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OfflineLacrossa
Cmdr of PiratePenguins

Registered: 09/20/05
Posts: 21
Loc: Mormonia
Last seen: 17 years, 3 months
sexy red contamination
    #4839061 - 10/22/05 07:40 PM (18 years, 5 months ago)

...aka "lipstick mold," aka "Geotrichum"  :shocked:

My friend's first grow effort appears to be contaminated. He inoc'd 12 jars with one 10cc syringe (admittedly, spreading it a bit thin) of PF classic. He followed sterilization procedures as best he could: 10% solution of bleach applied liberally; mass quantities of lysol sprayed and wiped inside "clean" room; 70% isopropyl alcohol wipe-down of syringe needle & counter-tops. He PC'd and didn't open the PC until inside the clean room. He wore disposable latex gloves, and a simple dust mask. After inoculation, all of his jars were fitted with synthetic filter discs and then placed in a TiT incubator. The TiT has been opened only three times to observe growth.

Here is one of his jars--

--Day 10 @ 85.6?F  [Notice the blurry red dots off-center]

He's not sure what to do. At least half of the jars show some sign of red, sometimes only as small as little flakes or streaks. After flipping through Stamets & searching the Shroomery, he thinks he's found a likely suspect.

From The Mushroom Cultivator: Geotrichum

"Mycelium whiting at first, often taking on a frosty appearance and then forming whiting balls of mycelium. With age, the mycelium becomes pinkish and then reddish due to the maturation of spores. Older colonies of this fungus fade to a dull orange... Commonly encountered in agar plates made from a soil infusion; otherwise rarely encountered in sterile culture. An occasional contaminant of mushroom beds (compost), Lipstick Mold inhibits primordia formation and development."

If it _is_ contamination, he's not sure from which vector it would have come. He placed one jar w/ substrate inside the clean room, and left it un-innoc'd--he just placed a filter disc on it. His intention was to see what might grow inside the jar if left unattended in the open air. So far (nearly 3 weeks), nothing has grown. He wonders about his syringe needle, however. It was new and sterilized, and he used an alcohol lamp to heat the needle before penetrating the subtrate for inoculation... but he did not do this every time.

Now he wants some more experienced opinions. What do you guys think?

Lac

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OfflineRogerRabbitM
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Re: sexy red contamination [Re: Lacrossa]
    #4839748 - 10/22/05 11:09 PM (18 years, 5 months ago)

The filter disks need to be sterilized with the grains. If you put the filter disk on later, that might be the vector.

Latex gloves should be washed with 70% alcohol once you put them on.

A dust mask will not stop bacteria leaving your mouth. Get a medical mask from a health supply store. It will be labeled, "stops 99% of the wearers exhaled bacteria".

Lysol will do very little to clean the air in a room. Lysol is a surface disinfectant. Use Oust or 10% bleach in a misting bottle to spray the air. You should still be using a glove box so you work with no air currents.

That temperature is too high. 80F should be considered the maximum incubation temperature. Due to thermogenesis, the temperature inside the jar is several degrees warmer than outside. Glass is also an insulator, further raising the temperature within. Higher temperatures favor molds, lower temperatures favor mushroom mycelium. Personally, I incubate on an open shelf at normal room temperature.

The needle needs to be flame sterilized between each jar.

What grain was used, and how did you protect it during inoculation? I suspect the grains became contaminated before the filter disks went on.
RR


--------------------
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semper in excretia sumus solim profundum variat

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OfflineLacrossa
Cmdr of PiratePenguins

Registered: 09/20/05
Posts: 21
Loc: Mormonia
Last seen: 17 years, 3 months
Re: sexy red contamination [Re: RogerRabbit]
    #4840264 - 10/23/05 01:43 AM (18 years, 5 months ago)

Thanks, Roger--appreciate the critique!

To clarify: these are 12 PFtek ?pint jars growing PF classic strain. Used standard PF cake formula of fine vermiculite, brown rice flour and water. Not very adventurous, roughly comparable to heterosexual missionary-style sex. :smile:

Filter discs were sterilized along with the substrates + jars in the pressure cooker. My friend placed small squares of aluminum foil over the discs/jars to prevent the discs from dampening during PC'ing (while acquainting himself with the pressure cooker, he noticed that the discs tended to become wet if left exposed, and he reasoned that this would defeat the filter).

