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Invisiblemetacohl
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Re: Lepista nuda [Re: mikesethnobotany]
    #15815968 - 02/15/12 11:35 PM (8 years, 8 months ago)

I just wanted to add that I found a specimen of Lepista Nuda today and will be doing a tissue sample to agar. I will post my results. The continued input of those who have attempted cultivation of this species is greatly appreciated.


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Re: Lepista nuda [Re: metacohl]
    #15815986 - 02/15/12 11:39 PM (8 years, 8 months ago)

Good news, and good luck metacohl!

They are a pretty species, aren't they?  I was surprised to find out that
something so lovely is edible. (and George Carlin told me, there is no blue food!)

I am really looking forward to what you see on agar.

Take care,

JD


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Re: Lepista nuda [Re: Javadog]
    #15817109 - 02/16/12 05:06 AM (8 years, 8 months ago)

P.S. I poured a sleeve of PDYA dishes this evening and made a xfer to
two of them from the leading edge of my latest (A agar -> normal agar)
L. nuda dish this evening.

Tomorrow I am going to pour a sleeve of PMEA tomorrow.

(Poo Malt Extract Agar)

These are just my attempts to see if this species grows better on
different agar formulas.  I want to a new to the point where I can
try Carsten's manure-with-the-grains idea.

Take care,

JD


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Re: Lepista nuda [Re: Javadog]
    #15817163 - 02/16/12 05:44 AM (8 years, 8 months ago)

I'm curious how the PMEA will perform. :smile: I guess there will be quite a difference, as I have seen good results on pure manure agar without other ingredients.

Generally, primary wood decomposers can utilize most nutrients they find plus they synthesize their own vitamins, while most secondary decomposers depend on a special set of nutrients, provided by microbes. Thats why MEA isn't good for each and every mushroom. It does contain some proteins and other nitrogen compounds, but mushrooms, which grow on leaf litter or manure often can't digest them.

So... just add manure... and keep an eye on the pot when boiling, manure tea tends to foam and boil over, leaving quite a mess on the stove.

Carsten


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Re: Lepista nuda [Re: bambus]
    #15817174 - 02/16/12 05:57 AM (8 years, 8 months ago)

Quote:

bambus said:
The agar I used was the corn-steep pre-mix from mushroompeople.
I also added a bit of H-manure to it when cloning this species.
The fully colonized grain jars show no "spots" whatsoever. The mycelium has a very uniform color to it. The growth is not rhizomophic at all, just light & fluffy.



Interesting that the corn steep agar does not show any color reaction at all. So far I have only seen L. nuda rhizomorphs in casing soil. They were extremely thin and didn't really deserve that name.

Carsten



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Re: Lepista nuda [Re: Mycelio]
    #15817654 - 02/16/12 10:29 AM (8 years, 8 months ago)

Paper on L. Nuda cultivation if anyone has access or wants to purchase.

http://www.pubhort.org/isms/11/1/v11_p1_a70.htm


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Edited by metacohl (02/16/12 10:30 AM)


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Re: Lepista nuda [Re: metacohl]
    #15817663 - 02/16/12 10:31 AM (8 years, 8 months ago)

*Edit* The majority of the papers I've skimmed say they are culturing on MEA or PDA, but below is some info on alternative mediums.

From another paper

"Mycelial growth of four strains of the wood blewit mushroom Lepista nuda on culture media with different organic supplements (coconut bagasse, sugarcane bagasse, oak sawdust), pH (6, 7, 8) and temperature (20°C, 25°C) was evaluated. Linear growth rate (Kt) on agar medium was measured. Biomass production of mycelia and phenolic content on oak liquid culture, under two different gaseous interchange treatments, culture bottles with filter (F) and culture bottles without filter (WF), also were observed. Results showed a significant influence of the supplements and strains on agar. The best growth responses were observed on oak agar medium at 20°C and 25°C (0.417 and 0.410 mm d-1, respectively). For both temperatures, in all treatments no differences significantly were observed. The highest Kt by L. nuda strains was registered at pH 7. On liquid culture, the F treatment significantly increased the biomass production (4.10 mg mL-1) by almost 1.8 times compared to the WF treatment (2.29 mg mL-1) at 13 days of incubation. Phenols concentration decreased in both treatments but in F treatment the phenols concentration (0.79 mMol L-1) decreased less than WF (0.62 mMol L-1). No correlation between the biomass production and phenolic content was observed."

GAITAN-HERNANDEZ, Rigoberto  y  BAEZ RODRIGUEZ, Iván. Mycelial growth of Lepista nuda native wild strains on culture media with different organic supplements. Rev. Mex. Mic [online]. 2008, vol.26, pp. 41-49. ISSN 0187-3180.

