The Ganoderma weberianum-resinaceum lineage: multilocus phylogenetic analysis and morphology confirm G. mexicanum and G. parvulum in the Neotropics

Abstract Many species of Ganoderma exhibit a high phenotypic plasticity. Hence, particularly among them, the morphological species concept remains difficult to apply, resulting in a currently confused taxonomy; as a consequence, the geographical distribution range of many species also remains very uncertain. One of the areas with a strong uncertainty, as far as morphological species concept is concerned, is the Neotropics. It is common that names of species described from other regions, mainly from northern temperate areas, have been applied to Neotropical species. The aim of the present study was to determine which species might lay behind the G. weberianum complex in the Neotropics, using morphological studies and phylogenetic inferences based on both single (ITS) and multilocus (ITS, rpb2, and tef1-α) sequences. The results indicated that G. weberianumsensu Steyaert, which is the usually accepted concept for this taxon, was absent from the Neotropics. In this area, G. weberianumsensu Steyaert encompassed at least two phylogenetic species, which are tentatively, for the time being, identified as belonging to G. mexicanum and G. parvulum. These two species could be distinguished morphologically, notably by the ornamentation or its absence on their chlamydospores. The results also showed that additional species from the Neotropics might still exist, including, e.g., G. perzonatum, but their circumscription remains uncertain until now because of the paucity of material available. Furthermore, it was found that the current concept of G. resinaceum embraced a complex of species.


Introduction
Ganoderma P. Karst. has always been considered as an extremely difficult group with many poorly circumscribed species, forming species complexes (Moncalvo and Ryvarden 1997). Early in the 20 th century, Lloyd (1905) already emphasized the excessively confused taxonomy of Ganoderma stating "these fungi have been described and named over and over again, until the literature has become an almost unfathomable maze of meaningless and conflicting names".
A century later, one can deduce that the situation has improved very little, if at all. Ryvarden (1991), for instance, still concluded that the taxonomic issue of the genus worldwide was very "chaotic". Hitherto, there is no comprehensive Ganoderma study and the absence of a world monograph contributed to "problems with species circumscriptions and identification", fide Moncalvo (2000).
Nowadays, about 220 species have been described in Ganoderma, over 400 taxa if one includes varieties, of which 167 apply to the so-called laccate species (Ryvarden 1991, Moncalvo andRyvarden 1997, Index Fungorum http://www.indexfungorum. org/names/names.asp). Nonetheless, estimations based on the identification of terminal clades shown by phylogenetic analysis of a large ITS sequence data set gave a range of 60-80 terminal clades or phylogenetic species within the "laccate" Ganoderma spp. and 10-30 within the "non-laccate" Ganoderma spp. (Moncalvo 2000). Over the past two decades, phylogenetic studies have tried to elucidate the status of certain species and to better circumscribe their geographic distribution (e.g., Moncalvo 2000, Wang et al. 2009, Yao et al. 2013, de Lima-Junior et al. 2014, Zhou et al. 2014, Hapuarachchi et al. 2015, Loyd et al. 2018. However, the real species number and their distribution range remain largely unknown (Moncalvo andRyvarden 1997, Moncalvo 2000).
Alliances of taxa, taxonomically informal but morphologically homogeneous and phylogenetically (variably) supported, also have been evidenced within Ganoderma (e.g., Moncalvo 2000, Hong andJung 2004). Moncalvo (2000), for instance, as a result of phylogenetic analyses based on, so far, the most comprehensive ITS DNA sequences data set, and morphological characters, identified three core groups (1-3) and a bunch of residual species of uncertain affinities. The three core groups were furthermore divided into several subgroups. The core group 1 included most of the laccate species, and was divided into G. curtisii, G. lucidum, G. resinaceum, and G. tropicum lineages (Moncalvo 2000).
The G. resinaceum lineage (subgroup 1.2, Moncalvo 2000) comprised species having laccate pileus, basidiospores with "extremely fine ornament" (Pegler and Young 1973), and chlamydospores formed in their basidiomes and in pure cultures on artificial media. In this lineage, Moncalvo (2000) mentioned "genetically isolated populations", from North and South America and the Old-World that could be equated to as many species or species complexes. Moncalvo (2000) also suggested that the G. weberianum complex would represent the tropical Asian "counterpart" of the northern temperate G. resinaceum complex.
