Reassessment of the generic limits for Hydnellum and Sarcodon (Thelephorales, Basidiomycota).

DNA sequences from the nuclear LSU and ITS regions were used for phylogenetic analyses of Thelephorales with a focus on the stipitate hydnoid genera Hydnellum and Sarcodon. Analyses showed that Hydnellum and Sarcodon are distinct genera but that the current division, based on basidioma texture, makes Sarcodon paraphyletic with respect to Hydnellum. In order to make genera monophyletic several species are moved from Sarcodon to Hydnellum and the following new combinations are made: Hydnellumamygdaliolens, H.fennicum, H.fuligineoviolaceum, H.fuscoindicum, H.glaucopus, H.joeides, H.lepidum, H.lundellii, H.martioflavum, H.scabrosum, H.underwoodii, and H.versipelle. Basidiospore size seems to separate the genera in most cases. Hydnellum species have basidiospore lengths in the range 4.45-6.95 µm while the corresponding range for Sarcodon is 7.4-9 µm. S.quercinofibulatus deviates from this pattern with an average spore length around 6 µm. Neotropical Sarcodon species represent a separate evolutionary lineage.

Introduction

The order is a distinctive lineage of , well-known for its almost ubiquitous ectomycorrhizal life style (Tedersoo et al. 2010). Several species have stipitate hydnoid basidiomata (Fig. 1). They have traditionally been divided into four genera, and with hyaline basidiospores, and and with yellow to brown tinted basidiospores (Maas Geesteranus 1975). In both cases the genera within each pair differ in basidioma structure, with and being hard and dry, and and forming softer, fleshier basidiomata. This difference in texture is, however, difficult to assess and a series of recent molecular phylogenetic analyses, as outlined below, have indicated that the traditional, morphology-based generic limits are equivocal.

Figure 1.

Fruiting bodies of and ABCD.

In a recent comprehensive study of stipitate hydnoid species from south-eastern North America, Baird et al. (2013) found that could not be separated from and the genera were hence combined into a more comprehensive . The same study suggested that the generic limits of and need reassessment.

Nitare and Högberg (2012) examined the Nordic species of and included a preliminary molecular phylogeny for the species accepted in . species were also included in non-published test runs and found to be nested among species. They concluded that revisions of limits of both genera were probably necessary. Miscevic (2013) expanded on the results in Nitare and Högberg (2012) by including more sequences for each species and by including a selection of species in published phylogenies. The results were in congruence with Baird et al. (2013) with regard to overall tree topology and again the conclusion was that the limits of and need further study. A recent phylogenetic overview of (Vizzini et al. 2016) and a study of from the Mediterranean region (Loizides et al. 2016) came to similar conclusions, although Vizzini et al. (2016) did not include sequences from several Neotropical species described by Grupe et al. (2015, 2016).

In this paper we analyse ITS and nuclear LSU sequences from a wide selection of species with a focus on and in order to resolve the relationship between these two genera. We also make some nomenclatural changes that follow from the revision of genus circumscriptions. We demonstrate that Neotropical species do not cluster with temperate and boreal species and may be warranted as one or more new genera with more data.

Methods

For the phylogenetic analyses we compiled two datasets. The first dataset consists of nuclear LSU sequences from most genera in and from a majority of the and species occurring in Europe. For our two target genera we chose only sequences generated for this study from recently collected basidiomata. We deliberately excluded sequences from specimens identified as or since these names seem to cover more than just two species and it is currently unclear how the names should be applied (Ainsworth et al. 2010). Since this study is positioned as a revision of the genus limits we were more interested in sequence quality control than a complete coverage of all species reported from Europe.

For our second dataset we chose a different strategy. Here we included ITS sequences from all and species represented among our own sequences and in GenBank as of December 1, 2018. The reason is that many species, and especially the recently described species from tropical regions, are only available as ITS sequences. However, we made no attempt to verify the identifications given in GenBank and do not endorse them as correct.

DNA was extracted from recent dried collections of basidiomata from North Europe. Voucher numbers, herbarium location, and GenBank numbers are given in Table 1. DNA extraction and PCR protocols follow Larsson et al (2018). Sequencing was either done in-house at University of Oslo, or as a commercial service by Macrogen Inc., South Korea. Assembly of chromatograms was done with Sequencher 5.2.4 (Gene Codes Co., Ann Arbor). Aligning was performed either manually using the editor in PAUP* 4.0a (Swofford 2002) or the software ALIVIEW 1.18 (Larsson 2014), or automatically utilising the L-INS-i strategy as implemented in MAFFT v. 7.017 (Katoh and Standley 2013), followed by manual adjustment.

