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Blue-green spored club-shaped fungus on seeds
Stephen Mifsud, 25-11-2016 13:54
Stephen MifsudHi, I found numerous seeds of Washingtonia robusta (a species of plam tree) on soil infected with a clustered fungus having a club-like shape (Synnemata?) initially white then covered by blue-green spores (reminds me the colour of Penicillium and blue cheese).

Microscopy later tonight, but do have any suggestions (Family / genus) ?

This was picked up from the Mal tese islands



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Jacques Fournier, 28-11-2016 09:47
Jacques Fournier
Re : Blue-green spored club-shaped fungus on seeds
Hi Stefen,
good to see that Ascofrance forum is back. Thanks Christian!

this looks like conidial stromata of a Xylaria. You should fing conidia at the white tips.
Make sure old mature stromata are not present around, they are black and less conspicuous but they would me more informative. Not so many species occur on palm seeds.
Cheers,
Jacques
Stephen Mifsud, 28-11-2016 12:27
Stephen Mifsud
Re : Blue-green spored club-shaped fungus on seeds
Dear Jacques. Thank you for your message. Yes the forum was down andI could not post further developments with the Id of this fungus. I have found typical penicilli structures making the fungus a much likely Penicillium sp. Still, I have not really pinned a satisfying species but P. vulpinum, P. clavigerum and P. claviforme looks to be fairly good hypothesis.

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Jacques Fournier, 28-11-2016 12:30
Jacques Fournier
Re : Blue-green spored club-shaped fungus on seeds
OK Stefen
you are right, definitely not a Xylaria!
Jacques
Stephen Mifsud, 28-11-2016 13:50
Stephen Mifsud
Re : Blue-green spored club-shaped fungus on seeds
Penicilliopsis palmicola looking to be the right direction. Trying to find more info.
Thanks.

Jason Karakehian, 30-11-2016 03:18
Jason Karakehian
Re : Blue-green spored club-shaped fungus on seeds
I have found something which looks very similar growing out of the soft spot of fruits of Hyphaene coriacea palm in Gorongosa National Park, Mozambique. I haven't yet had a chance to work it up though. It is great to see your post about your collection.
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Stephen Mifsud, 30-11-2016 18:33
Stephen Mifsud
Re : Blue-green spored club-shaped fungus on seeds
Yes you are quite right, must be very related. Unfortunately I have not found many useful literature to progress. Although the genus is right, I've opted for palmicola because the epithet suggest it is 'specific' to palms.  I've seen on the uncertain internet P. palmicola looking like my specimen and others which are yellow ?! Any literature from members of AscoFrance is welcomed.

 Lookin forward for you microscopy to compare. I have measurmenst and other photos, but quite useless withour refernces to work through the ID.
Jason Karakehian, 30-11-2016 18:55
Jason Karakehian
Re : Blue-green spored club-shaped fungus on seeds
Excellent, I hope to work on this in December and January, and I will definitely stay in touch with my observations! It grows well in culture and even forms smaller versions of those horn-like projections.  Best - Jason
Stephen Mifsud, 30-11-2016 20:06
Stephen Mifsud
Re : Blue-green spored club-shaped fungus on seeds
I am culturing some too (on PDA, Czapek, and YEA) but I have some airborne contaminations.... In fact this leads to another question, perhaps it is already covered somewhere else one the forum.

When it comes to cultivate fungi on media at home, what advise you can give me to keep as much as possible a sterile environment.  I would like to add some chemical to act as an antibacterial agent (what please?) while apart from bacterial contamination, I tend to have Penicillium sp. and Mucor sp. colonies. Just one or two but still it spoils the plate :-( Is there any specific chemical to restrict these unwanted contaminants?

