ARGHHH why so hard ???

[Guest electro]

B*****y H**l

the gnome down the road wants to know why he finds it so difficult to get mushies growing ....

He can grow anything else he wants .. but mushies try as he might he cant get right.

he would love to get a whole bunch of edibles growing for both himself and possibly as a side business (supplying restaraunts in the area n such) ....

he can get an entire jar colonised in just on 1 week, with brilliant white mycellium... but always something has to spoil it ...

either it smells slightly off & gets thrown out or tiny purple dots appear & it gets exctracted & the extracxt thrown away ... it is theory when contams are involved no eating stuff possibly detrimental to health.

It has been almost a 6 months the bl##dy bluegreen mould epidemic (strangely the absence of which coincides with the purchase of a pressurecooker), and so many jars Look perfect (& get declared a success before the chickens hatch).

- hence ever changing contamination rates noted on the site by electro.... the things look perfect.. sometimes even smell perfect .. but when they are broken up to be cased they dont quite smell like brillianbt mushies/ have a slight bacterial smell, have slight pink colouration (which could well be something in verm), have slight green patches (which could well just be the greenish edge of medium brown rice)....

could well be is not good enough though - especially when health is involved ...

the only mushies that have been grown are from others jars ... lol he can fruit them & grow very agressive happy myc - the myc just smells of it's co inhabitant.

Even liquid culture looks & smells fine./... until you look really close at the 3rd generation culture where you see slight purple dots beginning to appear ... then in the 4th gen where there is much more purple (especially when you heat it to 29c instead of 27 ... hrmmmm)...

how the hell do you manage to contam a liquid jar by when you are sucking liquid from one (through aluminium lid covered with metho) and injecting into another jar through another aluminium lid covered in metho ! ???

Is agar a whole bunch easier to keep sterile or something >?>>i assume it is solid & therefor able to have contam ( or rather the healthy mycellium) cut from it ????

Rev if ff is good to go does it have any agar equipment handy (scalpels, petri dishes with lids .. a small amount of agar & whatever u need to mix with it to get one started ???)

Is a glove box what the gnome down the road needs to get past the contam problems ???

I know he needs new liquid (or agar ) culture to start with .,.. pink mould is there... but to keep sterile.

He has asked me to put up a wanted list for new supplies:

ff- Agar

ff -Agar (petri) dishes

ff - filter disks

Enoki Culture

Oyster Culture

Shittake Culture/spore print

Field Mushroom (the cultivated variety) Culture/spore print

Help needed:

If anyone knows of any mycology workshops around the place, let me know please

also if anyone has a link for decent glove box (one that works .. not sifting through 30 different ideas, 28 of which wont work...)

any ideas for re visiting sterile procedure all infront of an oven that is on (though it is fan forced, which may be just blowing contams in ....

Thanks all for listening to this whinge, and thankyou even more if you can offer any ideas that might help ...

[smogs]

hmm so theres no contams until you case it?

and u cant find any contams in jars or liquid culture?

obviously something is going wrong with the casing.. what casing materials and methods have you tryed? have you tryed a less laboritorial apporoach and more natural ie mix jar with cowshit and straw and put it outside and water well in shade (un-stearalised)??

[mr b.caapi]

it sounds like ,if there is "brilliant white mycellium" filling the canister and then contaminants start then there is a problem with the sealing of the canister....

ive noticed that if a canister is picked up alot while it is colonising,it increases the risk of contaminants.

just a shot in the dark as im no expert..

[ 01. March 2004, 20:07: Message edited by: mr b.caapi ]

[Guest electro]

-->[QB] hmm so theres no contams until you case it?

No, there are contam in the jar that only show the day they are scheduled for opening

all looks perfect.. go to open.. find a 1mm spot of pink in every single jar as they are being examined prior to casing.

either that or all looks fine, then the lid is opened & a slight (but definatley present) bacterial smell wofts out rather than a pungent mushie smell...

--->u cant find any contams in jars or liquid culture?

Just did in the lc ... little spots that looked like dirt ... i still cant figure out how they got in there .. it was in sealed spice jars (half full to use internal air). the lid was tipped ever so slightly to allow injection inb or out.

obviously something is going wrong with the casing.. what casing materials and methods have you tryed?

