Hey, so I had a big spawn run and all of my bulk substrates got a bacterial contamination, or so it seems, sour smelling, almost chemically, no signs of visual contamination. I noticed the smell around day 3 of colonization. I'll explain my processes, and then I have a few questions. All of my agar work is done in front of a flow hood, as far as dishes getting contaminated I have a less that 2% contamination rate, so I highly doubt that has anything to do with it.
GRAIN PREP
So I fill up my cooker with wheat kernels, fill with hot water, scoop floaters, drain and fill 3 timesish until the water is mostly clear. Then I soak in cold water for 12-18 hours, then I cook until hydrated and spin them dry before bagging, like dry, dry, and completely cooled. Then I bag the grains 5lbs per 14a filter bag, I pre seal them before sterilizing. Then I load them in my sterilizer, get them up to 180f-190f and cook them for about 18 hours. Unload, let them fully cool, inoculate each bag with a ( non iso, BRF cloned culture on the 8th transfer currently) full agar dish cut into wedges. They are fully colonized by day 14.
SUBSTRATE PREP
I sterilizer my bulk substrate in a sort of modified insulated bucket Tek style bin, I boil water in the bin, wait until it's over 200f, dump in my bulk ingredients (780 grams coco + 225 grams verm + a handful of gypsum + a little more than 3 litres of water) let it sit for about 2 hours, mix, let it sit for another 2 hours or so, then mix again. The bin stays above 180f for like 12 hours, I usually have to keep mixing and waiting for 2-3 days before the substrate is back under 100F.
SPAWNING PROCEDURE
I mix my substrate and spawn using a mixer just because I have it and it's easier. Before I spawn to bulk I clean the mixer with vinegar, scrub, dry, and then H2O2, scrub and dry. I open air mix my substrate with the spawn, I keep the windows closed, but it's just regular household air, no filtration. I use 9-10lbs substrate per 5lb bag of spawn, which works out to about a 1:3-1:2 volumetric spawn to sub ratio. I put the spawned substrate into lined trays (24"L * 20"W * 3"D)which I then put inside a contractors garbage bags And no tie them closed, normally I cut slits in the bags and put 2 layers micropore over them. I let them colonize at 70f-75f they're almost always done by day 10-12, but I let them sit for about 4-5 days before I troducing to fruiting conditions.
I've had 4 successful runs with my current setup, no major contamination issues, lost a few substrates to trich, but less than 10%. This last run I did two things differently that I can think of.
1. I didn't slit the bags and micropore them, I only loosely tied the bags.
2. I let the grain bags sits fully colonized for two weeks before I spawned to bulk because of issues with my fruiting chamber.
So my questions are
1. Could bacteria be growing because my culture is just getting old, or is a 7th transfer from a clone not old enough for that to be happening?
2. Could the bacteria be growing because I didn't put GE holes even tho I loosely tied the bags and there was still probably a 1/2" Gap at the end.
3. Could the fully colonized bags sitting for two weeks be the culprit for bacterial growth?
I'm sure I can figure this out with a few more bulk runs, but if my culture is getting old, if rather just use a new culture than to waste my time and money on more grain bags and substrate. Thanks in advance for any answers!
Edited by 3SIXTY5 (08/30/20 12:10 PM)
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I don't use grain jars, I use 14a unicorn bags, I think you're confused.
it's atmospheric pressure sterilization, you don't need to use a PC to sterilizer grains, I have an AA41 PC, but when you're doing bulk grains the PCs aren't realistic. look up a drum sterilizer if you haven't heard of this before. The bags are pre sealed prior to going in the sterilizer I cook them for 18 hours and start the timer once the grains have reached 180F.
I just read Kizzles guide on identifying contamination and read that a lack of GE creates an anaerobic environment which is a breeding ground for bacteria, so I'm thinking that's probably my issue.
I'm more worried about an aging culture though, because that happened to me a year or two ago.
Thanks for your response, I've gotten enough views on the thread but only one response hopefully some other mfers chime in.
Edited by 3SIXTY5 (08/30/20 03:03 PM)
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