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InvisibleJoeMama1992
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Registered: 12/27/11
Posts: 901
Wild-type Claviceps paspali productivity
    #27881953 - 07/29/22 04:15 PM (1 year, 9 months ago)

I've been trying to find an answer to this question for years, but it never dawned on me to ask it at the mycology board until now :crazy2:

Basically, I just wanna know (and would love to see refs) what the generalized average alkaloid/peptide etc content is of Claviceps paspali found in the wild.

Am aware of UV mutation and all that, and people shooting for 2+g/liter strains, but anything to expedite the process from would-be seedhead to submerged culture is an enormous benefit to any and all interested in the concept.

Bearing in mind the fact that there will certainly be some non-producers and what will amount to "may as well be" non-producers:

What would those informed on the subject estimate the realistic yield of desirables be in g/liter of a freshly harvested and cleaned, non-isolate, mixed-strain submerged culture of C. paspali be?
0.25g-0.50g/liter too hopeful? Is 0g/liter the only realistic answer to this?


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InvisibleCreonAntigone
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Registered: 05/30/21
Posts: 2,971
Re: Wild-type Claviceps paspali productivity [Re: JoeMama1992]
    #27881993 - 07/29/22 04:42 PM (1 year, 9 months ago)

Impossible to predict because it is all based on strain differences, however we can get some general guesses by looking at other studies. You could hit on a lucky break and get a very good alkaloid producer right away, or one that is almost impotent, with a huge variation by region, etc.

You could probably quickly identify the most promising isolates yourself with a bit of chemistry. Probably the best approach would be brute force, to start many different strains and try to get a lucky break.

Be aware that they don't produce only lysergic alkaloids, C. Paspali is known to produce several indole tremorgenic alkaloids such as Paspaline, which are the truly toxic principle found in this ergot, as opposed to the alkaloids which tend to be the less toxic one (not being the ergopeptines but the water soluble alkaloids such as LSA).

Also, it will WILDLY vary by media type even in the same strain.

Quote:

0.25g-0.50g/liter too hopeful? Is 0g/liter the only realistic answer to this?





Let's take a look at the studies.

"Quantitative changes of the alkaloid complex in a submerged culture of Claviceps paspali".
Quote:

"A two-week fermentation in a synthetic medium with mannitol at 24 ± 1 °C resulted in an increased production of total alkaloids from the original value of 100–200 μg/mL to more than 2000 μg/mL."






2000 ug/ml at the high end. Here's a study showing the impact of mannitol from their culture, which was multi-strain and not an isolate. That was only when they perfected their media - by the way, might want to try and copy what they did - but early results were 100-500micrograms/ml.

Stevens and Hall is the famous strain people refer to, and that didn't perform much better. Looking it up, they tested the effects of different temperatures and media - some as low as 65ug/ml, but most were around 600-1300ug/ml.

My guess is, 500-1500micrograms per ml - but early attempts may be low, 50micrograms or less, and you'll need to improve and adjust the media.

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InvisibleJoeMama1992
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Registered: 12/27/11
Posts: 901
Re: Wild-type Claviceps paspali productivity [Re: CreonAntigone]
    #27882024 - 07/29/22 05:04 PM (1 year, 9 months ago)

Man, I hate to be typing this after the amount of work put into that post, but the curiosity here is with the use of mixed wild-type genetics, without isolation of a single strain. Sort of the paspali equivalent to a multispore inoculation of cubensis. The hope here is that the abundance of genetic diversity would almost negate the likelihood of non-producer genetics.

Still, it's likely that a focus towards single-strain experimentation is the ultimate outcome for all.

Edited by JoeMama1992 (07/29/22 05:07 PM)

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InvisibleCreonAntigone
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Re: Wild-type Claviceps paspali productivity [Re: JoeMama1992]
    #27882094 - 07/29/22 05:51 PM (1 year, 9 months ago)

Quote:

JoeMama1992 said:
Man, I hate to be typing this after the amount of work put into that post, but the curiosity here is with the use of mixed wild-type genetics, without isolation of a single strain. Sort of the paspali equivalent to a multispore inoculation of cubensis. The hope here is that the abundance of genetic diversity would almost negate the likelihood of non-producer genetics.

Still, it's likely that a focus towards single-strain experimentation is the ultimate outcome for all.




The problem here is you're trying to compare apples to oranges, claviceps are not mushrooms and not even in the same family of life, they are ascomycotes while psilocybe are basidiomycotes.

It seems logical to me that eventually you will have to isolate a strain. If you introduce multiple strains to a single media, they will eventually go together, and some strains will die off or merge with the others.

You can go ahead and try 'multispore' by grinding up sclerotia from multiple sources, which will contain a huge number of conidia, those will germinate on agar, maybe you will get a higher number of alkaloids, go ahead and try. The ascospores will not germinate on agar so don't bother with those (hard to locate anyway). I'm not sure what the outcome would be.

But please tell me, what is the repeatable strategy here for doing the same thing more than once? If you always inoculate by grinding the sclerotia and never doing isolates, you'll always need more and more sclerotia, it hardly seems efficient. The most efficient way to keep it going to ferment more batches is to derive inoculum from your last batch, which will eventually isolate into a strain, unless you have an endless batch of new infected paspalum grass (maybe you do?).

Really the studies seem to suggest it is not that hard to find a suitable isolate for C. Paspali, though.

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InvisibleCreonAntigone
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Registered: 05/30/21
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Re: Wild-type Claviceps paspali productivity [Re: CreonAntigone]
    #27882116 - 07/29/22 06:03 PM (1 year, 9 months ago)

But let me not be dismissive and say there is one way to do what you're doing: if you want to do multispore every single time with Claviceps, you can't use a laboratory alone because the sexual structures don't form in sterile conditions.

What you COULD do is the following: you could make many multispore isolates by grinding up sclerotia and fermenting, then take the best and more productive culture from these and introduce it back into the field where you found the paspalum grass. This has a chance of increasing the average alkaloid content of the next generation's sclerotia. You would be limited by the amount of grass available, also, it goes without saying you should not do it if there isn't any risk of harm to wildlife or humans who might eat the grass.

I did read a study that repeatedly exposed cultures to new grasses to produce new sclerotia, and it found that doing this repeatedly resulted in better-producing strains in the submerged cultures. What you'd have to do is grow grass in controlled conditions (or in a field) to do this, how else would you get the new spores?

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InvisibleJoeMama1992
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Registered: 12/27/11
Posts: 901
Re: Wild-type Claviceps paspali productivity [Re: CreonAntigone]
    #27904134 - 08/15/22 10:13 AM (1 year, 8 months ago)

Haven't been able to check in for a bit, but wanted to post just to say that i haven't abandoned the thread or topic, just busy as hell lately. Later today I plan to settle in and get into it, should be active then.

:thanx:

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