Abstract
Hydnum repando-orientale is an East Asian species closely related to H. boreorepandum and H. repandum; all three species produce edible mushrooms. We identified two ecological groups of H. repando-orientale in Japan: a temperate group occurring in Fagaceae-dominated forest at < 1200 m a.s.l. (ROF) and a subalpine group occurring in coniferous forest in highland at > 1900 m a.s.l. (ROC). We re-examined the taxonomy of the two ecological groups of H. repando-orientale using integrative approaches. Phylogenies of the two ecological groups and other related species were inferred from the internal transcribed spacer (ITS) and gene portions encoding the large subunit of nc rRNA (LSU), translation elongation factor-1 alpha (TEF1), RNA polymerase II largest subunit (RPB1), and RNA polymerase II second-largest subunit (RPB2). The concatenated phylogenetic tree separated the two ecological groups into well-supported sister clades. Also, species delimitations based on the topological congruence (GCPSR) and coalescent models (GMYC and BP&P) supported to separate the two ecological groups. Morphological analysis showed that ROC specimens had significantly larger basidiospores, compared with ROF specimens. Mon-mon mating tests using six ROF, three ROC, and three H. boreorepandum strains each showed independent incompatible groups, whereas one ROC strain showed compatibility with both ROC and ROF populations. Based on these results, we defined the ROC group as a new species, H. subalpinum. Because H. repando-orientale and H. subalpinum have smaller genetic divergence in nc rDNA and maintain slight sexual compatibility, they may have recently speciated in East Asia.
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Data availability
Voucher specimens and culture collections have been deposited in the Tottori University Mycological Herbarium (TUMH), Fungus/Mushroom Resource and Research Center, Faculty of Agriculture, Tottori University. The newly generated sequences have been submitted in INSD with the accession numbers listed in Tables 2 and S2. The alignments for phylogenetic analyses are provided in Supplementary Data. All other data generated or analyzed in this research available from the corresponding author on requests.
References
Abarenkov K, Tedersoo L, Nilsson RH et al (2010) PlutoF—a web based workbench for ecological and taxonomic research, with an online implementation for fungal ITS sequences. Evol Bioinforma 6:EBO.S6271. https://doi.org/10.4137/EBO.S6271
Agerer R, Kraigher H, Javornik B (1996) Identification of ectomycorrhizae of Hydnum rufescens on Norway spruce and the variability of the ITS region of H. rufescens and H. repandum (Basidiomycetes). Nova Hedwigia 63:183–194
Aime MC, Miller AN, Aoki T et al (2021) How to publish a new fungal species, or name, version 3.0. IMA Fungus 12:11. https://doi.org/10.1186/s43008-021-00063-1
Asahina Y (1939) Illustrations of Japanese cryptogams (in Japanese). Sanseido, Tokyo
Bengtsson-Palme J, Ryberg M, Hartmann M et al (2013) Improved software detection and extraction of ITS1 and ITS2 from ribosomal ITS sequences of fungi and other eukaryotes for analysis of environmental sequencing data. Methods Ecol Evol 4:914–919. https://doi.org/10.1111/2041-210X.12073
Bouckaert R, Vaughan TG, Barido-Sottani J et al (2019) BEAST 2.5: an advanced software platform for Bayesian evolutionary analysis. PLoS Comput Biol 15:e1006650
Cao B, Haelewaters D, Schoutteten N et al (2021a) Delimiting species in Basidiomycota: a review. Fungal Divers 109:181–237. https://doi.org/10.1007/s13225-021-00479-5
Cao T, Hu Y-P, Yu J-R et al (2021b) A phylogenetic overview of the Hydnaceae (Cantharellales, Basidiomycota) with new taxa from China. Stud Mycol 99:100121. https://doi.org/10.1016/j.simyco.2021.100121
Coker WC, Beers AH (1951) The stipitate Hydnums of the Eastern United States. Oxford University Press, London
Darriba D, Posada D, Kozlov AM et al (2020) ModelTest-NG: a new and scalable tool for the selection of DNA and protein evolutionary models. Mol Biol Evol 37:291–294. https://doi.org/10.1093/molbev/msz189
