New Cantharellus species from South Korea

Abstract In this third contribution involving new Cantharellus species from South Korea, two new species are introduced. In addition, we document a first report of the recently described Japanese Cantharellus anzutake outside of Japan based on identical ITS sequence data. Cantharellus citrinussp. nov. is introduced as a new member of subgenus Cinnabarini, to which the closely related Korean C. albovenosus and Chinese C. phloginus also belong. Cantharellus curvatussp. nov. is introduced as a new member of subgenus Parvocantharellus, in which the Korean C. koreanus was recently placed. The respective placements of the new taxa are significantly supported by a phylogenetic analysis of sequences from the transcription elongation factor (tef-1).


Introduction
In our previous contributions reporting the biodiversity of Cantharellus Adans.:Fr. in Korea (Antonín et al. 2017;Buyck et al. 2018), we have reviewed the still limited taxonomic knowledge on Cantharellus species in Asia. During the past two years two more new chanterelles have been described from Asia: C. anzutake W. Ogawa, N. Endo, M. Fukuda and A. Yamada from Japan (Ogawa et al. 2018) and C. hainanensis N.K. Zeng, Zhi Q. Liang & S. Jiang from China ). In the present paper, we describe two more new species from South Korea supported by morphological features and in particular by sequence data obtained for the transcription elongation factor (tef-1) gene. In addition, identical ITS sequence data also document the presence, in South Korea, of the recently described C. anzutake from Japan, a species belonging to subg. Cantharellus and based on a 100 base pair deletion in the internal transcribed spacer 1 (ITS1) of the rDNA (Ogawa et al. 2018). Obtained tef-1 sequence data from all of our recent collections of chanterelles in South Korea could not confirm the presence of any of the European or North American Cantharellus previously reported from South Korea (Park and Lee 1991;Kim 2004;Kim et al. 2006;Lee 2011) nor any of the chanterelles already described from India (Das et al. 2015;Kumari et al. 2011Kumari et al. , 2013, neighbouring China (Shao et al. 2011(Shao et al. , 2014(Shao et al. , 2016aTian et al. 2012;, Japan (Suhara and Kurogi 2015;Ogawa et al. 2018) or Malaysia (Corner 1966;Eyssartier et al. 2009)

Field work
Collections for this work were made during field trips of the last author in collaboration with colleagues from the National Institute of Forest Science (former Korea Forest Research Institute) in the margin of two larger inventory projects: "Diversity and molecular taxonomy of marasmielloid and gymnopoid fungi (Basidiomycota, Omphalotaceae) in South Korea" and "Phylogeny of litter decomposing fungi in South Korea". The various localities in which Cantharellus specimens were collected are shown below (Fig. 1).

Morphology
Macroscopic descriptions of collected specimens were based on fresh basidiomata. Colour abbreviations follow Kornerup and Wanscher (1983). Microscopic features were studied using dried material mounted in H 2 O, approximately 5% KOH, Melzer's reagent and Congo Red, using an Olympus BX-50 light microscope (Tokyo, Japan) at 1000× magnification. For the hymenophore, "L" refers to the number of whole gill folds, while "l" refers to the number of shorter gill folds between each pair of entire gill folds. For basidiospores, the factor E indicates the quotient of the length and width in any one basidiospore and Q is the mean of the E-values; the basidiospore values are based on 20 measurements in each collection. Specimens are preserved in the herbarium of the Moravian Museum, Brno, Czech Republic (BRNM) and duplicates deposited in the fungarium of the Natural History Museum in Paris, France (PC).

Taxon sampling, sequence data and phylogenetic analyses
Translation elongation factor 1-alpha (tef-1) sequence data were produced following Buyck et al. (2014) for the newly described species from dried materials: four sequences for four collections of Cantharellus citrinus sp. nov and one sequence from a collection of C. curvatus sp. nov. Additional tef-1 sequences were obtained for two previously described species: for two collections of C. koreanus Buyck, Antonín & V. Hofst. and for one collection of C. albovenosus Buyck, Antonín & V. Hofst. We introduced these newly produced tef-1 sequences in the alignment obtained by Antonín et al. (2017). Species of subg. Pseudocantharellus Eyssart. & Buyck were used as outgroup sequences. GenBank submissions numbers are given in Fig. 2. Alignment of sequence data was performed manually in MacClade (Maddison and Maddison 2003). Three independent searches for the most likely tree were conducted in PhyML v. 3.0 (Guindon and Gascuel 2003) to check for convergence toward the same likelihood value. These searches used the GTR evolutionary model (Abadi et al. 2019) with the proportion of invariable sites, the gamma shape parameter and the number of substitution categories estimated during the search. Branches were considered as significantly supported when maximum likelihood bootstrap support (ML-bs) was ≥ 70%.

