Could the bag of brown rice flour be a problem?  The grind seems super super dusty fine this time.  I would usually grind down Uncle Bens, but got lazy and used bagged up 'premium' brown rice flour.



Hard to tell much from the picture but the top of the cakes near the inoculation point is bacteria ridden but the bottom halves look OK.


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OfflineReverance
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Re: Bacteria inside the spore syringe ? [Re: Ps.NoName]
    #26818064 - 07/11/20 07:11 PM (3 years, 7 months ago)

I just did a bunch of plates, nocced them up with syringes from two seperate vendors. plates from vendor a were all bacterial. plates from vendor b. were fine. so yeah, Id love an answer to this if just to be more knowledgeable


Edited by Reverance (07/11/20 07:16 PM)


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Offlinedlj403
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Re: Bacteria inside the spore syringe ? [Re: Reverance]
    #26818138 - 07/11/20 07:46 PM (3 years, 7 months ago)

spore syringes are by definition unclean. you can shoot them direct and cross your fingers. if all the rest of your technique is spot on, plus you shake well and there's more spore than other crap, you're golden.

but the real world doesn't work that way. and even if you have success the first time out, the law of large numbers will eventually catch up with you.


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Offlinecallmedismail
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Re: Bacteria inside the spore syringe ? [Re: dlj403]
    #26818149 - 07/11/20 07:51 PM (3 years, 7 months ago)

This is why the most recent advice is all agar from day one, I think. I have had similar problems going MS > grain. Shitty.


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Invisiblemicelio
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Re: Bacteria inside the spore syringe ? [Re: Ps.NoName]
    #26818169 - 07/11/20 08:04 PM (3 years, 7 months ago)

What does it take to kill a bacterial spore? Petri dish timelapse




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OfflinePs.NoName
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Re: Bacteria inside the spore syringe ? [Re: micelio]
    #26818326 - 07/11/20 09:39 PM (3 years, 7 months ago)

I don't think I have just been lucky with not getting contams doing MS to brown rice PF jars.  Done this many times since 1999. Normally would make my own spore syringes from prints I took.  I was reasonably sterile in that process and never lost 100% of cakes to bacteria/mold.  The success rate with PF tek at least for me (i think most people?) has been very high.

I may try making a syringe from an old print one more time, use the last suspect bacterial vendor syringe, and do an not inoculated control jar to see if it stays healthy.

Just need a small batch to work really.  Handful of cakes produce enough to last a few years.  Don't trip much anymore.  Wanted to do them outdoors again but starting to get late in the good season for me to do so.  I think I still have time.

Last time outdoor.




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OfflineFailboat
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Re: Bacteria inside the spore syringe ? [Re: Ps.NoName]
    #26818773 - 07/12/20 06:08 AM (3 years, 7 months ago)

What is your procedure?


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OfflinePs.NoName
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Re: Bacteria inside the spore syringe ? [Re: Failboat]
    #26818983 - 07/12/20 09:19 AM (3 years, 7 months ago)

make pf cakes half pint jars 2:1:1.  Steam 60-90 minutes.

If doing outdoors I dig a spot in the mulch bed crumbled cakes.  cover with coco coir and then some straw on top.  Keep it moist  It took about 3 weeks and I had that killer flush in the pics above.  It is so hands off and so easy to do.  Worst part is making the cakes takes like an hour or two.


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OfflineFailboat
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Re: Bacteria inside the spore syringe ? [Re: Ps.NoName]
    #26819724 - 07/12/20 03:24 PM (3 years, 7 months ago)

So you just flamin the needle and squirting in open air through an open lid? Need all the details.


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InvisibleShenmue
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Registered: 12/21/18
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Re: Bacteria inside the spore syringe ? [Re: Failboat]
    #26819744 - 07/12/20 03:36 PM (3 years, 7 months ago)

I've been getting dirty spore syringes from my main spore vendor lately so I understand your frustration. I noticed cubes can usually handle a spore syringe to grain but for some reason panaeolus cyanescens can't handle it. Dont even attempt to try it with panaeolus cyanescens! I'm sure someone has done it but I tried 3 syringes with both wbs and ryeberries. Agar is definitely the way to go. It's easy to make and if you're lazy you can buy it already done. For now on I'm only using agar..


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OfflinePs.NoName
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Re: Bacteria inside the spore syringe ? [Re: Failboat]
    #26819795 - 07/12/20 04:11 PM (3 years, 7 months ago)

Quote:

Quirkmeister92 said:
So you just flamin the needle and squirting in open air through an open lid? Need all the details.




I do it in clean room with still air.  Needle flamed.  Same way I always have for like 20 years without issue.  First time in many years I needed to order spores and just seems quite suspicious that never really had trouble with losing 100% of cakes until I bought vendor made syringe instead of doing it myself.

I don't think its coming from the act of sticking the needle into the cake, but either in the substrate or spore syringe.  Doing a test where I will leave out one jar without it getting inoculated and I going to try a different old print from a different variety as maybe it aged better.  That way I will have vendor syringe, my syringe, and a control jar and see what happens.