Table 1.

Specimens sequenced or downloaded from GenBank. Herbarium acronyms follow Thiers. Sequences generated for this study are marked in bold.

Species	Voucher	Herb.	GenBank number
			ITS	LSU
Amaurodonaquicoeruleus Agerer	Agerer & Bougher	M	AM490944	AM490944
Amaurodonviridis (Alb. & Schwein.:Fr.) J.Schröt	KH Larsson 14947b	O	MK602707	MK602707
Bankerafuligineoalba (J.C.Schmidt:Fr.) Pouzar	E Larsson 400-13	GB	MK602708	MK602708
Bankeraviolascens (Alb. & Schwein.:Fr.) Pouzar	MV 130902	GB	MK602709	MK602709
Boletopsisleucomelaena (Pers.:Fr.) Fayod	M Krikorev 140912	GB	MK602710	MK602710
Hydnellumaurantiacum (Batsch:Fr.) P.Karst.	RG Carlsson 08-105	GB	MK602711	MK602711
Hydnellum aurantiacum	E Bendiksen 177-07	O	MK602712	MK602712
Hydnellum aurantiacum	O-F-295029	O	MK602713	MK602713
Hydnellumauratile (Britzelm.) Maas Geest.	O-F-294095	O	MK602714	MK602714
Hydnellum auratile	O-F-242763	O	MK602715	MK602715
Hydnellum auratile	J Nitare 110926	GB	MK602716	MK602716
Hydnellumcaeruleum (Hornem.:Fr.) P.Karst.	O-F-291490	O	MK602717	MK602717
Hydnellum caeruleum	E Bendiksen 575-11	O	MK602718	MK602718
Hydnellum caeruleum	E Bendiksen 584-11	O	MK602719	MK602719
Hydnellumcomplicatum Banker	REB 71		KC571711
Hydnellumconcrescens (Pers.) Banker	K(M)134463	K	EU784267
Hydnellumcristatum (G.F.Atk.) Stalpers	REB 169	TENN	JN135174
Hydnellumcumulatum K.A.Harrison	SE Westmoreland 69		AY569026
Hydnellumcyanopodium K.A.Harrison	SE Westmoreland 85		AY569027
Hydnellumdiabolus Banker	KAH 13873	MICH	AF351863
Hydnellumdianthifolium Loizides, Arnolds & P.-A.Moreau	ML61211HY		KX619419
Hydnellumearlianum Banker	REB 375	TENN	JN135179
Hydnellumferrugineum (Fr.:Fr.) P.Karst.	O-F-297319	O	MK602720	MK602720
Hydnellum ferrugineum	E Larsson 356-16	GB	MK602721	MK602721
Hydnellum ferrugineum	E Larsson 197-14	GB	MK602722	MK602722
Hydnellumferrugipes Coker	REB 176		KC571727
Hydnellumgeogenium (Fr.) Banker	O-F-66379	O	MK602723	MK602723
Hydnellum geogenium	O-F-296213	O	MK602724	MK602724
Hydnellum geogenium	E Bendiksen 526-11	O	MK602725	MK602725
Hydnellumgracilipes (P.Karst.) P.Karst.	E Larsson 219-11	GB	MK602726	MK602726
Hydnellum gracilipes	GB-0113779	GB	MK602727	MK602727
Hydnellummirabile (Fr.) P.Karst.	RG Carlsson 11-119	GB	MK602728	MK602728
Hydnellum mirabile	E Larsson 170-14	GB	MK602729	MK602729
Hydnellum mirabile	S Lund 140912	GB	MK602730	MK602730
Hydnellumpeckii Banker	S Svantesson 328	GB	MK602731	MK602731