Dettman JR, Jacobson DJ, Taylor JW (2003) A multilocus genealogical approach to phylogenetic species recognition in the model Eukaryote neurospora. Evolution 57:2703–2720. https://doi.org/10.1111/j.0014-3820.2003.tb01514.x
Dettman JR, Jacobson DJ, Taylor JW (2006) Multilocus sequence data reveal extensive phylogenetic species diversity within the Neurospora discreta complex. Mycologia 98:436–446. https://doi.org/10.1080/15572536.2006.11832678
Dobzhansky T (1940) Speciation as a stage in evolutionary divergence. Am Nat 74:312–321. https://doi.org/10.1086/280899
Donk MA (1956) The generic names proposed for Hymenomycetes V “Hydnaceae” (continuation). Taxon 5:69–80. https://doi.org/10.2307/1217317
Edler D, Klein J, Antonelli A, Silvestro D (2021) raxmlGUI 2.0: a graphical interface and toolkit for phylogenetic analyses using RAxML. Methods Ecol Evol 12:373–377. https://doi.org/10.1111/2041-210X.13512
Feng B, Wang XH, Ratkowsky D et al (2016) Multilocus phylogenetic analyses reveal unexpected abundant diversity and significant disjunct distribution pattern of the Hedgehog Mushrooms (Hydnum L.). Sci Rep 6:1–11. https://doi.org/10.1038/srep25586
Flouri T, Izquierdo-Carrasco F, Darriba D et al (2015) The phylogenetic likelihood library. Syst Biol 64:356–362. https://doi.org/10.1093/sysbio/syu084
Fujisawa T, Barraclough TG (2013) Delimiting species using single-locus data and the generalized mixed Yule coalescent approach: a revised method and evaluation on simulated data sets. Syst Biol 62:707–724. https://doi.org/10.1093/sysbio/syt033
Fujita MK, Leaché AD, Burbrink FT et al (2012) Coalescent-based species delimitation in an integrative taxonomy. Trends Ecol Evol 27:480–488. https://doi.org/10.1016/j.tree.2012.04.012
Garbelotto M, Gonthier P (2013) Biology, epidemiology, and control of Heterobasidion species worldwide. Annu Rev Phytopathol 51:39–59. https://doi.org/10.1146/annurev-phyto-082712-102225
Garbelotto M, Ratcliff A, Bruns TD et al (1996) Use of taxon-specific competitive-priming PCR to study host specificity, hybridization, and intergroup gene flow in intersterility groups of Heterobasidion annosum. Phytopathology 86:543–551. https://doi.org/10.1094/Phyto-86-543
Garbelotto M, Slaughter G, Popenuck T et al (1997) Secondary spread of Heterobasidion annosum in white fir root-disease centers. Can J For Res 27:766–773. https://doi.org/10.1139/x96-193
Gardes M, Bruns TD (1993) ITS primers with enhanced specificity for basidiomycetes – application to the identification of mycorrhizae and rusts. Mol Ecol 2:113–118. https://doi.org/10.1111/j.1365-294X.1993.tb00005.x
Grebenc T, Martín MP, Kraigher H (2009) Ribosomal ITS diversity among the European species of the genus Hydnum (Hydnaceae). An del Jardín Botánico Madrid 66:121–132. https://doi.org/10.3989/ajbm.2221
Hall D, Stuntz DE (1971) Pileate Hydnaceae of the Puget Sound Area. I. White-spored genera: Auriscalpium, Hericium, Dentinum and Phellodon. Mycologia 63:1099–1128. https://doi.org/10.1080/00275514.1971.12019214
Harder CB, LæssØe T, FrØslev TG et al (2013) A three-gene phylogeny of the Mycena pura complex reveals 11 phylogenetic species and shows ITS to be unreliable for species identification. Fungal Biol 117:764–775. https://doi.org/10.1016/j.funbio.2013.09.004
Harrington TJ, Mitchell DT (2002) Characterization of Dryas octopetala ectomycorrhizas from limestone karst vegetation, western Ireland. Can J Bot 80:970–982. https://doi.org/10.1139/b02-082
Harrison KA, Grund DW (1987) Preliminary keys to the terrestrial stipitate Hydnums of North America. Mycotaxon 28:419–426
Hughes KW, Petersen RH (2001) Apparent recombination or gene conversion in the ribosomal ITS region of a Flammulina (Fungi, Agaricales) hybrid. Mol Biol Evol 18:94–96. https://doi.org/10.1093/oxfordjournals.molbev.a003724
Hughes KW, Petersen RH, Lodge DJ et al (2013) Evolutionary consequences of putative intra-and interspecific hybridization in agaric fungi. Mycologia 105:1577–1594. https://doi.org/10.3852/13-041
