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Three new combinations of Butyriboletus (Boletaceae)

KUAN ZHAO1,2, GANG WU1, ROY E. HALLING3 & ZHU L. YANG1,* 1 Key Laboratory for Plant Diversity and Biogeography of East Asia, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming 650201, China 2 University of Chinese Academy of Sciences, Beijing 100049, China 3 Institute of Systematic Botany, New York Botanical Garden, Bronx, NY 10458, USA *e-mail: [email protected]

Abstract

Boletus peckii, B. pulchriceps and B. roseopurpureus, which were originally described from North America, are character- ized by a yellow tube layer that often bruises blue, a yellow reticulated especially at the apex, firm yellow-tinged flesh that often turns blue when exposed, smooth spores and an interwoven trichodermic . The phylogenetic positions of the three species are inferred by molecular phylogenetic analysis based on DNA sequences of four gene markers (ITS, nrLSU, tef1-α and rpb1). Both morphological features and molecular phylogenetic evidence indicate that these three boletes belong to Butyriboletus, and thus, should be transferred to the .

Introduction

Butyriboletus D. Arora & J.L. Frank (2014: 466), typified by appendiculatus Schaeff. (1774: 130), was recently established to accommodate the “butter boletes”, which were historically placed in Boletus sect. Appendiculati Konrad & Maubl. ex. Lannoy & Estadès (2001: 60) (Arora & Frank 2014). Later on, an additional four species, namely B. cepaeodoratus Taneyama & Har. Takah. (2013: 463), B. fuscoroseus Smotl. (1912: 47), B. roseogriseus Šutara et al. (2014: 7) and B. ventricosus Taneyama & Har. Takah. (2013: 461), were transferred to the genus (Vizzini 2014a). To date, Butyriboletus has harbored some 18 species, all distributed in the Holarctic regions (Zang 2006; Assyov 2012; Takahashi et al. 2013; Arora & Frank 2014; Li et al. 2014; Šutara et al. 2014). In recent phylogenetic analyses, three species, Boletus peckii Frost (1878: 45), B. pulchriceps Both, Bessette & R. Chapm. (2000: 143) and B. roseopurpureus Both, Bessette & Roody (2000: 150) nested in the same clade (“regius clade” in Nuhn et al. 2013; Clade 46 in Wu et al. 2014 ) with Bu. appendiculatus and its allies, with high support values, indicating that they all may belong to Butyriboletus. In this study, morphological comparisons and phylogenetic analyses based on four gene markers, i.e., internal transcribed spacer (ITS), nuclear ribosomal large subunit DNA (nrLSU), translation elongation factor 1-α (tef1-α) and the largest subunit of RNA polymerase II (rpb1), were conducted to confirm the taxonomic status and phylogenetic position of these species.

Materials and Methods

Sampling Specimens examined are deposited in the Buffalo Museum of Science (BUF), Philipps University Marburg (MB), the New York Botanical Garden (NY), University of Tennessee (TENN), Farlow Herbarium of Harvard University (FH) and the Herbarium of Cryptogams of Kunming Institute of Botany, Chinese Academy of Sciences (HKAS). The species appear in alphabetical order by species epithet. The generic name Boletus is abbreviated as “B.”, Butyriboletus as “Bu.”, as “C.”, while “” as “E.”

Accepted by Genevieve Gates: 7 Oct. 2015; published: 11 Nov. 2015 51 Morphological studies Macroscopic characters were from the previous descriptions of the species. For microscopic observation, the dried materials were sectioned and mounted in 5% KOH solution. Sections of the pileipellis were made halfway across the pileus radius. Pileipellis, basidia, , pleuro- and cheilocystidia were obtained using an Axioskop 40 microscope following the standard method described in previous studies (Li et al. 2009, 2011; Zhu et al. 2014, 2015; Wu et al. 2015). Microscopic features were from hand drawn by the first author. In the descriptions of basidiospores, the abbreviation [n/m/p] means n basidiospores measured from m basidiomata of p collections; Q is used to mean

“length/width ratio” of a spore in side view; Qm means average Q of all basidiospores ± sample standard deviation.