Hydnellum peckii	E Larsson 174-14	GB	MK602732	MK602732
Hydnellum peckii	E Bendiksen 567-11	O	MK602733	MK602733
Hydnellumpineticola K.A.Harrison	RB 94		KC571734
Hydnellumpiperatum Maas Geest.	REB 322	TENN	JN135173
Hydnellumregium K.A.Harrison	SE Westmoreland 93		AY569031
Hydnellumscleropodium K.A.Harrison	REB 3	TENN	JN135186
Hydnellumscrobiculatum (Fr.) P.Karst.	REB 78	TENN	JN135181
Hydnellumspongiosipes (Peck) Pouzar	REB 52	TENN	JN135184
Hydnellumsuaveolens (Scop.:Fr.) P.Karst.	E Larsson 139-09	GB	MK602734	MK602734
Hydnellum suaveolens	E Larsson 8-14	GB	MK602735	MK602735
Hydnellum suaveolens	S Svantesson 877	GB	MK602736	MK602736
Hydnellumsubsuccosum K.A.Harrison	REB 10	TENN	JN135178
Lenzitopsisdaii L.W.Zhou & Kõljalg	Yuan 2959	IFP	JN169799	JN169793
Lenzitopsisoxycedri Malençon & Bertault	KH Larsson 15304	GB	MK602774	MK602774
Odontiafibrosa (Berk. & M.A.Curtis) Kõljalg	TU115028	TU	MK602775	MK602775
Phellodon cf niger	E Larsson 35-14	GB	MK602782	MK602782
Phellodontomentosus (L.:Fr.) Banker	E Bendiksen 118-10	O	MK602781	MK602781
Pseudotomentellaflavovirens (Höhn. & Litsch.) Svrček	KH Larsson 16190	O	MK602780	MK602780
Sarcodonamygdaliolens Rubio Casas, Rubio Roldán & Català	SC 2011		JN376763
Sarcodonaspratus (Berk.) S.Ito			DQ448877
Sarcodonatroviridis (Morgan) Banker	REB 104	TENN	JN135190
Sarcodon atroviridis	REB 61		KC571768
Sarcodonbairdii A.C.Grupe & Vasco-Pal.	Vasco 990	HUA	KR698938
Sarcodoncolombiensis A.C.Grupe & Vasco-Pal.	Vasco 2084	HUA	KP972654
Sarcodonfennicus (P.Karst.) P.Karst.	S Westerberg 110909	GB	MK602739	MK602739
Sarcodon fennicus	O-F-242833	O	MK602738	MK602738
Sarcodon fennicus	O-F-204087	O	MK602737	MK602737
Sarcodonfuligineoviolaceus (Kalchbr.) Pat.	LA 120818	GB	MK602740	MK602740
Sarcodon fuligineoviolaceus	B Nylén 130918	GB	MK602741	MK602741
Sarcodon fuligineoviolaceus	A Molia 160-2011	O	MK602742	MK602742
Sarcodonfuscoindicus (K.A.Harrison) Maas Geest.	OSC 113622	OSC	EU669228
Sarcodonglaucopus Maas Geest. & Nannf.	RG Carlsson 13-060	GB	MK602743	MK602743
Sarcodon glaucopus	J Nitare 060916	GB	MK602744	MK602744
Sarcodon glaucopus	Å Edvinson 110926	GB	MK602745	MK602745
Sarcodonimbricatus (L.:Fr.) P.Karst.	S Svantesson 355	GB	MK602748	MK602748
Sarcodon imbricatus	J Rova 140829-2	GB	MK602746	MK602746
Sarcodon imbricatus	E Larsson 384-10	GB	MK602747	MK602747
Sarcodonjoeides (Pass.) Bataille	RG Carlsson 11-090	GB	MK602749	MK602749
Sarcodon joeides	K Hjortstam 17589	GB	MK602750	MK602750