Jason Karakehian, 30-11-2016 20:31
Jason Karakehian
Re : Blue-green spored club-shaped fungus on seeds
I think you might try acidifying your media but I don't know what pH to bring it to. I'm sure you can find out online. I believe that that will help cut down on bacterial growth. I have had Streptomycin and Chloramphenicol in my media which are both antibacterial agents. Unfortunately I don't know how to obtain these or how to prepare them as a friend gave me a bunch of vials of it and so I never learned! I just add a vial of each to my media after autoclaving and think nothing else of it. I'm sure however that more can be learned online. I have access to a lab and a laminar flow hood so I usually do most of my culture work there. I believe however, that if you are in an enclosed room with no air flow, you can lift the edge of the petri dish up just enough to insert a sterile probe. I don't know what your home set up is like but I assume that you are careful about sterile conditions with the plates and autoclaving your media in some kind of system. One thing you can do is get a clear plastic bin and cut holes in it big enough to insert your arms into, this will still the air inside enough to perhaps prevent contamination.  With ascos, you can turn a petri dish of media upside down and place a small damp piece of filter paper or paper towel on the inside of the lid and then place an apothecium or whatever on top of that. I usually wrap it in parafilm to keep the humidity in but I don't think it makes much difference - it is my own superstition. The spores will shoot upwards onto the media and in this way bacteria and other potential contaminants do not fall onto the media. And keep an eye on it - you don't want tons of ascospores building up. You can rotate the plate to let them accumulate on another section of the plate. When you have enough remove the fungus and wrap the plate in parafilm and watch them to see if they germinate specifically and not something else. Be careful to place only a portion of your apothecium and no mites! Also if the apo is on wood be careful not to include any other very tiny perithecia of other fungi as you will likely have contamination and not realize it unless you see morphologically different spores.  Perhaps you know most of this? I apologize if too much or not helpful.
  Also, in regards to our fungus, I found this online, have you seen this? http://mushroomobserver.org/observer/index_observation?q=j9Z. We can try to hunt down this literature if you have not seen this?
 I have more advise on culturing but too much to write all here. I also have an article that was in Fungi magazine about a home sterile hood from a clear plastic bin but I can't find the reference!!  Here's one I found but maybe this is too much for our purposes:
http://www.freshcapmushrooms.com/learn/keeping-it-clean-how-to-design-and-build-a-laminar-flow-hood
Jason Karakehian, 02-12-2016 13:04
Jason Karakehian
Re : Blue-green spored club-shaped fungus on seeds
Stephen, I have literature on our fungus that I would like to share, please send me your email address and I will email it to you there.  Best wishes - Jason
Jason Karakehian, 02-12-2016 13:15
Jason Karakehian
Re : Blue-green spored club-shaped fungus on seeds
Stephen, also, just rereading your question about culturing - I am not aware of any agent that would cut down on the growth of specific fungi that might be a contaminant in your culture. The only thing for now is good sterile technique and try to excise spots of contamination in your plates. A helpful article is: A simple technique for purifying fungal cultures contaminated with bacteria and mites by Ko, Kunimoto and Ko, Journal of Phytopathology Volume 149, Issue 9
September 2001
Pages 509–510.
Let me know if you can't get access to this and I will get it for you. I have a photocopy and no PDF.
Best - Jason
Stephen Mifsud, 04-12-2016 23:01
Stephen Mifsud
Re : Blue-green spored club-shaped fungus on seeds
Dear Jason,

I am indebted for your help, time and useful comments. I can't afford a laminar flow at home (space restrictions) but i try to keep sterile conditions as much as possible.  From experience I adopted the following practices. 

- When preparing media, I only make a small batch that will be used with a week. Storing media in fridge for a long period will unavoidably lead to contaminations.

- I place my prepared media in a top-sealed plastic bag and then this in a plastic container. This had reduced drasticallly contamination of stored media

- I do not have the facility of an autoclave but I don't think it is essential when preparing small batches.  I pour the boiled media in plates when still very hot and cover with aluminium foil while cooling

- Problems of contamination start after inocculation and standing at room temperature. I try to achieve sterility by keeping my tweezers and inoculation needles in alcohol and occassionally put them in a flame of a spirit burner. Still I get 2/5 plates contaminated with Penicilium sp. and Staphylococcus or Pseudomonas. I am confident taht adding a broad range antibiotic like streptomycin will cut down the bacterial growth, but I can't find to buy very easily.

- I found that placing the innoculated dishes in a closed contained while standing helps a bit, but I think that contamination occurs on the process of inocculation, when the plate is open and some spores or bacteria fall on the surface of the media. I never experiened mite contamination.

So that is my home setup, simple and not ideal but soon I shall be working in a mycological Lab with a Laminar flow and hopefully I would get better results.



Stephen Mifsud, 04-12-2016 23:22
Stephen Mifsud
Re : Blue-green spored club-shaped fungus on seeds
With regards our fungus, I would really appreciate further literature and my email is info@maltawildplants.com

I did some further research on this synnematous fungus. I have found some depictions of Penicilliopsis palmicola (Sarophorum palmicola) and I have more doubt that our fungus is this species because, if the source I found is reliable, the metulae and branches of this species are distinctly swollen.
http://www.bcrc.firdi.org.tw/fungi/fungal_detail.jsp?id=FU200802290000
Moreover, google images of P. palmicola brings a lot of yellow, long branched synnemata which is different from my specimen?!?!