There was some skepticism about the dust mask. Anything that can be picked up on aisle 6 of Home Depot probably isn't going to be HEPA-esque. :wink: Lysol was used as a spray-and-wipe surface disinfectant. Oust was also sprayed in the air of the (bath)room. My friend would love to use a glovebox, in fact has already purchased/ordered one, but hasn't been getting replies from the vendor (a Shroomery sponsor) about delivery. Building a flowhood is in the immediate future.

The water temperature in the TiT is technically 82?F, there's a heat sensor suction-cupped to a couple of the jars and they register a few degrees higher. Do you recommend simply lowering the water temperature to 80?F? Do you find much difference in colonization times?

The needle was NOT flame sterilized between each jar, which was a serious oversight. My friend says that perhaps only 1/3 of the jars had a flame-sterilized needle inserted... roughly the proportion that are free from red blemishes (so far).

Does it look like this is a "lipstick mold" contam?

Different jar, same thing. And now that it's in better light, the lipstick appears to have a greenish friend.  :tongue:

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Offlineiamgod
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Registered: 06/30/03
Posts: 263
Loc: Crooklyn, New York
Last seen: 16 years, 24 days
Re: sexy red contamination [Re: Lacrossa]
    #4840974 - 10/23/05 10:49 AM (18 years, 5 months ago)

That jar looks very wet. That doesn't look like green but if you keep it for a while it will probably develop. Are you sure about the contam-free virtue of your syringe?  :cthulhu:


--------------------
If what you seek is truth then drugs can not offer truth. Drugs can offer the truth of drugs.......Altered states. Truth is not an altered state of mind. ~ Big Headed Nice Guy Who Loves You And Your Dog

http://www.inlibertyandfreedom.com/Flash/Think_It_Over.swf

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OfflineLacrossa
Cmdr of PiratePenguins

Registered: 09/20/05
Posts: 21
Loc: Mormonia
Last seen: 17 years, 3 months
Re: sexy red contamination [Re: iamgod]
    #4842046 - 10/23/05 04:09 PM (18 years, 5 months ago)

The syringe was fine initially, but probably picked up some contams between jars.

My friend noticed a fairly intense smell from the TiT (when fanned) from about day 4 onward. What should healthy mushrooms smell like?

He took apart one of the cakes w/ red streaks last night, to find this--

A bit blurry from the picture, but clearly deep green contamination in the center.

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Offlineiamgod
just some dudesome where

Registered: 06/30/03
Posts: 263
Loc: Crooklyn, New York
Last seen: 16 years, 24 days
Re: sexy red contamination [Re: Lacrossa]
    #4842760 - 10/23/05 07:11 PM (18 years, 5 months ago)

Healthy mushrooms should smell nice and mushroomy. When you first grow Ps. cubensis you'll remember the distinct mushroomy smell forever. Also, in that cake pic, is it actually green mold, or is the cake bruising where you cut it?

Also, it is best not to handle contammed jars without at least proper ventilation and gloves.


--------------------
If what you seek is truth then drugs can not offer truth. Drugs can offer the truth of drugs.......Altered states. Truth is not an altered state of mind. ~ Big Headed Nice Guy Who Loves You And Your Dog

http://www.inlibertyandfreedom.com/Flash/Think_It_Over.swf

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OfflineLacrossa
Cmdr of PiratePenguins

Registered: 09/20/05
Posts: 21
Loc: Mormonia
Last seen: 17 years, 3 months
Re: sexy red contamination [Re: iamgod]
    #4842994 - 10/23/05 07:53 PM (18 years, 5 months ago)

Latex gloves and a fan were used to pull the cake out of the jar and into the sink. My friend's well aware of the health hazards posed by the questionable fun of growing mold colonies. :wink:

The green was quite distinct. Here's another, clearer picture taken a few seconds after it came out of the jar:

Down it went to a sliced death. A li'l spaghetti and tomato went with it.

The rest of the jars today are showing the same growth, so the whole batch is getting thrown out. Looking back at the inoculation/incubation period, it seems a good guess that the syringe needle was contaminated between jars and spread contamination once it was inserted. Generally the jars have the strongest mold growth in the bottom of the jar, roughly where the syringe was placed. Once the jars were all placed in the TiT, the bad ones probably cross-contaminated the rest.