Sarcodon joeides	J Nitare 110829	GB	MK602751	MK602751
Sarcodon joeides	REB 270		KC571772
Sarcodonlepidus Maas Geest.	E Grundel 110916	GB	MK602753	MK602753
Sarcodon lepidus	RG Carlsson 10-065	GB	MK602752	MK602752
Sarcodon lepidus	J Nitare 110829	GB	MK602754	MK602754
Sarcodonleucopus (Pers.) Maas Geest. & Nannf.	O-F-296944	O	MK602756	MK602756
Sarcodon leucopus	O-F-296099	O	MK602755	MK602755
Sarcodon leucopus	P Hedberg 080811	GB	MK602757	MK602757
Sarcodonlundellii Maas Geest. & Nannf.	L&A Stridvall 06-049	GB	MK602758	MK602758
Sarcodon lundellii	O-F-242639	O	MK602759	MK602759
Sarcodon lundellii	O-F-295814	O	MK602760	MK602760
Sarcodonmartioflavus (Snell, K.A.Harrison & H.A.C.Jacks.) Maas Geest.	A Delin 110804	GB	MK602763	MK602763
Sarcodon martioflavus	O-F-242435	O	MK602762	MK602762
Sarcodon martioflavus	O-F-242872	O	MK602761	MK602761
Sarcodonpakaraimensis A.C.Grupe & T.W.Henkel	T Henkel 9554	BRG	KM668103
Sarcodonpallidogriseus A.C.Grupe & Vasco-Pal.	Vasco 989	HUA	KR698939
Sarcodonportoricensis A.C.Grupe & T.J.Baroni	TG Baroni 8776	NY	KM668100
Sarcodonquercophilus A.C.Grupe & Lodge	CFMR-BZ-3833	NY	KM668101
Sarcodonquercinofibulatus Pérez-De-Greg., Macau & J.Carbó	JC 20090718-2		JX271818	MK602773
Sarcodonrufobrunneus A.C.Grupe & Vasco-Pal.	Vasco 1989	HUA	KR698937
Sarcodonscabripes (Peck.) Banker	REB 351	TENN	JN135191
Sarcodonscabrosus (Fr.) P.Karst.	O-F-295824	O	MK602764	MK602764
Sarcodon scabrosus	O-F-292320	O	MK602766	MK602766
Sarcodon scabrosus	O-F-360777	O	MK602765	MK602765
Sarcodonsquamosus (Schaeff.) Quél.	O-F-177452	O	MK602768	MK602768
Sarcodon squamosus	E Larsson 248-12	GB	MK602767	MK602767
Sarcodon squamosus	O-F-295554	O	MK602769	MK602769
Sarcodonumbilicatus A.C.Grupe, T.J.Baroni & Lodge	TJ Baroni 10201	NY	KM668102
Sarcodonunderwoodii Banker	REB 50		KC571781
Sarcodonversipellis (Fr.) Nikol.	RG Carlsson 13-057	GB	MK602771	MK602771
Sarcodon versipellis	RG Carlsson 11-085	GB	MK602772	MK602772
Sarcodon versipellis	E Bendiksen 164-07	O	MK602770	MK602770
Sistotremabrinkmannii (Bres.) J.Erikss.	KH Larsson 14078	GB	KF218967	KF218967
Steccherinumochraceum (J.F.Gmel.:Fr.) Gray	KH Larsson 11902	GB	JQ031130	JQ031130
Thelephoracaryophyllea (Schaeff.:Fr.) Pers.	E Larsson 89-09S	GB	MK602776	MK602776
Thelephoraterrestris Ehrh.:Fr.	E Larsson 295-13	GB	MK602777	MK602777
Tomentellastuposa (Link) Stalpers	Th-0764	O	MK602778	MK602778
Tomentellopsispulchella Kõljalg & Bernicchia	KH Larsson 16366	O	MK602779	MK602779