Molecular studies Protocols for DNA extraction, PCR, and sequencing followed those in Zhao et al. (2014a, b) and the references therein. Eighty sequences, including 53 newly generated in this study and 27 retrieved from GenBank, were used in the phylogenetic analyses. DNA sequences of four loci (ITS, nrLSU, tef1-α and rpb1) were aligned with MAFFT v6.8 (Katoh et al. 2005) and manually optimized in BioEdit v7.0.9 (Hall 1999). The four single-gene datasets were then analyzed independently using the Maximum Likelihood (ML) method to detect the topologies of the four genes before they are concatenated together. Due to no significant incongruence detected (BS>70%, Nuhn et al. 2013), the resulting four alignments were then concatenated using Phyutility (Smith & Dunn 2008). Unavailable sequences of the loci of a few species were treated as missing data. The datasets were then analyzed using RAxML v7.2.6 (Stamatakis 2006) and MrBayes v3.1.2 (Ronquist & Huelsenbeck 2003) for ML and Bayesian Inference (BI), respectively. For both BI and ML analyses, the substitution model suitable for the four gene partitions was individually determined using the Akaike Information Criterion (AIC) complemented in MrModeltest v2.3 (Nylander 2004). GTR+I+G was chosen as the best model for all the datasets. For ML analysis, all parameters were kept default and the supports were calculated using nonparametric bootstrapping with 1000 replicates. BI analysis using selected models and four chains were conducted by setting generations to three million and stoprule command with the value of stopval set to 0.01. Trees were sampled every 100 generations. The first 25% generations were discarded as burn-ins and posterior probabilities (PP) were then calculated from the posterior distribution of the retained Bayesian trees. To confirm the phylogenetic position of the three species, a preliminary analysis of a combined dataset (nrLSU, tef1-α and rpb1), containing the main genera of Boletaceae as indicated in Nuhn et al. (2013) and Wu et al. (2014), was performed. The GenBank accession numbers of the taxa are listed in Table S1. Based on this analysis, Caloboletus calopus (Pers.) Vizzini (2014b: 1), Exsudoporus frostii (J.L. Russell) Vizzini, Simonini & Gelardi (2014c: 1) and Exsudoporus sp. (HKAS 52661) are close to Butyriboletus, and thus, were selected as outgroups in our concise analysis, which includes the alignment of ITS, nrLSU, tef1-α and rpb1 of 10 species of Butyriboletus and the above mentioned three outgroup taxa.

Results

The result of our preliminary analysis showed that members of Butyriboletus formed an independent clade in the family Boletaceae (Fig. S1). This clade is sister to Exsudoporus, with high support values (BS = 98%). The concise dataset included 10 species of Butyriboletus, and the outgroup taxa Caloboletus calopus, Exsudoporus frostii and Exsudoporus sp. The alignment contained 2578 nucleotide sites (including gaps), consisting of 654, 810, 512 and 602 sites (including gaps) for ITS, nrLSU, tef1-α and rpb1, respectively. Only the tree inferred from the ML analysis is shown herein (Fig. 1), as the Bayesian analysis produced nearly identical estimates of tree topologies. In the phylogenetic analysis, Boletus peckii, B. pulchriceps and B. rosepurpureus were grouped together with the “butter boletes”, forming a monophyletic clade, with relatively high statistical supports (BS= 97%, PP=1.00) (Fig. 1). Morphologically, the three species mentioned above all have a yellow hymenophore, a yellow reticulate stipe especially at the apex, a yellowish flesh, a bluish color change, smooth spores and an interwoven trichodermic pileipellis (Fig. 2–4), which are in accordance with the characters of Butyriboletus (Smith & Thiers 1971; Bessette et al. 2000; Arora & Frank 2014). Thus, both morphology and molecular analysis supported the recognition of these species as members of Butyriboletus.

52 • Phytotaxa 234 (1) © 2015 Magnolia Press ZHAO ET AL. FIGURE 1. Maximum-Likelihood phylogenetic tree generated from combined analysis based on DNA sequences of 4 molecular markers (ITS, nrLSU, tef1-α and rpb1). Support values BS >50% for ML and PPs >0.95 from BI are indicated along nodes.