Imazeki R, Hongo T (1957) Coloured illustrations of fungi of Japan (in Japanese). Hoikusha, Osaka
Ito S (1955) Mycological flora of Japan, vol 2, no 4 (in Japanese). Yokendo, Tokyo
Katoh K, Rozewicki J, Yamada KD (2019) MAFFT online service: multiple sequence alignment, interactive sequence choice and visualization. Brief Bioinform 20:1160–1166. https://doi.org/10.1093/bib/bbx108
Kauserud H, Hofton TH, Sætre G-P (2007) Pronounced ecological separation between two closely related lineages of the polyporous fungus Gloeoporus taxicola. Mycol Res 111:778–786. https://doi.org/10.1016/j.mycres.2007.03.005
Kawamura S (1913) Illustrations of Japanese fungi, 2nd edn. The Bureau of Forestry, Ministry of Agriculture and Commerce, Tokyo
Kawamura S (1929) The Japanese fungi (in Japanese). Daichi-shoin, Tokyo
Kawamura S (1954) Icons of Japanese fungi (in Japanese), vol 4. Kazamashobo, Tokyo
Kikuhara N (1987) The Corticiales and Clavariales of Japan (in Japanese). Seichiken, Tokyo
Kornerup A, Wanscher JH (1978) Methuen handbook of colour, 3rd edn. Eyre Methuen, London
Košuthová A, Bergsten J, Westberg M, Wedin M (2020) Species delimitation in the cyanolichen genus Rostania. BMC Evol Biol 20:115. https://doi.org/10.1186/s12862-020-01681-w
Kretzer AM, Bruns TD (1999) Use of atp6 in fungal phylogenetics: an example from the Boletales. Mol Phylogenet Evol 13:483–492. https://doi.org/10.1006/mpev.1999.0680
Kumar S, Stecher G, Tamura K (2016) MEGA7: molecular evolutionary genetics analysis version 7.0 for bigger datasets. Mol Biol Evol 33:1870–1874. https://doi.org/10.1093/molbev/msw054
Lanfear R, Calcott B, Ho SYW, Guindon S (2012) PartitionFinder: combined selection of partitioning schemes and substitution models for phylogenetic analyses. Mol Biol Evol 29:1695–1701. https://doi.org/10.1093/molbev/mss020
Lanfear R, Frandsen PB, Wright AM et al (2017) PartitionFinder 2: new methods for selecting partitioned models of evolution for molecular and morphological phylogenetic analyses. Mol Biol Evol 34:772–773. https://doi.org/10.1093/molbev/msw260
Le Gac M, Giraud T (2008) Existence of a pattern of reproductive character displacement in Homobasidiomycota but not in Ascomycota. J Evol Biol 21:761–772. https://doi.org/10.1111/j.1420-9101.2008.01511.x
Li J, He X, Liu X-B et al (2017) Species clarification of oyster mushrooms in China and their DNA barcoding. Mycol Prog 16:191–203. https://doi.org/10.1007/s11557-016-1266-9
Linnaeus C (1753) Species plantarum: exhibentes plantas rite cognitas ad genera relatas, cum diferentiis specificis, nominibus trivialibus, synonymis selectis, locis natalibus, secundum systema sexuale digestas. Impensis Laurentii Salvii, Stockholm, Sweden. p 1178
Liu YJ, Whelen S, Hall BD (1999) Phylogenetic relationships among ascomycetes: evidence from an RNA polymerase II subunit. Mol Biol Evol 16:1799–1808. https://doi.org/10.1093/oxfordjournals.molbev.a026092
Liu YJ, Hodson MC, Hall BD (2006) Loss of the flagellum happened only once in the fungal lineage: phylogenetic structure of Kingdom Fungi inferred from RNA polymerase II subunit genes. BMC Evol Biol 6:74. https://doi.org/10.1186/1471-2148-6-74
Looney BP, Adamčík S, Matheny PB (2020) Coalescent-based delimitation and species-tree estimations reveal Appalachian origin and Neogene diversification in Russula subsection Roseinae. Mol Phylogenet Evol 147:106787. https://doi.org/10.1016/j.ympev.2020.106787
Lücking R, Aime MC, Robbertse B et al (2020) Unambiguous identification of fungi: where do we stand and how accurate and precise is fungal DNA barcoding? IMA Fungus 11:14. https://doi.org/10.1186/s43008-020-00033-z
Maas Geesteranus RA (1971) Hydnaceous fungi of the Eastern Old World. North-Holland Publishing Company, Amsterdam
Matheny PB, Wang Z, Binder M et al (2007) Contributions of rpb2 and tef1 to the phylogeny of mushrooms and allies (Basidiomycota, Fungi). Mol Phylogenet Evol 43:430–451. https://doi.org/10.1016/j.ympev.2006.08.024