In the phylogenetic analyses we assumed the following minimal partitions for the nrDNA region: ITS1, 5.8S, ITS2 and LSU (approximately 1200 bases of the 5’ end). Two datasets were analysed separately: an LSU dataset only including the LSU region, and an ITS dataset including ITS1, 5.8S and ITS2. We used the automated best-fit tests implemented in PAUP* 4.0a (Swofford 2002) to select optimal substitution models for each complete, non-partitioned dataset (PHYML) and optimal substitution model partitions for each minimal partition (BEAST). Models and partitions were chosen based on BIC score for the BEAST analysis and AICc score for the PHYML analysis. All tests were conducted using three substitution schemes and evaluated substitution models with equal and gamma-distributed among-site rate variation. The tests for the PHYML analysis also evaluated substitution models with invariant sites. The following partitions and models had the highest ranking, according to BIC: ITS1+ITS2 (GTR+G), 5.8S (K80+G), LSU (GTR+G). According to AICc the GTR+I+G model provided the best fit for both the ITS and the LSU datasets.

To generate Bayesian phylogenetic trees (BI) from the alignments we used BEAST 2.4.7 (Bouckaert et al. 2014). We prepared the xml-files for the BEAST 2 runs in BEAUTI 2.4.7 (Bouckaert et al. 2014). We set the substitution model to GTR+G for the LSU run. In the ITS run we set it to HKY+G for 5.8S, since it is the most similar model to K80+G available in the program. Test runs revealed convergence problems due to insufficient data for some substitution rates in the GTR+G model initially used for the ITS1+ITS2 partition, and it was hence changed to HKY+G. In the ITS run the substitution rate of both partitions were estimated independently. We set the trees of the minimal nrDNA partitions as linked in this analysis and the clock models as unlinked. A lognormal, relaxed clock model was assumed for each partition, as test runs had shown that all partitions had a coefficient of variation well above 0.1 (i.e. implying a relatively high rate variation among branches). The clock rate of each partition was estimated in the runs, using a lognormal prior with a mean set to one in real space. We set the growth rate prior to lognormal, with a mean of 5 and a standard deviation of 2. We ran the Markov Chain Monte Carlo (MCMC) chains of both datasets for 20 million generations with tree and parameter files sampled every 1,000 generations. The analyses all converged well in advance of the 10 % burn-in threshold, had ESS values well above 200 for all parameters, and chain mixing was found to be satisfactory as assessed in TRACER 1.6.0 (Rambaut et al. 2014). After discarding the burn-in trees, maximum clade credibility trees were identified by TREEANNOTATOR 2.4.7 (Bouckaert et al. 2014).

To generate Maximum Likelihood (ML) gene trees we used PHYML 3.1 (Guindon et al. 2010). We set the substitution model to GTR+I+G for both the ITS and LSU datasets. Tree topology search was conducted using NNI+SPR, with ten random starting trees. Non-parametric bootstrap analyses with 1000 replicates were performed on the resulting trees.

Results

Seventy-five specimens from the genera , , , , , , , , , , and , were sequenced for this study. In addition, 39 sequences were downloaded from public databases (GenBank, UNITE) including outgroup sequences of () and () included in the LSU dataset. The ITS analyses were rooted by the default method (BEAST) or left unrooted (PHYML).

The aligned LSU dataset consisted of 1443 nucleotide positions. After exclusion of ambiguous regions 1377 positions remained for the analyses. BI returned a tree where the focus genera and are distributed over two strongly supported clades. The larger of these clades includes the type of , , and an additional 17 species, all except one forming strongly supported terminal clades. Nine of these taxa are currently placed in . With a few exceptions the relationships within are not resolved. and are recovered as a strongly supported group; and are grouped with 0.97 posterior probability support; , , and form a subclade with 0.97 posterior probability support; and finally and form a strongly supported clade. The type of , , and three other species form the second main clade. The three sequences of cluster together but the clade is unsupported. and are recovered as sister clades but the support for this arrangement is weak.

For target taxa the ML tree is essentially similar to the BI tree with strong support for the similarly composed and clades (Fig. 2). As for the BI analysis the relationships among species within and are not resolved except for a weak to moderate support for grouping with and with . , , and also group together in the ML tree but without support. Again does not get support and is not separated from .

Figure 2.

Maximum likelihood analyses of LSU dataset for . Branches in bold have a posterior probability value of 1 in Bayesian inference and 100% bootstrap support in ML analysis, if not otherwise indicated by a figure. Lower support values on other branches are indicated by figures. and are used as outgroup (branch lengths shortened).

The aligned ITS dataset consisted of 1068 nucleotide positions of which 505 remained for the analyses after removal of ambiguous regions. Bayesian inference produced a tree with two strongly supported clades (Fig. 3). The smaller one, which we here informally call “Neosarcodon”, contains nine species, all with a distribution in the tropical and subtropical Americas. Remaining and taxa, including both type species, formed the other clade. Within the latter clade two subclades are visible, corresponding to the genera and , and with the same delimitation as in the LSU trees. Only the subclade has strong support. Within each larger clade several groups of taxa received moderate to strong support. The reader is referred to Fig. 2 for further details.

Figure 3.

Ultrametric default rooted BEAST tree of ITS dataset for and . Posterior probability values and bootstrap percent support from ML analysis are indicated by figures; na = not applicable.

The ML tree recovered the same two main clades with strong support but could not resolve the relationships within the larger / clade. In the ML tree the clade corresponding to in the LSU tree is correctly identified but not supported while the clade corresponding to appears polyphyletic.

Based on these results we hereby revise the limits of the two genera by moving a number of species from to . Consequently the genus description for must be emended while the genus description for can remain unaltered.

Taxonomy

P.Karst., Meddn Soc. Fauna Flora fenn. 5: 41 (1879).