G. stipitatum (Murrill) Murrill, G. subamboinense var. subamboinense, G. subamboinense var. laevisporum, G. subincrustatum Murrill, and G. vivianimercedianum M. Torres were re-examined. Strains examined during this study were deposited at CBS, CIRM-CF, and BCCM/MUCL. The formation of chlamydospores was examined after growing the strains on malt extract agar medium at 25 °C over four weeks according to previous results of Bazzalo and Wright (1982).
The microscopic observations procedure followed Decock et al. (2007). Specimen sections were mounted in 5% KOH solution. Melzer's reagent and cotton blue were used to test the amyloidity or dextrinoidity and cyanophyly of the microscopic structures, respectively. Microscopic characters were observed under a light microscope Axioscope 40 Carl Zeiss. Images were captured using Axio Vision 4 software on the same microscope. At least 30 structures of each mature specimen were measured. Basidiospores were measured without taking in account the apical umbo when not shrunk. Cuticular cells were measured from the middle part of the basidiome except in the case of some type materials, where only a fragment was received as loan. The 5% extremes of all microscopic measurements from each size range were given in parentheses and the arithmetic mean was provided in brackets. Color terms follow Kornerup and Wanscher (1963), and terms in descriptions are defined in Torres-Torres and .
The ITS data set was composed by 30 specimens/cultures, of which 29 originated from the Neotropics (Table 1). It was subdivided into three partitions: ITS1, 5.8S, ITS2. In this case, G. austroafricanum M.P.A. Coetzee, M.J. Wingf., Marinc. & Blanchette was selected as outgroup according to the results obtained by Coetzee et al. (2015).
All sequences were automatically aligned with MUSCLE (Robert 2004) and manually adjusted using PhyDe (Müller et al. 2010). PartitionFinder (Lanfear et al. 2012) was used to determine the best evolutionary model for each gene using the corrected Akaike information criterion (AICc). Maximum Likelihood (ML) analyses were conducted using RAxML 7.0.4 (Stamatakis 2006) and Bayesian Inference (BI) analyses with MrBayes v.3.2.2 (Ronquist and Huelsenbeck 2003). In the ML analysis, the default priors were used, including individual parameters for each partition, performing 1000 replicates under the GTRGAMMA model. BI analyses were run on CIPRES Science Gateway (Miller et al. 2010). Two independent runs, with 4,000,000 generations each, were carried out with a sampling frequency every 1000 generations and a burn-in of 25%. A 50% majority rule consensus tree with posterior probabilities (PP) was obtained. Convergence of the Markov chains to a stationary distribution was assessed by visual examination of the log likelihood values in the program Tracer v1.7.1 (Rambaut et al. 2018). Nodes were considered supported when bootstrap values (BS) were ≥ 75% and the PP was ≥ 0.85. The final alignments were deposited in TreeBASE (www.treebase.org), under accession ID: 24140 (http://purl.org/phylo/treebase/phylows/study/TB2:S24140). Bold names= newly generated sequences for this study.