Phylogeny
The final alignment of tef-1sequences included 837 characters. After the removal of three spliceosomal introns, the alignment used for phylogenetic analyses included 629 characters. The most likely tree (Fig. 2)  Diagnosis. Differs from its closest Asian and North American relatives in the variously colo ured but often bright lemon yellow pileus, similarly tinted stipe and smaller size, as well as in differences in sequence data produced for the transcription elongation factor (tef-1).
Holotype. South Korea. Description. Basidiomata dispersed in small groups or fascicles. Pileus 4-20 mm broad, convex, with involute margin when young, then plane or infundibuliform with depressed centre and inflexed to straight, smooth margin, irregularly undulate when old, hygrophanous, finely tomentose when very young, soon glabrescent and smooth or slightly rugulose, uniformly coloured, light yellow, orange yellow to light orange (3-4A6, 4-5A5-7), sometimes with greyish yellow tinge when old. Hymenophore composed of thick vein-like folds, sometimes strongly decurrent in a reticulate pattern on upper stipe, often not reaching the pileus margin, forking or with rare lamellulae, transversely anastomosed in between, white to whitish (3A2); edges concolorous. Stipe 4-22 × 1-3(-4) mm, slightly tapering towards base when young, then cylindrical, sometimes curved, finely pubescent when young, later glabrescent, smooth, concolorous with pileus or slightly paler. Context thin, yellowish, fibrillose-hollow and yellow- ish whitish in stipe when old, with a spicy apricot smell and mild taste. Spore print not obtained.
Habitat Remarks. The description is based on the type specimen, but examination of the other specimens shows that variation of morphological features includes a rather wide amplitude of the overall colour, which seems -based on identical tef1 sequences -to extend from entirely and predominantly pale lemon yellow to an overall deep orange. Collection from Jinan (VA 16.169, BRNM 825753, PC 0142467) differs from other collections of this species by an orange (5-6A7) pileus, light yellow to light orange (4-5A5) lamellae and a stipe more or less concolorous with the pileus.
This new species is here placed in Cantharellus subg. Cinnabarini (Fig. 2), a subgenus that comprises several species exhibiting a similarly wide colour range, e.g. the Malagasy C. variabilicolor Buyck & V. Hofst. (in Ariyawansa et al. 2015) or the North American C. cinnabarinus (Schwein.) Schwein. Cantharellus citrinus is here shown to be part of a well-supported clade composed of two other Asian species, the Chinese C. phloginus and Korean C. albovenosus. The latter two species are very different in general aspect, but, except for a single mutation in the coding part, the tef-1 sequences of both species are identical, even including the introns. Yet, their very different general habitus justifies us in our view that we should accept them as a separate species. The clade comprising these Asian species is sister to a clade composed of North American species.
Because of its very small overall size and comparable overall colour, C. citrinus could also easily be mistaken for some species in Cantharellus subg. Parvocantharellus Eyssart. & Buyck, in particular the European C. romagnesianus (= C. pseudominimus Eyssart. & Buyck, see Olariaga et al. 2015). Under the microscope, C. citrinus differs hardly from its Asian relatives and identification relies principally on field characters or sequence data. Diagnosis. Differs from the European C. romagnesianus in the distinctly smaller spores and shorter basidia (see Olariaga et al. 2016), the strongly veined hymenophore and sequence data obtained from the transcription elongation factor one alpha (tef-1).

Cantharellus anzutake
Habitat Remarks. This species is a typical member of Cantharellus subg. Cantharellus, and belongs to a group that is often referred to as the 'golden chanterelles' or the C. cibarius Fr. complex, representing the commercially most important chanterelles on the international market. This species complex is reputedly very difficult to identify, in particular because of the very variable field aspect of the various species involved (Olariaga et al. 2015(Olariaga et al. , 2016. Hence, positive identification frequently requires molecular sequence data. Our identification is here based on the high quality ITS sequence we obtained for VA 16.142 and which is identical to the one deposited for the C. anzutake holotype (GenBank LC085359, similarity 100% for 100% coverage); both these ITS differ from other yellow species of chanterelles described from Asia by a ca 100 bp deletion in the ITS1 region (Ogawa et al. 2018).
Because of the whitish hymenophore when young and the sometimes deep orange-yellow to cinnamon buff pileus surface, this species may be somewhat reminiscent of C. albovenosus. The latter species, however, has always a much brighter orange pileus and a more veined hymenophore that remains white, even with age, and it belongs in subgenus Cinnabarini (see Antonín et al. 2017). It is interesting to note that both Japanese and Korean specimens were collected near Pinus densiflora among possible host trees.