Type species.

(Scop.:Fr.) P.Karst. (1879)

Basionym.

Scop.:Fr. (1772)

Basidiomata with pileus and stipe, single or concrescent; pileus thin to thick, at first smooth and velutinous, when mature felted, fibrillose, scaly, ridged, or irregularly pitted and scrupose, mostly brownish but also with white, olive yellowish, orange, purplish or bluish colours, often concentrically zonate; stipe narrow to thick, solid, mostly short; hymenophore hydnoid, usually strongly decurrent; context from soft and brittle to corky or woody; hyphal system monomitic, septa with or without clamps, context hyphae inflated or not; cystidia lacking; basidia narrowly clavate, producing four sterigmata; basidiospores with irregular outline, more or less lobed, verrucose, brownish. Terrestrial, forming ectomycorrhiza with forest trees.

(Rubio Casas, Rubio Roldán & Català) E.Larss., K.H.Larss. & Kõljalg comb. nov.

830570

Rubio Casas, Rubio Roldán & Català, Boln Soc. Micol. Madrid 35: 44−45. 2011. Holotype: Spain, Tamajón, Barranco la Jara. L. Rubio-Casas & L. Rubio-Roldán, AH 42113.

(P.Karst.) E.Larss., K.H.Larss. & Kõljalg comb. nov.

830571

P.Karst., Bidr. Känn. Finl. Nat. Folk 37: 104. 1882. Type: not indicated (neotype: H, designated by Maas Geesteranus & Nannfeldt 1969: 406)

(Kalchbr.) E.Larss., K.H.Larss. & Kõljalg comb. nov.

830572

Kalchbr., in Fries, Hymenomyc. eur. (Upsaliae): 602. 1874. Holotype: Slovakia, Presovsky kraj, Olaszi. C. Kalchbrenner, UPS F-173546.

(K.A.Harrison) E.Larss., K.H.Larss. & Kõljalg comb. nov.

830573

K.A.Harrison, Can. J. Bot. 42: 1213. 1964. Holotype: USA, Washington, Olympic Nat. Park, A.H. Smith. MICH 10847.

(Maas Geest. & Nannf.) E.Larss., K.H.Larss. & Kõljalg comb. nov.

830574

Maas Geest. & Nannf., Svensk bot. Tidskr. 63: 407. 1969. Holotype: Sweden, Uppland, Börje par., J. Eriksson. UPS F-013955.

(Pass.) E.Larss., K.H.Larss. & Kõljalg comb. nov.

830575

Pass., Nuovo G. bot. ital. 4: 157. 1872. Holotype: Italy, Emilia-Romagna, Collecchio, G. Passerini. PAD.

(Maas Geest.) E. Larss., K.H.Larss. & Kõljalg comb. nov.

830576

Maas Geest., Verh. K. ned. Akad. Wet., tweede sect. 65: 105. 1975. Holotype: The Netherlands, Lochem, Ampsen, G. & H. Piepenbroek. L.

(Maas Geest. & Nannf.) E.Larss., K.H.Larss. & Kõljalg comb. nov.

830577

Maas Geest. & Nannf., Svensk bot. Tidskr. 63: 421. 1969. Type: Sweden, Uppland, Storvreta, S. Lundell & J.A. Nannfeldt, distributed in S. Lundell & J.A. Nannfeldt exs. suec. as number 252 (lectotype, designated here, UPS F-010975; MycoBank No.: MBT387081). The UPS herbarium has two copies of the exsiccate and the specimens of are registered as F-010975 and F-013956, respectively. From F-010975 an ITS2 sequence has been generated [GenBank MK753037] and this specimen is here selected as lectotype).

(Snell, K.A.Harrison & H.A.C.Jacks.) E.Larss., K.H.Larss. & Kõljalg comb. nov.

830578

Snell, K.A.Harrison & H.A.C.Jacks., Lloydia 25: 161. 1962. Holotype: Canada, Quebec, Ste Anne de la Pocatière, H.A.C. Jackson & W.H. Snell 13 Sep. 1954, BPI 259438.

(Fr.) E.Larss., K.H.Larss. & Kõljalg comb. nov.

830579

Fr., Anteckn. Sver. Ätl. Svamp.: 62. 1836. Type: not indicated (neotype: Sweden, Småland, Femsjö, S. Lundell, UPS F-013954, designated by Maas Geesteranus & Nannfeldt 1969: 426)

(Banker) E.Larss., K.H.Larss. & Kõljalg comb. nov.

