Observation 208129: Tylopilus P. Karst.
When: 2015-06-28
No herbarium specimen

Notes: A single, small but robust young specimen in ground under live Oak. Velvety dry cap = 5.5 cm., stem = 3.5 cm. above ground and 1.5 cm. buried in ground. Stem width = 3 cm. at bulbous base and 1.5 cm. at apex. Fresh whitish pore surface easily bruises brown. Pores = 2 mm. deep; 2 pores per mm. KOH on cap instantly red-orange.

Images

532777
IMG_9592.JPG
Copyright © 2015 Judi Thomas
532778
IMG_9590.JPG
Copyright © 2015 Judi Thomas
532779
IMG_9594.JPG
Copyright © 2015 Judi Thomas
532780
IMG_9611.JPG
Copyright © 2015 Judi Thomas
Brown reticulation over white stem near apex.
532781
IMG_9621.JPG
Copyright © 2015 Judi Thomas

Proposed Names

-21% (5)
Eye3
Used references: Kuo’s “Mushrooms of the Midwest” , MO observations
19% (4)
Recognized by sight: For the record (X-14-16), this mushroom was also posted in obs 208130, obs 208567, obs 209237, obs 244148 (to be sequenced), and obs 253128.
28% (4)
Eyes3
Recognized by sight

Please login to propose your own names and vote on existing names.

Eye3 = Observer’s choice
Eyes3 = Current consensus

Comments

Add Comment
Why would ITS be more ambiguous?
By: I. G. Safonov (IGSafonov)
2016-10-21 22:30:48 PDT (-0700)

I thought ITS is better at species resolution that LSU by virtue of having a better barcode gap. The LSU sequence for obs 201760 was a match for L. carminipes, B. pallidoroseus and B. bicolor sequences in the 99.4-99.6% range (if those ID’s are indeed correct). Hence I would expect ITS to favor one of those three. Am I missing something?

Sequences
By: I. G. Safonov (IGSafonov)
2016-10-21 21:33:25 PDT (-0700)

I just posted the sequence for obs 215764 in that obsie.

I’ve never posted to GenBank. I heard it was not at all straightforward. Are there step by step instruction for novices on how to submit sequences to GB? If not, and you can do it with your eyes closed, I can send you the data. I have a total of 20+ sequences for 20 or so bolete observations.

LSU
By: Alan Rockefeller (Alan Rockefeller)
2016-10-21 20:51:35 PDT (-0700)

About 210760, if the LSU was that ambiguous, it’s likely the ITS would be even moreso.

It would be a good idea to put your sequences into GenBank so the next time someone sequences this and runs BLAST they get directed to your photos.

Obs 215764 is odd, could you post the sequence you got for that in the observation?

Alan,
By: I. G. Safonov (IGSafonov)
2016-10-20 11:17:48 PDT (-0700)

Thank you for sharing your knowledge and for your advice.

It’s been a tough call between ITS and LSU for sequencing my bolete collections in the past for a few reasons. One of them has nothing to do with science – it the cost of doing business. I have many samples to sequence, but cannot afford running even the smallest panel of loci (n=2) for each sample.

The last batch of boletes I did was all LSU and I got mixed results. For instance, it worked beautifully for obs 217160 to show that it indeed belongs in Caloboletus, but is not any of the known species. On the other hand, it was too ambiguous with B. pallidoroseus, obs 210760, which is in line with your statement. Perhaps, ITS would have worked better in this case. I think part of the problem is that identification of species in bicolor and carminipes clades is challenging – I wonder how many sequences in GenBank are linked to bad ID’s for those groups as well as for something like the subvelutipes group. And then I have obs 215764, wherein sequencing LSU in theory should have worked well for placement into a genus, except that GenBank returned no meaningful hits. The only explanation I can think of is that no species belonging to this new (?) genus have been sequenced or entered into GenBank. What do you think?

Yes, both ITS and LSU are much easier to sequence, unlike the single-copy protein-coding genes. Even though TEF-1 or RPB’s have a better resolving power than the two ribosomal genes, I would be reluctant to sequence them right now. There is simply insufficient amount of data posted to GenBank at this time.

Now I need to go back to my list and re-evaluate my choice of loci for each of them.

nrLSU
By: Alan Rockefeller (Alan Rockefeller)
2016-10-20 04:49:20 PDT (-0700)

If you are just going to sequence one gene, nrITS is usually the best to do – however not with collections where the genus is unknown. The reason is that ITS is so variable that unless there is something really close in GenBank, you’ll get no useful data at all. I’ve had that happen to me twice with boletes – once with Gyroporus and once with Retiboletus. nrLSU would be a better choice because it’s more conserved, and you’d have a better chance of being able to see the genus. ITS varies too widely to directly compare all the boletes. This is why most bolete researchers use LSU, and why there are so many LSU bolete sequences in GenBank.

For small groups of boletes like porcini or butter boletes, ITS is the best choice because it’s most likely to give resolution at the species level.

Since LSU is a multi-copy gene, it’s about as easy to sequence as ITS.