Molecular phylogeny
The combined dataset contained 172 DNA sequences: 71 ITS, 50 rpb2, and 51 tef1-α. The final alignment comprised 526 bp in the ITS, 776 in the rpb2, and 1123 in the tef1-α. The concatenated data set (ITS + rpb2 + tef1-α) was 2425 bp long. From it, 23 ambiguous sites (12 from ITS1 and ITS2, 11 from tef1-α introns) were removed. The evolutionary models that best fit the individual dataset according to the AICc criterion were ITS1 = GTR+I+G, 5.8S = K80, ITS2 = GTR+I+G, rpb2 1 st = GTR+I, 2 nd = HKY+G, 3 rd codon positions = HKY+G, rpb2 intron= K80, tef1-α 1 st = GTR+I, 2 nd = HKY+G, 3 rd codon positions = GTR+G, and tef1-α intron = GTR+I. In BI analyses, the average standard deviation of split frequencies was 0.008100 in the concatenated data set and 0.008875 in the ITS data set. As far as our specimens from the Neotropics are concerned, the phylogenetic trees obtained from Bayesian (not shown) and Maximum likelihood inferences using the concatenated ( Fig. 1) and the ITS ( Fig.  2) data sets showed overall the same two clades, except for the unsupported branch of the specimen MUCL 43522 present in the concatenated ML and BI analyses, which collapsed in the ITS tree, and the placement of the specimen UMNFL100, G. subamboinense var. laevisporum, from Florida (Figs 1-2). The Ganoderma weberianum-resinaceum lineage was resolved with strong support (PP 1, BS 100%). It was divided into two major clades, I and II (Fig. 1). Clade I (PP 1, BS 98%) corresponded to the G. resinaceum clade as defined by Moncalvo (2000), with G. austroafricanum, from South Africa, located in a basal position. This was further subdivided in an unsupported clade A (PP 0.66, BS 58) and moderately supported clade B (PP 0.89, BS 82). Clade A included specimens all originated from the temperate area of the Northern Hemisphere and a specimen from the highlands of central Kenya (MUCL 57035). Clade A was structured into two subclades, A1 (PP 1, BS 100), with the Kenyan specimen (MUCL 57035) in basal position, and A2 (PP 1, BS 89), with a specimen from China (MUCL 46912) in basal position. Clade B brought together several specimens originated from both North and South America, distributed into three well-supported subclades that corresponded to G. sessile Murrill and G. polychromum (Copel.) Murrill (PP 1, BS 89; PP 0.99, BS 100), as defined by Loyd et al. (2018). Two specimens from Argentina, tentatively identified as G. sessile and G. platense Speg., formed together a third distant well-supported subclade (PP 1, BS 99).
Taking in account both single and multilocus phylogenetic analyses, we considered that our Neotropical specimens formed two related but distinct, well-supported terminal clades, C1.1 and C1.2 (Figs 1-2) that could be equated each to a phylogenetic species. The additional branch formed by the single specimen from Mexico also might be equated to a distinct phylogenetic species.

Morphological studies
From a morphological perspective, specimens in the phylogenetic species C1.1 and C1.2 were very similar, characterized by an overall reddish brown to violet brown pileal surface, light, cork-colored context, occasionally paler in the upper zone -described as not fully homogenous by Torres-Torres et al. (2012)-, with none to several (up to 4) dense, brown stripes or continuous lines of resinous deposits extending from the base of the context toward the margin. The cuticular cells were mainly cylindrical to clavate, apically rounded, regular, amyloid, and the basidiospores ovoid to broadly ovoid, with free to subfree pillars, and chlamydospores ("gasterospores" in Bazzalo and Wright 1982) in their context (Figs 3-4).
These chlamydospores were mostly subglobose in the context of the basidiomes and more variably shaped in pure culture on artificial media, thick-walled, hyaline to yellowish, and with dextrinoid content. In the specimens of C1.1, the chlamydospores were constantly, permanently smooth-walled ( Fig. 3D-F, I) whereas in specimens of C1.2, they were smooth becoming roughened on aging, with free to partially anastomosed fine ridges with a meridian orientation ( Fig. 4E-H, K). The Mexican specimen (Guzmán-Dávalos 9569, IBUG!) also presented chlamydospores in the context but they were punctuated, ornamented with thick pillars (Fig. 5).
In general, the morphology allowed the distinction of three morphotypes, which could be considered as three morphospecies. Each of these also corresponded to a phylogenetic species.

Taxonomic conclusions
The present study, using single and multilocus phylogenetic inferences combined with morphological and in vitro culture studies, concordantly revealed two species of Ganoderma in the G. weberianum sensu Steyaert lineage, spanning over the Neotropics. Furthermore, a specimen from Mexico, represented only by the ITS sequence (Guzmán-Dávalos 9569, Figs 2 & 5), also could be equated to a morphological and phylogenetic species, pending confirmation when additional material is available. However, none of these three species could be equated to G. weberianum sensu Moncalvo (Moncalvo 2000), which is restricted to tropical Asia.
Clade C1.1 contained the ex-type strain of G. subamboinense var. laevisporum; hence, it could correspond to this taxon. The sister clade C1.2 contained the ex-type strain of G. subamboinense var. subamboinense; thus, it could correspond to the typical variety. Furthermore, both the molecular and morphological data would warrant recognition of both varieties at species level.
