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Invisibleshymanta
Mad Scientist
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Registered: 01/27/05
Posts: 907
100% contamination rate!!
    #5942655 - 08/07/06 11:29 PM (17 years, 5 months ago)

After a year or so of refining my sterile technique and being very successful with it, I'm suddenly confounded.

In the last month I have managed to achieve a 100% contamination rate!!  With two casings going strong and having no problems what so ever, every jar of grain put in the incubator has contaminated with brown shit I've never seen before.  Four different grains.  Rye, oat, quinoa, and millet.  Every one eaten by some brown funk.

My grain prep this past month:
Soak over night >> rinse >> simmer >> rinse >> drain >> PC/60 mins >> inoculate in glove box >>  incubate >> contaminate :confused:

Anyone have any thoughts?


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OfflineYamidude
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Re: 100% contamination rate!! [Re: shymanta]
    #5942660 - 08/07/06 11:31 PM (17 years, 5 months ago)

vendor spores? (no need to list vendors name if they are btw)


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Invisibleshymanta
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Re: 100% contamination rate!! [Re: Yamidude]
    #5942670 - 08/07/06 11:35 PM (17 years, 5 months ago)

I doubt it. I would have considered this if I hadn't been working with at least 10 different prints.


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OfflineYamidude
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Re: 100% contamination rate!! [Re: shymanta]
    #5942686 - 08/07/06 11:40 PM (17 years, 5 months ago)

I was asking if the specimens you have came from a vendor or from someone else?


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InvisibleOmnicracker
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Registered: 09/18/05
Posts: 1,421
Loc: Wal-Merica
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Re: 100% contamination rate!! [Re: Yamidude]
    #5942717 - 08/07/06 11:48 PM (17 years, 5 months ago)

try PCing longer, especially with rye.

how hot is your incubator getting?


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




xxx..Learn Something..xxx


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Invisibleshymanta
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Re: 100% contamination rate!! [Re: shymanta]
    #5942727 - 08/07/06 11:51 PM (17 years, 5 months ago)

Oh, sorry. Spores were not from vendors. Some from trades, some prints I've taken. A couple were cultures I did purchase from a vendor.


Edited by shymanta (08/07/06 11:52 PM)


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InvisibleTippinthru
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Registered: 04/07/05
Posts: 1,131
Loc: "The Garden"...
Re: 100% contamination rate!! [Re: shymanta]
    #5942765 - 08/08/06 12:03 AM (17 years, 5 months ago)

Trade prints are always questionable.

Even, your own prints, if you don't have the techniques almost perfect.

Vendor syringes are the most RELIABLE.

Since, 100% went south, it could be that your are missing something somewhere in your process, that lets the nasties through the door.

Think on it. Detail your process in writing.

Inspect the list, and look for the error, in it.

Some small thing, can do you in.

Ponder what that could be.

Then fix it, and try again.

If that doesn't work, go over your list again, and fix EVERY possible entry point.


--------------------
Perfection is attained by slow degrees; it requires the hand of time...
[


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InvisibleOmnicracker
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Re: 100% contamination rate!! [Re: Tippinthru]
    #5942767 - 08/08/06 12:04 AM (17 years, 5 months ago)

what kind of filter lids are you using?


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xxx..Learn Something..xxx


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Invisibleshymanta
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Re: 100% contamination rate!! [Re: Omnicracker]
    #5942776 - 08/08/06 12:08 AM (17 years, 5 months ago)

Incubator is set to 84 for those who need it. Even the room temp jars are getting this shit.

I don't suspect them prints necessarily. Most I've used before with success.

Edit: lids are poly-fill and Tyvek. Here they are in more detail... http://www.shroomery.org/forums/showflat.php/Number/5561630#Post5561630


Edited by shymanta (08/08/06 12:12 AM)


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InvisibleTippinthru
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Re: 100% contamination rate!! [Re: shymanta]
    #5942800 - 08/08/06 12:14 AM (17 years, 5 months ago)

Something isn't right.

It has to be a major thing, involving every jar.

Because EVERY JAR contaminates.

Ponder your inoculation method.

And, all else every jar has in common.