If this is in Boletus s.str., ITS will do a great job and would be better than LSU. If it’s somewhere else, LSU would be better.

Terri,
By: I. G. Safonov (IGSafonov)
2016-10-20 00:13:25 PDT (-0700)

I understand what you mean. It’s true that multiple genes are necessary for building reliable phylogenetic relationships. This is the current gold standard in the field – at least a 3-gene “pack”. Obviously, the more, the better, but then it gets expensive and labor intensive.
I know that ITS has its shortcomings, like the inability to resolve closely related species. Roy Halling thinks it’s the least informative locus for the Boletaceae phylogeny in general, but at the same time Arora and Frank had good luck with it for their 2014 Butter Bolete study and Brun’s seminal 1997 work on Suillus was done with ITS alone. Perhaps, it’s case specific. There are probably other examples to bolster pros and cons of ITS use; however, it seems true that ITS is falling out of favor for boletes, as fewer boletelogists use it in conjunction with other loci, while it continues to work great for agarics and other fungi. Yet, there is a lot that’s going for ITS right now, and the paper I referenced does a nice job explaining why ITS is here to stay as the barcode gene till a better substitute is found to supplant it.
The bottom line is that what I am doing here, specifically for this obsie (and other uknown boletes), is “fingerprinting”. The immediate goal is to place this bolete into a genus. ITS should tell us readily if this is a Tylopilus or if it’s B. pallidus or neither. I also think that this bolete is sufficiently unique that a GenBank search may come back empty-handed. I had this happen on at least a couple of occasions in the past for boletes I couldn’t ID, e.g. obs 179586 and obs 215764. :-)

Igor,
By: Terri Clements/Donna Fulton (pinonbistro)
2016-10-19 22:33:13 PDT (-0700)

I was just curious as I know from our experience with morels that ITS is not sufficient in many cases to resolve to species. TEF-1, RPB1 and RBP2 are needed. I wondered about boletes so I checked out some Tylopilus spp on GenBank and it seems that RPB1, LSU and TEF-1 seem to be used most often for recent submissions. Of course that’s just a small sample.

Terri

Terri,
By: I. G. Safonov (IGSafonov)
2016-10-19 18:11:47 PDT (-0700)

I am going to go with nrITS for starters, an obvious choice. By the way, this locus is officially the universal bar code for fungi:

http://www.pnas.org/content/109/16/6241.full.pdf.

Besides, there’s been a lot of ITS data deposited in GenBank, more than for any other gene. Even if there is no match, at least there is a good chance we can find the closest relative, if any.

Igor, what markers will you sequence? Terri
By: Terri Clements/Donna Fulton (pinonbistro)
2016-10-19 14:34:22 PDT (-0700)
That’s great news, Igor.
By: Judi T. (AvidAmateur)
2016-10-19 07:35:18 PDT (-0700)

I’m so excited at the prospect of finding out where these specimens fit into the overall scheme of things. Thank you so much for facilitating what’s sure to be a great learning experience for all of us.

Tylopilus?
By: I. G. Safonov (IGSafonov)
2016-10-18 19:29:29 PDT (-0700)

I see the consensus shifted to Tylopilus, a move probably inspired by the stocky stature of this mushroom and the olivaceous staining of the stipe. Well, we shall see ’bout that. :-) I just heard back from the DNA gent — sequencing shall be done in November.

Terri, that’s most kind of you.
By: Judi T. (AvidAmateur)
2016-10-18 18:49:36 PDT (-0700)

I’d love to see that paper. I haven’t had time to search the internet. I’m sure there must be some “DNA 101” basic information out there. So much to learn, so little time:) Thanks so much.

Hi Judi
By: Terri Clements/Donna Fulton (pinonbistro)
2016-10-18 18:05:37 PDT (-0700)

If you’re interested I could email you a pdf of a 2012 paper on morels that addresses some of your DNA sequencing questions. Our club has started to sequence some of our finds, starting with morels, and from our experience some of the key factors include the quality of the specimen being sequenced (age and overheating degrade DNA for example), the quality of the data on GenBank (for example there have been many recent studies of morels resulting in many recent submissions to GenBank so there is good data to compare with), and which DNA markers are useful in differentiating species in a given genus (this is the subject of the pdf).

Terri

HI Igor, I looked at your ob # 250592.
By: Judi T. (AvidAmateur)
2016-10-16 18:58:56 PDT (-0700)

One thing that stands out is the reticulation on your specimen is more apparent and appears to cover a larger portion of the stipe than in any of my specimens. Also the stipe in your ob seems more slender and relatively equal over its length, whereas the specimens that I keep finding here have what I call a very robust appearance and are quite bulbous at some point, most often toward the base. Another distinctive feature I have come to recognize among my oddball specimens is a markedly “fat”, overstuffed-cushion-like cap. I can’t tell from the angle of your pictures if this is true for your specimen or not. Finally, the stipe flesh of my specimens always bruise a brownish-pink when sliced. Do you recall if that was the case with your specimen?