Niskanen T, Liimatainen K, Nuytinck J et al (2018) Identifying and naming the currently known diversity of the genus Hydnum, with an emphasis on European and north American taxa. Mycologia 110:890–918. https://doi.org/10.1080/00275514.2018.1477004
Noor MAF (1999) Reinforcement and other consequences of sympatry. Heredity 83:503–508. https://doi.org/10.1038/sj.hdy.6886320
Nosil P, Vines TH, Funk DJ (2005) Reproductive isolation caused by natural selection against immigrants from divergent habitats. Evolution 59:705–719. https://doi.org/10.1111/j.0014-3820.2005.tb01747.x
Olariaga I, Grebenc T, Salcedo I, Martín MP (2012) Two new species of Hydnum with ovoid basidiospores: H. ovoideisporum and H. vesterholtii. Mycologia 104:1443–1455. https://doi.org/10.3852/11-378
Otrosina WJ, Garbelotto M (2010) Heterobasidion occidentale sp. nov. and Heterobasidion irregulare nom. nov.: a disposition of North American Heterobasidion biological species. Fungal Biol 114:16–25. https://doi.org/10.1016/j.mycres.2009.09.001
Paradis E, Schliep K (2019) ape 5.0: an environment for modern phylogenetics and evolutionary analyses in R. Bioinformatics 35:526–528. https://doi.org/10.1093/bioinformatics/bty633
Petersen RH, Hughes KW, Redhead SA et al (1999) Mating systems in the Xerulaceae (Agaricales, Basidiomycotina): Flammulina. Mycoscience 40:411–426. https://doi.org/10.1007/BF02464396
Phillips R (2005) Mushrooms and other fungi of North America. Firefly Books, New York
Pons J, Barraclough TG, Gomez-Zurita J et al (2006) Sequence-based species delimitation for the DNA taxonomy of undescribed insects. Syst Biol 55:595–609. https://doi.org/10.1080/10635150600852011
R Core Team (2021) R: A language and environment for statistical computing. https://www.r-project.org/. Accessed 1 Aug 2021
Rambaut A, Drummond AJ, Xie D et al (2018) Posterior summarization in Bayesian phylogenetics using Tracer 1.7. Syst Biol 67:901–904. https://doi.org/10.1093/sysbio/syy032
Rannala B, Yang Z (2003) Bayes estimation of species divergence times and ancestral population sizes using DNA sequences from multiple loci. Genetics 164:1645–1656. https://doi.org/10.1093/genetics/164.4.1645
Rea C (1922) British Basidiomycetae: a handbook to the larger British Fungi. Cambridge University Press, France
Ripková S, Hughes K, Adamčík S et al (2010) The delimitation of Flammulina fennae. Mycol Prog 9:469–484. https://doi.org/10.1007/s11557-009-0654-9
Roberts P, Evans S (2011) The book of fungi. University of Chicago Press, Illinois
Ronquist F, Teslenko M, van der Mark P et al (2012) MrBayes 3.2: efficient Bayesian phylogenetic inference and model choice across a large model space. Syst Biol 61:539–542. https://doi.org/10.1093/sysbio/sys029
Sillo F, Gonthier P, Lockman B et al (2019) Molecular analyses identify hybridization-mediated nuclear evolution in newly discovered fungal hybrids. Ecol Evol 9:6588–6605. https://doi.org/10.1002/ece3.5238
Stamatakis A (2014) RAxML version 8: a tool for phylogenetic analysis and post-analysis of large phylogenies. Bioinformatics 30:1312–1313. https://doi.org/10.1093/bioinformatics/btu033
Sugawara R, Yamada A, Kawai M et al (2019) Establishment of monokaryotic and dikaryotic isolates of hedgehog mushrooms (Hydnum repandum and related species) from basidiospores. Mycoscience 60:201–209. https://doi.org/10.1016/j.myc.2019.02.007
Sugawara R, Maekawa N, Sotome K et al (2022a) Systematic revision of Hydnum species in Japan. Mycologia 114:413–452. https://doi.org/10.1080/00275514.2021.2024407
Sugawara R, Shirasuka N, Yamamoto T et al (2022b) Two new species of Sistotrema s.l. (Cantharellales) from Japan with descriptions of their ectomycorrhizae. Mycoscience 63:102–117. https://doi.org/10.47371/mycosci.2022.02.003
Swenie RA, Baroni TJ, Matheny PB (2018) Six new species and reports of Hydnum (Cantharellales) from eastern North America. MycoKeys 72:35–72. https://doi.org/10.3897/mycokeys.42.27369
Taylor JW, Jacobson DJ, Kroken S et al (2000) Phylogenetic species recognition and species concepts in fungi. Fungal Genet Biol. https://doi.org/10.1006/fgbi.2000.1228
