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OfflineSharpstuff


Registered: 05/04/11
Posts: 510
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Last seen: 7 years, 9 months
Chitosan in Agar / Slants * 1
    #21845263 - 06/23/15 09:33 AM (8 years, 9 months ago)

Hey guys been looking into Chitosan a lot lately and have found that it is fungistatic and bacteristatic meaning it inhibits the germination of fungi/bacteria but does not inhibit the growth of the organism if already germinated. If this was to be added to a mix of agar do you think it would help inhibit contams but favor the already live tissue?

Chitosan is sourced from crustaceans but can also be sourced from fungi by deacytalating chitin.

Wondering if anyone has tried this ? I've read hydrogen peroxide is also fungistatic hence why some people use it but have also read it inhibits the growth of fungi.

Would love to hear your thoughts!

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InvisibleFantastic Mr. Fox
Grass Seed Connoisseur


Registered: 02/12/15
Posts: 3,227
Loc: Flag
Re: Chitosan in Agar / Slants [Re: Sharpstuff]
    #21845354 - 06/23/15 09:59 AM (8 years, 9 months ago)

chitosan is relatively insoluble in water, it has to be dissolved by dilute acids.


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OfflineSharpstuff


Registered: 05/04/11
Posts: 510
Loc: Australia Flag
Last seen: 7 years, 9 months
Re: Chitosan in Agar / Slants [Re: Fantastic Mr. Fox]
    #21845405 - 06/23/15 10:11 AM (8 years, 9 months ago)

Cheers but not a very constructive reply.

-You can get water soluble derivatives.
-Alot of fungi do well in slightly acidic environments.

Does anyone on the shroomery actually contribute or do they just shoot stuff down.

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InvisibleFantastic Mr. Fox
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Registered: 02/12/15
Posts: 3,227
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Re: Chitosan in Agar / Slants [Re: Sharpstuff]
    #21845429 - 06/23/15 10:17 AM (8 years, 9 months ago)

:lol:

Really?
I wasnt trying to be a dick.
Just keeping it blunt..

:goodluck:


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Offlinematsc
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Re: Chitosan in Agar / Slants [Re: Fantastic Mr. Fox]
    #21845855 - 06/23/15 12:35 PM (8 years, 9 months ago)

Not sure about on a plate, but in plants it acts as a protectant not on its own, but by stimulating the plants own immune system. Basically the plant goes "oh god a fungus!" and ramps up its antifungal/antipathogen pathways, so that when a real nasty comes along the defenses are already in place.

On an agar plate, I have no idea at all how it would behave. Worth an experiment I suppose.


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InvisibleFantastic Mr. Fox
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Registered: 02/12/15
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Re: Chitosan in Agar / Slants [Re: matsc]
    #21845992 - 06/23/15 01:09 PM (8 years, 9 months ago)

IMO Its more work than its worth..
They're other additives.. i.e Gentamicin;which is an aminoglycoside antibiotic/bactericidal antibiotic.
It has successfully shown it's place in our hobby,

There's also chloramphenicol, which is usually used in conjunction with gentamicin.

Chitosan has no history of being used for mycological work.. As far as I know. (link/correct me if I'm wrong)


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Giving is all we have, for we're just grateful to be alive

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Offlinematsc
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Re: Chitosan in Agar / Slants [Re: Fantastic Mr. Fox]
    #21846210 - 06/23/15 02:09 PM (8 years, 9 months ago)

http://www.sciencedirect.com/science/article/pii/S0168160599000069
Roller, S., and N. Covill. "The Antifungal Properties of Chitosan in Laboratory Media and Apple Juice." International Journal of Food Microbiology 47.1-2 (1999): 67-77. Web.

http://onlinelibrary.wiley.com/doi/10.1002/app.33612/abstract
Ren, Jianming, Jingli Liu, Rongchun Li, Fang Dong, and Zhanyong Guo. "Antifungal Properties of Chitosan Salts in Laboratory Media." Journal of Applied Polymer Science J. Appl. Polym. Sci. 124.3 (2011): 2501-507. Web.

Surprisingly, it looks like chitosan really does have some antibiotic properties. Its a bit fuzzy on the how and why, but the results are interesting if nothing else.

Edit:
Heres one more-
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC91788/
Rhoades, J., and S. Roller. "Antimicrobial Actions of Degraded and Native Chitosan against Spoilage Organisms in Laboratory Media and Foods." Applied and Environmental Microbiology 66.1 (2000): 80-86. Web.


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Edited by matsc (06/23/15 02:19 PM)

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InvisibleFantastic Mr. Fox
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Registered: 02/12/15
Posts: 3,227
Loc: Flag
Re: Chitosan in Agar / Slants [Re: matsc]
    #21846233 - 06/23/15 02:18 PM (8 years, 9 months ago)

Thanks for the links. :thumbup:

I still stand my ground..
All youve shown me is that Chitosan is good as a fungicide/perservative for certain fruits & vegetables. But I dont see it aiding in our hobby, due to the fact that its a fungicide... Last I checked we were trying to cultivate/fruit fungi, not kill it. :lol:

Nice links, nonetheless


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Offlinematsc
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Re: Chitosan in Agar / Slants [Re: Fantastic Mr. Fox]
    #21846238 - 06/23/15 02:20 PM (8 years, 9 months ago)

Yeah again, no clue how this would react in the context of intentionally cultured fungi. I may be able to dig up an article somewhere but my lunch break is almost up.