50/50 actually seems to make the slight bacterial smells regress, but it helpt the pink dot mould (that appears a little later on)

Tried innoculating dung with the only non contaminated 1.5 kg jar of grain .... it got too wet at the bottom & rotted. The colonised straw was moved outside and was quickly eaten by snails (there one day, slug trails and nothing else the next).

re picking up the canister - they get innoculated then sit in their hot box (fanned through a filter) for a week then are checked...

Also filter discs have no tbee unsed .. cotton stretched over tiny opennings has been tried, as have thick layers or verm to protect .... contam still gets in.

I think the main prob of the gnome downb the road is using grain that it too wet & not cooking it long enough .. but with the liquid culture infected now it makes it a very big pain ....

i think he needs to learn to agar culturing (also does agar culture last longer in the fridge ?)

[ 02. March 2004, 05:36: Message edited by: electro ]

[mescalito]

I'm no expert....but I would be going back to a spore syringe and go from there.

There is either a colour from the lid/seal or vermiculite showing up,or the shroom food is not sterilised fully.

[Guest electro]

yea, i think culture is where it is falling down..

will try agar when i can get some petri dishes & test tubes from rev

[spiders]

When i started everythign always contaminated as well. I just had to become absolutely paranoid with sterility.

I made a glove box which i spray with a mist of h202 or peroxide before use. I sterilise everything with rubbing alcohol. I pc everything.

The thing that causes contamiantion in 9 out of 10 circumstances is air contact. You dont want any spore print to be exposed to air.

use a small pc'd jar with a hole in the lid. In a glove box, scraped some spores into the jar, seal and then insert your sterile syringes in the hole in the lid. Do everything in a glove box!!!

[mescalito]

Anyone gotta link to easy glove box plans on the net?

[visualfx]

http://interneurone.net:443/gallery/upload...ad/BENQ0008.JPG

i think this image speaks for its self...

[mesq]

*looks at the glovebox*

*ahem* :D

Looks familiar.

[mesq]

Electro...

You didnt respond to your mail...

Umm

Can you email me with something other than your telstra one... and I can help you... I think you should stop throwing things away sometimes and see what happens... by the way... you can also save mycelium from a cake by agar culture..

[Guest electro]

sorry mesq, that account gets around 200 emails a day (all spam) so i try not to check it too regularly.

Also re the glove box ... so a fan with a filter is not necessary ?

Is 3% peroxide enough, or is 40% recomended for use in glove boxes for killing spores ?

[ 03. March 2004, 16:36: Message edited by: electro ]

[visualfx]

Mesqualero:

*looks at the glovebox*

*ahem* :D

Looks familiar.

just passing on info...not claiming credit!

[smogs]

if using a spray to kill stuff in a glove box wouldnt it be betetr to use something that kills bacteria as well? ie 10% bleach solution or lysol spray.

[Bolwarra]

I thought bleach was Peroxide? And that a 3% solution (as electro said) would be enough to kill mold spores in a substrate? But wouldn't effect already established contams.

[occidentalis]

Normal laundry/household bleach is usually sodium hypochlorite and hydroxide. Hair bleach is peroxide. I'd go for an ethanol spray for a glove box, such as the horribly perfumed glen 20.

[Beaker]

electro:

sorry mesq, that account gets around 200 emails a day (all spam) so i try not to check it too regularly.

Also re the glove box ... so a fan with a filter is not necessary ?

Is 3% peroxide enough, or is 40% recomended for use in glove boxes for killing spores ?

I wouldn't bother with a filter and fan.. had no problems without it..

As for preparing the glovebox... I firstly wash out the inside with a strong bleach solution thouroughly... I then seal it with all of the goodies inside and leave it for about 15mins..

I then put my hands in, grab the can of Glen20 (which I now can't stand the smell of.... think I'll go 'country fresh' ) and spray everything until it is a little foggy.... wait about 20 secs and start work...(unless it's with a print... then I wait a little longer)

I wouldn't use peroxide because its too expensive to be using all the time... White King hasn't let me down so far...

anyhoo...some of you might think that this is a bit of overkill, but that's what I do... and it works.. no contams... ever..

