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Solipsis
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Ideas on harvesting Metarhizium mold spores 2
#26334899 - 11/21/19 03:38 AM (4 years, 2 months ago) |
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Harvesting mold? Who would want to do that?
Metarhizium brunneum is a biological pesticide I want to investigate and use, I have it on agar and am seeing the first sporulation developing.
To not have a big sporeload blown through my lab, I guess I should inoculate something with a not yet sporulating agar inoculant.
Does anyone have tips on how to grow and collect the most spores in the cleanest way? For all us "Trich farmers", now might be a time to use our experience for good.. xD
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DutchMyco
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Re: Ideas on harvesting Metarhizium mold spores [Re: Solipsis]
#26341992 - 11/24/19 09:21 AM (4 years, 2 months ago) |
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Nice experiment!
Depending on how much you want or need, something like this might work for a home type lab. Make or find shallow plates/jars with a lid with injection port. Inoculate with you culture and wait till you'll have enough sporulation, then inject with sterile water (possibly saline or with Tween 80)and flood the plate. Then suck up the spore solution, trough a cotton/glaswool filter to filter out as much of the hyphal pieces as possible. Might be able to centrifuge your spores if you want to dry them, or keep suspended if you can keep it from germinating.
There was a topic not that long before here about using static to get clean spores from a wild mushroom, might work as well with a plate full of mold, might also flood your entire lab with spores though.
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Heruuka
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Re: Ideas on harvesting Metarhizium mold spores [Re: Solipsis] 1
#26342090 - 11/24/19 10:21 AM (4 years, 2 months ago) |
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How about collecting what's on the lid of the plate (assuming you're incubating upside down)? can we get a picture to know what we're dealing with?
How much are you wanting to recover, a mg? 10 mg? 100 mg?
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Solipsis
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Re: Ideas on harvesting Metarhizium mold spores [Re: Heruuka]
#26356746 - 12/02/19 06:33 AM (4 years, 2 months ago) |
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Thanks!
Ah yes, idk what the saline does exactly but i have read about it being used in research for spore suspension and tween too. (I always wonder how soft green soap compares to the tweens, but hey i have tween-80 so..)
The injection of water etc sounds a bit like an LI method and i guess it could work. Just not sure what the more efficient way is of converting , say a 350 mL filterbox with grains, into as many spores as possible. But I guess it's just a matter of time and it will basically become more and more spores until its consumed right?
I also don't know how many spores are in a mg... trying to find this info now. [10 mg = ~ 4.7 × 108 spores]
I see synthetic oil is used if one wants to have the pesticide float on water against e.g. mosquitos, but i would want to use it applied to soil of potted plants and experiment with it in a mushroom grow. And actually should set up a proper experiment to see if Metarhizium can easily become a contam, maybe consuming metabolite exudate off of your myc? Not sure how something like Trich gets a hold exactly?
Static is also a cool idea lol. But would definitely still try to applied in a confined space.
 
i do not incubate upside down, I don't have condensation the effect of which is avoided that way, I'd say.
Collecting what is on the lid is an interesting idea but it seems less practical than getting them into suspension right away without opening, even or esp in a flowhood.
It would be cool to know what suppresses germination of the spores until the suspension is applied or maybe its not really necessary stored at 3C.
Say you would want to apply MET for mushroom cultivation, I think Bti is sprayed on myc blocks but maybe it would also be cool to make a gnat trap by attracting with some smell they really like best (involving wet soil i guess) ambushing with MET.. However wet soil might complicate things if it starts growing other molds.. :\ I wish there was an easy way with a smell that gets gnats extremely excited that can be used pure without having it made in vivo by soil organisms...
Hopefully instead its enough to have a quite limited amount of sporeload in the air of your growroom if it is first tested not to be a significant contam threat.
Edited by Solipsis (12/02/19 06:47 AM)
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Jawn876

Registered: 09/12/18
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Re: Ideas on harvesting Metarhizium mold spores [Re: Solipsis]
#26361757 - 12/04/19 08:04 PM (4 years, 2 months ago) |
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If youre looking for an attractive smell for insects, you might be able to use other fungi. Lots of fungi produce octenol (https://en.wikipedia.org/wiki/1-Octen-3-ol) I think Tradd Cotter has some idea using fungal produced octenol for a mosquito trap. I dunno if that helps.