Taxonomy

Butyriboletus peckii (Frost) Kuan Zhao, Zhu L. Yang & Halling, comb. nov. Fig. 2a–d MycoBank: MB 812757

Basionym: Boletus peckii Frost, Ann. Rep. N.Y. St. Mus. Nat. Hist. 29: 45, 1878

Three new combinations of Butyriboletus Phytotaxa 234 (1) © 2015 Magnolia Press • 53 Pileus 2.5–9.5 cm in diameter, convex, becoming broadly convex to nearly plane in age; surface dry, finely tomentose, deep red to dull red or rose red, fading to brownish rose to dull brown or buffy brown from the disc outward, margin retain its color longer than the disc; context whitish to pale yellow, bluing slightly when exposed. Hymenophore surface at first bright yellow to golden yellow, becoming dingy olive yellow to brownish yellow in age, bluing then slowly staining brownish when bruised; pores circular to somewhat elongate, 2–3 per mm; tubes 4–15 mm deep. Stipe 4–9 × 1–1.6 cm, mostly equal, gradually broadening towards base, dry, solid, minutely and densely pruinose, rosy red to dark red or purplish red with yellow at the apex, typically reticulate nearly overall or at least on the upper portion, reticulation usually distinct; basal mycelium whitish to pale yellow. Odor not distinct. Taste unpleasant, typically bitter or somewhat tart (Smith & Thiers 1971; Bessette et al. 2000).

FIGURE 2. Microscopic features of Butyriboletus peckii (from TENN 027510). a. Basidiospores. b. Pleurocystidia and cheilocystidia; c. Basidia and a pleurocystidium; d. Pileipellis. Bars: a–c= 10 μm; d = 20 μm.

Basidiospores [40/1/1] 9–13 (14) × 3.5–5 μm [Q = (2.40) 2.44–3.25 (3.67), Qm =2.83 ± 0.31], subfusiform, subhyaline in KOH. Hymenophoral trama boletoid. Basidia 24–35 × 7–10 μm, clavate, hyaline in KOH, 4-spored. Pleurocystidia 26–38 × 8–12 μm, smooth and thin-walled, hyaline in KOH, narrowly lageniform, lageniform to fusoid- ventricose; cheilocystidia similar to pleurocystidia in shape but smaller. Pileipellis composed of interwoven hyphae 4–6 μm in diameter, smooth, thin-walled, hyaline. Clamp connections absent. Habitat: Scattered or in groups on the ground in broadleaf forests and mixed woods, especially with and . Known distribution: Known from eastern North America. Material examined: USA. Tennessee, Sevier, Gatlinburg, 06 Aug 1963, TENN 027510.

54 • Phytotaxa 234 (1) © 2015 Magnolia Press ZHAO ET AL. Notes: Boletus peckii was transferred to the genus Caloboletus for its bitter taste (Vizzini 2014b). However, recent phylogenetic analyses indicated that this species nested with Bu. appendiculatus and its allies (“regius Clade” in Nuhn et al. 2013; Clade 46 in Wu et al. 2014; this study); Morphologically, its context turns blue when exposed and then becomes light red (Smith & Thiers 1971; Bessette et al. 2000), which is also different from the bluish color change in Caloboletus (Zhao et al. 2014a).

Butyriboletus pulchriceps (Both, Bessette & R. Chapm.) Kuan Zhao, Zhu L. Yang & Halling, comb. nov. Fig. 3a–d MycoBank: MB 812758

Basionym: Boletus pulchriceps Both, Bessette & R. Chapm, North American Boletes, A Color Guide to the Fleshy Pored Mushrooms (Syracuse): 143, 2000

FIGURE 3. Microscopic features of Butyriboletus pulchriceps (from the type). a. Basidiospores. b. Pleurocystidia and cheilocystidia; c. Basidia and a pleurocystidium; d. Pileipellis. Bars: a–c= 10 μm; d = 20 μm.

Pileus 8–15 cm in diameter, hemispherical to convex, margin strongly incurved and wavy when young; surface dry, appressed-tomentose but sub-shiny to the naked eye, pale pink over the disc, deeper pink over the marginal areas, fading in age to ; context pale yellow, deep yellow above tubes, bluing weakly when exposed but soon fading to whitish. Hymenophore surface yellow at first, brownish olive when mature, bluing weakly or unchanging when

Three new combinations of Butyriboletus Phytotaxa 234 (1) © 2015 Magnolia Press • 55 injured; pores angular, 1–2 per mm; tubes 8–17 mm long, concolorous with the surface, not bluing or only weakly bluing when bruised. Stipe 6.5–7.5 × 1–2 cm, tapered upwards, reticulated over the upper half to two-thirds, the reticulation not strongly raised, with simple, narrow and elongated meshes, the meshes white, the interspaces finely pruinose, yellow near the apex, white with a very pale flush of pink downward; context pale yellow throughout, not bluing when exposed; basal mycelium white. Odor nutty, sweet. Taste not distinct (Bessette et al. 2000).