Cheers

Beaker

[ 04. March 2004, 11:03: Message edited by: Beaker ]

[Rev]

Sounds like you have dirty innoculum or your p/c isnt being worked hard enough.

Getting clean inoculum is the biggest first hurdle

If you have crytic contamination - hiding amongst the seemingly healthy mycelium you ar doomed from the beginnings.

I find cultures most often grow too fast

To slow down either add a little too much peroxide or cut the mix to a 1/4 or 10%.

Every now and then i transfer the wedge to underneath another wedge and let the mycelium grow through leaving any crud behind. This is particularly important if you want liquid myc.

I like to take a syringe full of sterile water and just lifting the plate enought to pass the needle over the mycelium start scrapig the sufarce , squirt a little water on and such it up, repeat if necessary. Test it and if clean use to innoculate honey water. Draw it all up, test agin, go to grain. Other option is get one good jar then inject sterile water scrap n suck just the same and repeat.... instant mycelial water.

Its important to test you innoculum at each step before moving on. Beacuse this is time consuming make as much in one sitiing as you can - it keeps well at room temp out of direct sunlight.

Also there seems to be an inverse relationship between contam and aeration. The more oxygen in the mix the less it contams. Another reason i like polyfill

Really hot weather can cause an outbreak of yellowish mycelium and black pin moulds (possibly the same thing)

Agar can make things very much easier especially in conjunction with Peroxide. Before i bought my laminar flow i never had a glovebox, i did my inoculations in the kitchen on top of the chest freezer. Sure i had contam but mostly via improper sterilisation of shitty grain

I used agar to grain but you may very well use peroxide agar to peroxide grain.

You make agar as usual and grain as usual (these days i use polyfill jars). You ad 1 to 3 mL of 3% peroxide per litre of agar just immediately prior to pouring.

And you add Peroxide to the grain spawn after sterilisation as well. Just a bit - the exact amount escapes me - but you just want to get a bit in there and mix it about thoroughly. It protects the grain from any odd stray spore tha gets in

I used the double wedge transfer technique. 9 out of 10 worked

These days is run my experiments on polyfill jars with mycosyringes done under the flowhood. Most time is just inoculate in the open though on the benchtop with just a lighter or spiritlamp to flame the tip. o/wise i have to fire up the laminar and let it run 20 mins before using.

Contam is very low and almost always traceable to the parent mycoculture or shit grain.

The other thing i do is after a long time of presoaking grain and bleaching before cooking in a suacepan etc. now i just put cook it in a rice cooker at teh ratio for the grain and walk away

Perfect

the prolonged cooking kills off most of the contam and the set water grain ratio stops it from splitting

Ryegrass, milllet, wheat, rice , sorghum and mixes - its all good. Rice cookers are the shit

$25 if you look in the right place. Add gypsum to stop grains clumping

Electro get to know your contaminants.

Id eat shrooms with a touch of vertcillium or trich no problems. Its other things you want to worry about.

Often i just dig out half contaminated jars and mix the good bits in with mulch, often the beasties outddors, the rain and sunshine clean it up and away it goes. Never gotten sick of these mushrooms. Also Outdoor fruited mushrooms can actually give cleaner prints than many indoors. The UV, rain and fresh air dont seem to cause the proliferation of blue and green moulds you get with humanity.

FF has got

oyster, shimeji, luminescent panellus, king stroph and shiitake mycosyringes @ 12 each

cultures of thes plus enoki for $27.50 (in duplicate)

Im not selling buttons for the moment cos the are so damn easy to clone and so accessible - try it out. dont be worried it the explants bruise brown , they often refuzz in a bout a week.

Agar, plates etc no problem talk to me at

[email protected]

Filter patches? till i find a source for Tyvek and try it out im sticking with polyfill

www.fungifun.com

When i find the right source FF will

get some filter patches in.

[ 01. April 2004, 19:33: Message edited by: reville ]

[Guest electro]

Rev:

Firstly, congrats on the bub... i hope life is treating all three of you guys well

secondly, I am playing with agar at the moment, it seems very cool... so much easier to spot contams (only 1 out of 6 plates contamed, and it was the one with a gaping hole in the alfoil lid - i wonder where that contam came from lol).