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OfflinePs.NoName
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Re: Bacteria inside the spore syringe ? [Re: Shenmue]
    #26819804 - 07/12/20 04:15 PM (3 years, 7 months ago)

Quote:

Shenmue said:
I've been getting dirty spore syringes from my main spore vendor lately so I understand your frustration. I noticed cubes can usually handle a spore syringe to grain but for some reason panaeolus cyanescens can't handle it. Dont even attempt to try it with panaeolus cyanescens! I'm sure someone has done it but I tried 3 syringes with both wbs and ryeberries. Agar is definitely the way to go. It's easy to make and if you're lazy you can buy it already done. For now on I'm only using agar..




I was able to do ovoid into a PF cake and it took pretty well.  I just mixed in some sawdust into the PF formula and cut back some on the brown rice.

Then broke up the PF cakes into wood chips last summer.  They colonized a ton of wood chips but saddly no fruits this spring.  Not sure if the conditions where not good for the ovoid or they are just not ready yet to fruit.  Fingers crossed for maybe autumn fruits and next spring.  Gotta work on some ground cover plants in the wood chip bed.


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OfflineFailboat
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Re: Bacteria inside the spore syringe ? [Re: Ps.NoName]
    #26819823 - 07/12/20 04:29 PM (3 years, 7 months ago)

Clean room with still air? I doubt it's as clean or as still as you think it to be. If you're not using a SAB then I would expect more failure than success. Use a SAB and then clean up your culture.


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OfflinePs.NoName
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Re: Bacteria inside the spore syringe [Re: Failboat]
    #26819874 - 07/12/20 05:10 PM (3 years, 7 months ago)

Quote:

Quirkmeister92 said:
Clean room with still air? I doubt it's as clean or as still as you think it to be. If you're not using a SAB then I would expect more failure than success. Use a SAB and then clean up your culture.




My experience over countless grows in the last 20 years says different but I will setup a SAB for the next time just to eliminate possibilities.  Better to do overdo sterility I suppose than cut corners. But I did not get lucky every single time in the last 20 years... just no way.

Used to live in a house with dirty roomates and I would innoc jars in my bedroom open air alcohol wipe down needle not knowing better and It pretty much always worked.  Not that I am recommending it.  Now i'm grown up with my own house that stays clean and the room I do it in is extra clean.


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Offlinemeowjinx
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Re: Bacteria inside the spore syringe [Re: Ps.NoName]
    #26819890 - 07/12/20 05:18 PM (3 years, 7 months ago)

You could just make agar and test it on there. It wouldn't cost much and you'd get a definitive answer on whether it's one of the syringes of the substrate and you'll get an answer faster than if you leave a jar alone


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OfflineLogicaL ChaosM
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Re: Bacteria inside the spore syringe ? [Re: Ps.NoName]
    #26820015 - 07/12/20 06:54 PM (3 years, 7 months ago)

U made your jar substrate too wet. Use less water and use half pint wide mouths, way better than regular mouths.


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OfflinePs.NoName
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Re: Bacteria inside the spore syringe ? [Re: LogicaL Chaos]
    #26820082 - 07/12/20 07:54 PM (3 years, 7 months ago)

Quote:

LogicaL Chaos said:
U made your jar substrate too wet. Use less water and use half pint wide mouths, way better than regular mouths.



Used the same substrate mix amounts as usual 2:1:1, but the micro ground dusty fine brown rice flour compared to me coffee grinding uncle ben's did seem to change feel of the substrate mix.  I will try again with less water.

Been using the same half pint jars forever.  Did not even realize they come in two different shapes.  I will see if the stores around me has wider ones.

never made agar and not sure if I want to start messing with it.  I'm probably being stubborn sticking to old school PF jars, but just frustrating the method have always done is failing.

thanks everyone for your time.


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OfflineFailboat
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Re: Bacteria inside the spore syringe ? [Re: Ps.NoName]
    #26820173 - 07/12/20 09:27 PM (3 years, 7 months ago)

Agar is easy and very worth while
:costanza:


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OfflineReverance
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Re: Bacteria inside the spore syringe ? [Re: Failboat]
    #26820232 - 07/12/20 10:26 PM (3 years, 7 months ago)

I have some plates that have bad bacterial contamination. any tips or tricks for transfering away from this mess? By that I mean what if the mycelium is sitting on the bacteria? could dilute peroxide help?


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Offlinemeowjinx
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Re: Bacteria inside the spore syringe ? [Re: Reverance]
    #26820242 - 07/12/20 10:37 PM (3 years, 7 months ago)

Do you have pictures of this curiosity?

Maybe you could cut as shallow a wedge as possible if the bacteria have somehow burrowed themselves in the agar under the myc. I've been able to basically "pry" pieces of myc off of plates before (not intentionally, but in your case it might be something you could try)

Maybe after you cut the myc out try streaking it a bit across the transfer plate to see if an uncontaminated section develops. Then maybe do one or two more transfers until you're confident that the myc has been isolated

Unless the bacteria is a fungal parasite of some sort, maybe then you'd have to start from scratch


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