Basidiospores [40/1/1] (10) 11–14 × 3–4.5 (5) μm [Q = (2.75) 2.88–3.83 (4.00), Qm =3.31 ± 0.32], subfusiform, subhyaline in KOH. Basidia 24–34 × 6–10 μm, clavate, hyaline in KOH, 4-spored. Hymenophoral trama boletoid. Pleurocystidia 22–40 × 6–8 μm, narrowly lageniform to lageniform, sometimes narrowly clavate, smooth and thin- walled, hyaline in KOH; cheilocystidia 30–44 × 9–13 μm, similar to pleurocystidia in shape. Pileipellis composed of interwoven hyphae 4–6 μm in diameter, smooth, thin-walled, hyaline; end cells not differentiated. Clamp connections absent. Habitat: Solitary, scattered, or in groups on the ground under oak. Known distribution: Known from Arizona, southwest of the United States. Material examined: USA. Arizona, Cochise County, Prope Chiricahua Mountains, Aug 1991, R. Chapman 0945 (BUF, type). Notes: Butyriboletus pulchriceps is closely related to Bu. peckii. However, Bu. pulchriceps has a pink to pinkish yellow pileal surface and has no distinctive taste, whereas the pileal surface of Bu. peckii is rose red to deep red and it has a bitter taste (Smith &Theirs 1971; Bessette et al. 2000). Butyriboletus pulchriceps is similar to Bu. roseopurpureus and B. speciosus. However, the pileal surfaces of the latter two species are bright red or purplish red, not pink to pinkish yellow. Furthermore, the context of Bu. roseopurpureus and B. speciosus will turn blue rapidly and obviously when injured, whereas that of Bu. pulchriceps blues weakly (Smith & Thiers 1971; Bessette et al. 2000).

Butyriboletus roseopurpureus (Both, Bessette & Roody) Kuan Zhao, Zhu L. Yang & Halling, comb. nov. Fig. 4a–d MycoBank: MB 812759

Basionym: Boletus roseopurpureus Both, Bessette & Roody, North American Boletes, A Color Guide to the Fleshy Pored Mushrooms (Syracuse): 150, 2000

Pileus 7–15 cm in diameter, convex, becoming broadly convex to nearly plane in age, margin incurved at first, later decurved; surface viscid-tacky in wet weather but drying quickly, velvety-tomentose at first, becoming appressed- tomentose to fibrillose, the fibrils darker, narrow; a striking pinkish purple when fresh, becoming dark purplish pink to dark purplish red, at times somewhat mottled in these colors, the marginal areas becoming grayish in age; context pale yellow, instantly dark blue when exposed but soon fading to pale slate color. Hymenophore surface at first lemon yellow or more golden, becoming yellowish olive to greenish olive at maturity, instantly dark blue to greenish blue when bruised, soon fading to pale greenish blue or greenish gray; pores circular, 1–2 per mm; tubes 5–10 mm deep, concolorous with the surface, bluing rapidly when injured. Stipe 4.5–8.5 × 2–4 cm, equal or slightly tapered downwards, reticulated near the entire length, or at least over the upper two-thirds, and the lower third appressed-tomentose, the reticultion in part strongly raised; bright yellow over most of the length of the stipe, occasionally with burgundy red areas near the base; surface instantly staining dark blue when bruised; context deep yellow, in basal areas burgundy red in some specimens, instantly bluing when exposed but soon fading to pale slate color; basal mycelium white. Odor fragrant. Taste very sour, like lemon (Bessette et al. 2000).

Basidiospores [60/2/2] (9) 10–13 × (3) 3.5–5 μm [Q = 2.20–2.88 (3.00), Qm =2.59 ± 0.24], subfusiform, subhyaline in KOH. Basidia 24–35 × 6–10 μm, clavate, hyaline in KOH, 4-spored. Hymenophoral trama boletoid. Pleurocystidia 32–44 × 10–15 μm, smooth and thin-walled, hyaline in KOH, lageniform to fusoid-ventricose; cheilocystidia similar to pleurocystidia in shape but smaller. Pileipellis composed of interwoven hyphae 4–6 μm in diameter, smooth, thin- walled, hyaline; end cells not differentiated. Clamp connections absent. Habitat: Solitary or in small groups in mixed woods of oak, beech, hemlock and maple. Known distribution: Known from eastern North America. Materials examined: USA. New York, Erie County, Ridge Park, 27 July 1995, E.E. Both 3765 (BUF, type); Tennessee, Great Smoky Mountains National Park, 24 Jul 2008, DLJ-TN-08-17 (NY 01042656).