Ganoderma subamboinense was originally described by Hennings (1904) as Fomes subamboinense Henn. The type specimen originated from Brazil. Bazzalo and Wright (1982) accepted the species and distinguished a var. laevisporum from the typical variety. Both varieties were characterized by the presence of chlamydospores in their context, which were rough-walled with "veins anastomosing to form a sort of reticulum" in the typical variety, and smooth-walled in var. laevisporum, what was later confirmed by Gottlieb and Wright (1999). Ryvarden (2000), based on presumed morphological resemblance, reduced G. subamboinense s.l. (both varieties) as a synonym of G. multiplicatum (Mont.) Pat. Similarities between G. subamboinense var. laevisporum and G. multiplicatum var. vitalii Steyaert were previously reported (Bazzalo and Wright 1982). Inversely, Torres-Torres et al. (2012) recognized G. multiplicatum as an independent species that could be differentiated from G. subamboinense s.l., e.g., in having apically irregular cuticular cells, with many protuberances. Steyaert (1980) had already characterized G. multiplicatum var. vitalii with "mostly irregular" cuticular cells.
The revision of a fragment of the holotype of G. subamboinense var. subamboinense (S F15183!) ( Fig. 4I-K) and of the holotype of G. subamboinense var. laevisporum (BAFC 25525!) ( Fig. 3G-I) confirmed the previous observations (Bazzalo andWright 1982, Gottlieb andWright 1999). Both varieties are mainly characterized by a pale context with resinous incrustations or thin resinous brown bands, stretching from base towards the margin, cylindrical to clavate, apically regular, amyloid cuticular cells, ovoid to broadly ovoid basidiospores with free to subfree pillars, and chlamydospores in their context. The chlamydospores were striated in the typical variety and smooth-walled in var. laevisporum.
The study of the holotype of G. multiplicatum (K 123639!), originating from French Guiana, confirmed irregular cuticular cells with both lateral and apical protuberances, distinct from those of both varieties of G. subamboinense. Phylogenetic analyses inferred from an ITS data set (Bolaños et al. 2016) or a combined ITS-LSU data set (de Lima-Junior et al. 2014) also showed that G. subamboinense var. laevisporum (ITS sequence from the ex-type culture ATCC 52419) and G. multiplicatum (that should be considered as sensu auctores) formed two distinct clades, in two distant lineages. Hence, the synonymy of G. subamboinense and G. multiplicatum, as suggested by Ryvarden (2000), here also is rejected.
Based on these observations and inversely to the previous conclusions of Torres-Torres et al. (2012, 2015), we did not found any consistent morphological difference between G. mexicanum and G. sessiliforme. Furthermore, the type specimen of both names originated from neighboring localities and, probably, related ecosystems. Therefore, G. sessiliforme and G. mexicanum are here considered as synonyms, the latter epithet (Patouillard 1898) having priority. However, the status and affinities of G. mexicanum are uncertain. Patouillard (1898) suggested that G. mexicanum was a sessile form of G. lucidum but with smooth ("lisses"), ovoid basidiospores. There is also a typewritten, undated note from R. Singer in the type specimen folder emphasizing "This is merely Ganoderma sessile Murr.", and then pencil corrected "same as [G. sessile]". Previously, Murrill (1902) also pointed out similarities between G. sessiliforme and G. sessile. Loyd et al. (2018) also suggested that G. sessiliforme might represent a synonym of G. sessile.
Nevertheless, G. sessile has distinctly larger basidiospores, 11.2-14.4 (-16.4) × 7.2-8.8 μm (fide Torres-Torres et al. 2015) and a duplex and spongy context , both features which would justify distinguishing these species. Gottlieb and Wright (1999) and Gottlieb et al. (2000) previously compared G. sessiliforme with G. subamboinense var. laevisporum. They argued that G. sessiliforme differed from G. subamboinense var. laevisporum by the size (up to 11 mm long) and ornamentation ("semirugose" under the SEM) of its basidiospores, and the lack of chlamydospores, both features that do not stand (cf. above). The characters of G. mexicanum, especially the light-colored context with resinous incrustations, the basidiospores size, and the presence of smooth chlamydospores, overall remind much those of G. subamboinense var. laevisporum and of our specimens from the clade C.1.1 but for the cuticular cells. The cuticular cells are more clavate and shorter, 25-38 μm in G. mexicanum compared to those of G. subamboinense var. laevisporum and specimens from C1.1, 30-50 μm (Figs 3G & 6C-D).