MET works well too. I was once living in a van that got infested with fire ants. I put some M. anisopliae colonized grains on some concentrated spots in the van and within a week I was ant-free. I lived in there and had all my food in there so bombing them out with poison wasn't an option.
I love your work btw. you have a great thought process
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Solipsis
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Re: Ideas on harvesting Metarhizium mold spores [Re: Jawn876]
#26370212 - 12/09/19 05:01 AM (4 years, 2 months ago) |
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Hmm seems like octenol does attract mosquitos and such, but repels gnats:
https://bioone.org/journalArticle/Download?fullDOI=10.1603%2FEC11066
I'm curious if MET's odor attracts or repels gnats and what strategies may have evolved between them. I'm very interested in MET, but from what I understand tolerance/resistance is developed in some insects... might mean a dual pesticide is best but we'll see later.. 
It would just be good to have either a synthetic odor in small quantities (slow release) or use some sort of mold or other fungus that is basically harmless but does attract the gnats. In general i would prefer to not use actual damp soil or random molds when a grow of edibles is involved haha.
Thanks very much.. =D Thats cool about the MET and ants (i didnt know they could be targeted as well i thought MET was meant to be more selective in what insects it kills).. Beauvaria sounds good for ants as well, I knew ppl that developed products of it.
Just using colonized grains seems interesting too and i have some with met that are ready about now.. Ideally though i would try to apply the spores to soil to have it more contained, but i guess the insects can infect each other anyway? Idk if the spores last long in soil cause from what I understand Bti doesn't really last in moist soil.
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Jawn876

Registered: 09/12/18
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Re: Ideas on harvesting Metarhizium mold spores [Re: Solipsis]
#26376050 - 12/11/19 08:40 PM (4 years, 2 months ago) |
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Oh right on. I was under the impression that octenol is what attracted gnats into the gills of oysters and such. Thanks for the correction. Do you know what chemical(s) make oyster mushrooms so attractive to them?
Interesting, I wonder if I have a mislabeled culture. Mine is more purpleish instead of greenish. The mycelium has the same form though. The friend I obtained the culture from said it was supposed to be the MET 52 strain but now I am uncertain. Perhaps another Metarhizium sp? Do you know how to identify it? or must it be sequenced?
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Solipsis
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Re: Ideas on harvesting Metarhizium mold spores [Re: Jawn876]
#26382467 - 12/15/19 12:08 AM (4 years, 2 months ago) |
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I tried to find the conidia and spores under microscope.. not sure if i checked all magnification but a friend and I were unable to ID what i have or at least what ended up growing on my grains. I should check again from agar plate just in case the grain contaminated with another mold lol.
I think i have a photo from the microscope but im a bit lazy and tired now. Basically the spore shape of what i saw is off.
On agar it looked OK and like it should, it first went yellow as you can see in the left pic - and then green which is not shown and the pic on the right is not mine. I tried to infect a little bug but both the grain and the bug escaped while i was away lol.
Soo yeah i will check with a microscope directly from agar and will try again to infect a bug or a number of bugs.. this time i def should infect a species i know it should kill or at least i want to use it for so trying to get some gnats going in a jar..
Its kinda unclear to me which species this strain should be able to kill exactly, is there a list?
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Jawn876

Registered: 09/12/18
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Re: Ideas on harvesting Metarhizium mold spores [Re: Solipsis]
#26401005 - 12/25/19 09:56 PM (4 years, 1 month ago) |
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https://www3.epa.gov/pesticides/chem_search/reg_actions/registration/fs_PC-029056_18-Apr-11.pdf This is the US EPA's sheet on it, "Metarhizium anisopliae strain F52 can infect larvae and adults of many insects, but is labeled for use on mites, thrips, ticks, whiteflies, and weevils" also "Submitted studies show that Metarhizium anisopliae strain F52 is not harmful to earthworms or to such beneficial insects as lady beetles, green lacewings, parasitic wasps, honey bee larvae, and honey bee adults." However no reference for 'submitted studies'
https://en.wikipedia.org/wiki/Metarhizium_brunneum
"M. brunneum has been isolated from Coleoptera, Lepidoptera, Diptera and soil samples, but a commercially developed isolate (below) has proved virulent against Hemiptera and Thysanoptera." -didn't realize This species used to be M. anisopliae, makes me really wonder what I have...