56 • Phytotaxa 234 (1) © 2015 Magnolia Press ZHAO ET AL. FIGURE 4. Microscopic features of Butyriboletus roseopurpureus (from NY 01042656). a. Basidiospores. b. Pleurocystidia and cheilocystidia; c. Basidia and a pleurocystidium; d. Pileipellis. Bars: a–c= 10 μm; d = 20 μm.

Notes: Bessette et al. (2000) separated B. roseopurpureus from B. speciosus (1874: 2) on account of its pileal surface with purplish tone and the smaller spores. However, according to a recent study (Arora & Frank 2014), the ITS sequences obtained from several collections of Bu. roseopurpureus are identical to those of Boletus speciosus collections from TENN sequenced by Li et al. (2014). Due to the lack of well dried specimens and sequences of tef1-α and rpb1 of B. speciosus for a multi-gene analysis, its systematic position can only be settled in the future.

Acknowledgments

We are very grateful to Kathryn H. Leacock at the Buffalo Museum of Science (BUF), Degreef Jérôme at the National Botanical Garden of Belgium (BR), Gerhard W. Kost at Philipps University Marburg (MB), B. Eugene Wofford at University of Tennessee (TENN) and Genevieve Lewis-Gentry at Farlow Herbarium of Harvard University (FH) for providing collections on loan. This study was supported by the Funds for International Cooperation and Exchange of the National Natural Science Foundation of China (No. 31210103919) and the CAS/SAFEA International Partnership Program for Creative Research Teams.

Three new combinations of Butyriboletus Phytotaxa 234 (1) © 2015 Magnolia Press • 57

and rpb1 KJ619481 KT002621 KF030359 KT002622 KT002623 - KT002625 KT002626 KT002627 KT002628 KF739741 KJ184565 KT002629 KF030372 - KT002630 KT002632 KT002631 KJ184560 KT002624 KF112512 Bu. appendiculatus Bu. -α tef1 KJ619472 KT002633 JQ327025 KT002634 KT002635 JQ327026 KT002637 KT002638 KT002639 KT002640 KF739779 KJ184571 KT002641 KF030410 - KT002642 KT002644 KT002643 KJ184566 KT002636 KF112169 GenBank Accessions GenBank nrLSU KJ605677 KT002609 AF456837 KT002610 KT002611 JQ326999 KT002613 KT002614 KT002615 KT002616 KF739665 KJ184559 KT002617 KF030262 KC184467 KT002618 KT002620 KT002619 KJ184554 KT002612 KF112339 ITS KJ605668 KT002598 KJ419923 KT002599 KT002600 - KT002602 KT002603 KT002604 KT002605 KJ909519 KJ909517 KT002606 KC184464 KC184466 KT002607 KT002608 KJ909521 KJ605655 KT002601 KM388726 Belgium Location Zoniënwoud, Belgium Meise, Germany Connecticut, USA USA Tennessee, Eprave, Belgium Meise, Belgium Arizona, USA Germany China Yunnan, Kunming, China Yunnan, Dali, USA York, New USA York, New USA Virginia, West Germany China Yunnan, Yulong, China Yunnan, Yulong, Montenau, Belgium USA Tennessee, China Yunnan, Kunming, Collector /Identifier B. Vanholen B. Vanholen - Flavius Popa R. E. Halling - de Kerckhove O. Van Lachapelle J. R. Chapman Flavius Popa Yang Zhu L. Wu G. E.E. Both M. Binder Taylor, A. N. Siegel Flavius Popa B. Feng J. Hao Y. A. De Kesel SAT Li Y.C. Isolate/Voucher /strain Isolate/Voucher BR 50200892955-50 BR 50200893390-25 Bap1* MB 000286 00013631 NY 3959 BR 50201618465-02 BR 50201533559-51 R. Chapman 0945 MB 000287 HKAS 54099 HKAS 63593 E.E. Both 3765 MB 06-059* JLF 2566 MB 000260 HKAS 57503 HKAS 68010 BR 50201590638-05 TENN 067311 HKAS 52661 sp. Specimens used in molecular phylogenetic studies and their GenBank accession numbers. roseopurpureus. . Species Butyriboletus appendiculatus Butyriboletus appendiculatus Butyriboletus appendiculatus Butyriboletus appendiculatus Butyriboletus brunneus Butyriboletus peckii Butyriboletus pseudoregius Butyriboletus pseudoregius Butyriboletus pulchriceps Butyriboletus regius Butyriboletus roseoflavus Butyriboletus roseoflavus Butyriboletus roseopurpureus Butyriboletus roseopurpureus Butyriboletus roseopurpureus Butyriboletus subappendiculatus Butyriboletus yicibus Butyriboletus yicibus Caloboletus calopus Exsudoporus frostii Exsudoporus Accessions numbers in boldface indicate newly generated sequences. An asterisk (*) at the isolate number emphasizes that the sequences were represented by two strains for strains two by represented were sequences the that emphasizes number isolate the at (*) asterisk An sequences. generated newly indicate boldface in numbers Accessions Bu TABLE 1. TABLE