Chlamydospores were not reported in the literature for G. parvulum nor for G. stipitatum (Steyaert 1972, 1980, Gottlieb and Wright 1999, Ryvarden 2000 fig. 22d). However, their fig. 8, which is captioned as type of G. parvulum, actually corresponds to the type of G. stipitatum (NY 985678!). The voucher specimen from which these punctuated chlamydospores were observed remained uncertain. Nonetheless, our study of the type of G. parvulum and G. stipitatum revealed scattered chlamydospores in the context of both. These chlamydospores were smooth-walled or also ornamented with anastomosed ridges (Fig. 8D-E).
The likely immaturity of the type of G. parvulum and G. stipitatum, to a certain extent, could prevent definitive taxonomic interpretations. Notwithstanding, we would follow Ryvarden (2000Ryvarden ( , 2004 in considering that these two epithets represent a single species. Furthermore, the main macro-and microscopic characteristics of G. stipitatum and to a lesser extent, of G. parvulum, as described above, overall, also correspond to those of G. subamboinense var. subamboinense. Therefore, G. parvulum, G. stipitatum, and G. subamboinense could be considered synonymous. In this case, the epithet parvulum  (Murrill 1902) has priority over subamboinense (Hennings 1904) and stipitatum (basionym: Fomes stipitatus Murrill 1903), contrary to the conclusion of Ryvarden (2004). Steyaert (1962Steyaert ( , 1980, Bazzalo and Wright (1982), and Gottlieb and Wright (1999) recognized G. bibadiostriatum as a distinct species. Ganoderma bibadiostriatum was characterized by a distinctly brown (fide Bazzalo and Wright 1982, or cinnamon, fide Steyaert 1962, 1980 context and basidiospores 7.0-11.0 × 5.5-7.5 μm, averaging 9.3 × 6.5 μm (fide Steyaert 1980), or 9-11 × 6-8 μm (fide Gottlieb and Wright 1999). These features differed from those of G. parvulum. Through phylogenetic inferences based on ITS and LSU, de Lima-Junior et al. (2014) showed that Brazilian specimens identified as either G. parvulum or G. stipitatum (identifications that should be considered as sensu auctores) were gathered into a single clade, which nested within the G. tropicum clade sensu Moncalvo (2000), and were unrelated to G. subamboinense var. laevisporum, hence unrelated to the G. weberianum-resinaceum lineage. Therefore, contrary to Ryvarden (2000Ryvarden ( , 2004 but following Gottlieb and Wright (1999), we also rejected the synonymy of G. bibadiostriatum with G. parvulum and G. stipitatum. We suggest that the identity of the G. parvulum-G. stipitatum clade shown by de Lima-Junior et al. (2014) should be re-evaluated, and that it might well represent G. bibadiostriatum.
The status and affinities of G. perzonatum have been debated and still are uncertain. Ganoderma perzonatum was described from Cuba (Murrill 1908). Moncalvo and Ryvarden (1997) first related it to G. parvulum. Previously, Steyaert in 1962 andWright in 1967, in two notes joint to the type specimen of G. stipitatum (NY 985678!) and to a second specimen annotated "probable type" [of G. stipitatum] (NY 985716!) also informally suggested that G. perzonatum and G. parvulum were synonymous. However, later on, Ryvarden (2004) retained the species that he associated to the G. resinaceum complex. The revision of the type specimen (NY 985702!) (Fig. 8F-I) confirmed the main features (Ryvarden 2004): a sessile, dimidiate habit with superposed pilei, a pale corky context with dark, resinous streaks, cuticular cells up to 100 μm long, and basidiospores 8.5-9.5 (-10) × 6-7 μm. Furthermore, chlamydospores with smooth or then ornamented with fine longitudinal ridges ( Fig. 8H-I), also were observed, a feature previously unnoticed. These characteristics brought it back to G. parvulum, as first suggested by Moncalvo and Ryvarden (1997).