Slide 18 on this presentation as well: https://www.abim.ch/fileadmin/abim/documents/presentations2008/session5/4_Pradier_ABIM-2008.pdf apparently works on ticks and mites too which aren't insects but arachnids
I'm not sure a list exists that is entirely comprehensive and detailed as to what it can/can't kill but it seems to be pretty generalist. Any progress on your culture?
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Shroomei
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Re: Ideas on harvesting Metarhizium mold spores [Re: Solipsis]
#26401478 - 12/26/19 11:10 AM (4 years, 1 month ago) |
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I have to admit that I often have clean agar plates with green mold. The spores seem to be pretty hydrophobic. Without surfactanct there will be no suspension. Prepare to play around with Tween.
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Solipsis
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Re: Ideas on harvesting Metarhizium mold spores [Re: Shroomei]
#26414029 - 01/03/20 10:10 AM (4 years, 1 month ago) |
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Well not really progress yet.. i will soon work together again with a friend and check spores from agar and cross-check them because what i received definitely looks like the real deal, its just the spores taken from the spawn that look off.
In the meanwhile i will also be getting Beauveria spores, idk if those will help with spider mites but we'll see.
shroomei: definitely, i think by far most spores will prove to be hydrophobic and it makes sense?
in another thread someone said to avoid tween for spore suspensions and germination but idk why.. i will need to find a surfactant which does bring them into suspension but does not permanently disable the spores from germination somehow.
I have ideas about how and why but hmm we'll see. There will be experimentation either way.
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Shroomei
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Re: Ideas on harvesting Metarhizium mold spores [Re: Solipsis]
#26416096 - 01/04/20 02:11 PM (4 years, 1 month ago) |
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Quote:
Solipsis said: ....
shroomei: definitely, i think by far most spores will prove to be hydrophobic and it makes sense?
in another thread someone said to avoid tween for spore suspensions and germination but idk why.. i will need to find a surfactant which does bring them into suspension but does not permanently disable the spores from germination somehow.
I have ideas about how and why but hmm we'll see. There will be experimentation either way.
The Tweens are really the favorite surfactant in biology and pharmaceutics because they often do not harm biological function. However there are publications that they will impair germination of some spores while others are completely fine with it. There is only trial and error.
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LizardWizard
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Re: Ideas on harvesting Metarhizium mold spores [Re: Shroomei]
#26416908 - 01/05/20 12:39 AM (4 years, 1 month ago) |
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IIRC, Beauveria Bassiana stays active in soil for a few months, having a half-life in soil of about 6 weeks.
Dry collection of spores seems to be the most efficient way to me, since moisture will likely trigger germination.
Storage, accoring to the packets in my fridge, should be done cool, since room temp storage will keep things fresh for about a year, while at 4°C it will remain viable for two years.
IME, both Beauveria Bassiana and Metarhizium Anisopliae have good but not total killing action on susceptible pests. It works better when combined with beneficial predatory insects. That being said, controlling populations into seeing about or less than 1 gnat a week / m² is definitely possible, and at that rate the pest won't cause significant harm.
Surfactants are not needed to mix spores into water. Wetting the spores with a small amount of water and mixing it in by hand to create a paste, you can mix the spore paste into a larger volume of water. A common application rate is 1g/liter; to eradicate gnats one should treat both soil and live gnats with this mixture (all gnats should be hit with spray droplets), also, all other suitable environments for the gnats (potted plants, bags of soil, ...) should also be treated to avoid re-infestation.
-------------------- The best things in life can be smelled on one's fingers.
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