...which brings another good question up. How much should one count on the filter discs? Either all were contaminated from the syringe needle (which was definitely burned-red between several jars), or a few were and the discs (+the dry layer of vermiculite on top of the substrate) failed to stop spores from entering. And, it would seem if you can smell anything coming out of the jars, the discs must not be filtering -that- much. These discs were purchased from Myco Supply, a Shroomery sponsor.

iamgod, you mentioned that the jars looked very wet. They were wet, a liberal amount more water was used than in the standard PFtek since the vermiculite was of the finer variety. My friend basically saturated the verm with water until it was submerged, then strained it for ~20 minutes before mixing with BRF. More water = more contams?

thanks for the feedback guys, it helps a lot!

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OfflineRogerRabbitM
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Re: sexy red contamination [Re: Lacrossa]
    #4843074 - 10/23/05 08:10 PM (18 years, 5 months ago)

I would recommend following the recipie to the letter. There is no need to add extra water. In fact, with less moisture, any substrate will colonize faster. You can always dunk at full colonization to replenesh any lost/used water. Also, too wet tends to lead to an aneoribic core, which then leads to bacterial contamination, which is the foul smell you noticed.
RR


--------------------
Download Let's Grow Mushrooms



semper in excretia sumus solim profundum variat

"I've never had a failed experiment.  I've only discovered 10,000 methods which do not work."
Thomas Edison

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Offlineiamgod
just some dudesome where

Registered: 06/30/03
Posts: 263
Loc: Crooklyn, New York
Last seen: 16 years, 24 days
Re: sexy red contamination [Re: RogerRabbit]
    #4843385 - 10/23/05 09:14 PM (18 years, 5 months ago)

Like RogerRabbit said, you want to keep your substrate airy. The more compact and wet it is, the harder it will be to colonize.


--------------------
If what you seek is truth then drugs can not offer truth. Drugs can offer the truth of drugs.......Altered states. Truth is not an altered state of mind. ~ Big Headed Nice Guy Who Loves You And Your Dog

http://www.inlibertyandfreedom.com/Flash/Think_It_Over.swf

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OfflineLacrossa
Cmdr of PiratePenguins

Registered: 09/20/05
Posts: 21
Loc: Mormonia
Last seen: 17 years, 3 months
Re: sexy red contamination [Re: RogerRabbit]
    #4844303 - 10/24/05 12:32 AM (18 years, 5 months ago)

Got it, will be much more careful of following the recipe exactly. The logic was that more water would be more helpful to the mycelium, but it looks like it's not that simple.

How much protection do those filter discs give? Once inoc'd, the filter discs were placed back on, and the jars were moved into the TiT but were exposed to open air (moving jars + filter physically to the tub). Is there a danger when you're fanning out the TiT?

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OfflineRogerRabbitM
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Re: sexy red contamination [Re: Lacrossa]
    #4844466 - 10/24/05 01:48 AM (18 years, 5 months ago)

No. In fact, the synthetic filter disks are not necessary with the dry vermiculite barrier in a pf jar. I'd dump the TiT and just incubate on an open shelf. If it's cold in your room, just run a small space heater. Jars will colonize just fine at room temp. It may take a few days longer than at 80, but contaminants will be lower. I haven't used an incubator in years. There is also no reason to worry about keeping them in the dark as they colonize. A few early pins won't hurt a thing. Birth a few days after 100% colonization.
RR


--------------------
Download Let's Grow Mushrooms



semper in excretia sumus solim profundum variat

"I've never had a failed experiment.  I've only discovered 10,000 methods which do not work."
Thomas Edison

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Offlinehyphae
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Re: sexy red contamination [Re: Lacrossa]
    #4892645 - 11/04/05 05:59 AM (18 years, 4 months ago)

I found Jars colonize quicker in total darkness not only that but it's just good mycosense to keep jars away from open sporeloads. You can put the jars back into the box they come in which works nicely. Also It's to your benefit to incubate at optimum temps not just because they will colonate faster but you'll also utilize the pinning trigger "temp drop" and by keeping them in the dark you can utilize another pinning trigger "light" now we all know neither is totally necessary but we also know they are pinning triggers as well as lowereing CO2 concentrations and full substrate colonization, using all these triggers in conjuction (at the same time) will give you better pinsets consistantly, so it's up to you, you can be just fine or you can go for those killer pinsets but remember parameters are there for a reason (YIELD!) :wink: GL


--------------------
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"We all have priorities. I used a closet once setup a nice little lab trouble was all the shit that was in there ended up in the bedroom that pissed off the GF then I ended up dumping her as she was getting in the way of my sterile culture technique! Ya I got priorities too!!!"

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