58 • Phytotaxa 234 (1) © 2015 Magnolia Press ZHAO ET AL. References

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Supporting Information

TABLE S1 Specimens used in the phylogeny of Boletaceae and their GenBank accession numbers (apart from species already included in Table 1). Isolate/Voucher GenBank Accessions Species Location /strain nrLSU tef1-α rpb1 Aureoboletus gentilis MG372a Brancciano, Lazio, Italy KF112344 KF134014 KF112557 Austroboletus fusisporus HKAS75207 China JX889720 JX889718 JX889721 Baorangia rubelloides MB 07-001 Chestnut Ridge Park, NY, USA KF030370 KF030405 KF030370 Boletellus projectellus AFTOL-713 MA, U.S.A. AY684158 AY879116 AY662660 merulioides AFTOL-ID 575 MA, U.S.A. AY684153 DQ056287 DQ435803 Be3 Bavaria, Germany KF030282 GU187682 GU187444 Boletus pallidus 179/97 Bavaria, Germany AF457409 KF030424 KF030396 Boletus sp. HKAS55440 Deqin, Yunnan, China KF112315 KF112145 KF112499 Boletus sp. HKAS59660 Yunnan, China KF112358 KF112153 KF112503 Boletus sp. HKAS76661 Nanyang, Henan, China KF112342 KF112205 - Borofutus dhakanus HKAS73789 Gazipur, Bangladesh JQ928616 JQ928576 JQ928586 castanella MB 03-053 Massachussetts DQ867117 KF030421 KF030382 Buchwaldoboletus lignicola Pul1 Maindreieck, Germany JQ326997 JQ327040 - piperatus MB04-001 Massachusetts, USA DQ534648 GU187690 GU187453 Crocinoboletus laetissimus HKAS59701 Chuxiong, Yunnan, China KF112436 - - Crocinoboletus rufo-aureus HKAS53424 Chenzhou, Hunan, China KF112435 KF112206 KF112533 Cyanoboletus pulverulentus 9606 West Newton, MA, USA KF030313 KF030418 KF030364 Fistulinella prunicolor REH9502 Fraser Island, Qld, Australia JX889648 JX889690 - Gymnogaster boletoides REH9455 SE Qld, Australia JX889673 JX889683 - lividus REG Gl1 Bavaria, Germany AF098378 GU187701 GU187461 Harrya chromapes ND4 North Carolina, USA JX889664 JX889704 - Heimioporus japonicus HKAS52237 Chuxiong, Yunnan, China KF112347 KF112228 KF112618 impolitum Bim1 Bavaria, Germany AF139715 JQ327034 - Imleria badia MB 03-098a Rutland, MA, USA KF030355 KF030423 KF030393 Lanmaoa carminipes MB 06-061 Erie Co., NY, USA JQ327001 JQ327022 KF030363 Leccinellum corsicum Buf 4507 - KF030347 KF030435 KF030389 variicolor HKAS57758 Lijiang, Yunnan, China KF112445 KF112251 KF112591 ...continued on next page