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Perfection is attained by slow degrees; it requires the hand of time...
[


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Offlinexaxphaanes
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Re: 100% contamination rate!! [Re: shymanta]
    #5942806 - 08/08/06 12:15 AM (17 years, 5 months ago)

it sounds like you are having the same problem i was having i also simmered after soaking and the jars would be very wet even if they didn't really look wet they still contamed i suggest that you up the pc time and don't simmer use agar's(6tango) tek worked wonders for me hope you will have the same man.


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"Anything i say is fictional"
  what you should look for in manure


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OfflineYamidude
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Re: 100% contamination rate!! [Re: xaxphaanes]
    #5942817 - 08/08/06 12:18 AM (17 years, 5 months ago)

How long are you simmering? Are you simmering or boiling?


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Invisibleshymanta
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Re: 100% contamination rate!! [Re: Tippinthru]
    #5942823 - 08/08/06 12:20 AM (17 years, 5 months ago)

I really like your reply. Great scientific thinking. High five! I'll write my next one as I go and study it. Oysters are next. I really want to try our some straw logs.

I kinda suspect my glove box. I'm thinking about building a clean room equip with hepa filtration...


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Invisibleshymanta
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Re: 100% contamination rate!! [Re: Yamidude]
    #5942847 - 08/08/06 12:26 AM (17 years, 5 months ago)

Quote:

Yamidude said:
How long are you simmering? Are you simmering or boiling?




I simmer (no boiling) until just before kernels begin splitting. About 20 mins.


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Invisibleshymanta
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Re: 100% contamination rate!! [Re: xaxphaanes]
    #5942856 - 08/08/06 12:28 AM (17 years, 5 months ago)

Xaxphaanes, can you post a link to agar's tek?


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InvisibleTippinthru
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Registered: 04/07/05
Posts: 1,131
Loc: "The Garden"...
Re: 100% contamination rate!! [Re: shymanta]
    #5942878 - 08/08/06 12:36 AM (17 years, 5 months ago)

Quote:

shymanta said:
Quote:

Yamidude said:
How long are you simmering? Are you simmering or boiling?




I simmer (no boiling) until just before kernels begin splitting. About 20 mins.




Jar content moisture is a common point.

ReadTHIS POST

It might help you out.

Straight soaking is "nearly" fool-proof.

Simmering is like cooking.

Not long enough, and it doesn't get cooked.
(to dry)

To long, and it is over-cooked.
(burst grains)

With your method, you never know how it is, until much later.

Straight soaks give you a wide degree of latitude.

Simmering has a VERY narrow margin, of error.


--------------------
Perfection is attained by slow degrees; it requires the hand of time...
[


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Offlinetrentallica
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Re: 100% contamination rate!! [Re: shymanta]
    #5942879 - 08/08/06 12:37 AM (17 years, 5 months ago)

all jars are affected. so it has to be something that all have in common. if the jars are sealed up then even the most nastey incubator should not be an issue. you said that you have spent a long time getting this down, so i dont think its that. this leaves your prep area, but even then the pc should zap that problem. maybe the pc is just not working as well as it use to?? i used my for 45 min with WBS and they all want to shit. the next time i just took my jars to work, and used an autoclave. next time i use the pc for WBS jars ill try 1 1/2 hours.

just a thought


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OfflineYamidude
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Re: 100% contamination rate!! [Re: trentallica]
    #5942888 - 08/08/06 12:39 AM (17 years, 5 months ago)

next time make a control jar..


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Offlinekristen
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Registered: 04/19/06
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Re: 100% contamination rate!! [Re: Yamidude]
    #5943141 - 08/08/06 02:44 AM (17 years, 5 months ago)

Anyone for inoculation procedure? Maybe heavily contaminated air?


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OfflineLily_Morgan
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Registered: 07/05/06
Posts: 437
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Re: 100% contamination rate!! [Re: kristen]
    #5943156 - 08/08/06 02:52 AM (17 years, 5 months ago)

Quote:

kristen said:
Anyone for inoculation procedure? Maybe heavily contaminated air?




That's actually exactly what I was thinking.  He must have some nasty crap in the vents of his living area.  Possibly have the air ducts flushed, and keep a hepa air filter in the room where the inoculation is being performed. :thumbup:


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