830580

Banker, Mem. Torrey bot. Club 12: 147. 1906. Holotype: USA, Connecticut, NY 776131.

(Fr.) E.Larss., K.H.Larss. & Kõljalg comb. nov.

830581

Fr., Öfvers. K. Svensk. Vetensk.-Akad. Förhandl. 18(1): 31. 1861. Type: not indicated (neotype: Sweden, Uppland, Danmark par., J. Eriksson & H. Nilsson, UPS F-013958, designated by Maas Geesteranus & Nannfeldt 1969: 430)

Quél. ex P.Karst., Revue mycol., Toulouse 3 (no. 9): 20 (1881).

(L.:Fr.) P.Karst. (1881)

L.:Fr. (1753).

Basidiomata with pileus and stipe, single or concrescent; pileus thin to thick, at first smooth and velutinous, when mature smooth or scaly, brownish; stipe thick, solid, mostly short; hymenophore hydnoid, usually strongly decurrent; context soft and brittle; hyphal system monomitic, septa with clamps, context hyphae inflated; cystidia lacking; basidia narrowly clavate, producing four sterigmata; basidiospores with irregular outline, more or less lobed, verrucose, brownish. Terrestrial, forming ectomycorrhiza with forest trees.

Discussion

In this paper we show that the current morphology-based concepts of and do not correspond to monophyletic subgroups within the . The characters traditionally used to separate the two genera do not reflect true relationships. These characters, however, are vague and open to subjectivity; hence it is not surprising that they have now been shown to be unreliable. Maas Geesteranus (1975) pointed to the context structure and consistency as the main differentiating character. For he describes the context as “... fibrillose, soft or tough, corky to woody, more or less duplex, zoned, ...” and hyphae are said to be “...usually not inflating ...”. In the same structures are described as “... fleshy, brittle, soft or firm (never corky or woody), not duplex, not zoned ...” and “...hyphae inflating ...”. While these morphological characteristics remain true for , the corresponding descriptions for had to be emended.

Instead of context structure it seems that average basidiospore size may in most cases offer a possibility to separate a species from one belonging to . Table 2 summarizes basidiospore measurements from the literature. Average basidiospore lengths in fall between 4.45 and 6.95 µm while the same figures for are 7.4 and 9 µm, ornamentation excluded. However, clearly deviates from this pattern. According to measurements in the protologue (Pérez-de-Gregorio et al. 2011) and in Vizzini et al. (2013) average basidiospore length was measured to 6.95 and 7.0, respectively, but then included the ornamentation. Measurements excluding ornamentation would be approximately 1 µm less. Clearly, for basidiospore length alone will not be decisive for genus placement.

Table 2.

Basidiospore measurements for and from the literature. Sources: B = Baird et al. (2013), M = Maas Geesteranus (1975), J = Johannesson et al. (1999). All measurements exclude ornamentation. For species treated in this paper names follow our new classification. For other species names are according to cited authors.