If you have some chitosan and want to play around with it, by all means do so, and report your findings. But I wouldnt spend any money on it if its just an idea.


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OfflineMandarinfish

Registered: 01/27/15
Posts: 1,365
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Re: Chitosan in Agar / Slants [Re: Sharpstuff]
    #21886712 - 07/02/15 07:58 AM (8 years, 8 months ago)

.

Edited by Mandarinfish (07/15/20 10:43 PM)

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OfflineSharpstuff


Registered: 05/04/11
Posts: 510
Loc: Australia Flag
Last seen: 7 years, 9 months
Re: Chitosan in Agar / Slants [Re: Fantastic Mr. Fox]
    #21899610 - 07/05/15 03:20 AM (8 years, 8 months ago)

Quote:

Fantastic Mr. Fox said:
IMO Its more work than its worth..
They're other additives.. i.e Gentamicin;which is an aminoglycoside antibiotic/bactericidal antibiotic.
It has successfully shown it's place in our hobby,

There's also chloramphenicol, which is usually used in conjunction with gentamicin.

Chitosan has no history of being used for mycological work.. As far as I know. (link/correct me if I'm wrong)




Chitosan would much cheaper than antibiotics and is quite readily available and does not bring the impacts the over-use of antibiotics can bring. IMO Chitosan would be MUCH more viable if it worked successfully.

I posted this in advanced mycology with hopes of either I or someone else experimenting with the Chitosan. I could also not find it being used in mycology before so why wouldn't you want to experiment? It could potentially have uses beyond being used in agar.

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InvisibleFantastic Mr. Fox
Grass Seed Connoisseur


Registered: 02/12/15
Posts: 3,227
Loc: Flag
Re: Chitosan in Agar / Slants [Re: Mandarinfish]
    #21899615 - 07/05/15 03:25 AM (8 years, 8 months ago)

Quote:

Mandarinfish said:
I hope your grows don't end up smelling like shrimps. :laugh2:



:rofl:


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Giving is all we have, for we're just grateful to be alive

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OfflineSharpstuff


Registered: 05/04/11
Posts: 510
Loc: Australia Flag
Last seen: 7 years, 9 months
Re: Chitosan in Agar / Slants [Re: Fantastic Mr. Fox]
    #21899668 - 07/05/15 04:20 AM (8 years, 8 months ago)

Please go to the pub if you do not wish to contribute anything of value.

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InvisibleFantastic Mr. Fox
Grass Seed Connoisseur


Registered: 02/12/15
Posts: 3,227
Loc: Flag
Re: Chitosan in Agar / Slants [Re: Sharpstuff]
    #21899672 - 07/05/15 04:23 AM (8 years, 8 months ago)

Bishh's be bitter,
"Yes, sir"


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Giving is all we have, for we're just grateful to be alive

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InvisibleToadstool5
A Registered Mycophile
Male User Gallery


Registered: 01/22/15
Posts: 1,359
Loc: The Golden State
Re: Chitosan in Agar / Slants [Re: Fantastic Mr. Fox]
    #21907940 - 07/07/15 12:14 AM (8 years, 8 months ago)

It is my understanding that some fungi and bacteria hate acidity due to protonation interfering with the adsorption of certain essential anions and the release of certain essential cations.

The chitosan has amine groups N-H and substituted amino groups R1-NH-R2 that release the hydrogen into its ionic form and interfere with ionic transmission of the microbes' membranes. It also chelates and forms complexes with certain ions rendering them immobile.

Perhaps living mycelium creates an electrical charge that can be manipulated to induce dielectricity of the chitosan through enzymatic responses or a nervous system to make certain atoms rearrange and allow passive diffusion of ions into or out of the chitosan or to create immobility of other undesirable atoms. This rearrangement from dielectrical excitation could also cause a release of hydronium ions from the N-H bonds in an attempt to protonate nearby contaminates in an effort of self-defense like the peroxides released by mycelium. Perhaps this is a complex Reduction/Oxidation mechanism for ion transport and self-defense we do not understand yet? If reduction does not render a microbe dead then perhaps oxidative damage is the second line of attack?

Honestly if you can figure out how to apply it to agar, liquid culture, or substrate and it has the same antibiotic strength of hydrogen peroxide while preventing oxidative damage (or protonagenic damage for that matter) to the living mycelium transferred to the media treated with CS or CS-Salts...