Tedersoo L, Jairus T, Horton BM et al (2008) Strong host preference of ectomycorrhizal fungi in a Tasmanian wet sclerophyll forest as revealed by DNA barcoding and taxon-specific primers. New Phytol 180:479–490. https://doi.org/10.1111/j.1469-8137.2008.02561.x
Thomas E, Fujisawa T, Barraclough T (2021) splits: SPecies’ LImits by threshold statistics. R package version 1.0-20/r56. https://r-forge.r-project.org/projects/splits/%0A. Accessed 1 Jun 2022
Thompson JD, Higgins DG, Gibson TJ (1994) CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22:4673–4680. https://doi.org/10.1093/nar/22.22.4673
Wang PM, Liu XB, Dai YC et al (2018) Phylogeny and species delimitation of Flammulina: taxonomic status of winter mushroom in East Asia and a new European species identified using an integrated approach. Mycol Prog 17:1013–1030. https://doi.org/10.1007/s11557-018-1409-2
Yamada A, Katsuya K (1995) Mycorrhizal association of isolates from sporocarps and ectomycorrhizas with Pinus densiflora seedlings. Mycoscience 36:315–323. https://doi.org/10.1007/BF02268607
Yanaga K (2015) Taxonomic study of the order Cantharellales (Basidiomycota) in Japan (in Japanese). Dissertation, United Graduate School of Agricultural Sciences. https://repository.lib.tottori-u.ac.jp/5046. Accessed 1 Aug 2021
Yanaga K, Sotome K, Ushijima S, Maekawa N (2015) Hydnum species producing whitish basidiomata in Japan. Mycoscience 56:434–442. https://doi.org/10.1016/j.myc.2015.01.001
Yang Z (2002) Likelihood and Bayes estimation of ancestral population sizes in Hominoids using data from multiple loci. Genetics 162:1811–1823. https://doi.org/10.1093/genetics/162.4.1811
Yang Z (2015) The BPP program for species tree estimation and species delimitation. Curr Zool 61:854–865. https://doi.org/10.1093/czoolo/61.5.854
Yang Z, Rannala B (2010) Bayesian species delimitation using multilocus sequence data. Proc Natl Acad Sci 107:9264–9269. https://doi.org/10.1073/pnas.0913022107
Yang Z, Rannala B (2014) Unguided species delimitation using DNA sequence data from multiple loci. Mol Biol Evol 31:3125–3135. https://doi.org/10.1093/molbev/msu279
Yasuda A (1913) Kinrui-zakki 19 (in Japanese). Bot Mag Tokyo 27:339–340
Acknowledgements
We thank Dr. K. Hosaka (Department of Botany, Division of Fungi and Algae, National Museum of Nature and Science, Tokyo) for the loan of specimens from the National Museum of Nature and Science (TNS), the Nagano Prefectural Government, the ministry of the Environment, and the Forestry Agency of Japan for the permissions of field research in special protection zones. We also thank A. Koyama and M. Shishikura for donation of the basidiomata collections and digital photographs. We thank the DNA Data Bank of Japan for nucleotide sequence submission and Fasmac Co., Ltd for technical support regarding DNA sequencing. This research was financially supported by JSPS KAKENHI Grant Number JP20J20884 (Ryo Sugawara) from Japan Society for the Promotion of Science.
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This research was supported by JSPS KAKENHI Grant Number JP20J20884 (Ryo Sugawara) from Japan Society for the Promotion of Science.
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Ryo Sugawara, Naoki Endo, and Akira Nakagiri contributed to the study conception and design. Materials’ preparation was performed by Ryo Sugawara, Naoki Endo, Wataru Aoki, and Akiyoshi Yamada. Data collection and analyses were performed by Ryo Sugawara. The original draft of the manuscript was written by Ryo Sugawara and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.
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Sugawara, R., Aoki, W., Yamada, A. et al. Ecological speciation of Japanese hedgehog mushroom: Hydnum subalpinum sp. nov. is distinguished from its sister species H. repando-orientale by means of integrative taxonomy. Mycol Progress 21, 94 (2022). https://doi.org/10.1007/s11557-022-01844-7
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DOI: https://doi.org/10.1007/s11557-022-01844-7