Species	Measurements	Mean length
Hydnellumaurantiacum (M)	(5.8−)6−6.7 × (4−)4.3−4.9	6.35
Hydnellumauratile (M)	4.9−5.8 × 3.6−4.5	5.35
Hydnellumcaeruleum (M)	5.4−6(−6.3) × 3.4−4.3	5.70
Hydnellumcompactum (Pers.:Fr.) P.Karst. (M)	5.4−6.3 × 3.6−4.5	5.85
Hydnellumcomplicatum (B)	4−5 × 3−5	4.50
Hydnellumconcrescens (M)	5.4−6.1 × (3.6−)4−4.5	5.75
Hydnellumcristatum (B)	5−6 × 4−5	5.50
Hydnellumcruentum K.A.Harrison (B)	4−5 × 3−4	4.50
Hydnellumcumulatum (M)	4.3−5.6 × 3.6−4.3	4,95
Hydnellumdiabolus (B)	6−7 × 5−6	6.50
Hydnellumearlianum (B)	5−6 × 4−5	5.50
Hydnellumfennicum (M)	6.3−7.6 × 4.5−5.2	6.95
Hydnellumferrugineum (M)	(5.4−)5.8−6.3 × 3.6−4.5	6.05
Hydnellumferrugipes (B)	5−7 × 5−6	6.00
Hydnellumfuligineoviolaceum (M)	5.4−6.5 × 4−4.7(−5.4)	5.95
Hydnellumgeogenium (M)	4.5−5.2 × 3.1−3.6	4.85
Hydnellumglaucopus (M)	(5−)5.4−5.8(−6.3) × (3.6−)4−4.5	5.60
Hydnellumgracilipes (M)	4.3−4.6 × 2.7−3.6	4.45
Hydnellumjoeides (M)	5.4−5.8 × 3.6−4.2	5.60
Hydnellumlepidum (M)	5.8−6.3 × 3.6−4.3	6.05
Hydnellumlundellii (M)	4.9−5.8 × 3.6−4.2	5.35
Hydnellummartioflavum (M)	5−6.3 × 3.6−4.5	5.65
Hydnellumpeckii (M)	4.9−5.4 × 3.8−4	5.15
Hydnellumpineticola (B)	5−7 × 4−6	6.00
Hydnellumpiperatum (B)	4−6 × 4−5	5.00
Hydnellumscabrosum (M)	(5.4−)6.3−7.3 × (3.6−)4−5	6.80
Hydnellumscleropodium (B)	4−6 × 3−4	5.00
Hydnellumspongiosipes (B)	6−7 × 5−6	6.50
Hydnellumsuaveolens (M)	4−5 × 3−3.6	4.50
Hydnellumsubsuccosum (B)	5−6 × 4−6	5.50
Hydnellumversipelle (M)	4.5−5.5 × 3.5−4.5	5.00
Hydnellumunderwoodii (B)	5−7 × 5−6	6.00
Sarcodonatroviridis (B)	8−9 × 7−8	8.50
Sarcodonexcentricus R.E.Baird (B)	8−9 × 6−8	8.50
Sarcodonharrisonii R.E.Baird (B)	7−9 × 6−8	8.00
Sarcodonleucopus (M)	(6.7−)7.2−7.6(−9) × 4.5−5.6	7.40
Sarcodonimbricatus (M)	7.2−8.2 × 4.9−5.4	7.70
Sarcodonscabripes (B)	8−10 × 7−9	9.00
Sarcodonsquamosus (J)	7.2−8.2 × 4.9−5.4	7.70

Not all sequences from species described as spp. were recovered within either or . In our ITS-only analyses nine species formed a well-supported clade of their own, separated from sensu stricto and (Fig. 3). This clade, here informally called “Neosarcodon”, contains species collected in tropical and subtropical regions of the Western Hemisphere and may represent one or several distinct genera. However, further analyses based on an expanded dataset using more conservative molecular markers would be required to definitely identify any new higher taxa in the group.

The failure to generate support for and in the ITS-only analyses reflects the large genetical distances present among the species within this marker. Our general experience with the ITS region for thelephoralean target genera is that species are extremely well separated and the internal variation surprisingly low, even when a large number of specimens from both Europe and America are considered. On the other hand, the genetical difference among species is moderate to high, making alignments difficult and prone to ambiguities. In our ITS analyses we chose to remove ambiguous regions, thus halving the number of nucleotide positions suggested by automatic alignment through MAFFT. This seems to have affected the ML analyses most. However, the ITS analyses only served to position neotropical species and the results clearly show that they belong to a separate lineage.

Otto (1997) suggested that is a later synonym of and that the species we now call should be named (Schaeff.) Banker. The name change is based on a reinterpretation of Batsch’s original illustration, which, according to Otto, clearly shows the same species as . In phylogenetic analyses and are sister taxa and during our study we have sequenced several specimens identified as that turned out to be . Thus separating these species can be hazardous and to interpret illustrations must be even harder. We currently do not accept this unfortunate name change.

The present study will serve as the basis for further exploration of species limits within and . As has been demonstrated for the genera, many species interpretations are in need of revision. Over the years we have found numerous specimen misidentifications as well as specimens that could not be assigned to pre-existing names. A closer inspection of the ITS tree in Fig. 3, where we let the terminals retain the identifications given in GenBank, shows some examples. The American sequence of (KC571772) does not cluster with the European representative of the same species (MK602751) and the American sequence named seems to be identical to what is in Europe called . Considering that many stipitate hydnoid species are red-listed and used as indicators of forests in need of conservation (Ainsworth 2005, Nitare 2019), it is of utmost importance to sort out the taxonomy of these species.