I would switch over immediately. :shrug:


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If you do not know where the mushroom products you are consuming are grown, think twice before eating them. :badshroom:
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InvisibleToadstool5
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Registered: 01/22/15
Posts: 1,359
Loc: The Golden State
Re: Chitosan in Agar / Slants [Re: Toadstool5]
    #21907967 - 07/07/15 12:25 AM (8 years, 8 months ago)

On a personal note, good to see you again sharpstuff! I just realized i know you from our discussions on the ability of tilandsia species to be used as an air-filtration system due to its lack of soil and ability to chelate and metabolize particulate pollution.

You always have such ambitious ideas and i totally dig  it man :thumbup:


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If you do not know where the mushroom products you are consuming are grown, think twice before eating them. :badshroom:
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OfflineSharpstuff


Registered: 05/04/11
Posts: 510
Loc: Australia Flag
Last seen: 7 years, 9 months
Re: Chitosan in Agar / Slants [Re: Toadstool5]
    #21912454 - 07/07/15 10:59 PM (8 years, 8 months ago)

Thanks toadstool!

Well I do enjoy exploring new ways to do things that's what science is all about after all. Thanks for the lengthy explanation on the chitosan. I have access to water soluble chitosan-oligosaccharides which can be used without an acidic solute possibly preventing that immobilization if it does occur. I will hopefully be testing out various derivites of the chitosan to see what works best (if at all). I can't see it oxygenating the mycelium like Hydrogen peroxide does but does have similar properties so in theory it sounds like it would work quite well. Its also safer to handle them Hydrogen peroxide (large concentrates anyways)

Will keep this thread updated!

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InvisibleToadstool5
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Registered: 01/22/15
Posts: 1,359
Loc: The Golden State
Re: Chitosan in Agar / Slants [Re: Sharpstuff]
    #21912767 - 07/07/15 11:59 PM (8 years, 8 months ago)

Please do keep us updated! I cant offer much advise on which substituted amino salts to try or how you would synthesize them without reacting the hydroxyl groups but i did find evidence that it will not kill the fungal cultures completely.

Quote:

3.2.1. Radial growth
In culture media amended with chitosan, CC and CS had higher (P ≤ 0.05) inhibitory effect on the average colony radius with respect to control at 96 h (Table 2). None of the chitosans completely inhibited the fungus, so the effect is fungistatic and not fungicidal. In previous studies, Plascencia-Jatomea et al. (2003)
found that medium molecular weight chitosan (3 and 5 g/L) inhibited the radial growth of A. niger by 73% compared with the control, without observing complete inhibition. Similarly, it has been reported that chitosan and its derivatives are not able to inhibit 100% the growth of fungi as B. cinerea, Fusarium oxysporum and R. stolonifer (Guo et al., 2006; Hernández-Lazaurdo et al., 2008;
Rabea et al., 2006).

In this study both chitosans have a similar %DD, which may explain having similar fungistatic activity, as the %DD shows the proportion of free amino groups in the polymer that are directly related to the antimicrobial activity of chitosan.




Chitosan composite films: Thermal, structural, mechanical and antifungal properties


A.P. Martínez-Camachoa, M.O. Cortez-Rochaa, J.M. Ezquerra-Brauer a, A.Z. Graciano-Verdugo b,
F. Rodriguez-Félixa, M.M. Castillo-Ortegac, M.S. Yépiz-Gómez a, M. Plascencia-Jatomeaa,∗

It appears the fungistatic properties directly relate to the amino groups protonation of contams and effect different fungi to different extents. If you can manipulate the available amino groups to the correct strength of fungistatic activity it would be perfect. It is non-selective for gram positive and negative bacteria and would decimate a spore trying to establish itself while only weakening your culture.

Quote:

By analyzing the morphology of A. niger in the apical growth phase, a decrease was observed (P ≤ 0.05) in the average diameter of the hyphae (Table 4) developed in direct contact with chitosan plasticized films (pCSS and pCCS), which may be caused for changes in membrane permeability due to the pressure and strain of the fungal cell wall. Previous studies reported that, added in PDA agar, chitosan causes deformation and damage on the surface of the hyphae of A. niger, besides an increase in the average diameter of the hyphae, which augments with increasing concentration of polymer
(Plascencia-Jatomea et al., 2003).




It does cause damage and morphology changes in hyphae that are in direct contact of it. So just like hydrogen peroxide you should only use it when needed and with moderation.

What if, and this is speculation, the species you hope to cultivate in agar treated with chitosan has the enzymes needed to depolymerize chitosan into chitin and isnt effected much?

I would also love to cast an acetic-CS film, deacetylate it through NaOH, and then adhere it to the inside of a SAB. Imagine adding sorbitol to make a longer polymer and then plasticize it in a cast to make a SAB that is comprised of 100% CS-Plastic! :awesomenod:

I am going to try that someday when i have the funding and take swab samples from the insides of the CS-Plastic vs regular plastic to grow them out on agar and see what it cuts back and how much it cuts the levels.


--------------------
If you do not know where the mushroom products you are consuming are grown, think twice before eating them. :badshroom:
- Paul Stamets

AMU Teks :mushroom2: Stro's Write Ups

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