I could not open the link of mushroom observer that you provided (it opens a non-rlated page) but my samples are definitely the same as these:

http://mushroomobserver.org/58980?q=nB7
http://mushroomobserver.org/221873?q=nB7

or actually all of these:
http://mushroomobserver.org/species_list/show_species_list/611

Interestingly all collections are from South America:



Regarding literature:


The following paper gives a good introduction on the genus but I could not find the full paper:
https://www.jstor.org/stable/3761788?seq=1#page_scan_tab_contents

Another very important paper with keys is this paper:
http://link.springer.com/chapter/10.1007%2F978-1-4757-1856-0_31

If you can find it would be very useful



Another thing I did was to check what species are ther in the genus Sarophorum and Penicilliopsis. In the former I only found two species (S. palmicola and S. ledermannii) while for the latter there were a dozen:

Penicilliopsis Penicilliopsis Solms, Annales du Jardin Botanique de Buitenzorg 6: 53 (1887) [MB#3806]
Penicilliopsis africana Penicilliopsis africana Samson & Seifert, Advances in Penicillium and Aspergillus Systematics: 408 (1985) [MB#114759]
Penicilliopsis bambusae Penicilliopsis bambusae Nag Raj & Govindu (1970) [MB#319256]
Penicilliopsis brasiliensis Penicilliopsis brasiliensis Möller, Botanische Mittheilungen aus den Tropen 9: 293 (1901) [MB#141876]
Penicilliopsis clavariaeformis Penicilliopsis clavariaeformis Solms (1887) [MB#274252]
Penicilliopsis clavariiformis Penicilliopsis clavariiformis Solms, Annales du Jardin Botanique de Buitenzorg 6: 53 (1886) [MB#120178]
Penicilliopsis dichotoma Penicilliopsis dichotoma Hauman, Bulletin de la Société Botanique Belgique 69: 113 (1936) [MB#141964]
Penicilliopsis dybowskii Penicilliopsis dybowskii Pat., Bull. Soc. Mycol. France: 54 (1891) [MB#142485]
Penicilliopsis dybowskii var. dybowskii Penicilliopsis dybowskii var. dybowskii [MB#421814]
Penicilliopsis dybowskii var. macrospora Penicilliopsis dybowskii var. macrospora Beeli, Bulletin de la Société Royale de Botanique de Belgique 59 (2): 160 (1927) [MB#141715]
Penicilliopsis flavidus Penicilliopsis flavidus (Berk. & Broome) A.H.S. Br. [MB#124185]
Penicilliopsis juruensis Penicilliopsis juruensis Henn., Hedwigia 44: 59 (1905) [MB#142319]
Penicilliopsis ledermannii Penicilliopsis ledermannii (Syd. & P. Syd.) Hauman, Bulletin de la Société Botanique Belgique 69: 111 (1936) [MB#269153]
Penicilliopsis longissima Penicilliopsis longissima Hauman, Bulletin de la Société Botanique Belgique 69: 110 (1936) [MB#142089]
Penicilliopsis microsequoia Penicilliopsis microsequoia Hauman, Bulletin de la Société Botanique Belgique 69: 112 (1936) [MB#142431]
Penicilliopsis palmicola Penicilliopsis palmicola Henn., Hedwigia 43: 352 (1904) [MB#247191]
Penicilliopsis pseudocordyceps Penicilliopsis pseudocordyceps H.M. Hsieh & Y.M. Ju, Mycologia 94 (3): 541 (2002) [MB#484663]
Penicilliopsis togoensis Penicilliopsis togoensis Henn., Botanische Jahrbücher für Systematik Pflanzengeschichte und Pflanzengeographie 30: 40 (1901) [MB#246624]


I think I've written a lot today but we keep in touch and hope to find a name for this mysterious fungus

Jason Karakehian, 05-12-2016 00:01
Jason Karakehian
Re : Blue-green spored club-shaped fungus on seeds
Hi Stephen,
  The only thing I can think of here from what you describe is that you might try flaming your needles and tools for every transfer. I would definitely try acidifying the media if you can. Also, did you mention whether or not you wrapped plates in parafilm? I think that this cuts down on some contamination and dehydration. It also prevents me from carelessly picking up a plate by the lid and leaving the bottom on the countertop exposed to the open air! I would always be on the lookout for mites - keep checking those contaminated plates. The mites are so pernicious, they get into everything, including plates wrapped tightly in parafilm - they are, as a colleague once said, the closest thing to evidence for spontaneous generation! I myself have recently lost many plates of hard-gained cultures due to mite infestations. I am glad that you will soon have access to a laminar flow hood.  Kind regards - Jason
Jason Karakehian, 05-12-2016 00:07
Jason Karakehian
Re : Blue-green spored club-shaped fungus on seeds
Hi Stephen,
   Those Mushroom observer links that you provided I haven't seen, but definitely on the same track. I'm sorry those links didn't work. I will retry.
   I will write to you now using the email address you sent. Best - Jason
Joey JTan, 05-12-2016 00:19
Re : Blue-green spored club-shaped fungus on seeds
I tend to keep media plates at room temperature; I find storing them in a fridge invariably leads to lots of condensation, which can exacerbate contamination.  I also let plates dry out a little after pouring in a laminar flow hood, again to reduce condensation.  If you have a little bit of bacteria or contaminating fungi in your plate, condensation and free water on the agar surface can spread it all around the plate and ruin your chances of an axenic culture. 