60 • Phytotaxa 234 (1) © 2015 Magnolia Press ZHAO ET AL. TABLE S1. (Continued) Isolate/Voucher GenBank Accessions Species Location /strain nrLSU tef1-α rpb1 Mucilopilus castaneiceps HKAS75045 Nujiang, Yunnan, China KF112382 KF112211 - Neoboletus luridiformis AT2001087 Berkshire, England, UK JQ326995 JQ327023 - Neoboletus magnificus HKAS54096 Kunming, Yunnan, China KF112324 KF112149 KF112495 Octaviania tasmanica OSC132097 Avre River Picnic Area, Australia JN378494 JN378435 - Paragyrodon sphaerosporus MB06-066 Iowa, USA GU187593 GU187737 - Parvixerocomus aokii HKAS59812 Wanling, Hainan, China KF112378 KF112266 KF112597 Paxillus vernalis AFTOL-ID 715 China AY645059 DQ457629 - Phlebopus portentosus Ph1 Africa AF336260 FJ536679 FJ536646 pelletieri Pp1 Bavaria, Germany AF456818 JQ327036 KF030390 Porphyrellus brunneus REH9508 Fraser Island, Qld, Australia JX889646 JX889688 - Porphyrellus holophaeus HKAS50508 Ninger, Yunnan, China KF112465 KF112244 KF112553 Porphyrellus porphyrosporus AFTOL-1779 Walhalla, Bavaria, Germany DQ534643 GU187734 GU187475 Pseudoboletus parasiticus Xpa1 Bavaria, Germany AF050646 KF030443 KF030394 Pulveroboletus aff. ravenelii HKAS53351 Sanming, Fujian, China KF112406 KF112261 KF112542 Retiboletus griseus HKAS63590 Dali, Yunnan, China KF112417 KF112178 KF112537 Royoungia boletoides REH8774 Atherton, Qld, Australia JX889660 JX889701 - Rubroboletus latisporus HKAS 80358 Chongqing, China KP055023 KP055020 KP055026 Rubroboletus sinicus HKAS 56304 Deqin, Yunnan, China KJ605673 KJ619483 KJ619482 Rugiboletus extremiorientalis HKAS63635 Chuxiong, Yunnan, China KF112403 KF112198 KF112535 Solioccasus polychromus REH9417 Australia JQ287642 JQ287644 - Spongiforma thailandica DED 7873 Thailand EU685108 KF030436 KF030387 Strobilomyces floccopus AFTOL-716 MA, USA AY684155 AY883428 - Suillellus aff. amygdalinus HKAS57262 Qamdo, Tibet, China KF112316 KF112174 KF112501 Suillellus amygdalinus 112605ba Mendocino Co., CA, USA JQ326996 JQ327024 KF030360 HKAS54926 Marburg, Germany KF112411 HQ326866 KF112575 Tylopilus sp. HKAS46334 Dêqên, Yunnan, China KF112471 KF112271 KF112581 Tylopilus sp. HKAS50229 Xishuangbanna, Yunnan, China KF112423 KF112216 KF112574 Tylopilus sp. HKAS50281 Xishuangbanna, Yunnan, China KF112451 KF112284 - Tylopilus virens HKAS76678 Liangshan Yi, Sichuan, China KF112438 KF112272 KF112582 Veloporphyrellus aff. velatus HKAS57490 Lijiang, Yunnan, China KF112380 KF112209 KF112555 Xanthoconium stramineum 3518 Lake Mize, Gainsville, FL, USA KF030353 KF030428 KF030386 Xerocomellus aff. rubellus MB 03-033 Worcester, MA, USA KF030294 KF030419 KF030371 Xerocomellus chrysenteron Xch1 Bavaria, Germany AF050647 KF030415 KF030365 Xerocomus aff. macrobii HKAS56280 Chuxiong, Yunnan, China KF112418 KF112265 KF112541 Xs1 Bavaria, Germany AF139716 JQ327035 KF030391 Zangia roseola HKAS51137 Yunnan, China HQ326949 HQ326877 -

Three new combinations of Butyriboletus Phytotaxa 234 (1) © 2015 Magnolia Press • 61 FIGURE S1 Maximum-Likelihood phylogenetic tree generated from the combined dataset (nrLSU, tef1-α and rpb1). BS support values >50% for ML are indicated along branches

62 • Phytotaxa 234 (1) © 2015 Magnolia Press ZHAO ET AL.