Growth parameters of Australian varieties

http://www.hort.purdue.edu/newcrop/proceedings1996/v3-476.html

*Edit* And another about cloning from wild specimens

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1388993/pdf/hw4288.pdf

"Some saprophytic soil basidiomycetes may not be recovered by the method described here. Warcup (49) collected soil be- neath fruiting bodies of some soil basidiomycetes, such as Agaricus arvensis (Psalliota arvensis), Lepista nuda (Tricholoma nudum), and Marasmius oreades, and found that they will grow out from undisturbed soil blocks or in soil plates. During this study, soil was collected below fruiting bodies of Lepista nuda. This species could not be recovered from washed particles plated on lignin-guaiacol-benomyl agar but was isolated from hyphae growing out of small soil crumbs sprinkled on water agar. Thus, it seems that the particle-washing technique should be combined with other methods such as the soil plate method (48) or the soil crumb or sprinkle plate methods (3, 47) for a more complete estimate of the diversity of saprophytic basid- iomycetes."


Edited by metacohl (02/16/12 11:09 AM)


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Re: Lepista nuda [Re: metacohl]
    #15818890 - 02/16/12 03:51 PM (8 years, 8 months ago)

Very interesting.  Thanks for sharing.

Is anyone interested in explaining the term "Kt" in the phrase:
    "The highest Kt by L. nuda strains..."

Also by "Filtered" they are referring to filtered GE vs. open ports?

BTW, I am looking at the Cornsteep Dextrose Agar from MP, referred to
above, and I wanted to confirm that it is an agar mix that just needs
water.  This must be the case as it is cheaper than plain agar-agar.

FWIW, what are your favorite, non-chinese grocery, agar sources?

Take care all,

JD


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Re: Lepista nuda [Re: Javadog]
    #15820912 - 02/16/12 11:23 PM (8 years, 8 months ago)

Quote:

Javadog said:
Is anyone interested in explaining the term "Kt" in the phrase:
    "The highest Kt by L. nuda strains..."
JD




I believe they are defining the growth rate as Kt

"Linear growth rate (Kt) on agar "


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Re: Lepista nuda [Re: metacohl]
    #15822358 - 02/17/12 08:44 AM (8 years, 8 months ago)

A growth rate of 0.417mm/day is pretty slow anyway. I got more than three times that speed on compost with spent oyster sub, even without grain.

Thanks for adding those links and abstracts, metacohl!

Carsten


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Re: Lepista nuda [Re: Mycelio]
    #15822504 - 02/17/12 10:19 AM (8 years, 8 months ago)

Quote:

Mycelio said:
A growth rate of 0.417mm/day is pretty slow anyway. I got more than three times that speed on compost with spent oyster sub, even without grain.

Thanks for adding those links and abstracts, metacohl!

Carsten




I believe those measurements are from growth on agar plates? I think that adding some sort of compost or spent substrate could definitely improve the growth, they seem to be secondary decomposers?

So of my two tissue cultures from my wild specimen, only one seems to have been successful. My stipe tissue culture almost immediately started to liquify after I put in on a dish and now seems to be enveloped by some bacterial growth. While the dish on to which I scraped colonized soil fragments has some amazing fiborous, rhizomorphic growth stretching across sections of the dish. Literally some of the rhizomorphs are > 3mm long. Looks very similar to some of the macro photos I have seen of L. Nuda mycellium. I dont think my camera could capture them at the moment but Ill try to post some images later.

*edit* The growth is very reminiscent of this photo from mushroomexpert
L.Nuda Mycelium colonizing bacteria


Edited by metacohl (02/17/12 10:23 AM)


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Re: Lepista nuda [Re: metacohl]
    #15822766 - 02/17/12 12:16 PM (8 years, 8 months ago)

Here are some pics of my soil sample plate... and my disclaimer.. I apologize for the quality of the photos as this is my first time taking pictures of a petridish and I am still learning how to use my camera phone for macro photography.

Also... this was literally the first agar dish I have ever poured... and I didn't let it cool long enough before attempting to wrap it in parafilm....so the media splashed and bubbled on the lid...:facepalm3:




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Re: Lepista nuda [Re: metacohl]
    #15822802 - 02/17/12 12:27 PM (8 years, 8 months ago)

I guess my question is how do we know that is l. nuda growing on the plate.  Dirt could have dozens of things growing in it at the same time and who knows which one took off on the agar.  Not to be a downer, but I would just be very suspect of this culture being what you want.

They sell l. nuda at a grocery store near me, I have no idea the source but they had them a few weeks ago.  I am going soon and buy one(they are like $35-45 a lb) and see if I can clone.

Trout


--------------------
I need tropical cultures, ABM, V.v....!!!!