A fungus such as this should be fairly easy to culture assuming your media is sterile.  Less is often more, so simply touching the conidiophores with a heated-and-cooled pin and streaking it on the plate or doing a three-point inoculation should be more successful than using a lot of material.  Avoid opening the plate too much, just lift one side 1-2 cm to allow you to place your pin/needle inside.    

Very nice fungus!  I hope you can get clean cultures, especailly if it is an unsequenced species.
Jason Karakehian, 05-12-2016 01:31
Jason Karakehian
Re : Blue-green spored club-shaped fungus on seeds
Yes, that's excellent advise I think. I also like to store them at room temp. I also like to leave the lids open a crack after pouring the media in the laminar flow hood to let the steam out. When there is no fogging of the inside lids I know that they are cool enough to close and put away.  Best - Jason
Stephen Mifsud, 05-12-2016 07:25
Stephen Mifsud
Re : Blue-green spored club-shaped fungus on seeds
I perfectly agree and I will try to put in a sterile bag, in a closed contained at room temp because condemsation and water inside the plate is a nuisance. I'll try to reculture today. The ones I did are not coming good :-(

Stephen Mifsud, 05-12-2016 07:28
Stephen Mifsud
Re : Blue-green spored club-shaped fungus on seeds
Some more photo in habitus
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Viktorie Halasu, 05-12-2016 07:29
Viktorie Halasu
Re : Blue-green spored club-shaped fungus on seeds
Hello,
that paper from Mycologia is online on cyberliber: http://www.cybertruffle.org.uk/cyberliber/59350/0094/003/0539.htm
Viktorie
Danny Newman, 16-03-2018 02:23
Danny Newman
Re : Blue-green spored club-shaped fungus on seeds
Hello All,


I am excited to see such interest in these palm seed parasite fungi. They have been a favorite of mine for many years. I am the creator of the species list linked in a previous comment (http://mushroomobserver.org/species_list/show_species_list/611).


It has been very interesting to see that this also occurs in the old world. Jason's Mozambique material, this collection, and another from Hawaii currently going by the name Penicillium palmae (http://mushroomobserver.org/230574) are the only three I know of from outside the Neotropics with this characteristic abundant blue conidia. How many species are at work here remains a mystery; perhaps one cosmopolitan one, perhaps several specialized ones.


Below is a link to what for me has been one of the best (and only) resources on this group of fungi:


https://drive.google.com/open?id=0B6XMTmQ88ozaM28xRlRWN09xVTQ


This is the first time I have seen a collection accompanied by microscopy, so it should hopefully be possible to move closer to an identification.


-Danny

Stephen Mifsud, 17-03-2018 05:47
Stephen Mifsud
Re : Blue-green spored club-shaped fungus on seeds
Dear Danny,
I am also glad that someone out there is also studying this fungus. This year I have seen it again in one of the two locations I know it from here in Gozo. and tried to follow it but I had no time to do the microscopy. However, I  had successful isolates on culture media producing outstanding colonies with the production of synemata especially on MEA. As I said earlier, it is always growing on Washingtonia palms. The production of red dye is interesting too. At 15C colony growth is very very slow. I do not have an incubator. I think it is still in my LAB and I can send you some more data and if in a good state, even samples. I have not been in the Lab for a month now :-(



For a moment I understood you described Penicillium palmae, so I ask if you have the protologue of this species, please: Penicillium palmae Samson, Stolk & Frisvad, Studies in Mycology 31: 135 (1989)

Like me, you are not attributing this Washingtonia seed fungus with any species covered by Samson and Seifort? The closest I got was Sarophorum palmicola, but I know it is not that species and so you are suggesting to name this as Penicillium palmae, right?

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Danny Newman, 25-07-2018 02:44
Danny Newman
Re : Blue-green spored club-shaped fungus on seeds
Re: Stephen et al.,

The conversation on this and other, possibly-related palm seed parasites continues at https://mushroomobserver.org/262509.  I hope you will join us there.  MO is in great need of more ascomycetologists.