I like the idea about forcing myc to grow through from the bottom to the top of new plates - i take it the most agressive myc gets to the top first, where you sample it with a blade or biopsy punch & reclone onto new plates ? PS where do you get biopsy punches - im sure they woukld be good for cloning.

Re contam record & this thread - My prior contams i am sure came from my dodgy attempts to make dextrose liquid culture - alfoil lids that sit ontop of jars rather than being clamped closed while sitting in a grubby bacteria ridden fridge were a recipee for disaster - lol and all my eggs got placed into that ione basket to make more...

ALso a glovebox used in conjunction with agar seems to be working wonders, though time will tell with the first rice cake innoculated with a slice of colonised agar yeaterday

(whoever told me to turn down the heat once the pc started hissing - thankyou ... the noise is so much more tolerable now and it is no longer running out of water ... the things you miss when your pc is bought from the markets and instructions are written in chinese lol.)

I have tracked down most everything i (think) i need for my experiments except for more edible cultures and some screwcap test tubes (to make slants from)

I am definatley up for some shiitake mycosyringes @ 11 each aswell as an enoki culture for $27.50 (in duplicate)

Pm me with your/ff bank account details & i will direct transfer the money for the cultures (and test tubes if you have any) when i get paid wednesday week.

re " im sticking with polyfill"

Is polyfill really any safer than sterlilised verm layers (sandwidched between bleached alfoil layers) ontop of the jar ? or is it just more ecenomical (also are you filling metal tubing with this & welding into holes in lids or are you just placing some in holes in lids ?)

[Rev]

You may have to experiment with the rice

on its own it may lack the desired structure though it will be functional.

Open structured and hard items like Perlite,Verm, rye grass seed, millet all seem like good experimental mixes

Im testing some myc on rye grass cakes to see if my suspiscionsare supported (that ill make good cakes)

millet does but has a slighly higher contam rate than rye grass seed

I have tracked down most everything i (think) i need for my experiments except for more edible cultures and some screwcap test tubes (to make slants from)

I am definatley up for some shiitake mycosyringes

ready after next week, we have shimeji now. please see elm oyster post to see the shrooms

Enoki and other cultures for $27.50 (in duplicate)

these are made to order so please give at least 3 weeks delivery time (3 to 5 weeks depending on species)

Pm me with your/ff bank account details

check your PM

re Is polyfill really any safer than sterlilised verm layers (sandwidched between bleached alfoil layers) ontop of the jar ?

that sound too complex for simple minds like me its the simplicity, the economy and its dual nature as injection port AND filter patchg that atracts me. The results are very good

contam is down colonisation rates are up

im happy

(also are you filling metal tubing with this & welding into holes in lids or are you just placing some in holes in lids ?)

que? again thats all too hard

Its a hammer, a hole punch and a metal lid

or/

A drill, and a plastic lid

hole goes in, polyfill gets threaded though the hole..done (and reusable)

www.fungifun.com

[ 01. April 2004, 19:57: Message edited by: reville ]

[smogs]

polyfill is mianly for when your using whole grain substraight... so you can leave a big gap at the top and u can shake it up to mix it up.

speeds it up heaps!

[mesq]

 

quote:

---

Originally posted by electro:

 

[QB]

 

Re contam record & this thread - My prior contams i am sure came from my dodgy attempts to make dextrose liquid culture - alfoil lids that sit ontop of jars rather than being clamped closed while sitting in a grubby bacteria ridden fridge were a recipee for disaster

 

---

I used Band aids over the air exchange hole and alfoi on top for my Liquid mycelium jars they withstand PCing and form a sort of filter..

Liquid mycelium is really easy to contaminate too...as you no doubt already know..

[sakura]

I have very few probs with my liquid cultures...

I always use a little h2o2 in the solution if I'm starting from tissue culture or myc and have a 0% failure rate with these (I also dunk the tissue culture in 3% h2o2 for a few secs before I introduce it to the jar).

If I'm starting from spore, I don't use syringes, but scrape a little into each jar with a sterile scalpel using oven tek (figure if I have a contam in the print and I make a syringe I'll stuff the whole lot up, but if I use spore I have a good chance of success in at least one of the jars...).