Well things don't always look as they are and things can be misread and mistaken for what they realy are so don't read too much into what I say since I might be mistaken myself. And remember I rarely ever give a definate answer.


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Re: Lepista nuda [Re: trout]
    #15822872 - 02/17/12 12:47 PM (8 years, 8 months ago)

Quote:

trout said:
I guess my question is how do we know that is l. nuda growing on the plate.  Dirt could have dozens of things growing in it at the same time and who knows which one took off on the agar.  Not to be a downer, but I would just be very suspect of this culture being what you want.





Obviously no one could say with 100% certainty at this point that it is L. Nuda. I think that the fact the soil was 1) Visibly colonized by L. Nuda mycelium. 2) Among the different types of growth on the plate, 1 pattern matches the pictures of L. Nuda rhizomorphs online. 3) I would just be very suspect of any other filamentous fungi growing that much more quickly than the L. Nuda mycelium already colonizing the soil. All contribute to a good chance that it is L. Nuda.

Though once a transfer is made, the lilac color and blue staining of PDYA should make it idiot proof to identify.....


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Edited by metacohl (02/17/12 12:49 PM)


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Re: Lepista nuda [Re: metacohl]
    #15822983 - 02/17/12 01:19 PM (8 years, 8 months ago)

The picture by George Barron shows magnified single hyphae, not rhizomorphs. Sure there is some similarity to what I can see in your second pic, but you better don't put too much trust in mycelium from soil samples. Colonizing L. nuda mycelium does not sterilize the soil it grows in plus lots of soil fungi can grow like this, for example ink caps.

Carsten


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Re: Lepista nuda [Re: Mycelio]
    #15823077 - 02/17/12 01:42 PM (8 years, 8 months ago)

Quote:

Mycelio said:
The picture by George Barron shows magnified single hyphae, not rhizomorphs. Sure there is some similarity to what I can see in your second pic, but you better don't put too much trust in mycelium from soil samples. Colonizing L. nuda mycelium does not sterilize the soil it grows in plus lots of soil fungi can grow like this, for example ink caps.

Carsten




I did not realize the Barron picture was of single hyphae, thank you for clarifying that. Like I said, obviously it is impossible to be sure right now. Perhaps I am over estimating the likelihood of it being L. Nuda. I am hoping that the color of the pure culture will help to identify.

Also, I was not clear on whether L. Nuda as a species stains PDYA blue or just the isolate in the photograph you posted. Can you clarify this?


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Re: Lepista nuda [Re: metacohl]
    #15823171 - 02/17/12 02:11 PM (8 years, 8 months ago)

I'm not sure, but as I read it often, I guess all L. nuda strains do that. Perhaps it is a reaction between fungal enzymes and starch.

@Trout
You may find a less aromatic Lepista saeva instead.
http://www.shroomery.org/forums/showflat.php/Number/14659744

Carsten


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Re: Lepista nuda [Re: Mycelio]
    #15823196 - 02/17/12 02:17 PM (8 years, 8 months ago)

Quote:

Mycelio said:
I'm not sure, but as I read it often, I guess all L. nuda strains do that. Perhaps it is a reaction between fungal enzymes and starch.
Carsten




Thanks, that is what I was thinking as well. Doesn't seem to far fetched to me that a lavender mushroom might have a blue metabolite. :smile:


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Re: Lepista nuda [Re: metacohl]
    #15827273 - 02/18/12 12:50 PM (8 years, 8 months ago)

Look closer, we see brown staining around the mycelium on MEA and no color change on corn steep agar. This looks pretty much like the common chemical test for starch. In this case the iodine may be replaced by secreted amylase. L. nuda shall produce lots of those starch degrading enzymes according to this paper:
'Mycelial Culture Conditions of Lepista nuda and Extracellular Enzyme Activity, Sang-Dae Kim, Ji-Hye Kim, Jong-Bong Kim, Yeong-Hwan Han'
http://www.msk.or.kr/jsp/downloadPDF1.jsp?paperSeq=2271&fileName=41(3)-4(p.164-167).pdf
Can't read the korean text, but the tables on page 3 are interesting, especially the comparison of various sources of carbon and nitrogen.

Carsten


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Re: Lepista nuda [Re: Mycelio]
    #15827483 - 02/18/12 01:46 PM (8 years, 8 months ago)

Two days on Javadogs own WPDWYA* agar and everything looks better so far:


I was hoping to make my PMEA tonight, but I am being dragged to
HELL^H^H^H^H the in-laws today.  Ah well.

Take care,

JD

* Wrong Potato Dextrose Wrong Yeast Agar  ;0)


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Edited by Javadog (02/18/12 02:10 PM)


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