Whether or not these differences I notice are natural variations of the same species (perhaps as a result of habitat, climate differences or age) or are indicative of a different species, I certainly don’t have the experience to answer. It does raise a few questions in my mind, however: How far can variations within a species or species group extend before one must consider the specimens to be an entirely separate species? Is that something that can be answered by DNA sequences? For example, if you sequence several specimens in the Pluteus cervinus group, will there be slight variations in the DNA sequences? If so how many variations can be “tolerated” before a distinctly separate species is identified? I’m afraid I don’t even have enough knowledge about how to interpret DNA sequences to even ask the questions that are in my mind in any intelligent way:( I hope you can understand what I am trying to ask.

I hope we haven’t opened Pandora’s Box here while we are trying to identify these specimens that don’t seem to fit neatly into any described species. Variations upon variations upon variations. An evolved new species from one or more of these variations, etc., etc., etc. At any rate, hopefully we will have an answer by next July when they all start popping up again:)

Hello, Judy:-
By: I. G. Safonov (IGSafonov)
2016-10-16 15:50:47 PDT (-0700)

Thanks for the exhaustive info. I checked out obs 245089 and saw that I had actually voted “promising” for your B. pallidus proposal. That one is definitely more consistent with the species concept. Though the stipe lacks any bona fide reticulation below the apex, it’s is not smooth either – there’s definitely some coarse longitudinal ribbing. Sooo, maybe they are all some anomalous form of B. pallidus. I guess we’ll just have to wait for those DNA results to come back one day. I have most of my samples ready to ship, but I am still waiting for the DNA gent to get back to me with a response to my request.
Judy, I just realized that I might have recently picked a weird B. pallidus, too. Check out my obs 250592. That was my original diagnosis but then I changed it to Xerocomus. I had the sample in my possession for a couple of days, but the ditched it. :-( I think in many ways it’s very similar to your obs 245089. What do you think?

Terri and Donna, I did not taste
By: Judi T. (AvidAmateur)
2016-10-16 09:15:18 PDT (-0700)

this specimen. I have many oddball food/drug sensitivities (a few which have caused anaphylactic reactions), so I am super cautious about everything I put in my mouth. Unfortunately, that leaves me with just my senses of sight and smell when it comes to mushroom identification.

Good morning, Igor. Yes, I can confirm
By: Judi T. (AvidAmateur)
2016-10-16 08:22:00 PDT (-0700)

that all the obs listed under your Boletaceae proposal came from the same site. I refer to this area lovingly as the Bolete Farm. All the specimens in the obs under consideration came from the same suburban residential property, under a mature Pin Oak (estimated to be 3 + stories tall and 35-45 years old based on the age of this residential development. Interestingly, all of the specimens were found within an approximate 8’ X 8’ area under this tree, although the root zone (judging from the size of the canopy) would be much larger than that.

Another point of interest, I have an additional ob (# 245089) from this same site that was more clearly identifiable as Boletus pallidus, which Kuo has described as highly variable in appearance and possibly part of a species group.

Thank you for your continued interest in this cryptic mushroom.

Judy,
By: I. G. Safonov (IGSafonov)
2016-10-15 18:32:26 PDT (-0700)

Could you please confirm that all 6 observations I’ve listed under my Boletaceae proposal came from the same site and represent the same species? They look the same to me in the pix. The collection that will get sequenced (ITS locus) is obs 244148.

Judi, did you taste it?
By: Terri Clements/Donna Fulton (pinonbistro)
2016-10-15 18:26:11 PDT (-0700)
Place your bets, everyone. IG is
By: Judi T. (AvidAmateur)
2016-10-15 17:22:16 PDT (-0700)

working on getting a definitive ID through DNA on this mushroom whose identity has confounded us all for 2 years now … always popping up in the same exact place season after season.

Thanks so much, IG.
By: Judi T. (AvidAmateur)
2015-06-29 17:08:26 PDT (-0700)

Appreciate your help!

Thanks, Judy
By: I. G. Safonov (IGSafonov)
2015-06-29 15:44:36 PDT (-0700)

No need to worry. It’s all fixed now. :-)

IG, Not sure where to go from here. Got rid of the mixed collection heading and back .
By: Judi T. (AvidAmateur)
2015-06-29 12:45:11 PDT (-0700)

to where I started (without the extra pics). Guess you need to propose your suggested name again if that is what you want to do. THNX again!

Thanks, IG. For some reason the photos from my
By: Judi T. (AvidAmateur)
2015-06-29 12:40:40 PDT (-0700)

previous ob remained on the page (hidden from my view) when I called up aq new page to post my next observation. There were a ton of photos on there, for sure. Thanks for catching that. Sorry for the confusion.

Wow! What a mix up. Don’t know if the photo uploading mix-up was my computer or MO.
By: Judi T. (AvidAmateur)
2015-06-29 12:33:02 PDT (-0700)

Will delete this ob and correct the photo uploads. So sorry!

Created: 2015-06-29 12:08:54 PDT (-0700)
Last modified: 2016-10-22 10:34:14 PDT (-0700)
Viewed: 358 times, last viewed: 2016-10-27 22:40:58 PDT (-0700)
Show Log