However, G. perzonatum would differ from G. parvulum in having larger, sessile, dimidiate basidiomes and markedly cylindrical, longer cuticular cells. A specimen originating from the type locality of G. perzonatum (MUCL 43522, La Havana, Cuba, Fig. 9) shared these characters. It also produced striated chlamydospores, both in the context of the basidiome and in pure culture, similar to those of G. parvulum. However, this specimen formed a short, isolated branch, basal to the C1.2 clade, in phylogenetic inferences of the combined data set (Fig. 1). Ganoderma perzonatum remains of uncertain interpretation. It could be included, for the time being, in the concept of G. parvulum (hence G. parvulum s.l.).
The context of both G. mexicanum and G. parvulum was light-colored, usually very pale toward the crust and darker just above the tubes, with none to several brown resinous incrustations or resinous bands variably stretching through the context from the base to the margin. The context in G. parvulum sometimes showed yellow, scattered spots and a thin yellow line just below the crust. Both species have chlamydospores in their context and in pure culture on artificial media. There are not many morphological characters to differentiate them except for the ornamentation of their chlamydospores. However, chlamydospores are sometimes very scarce and difficult to observe in the basidiome. Nonetheless, they are always present, and frequent, in pure culture on artificial media.
The basidiospores were, on average, marginally wider in G. mexicanum in comparison to those of G. parvulum, viz. on average 8.6 × 6.4 μm or 9.0 × 6.0 μm, respectively. The cuticular cells were cylindrical to claviform, occasionally with 1-2 short lateral branches, strongly amyloid, usually smooth or with a fine apical granulation, which was more consistently present in G. parvulum. The cuticular cells also were marginally longer in G. parvulum (up to 100 μm long) compared to those of G. mexicanum (up to 65 μm long).
The distribution ranges and ecologies of both species are still little known. Ganoderma parvulum, as here interpreted, had been observed from the Brazilian Amazon, Colombia, and French Guiana in South America, Costa Rica and Nicaragua in Mesoamerica, and up to Cuba in the Caribbean. Loyd et al. (2017Loyd et al. ( , 2018 reported G. cf. weberianum from the subtropical southern Florida (USA) on the basis of two specimens (UMNFL 32 and UMNFL 100), which DNA sequences, nevertheless, were deposited in GenBank under G. subamboinense var. laevisporum. Loyd et al. (2018) described striated chlamydospores in the context of these specimens, which points toward G. parvulum. Our multilocus phylogenetic inferences showed that these Florida specimens nested within the G. parvulum clade (Fig. 1). However, there was incongruence between the topology resulting from the multilocus-based phylogenies and the ITS-based inferences (Fig. 2) regarding the position of UMNFL 100. The ITS sequences of this showed a change in three nucleotide positions, that could represent a misreading of the sequencer. Notwithstanding, these reports extend the distribution range of G. parvulum northerly to the subtropical, south-eastern USA. This ample distribution would imply a broad ecological range, but also could encompass a hidden diversity.
In French Guiana, G. parvulum has been observed at the Nouragues Nature Reserve (~4°04′18″N, 52°43′57″W), a spot of primary, very humid (3000 mm of rain / year), tropical rainforest characteristic of the Guianas shield, which belongs to the larger Amazonian rain forest phytochorion. Locally, this species was uncommon; three basidiomes only were observed during six, two-to three-weeks long surveys of polypores. These three specimens were found emerging from dead, fallen trunks. In French Guiana, it has been observed also once in an anthropic, semi-urban environment (culture BRFM 1043, voucher specimen and data on the substrate and host unavailable). The type specimen of G. parvulum, originating from Brazil, was also, most likely, collected in the same phytochorion. In Cuba, Greater Antilles, the species has been observed mostly in anthropic, urban or semi-urban environments (cf. list of specimens examined).
Ganoderma mexicanum, as here interpreted, has been observed from Argentina, Brazil, Martinique (Lesser Antilles), and Mexico. In Mexico, the species is known from a rather restricted area of the Morelos State, which is the type locality of G. mexicanum and G. sessiliforme, and of a third additional specimen collected in secondary tropical forest with Quercus sp. (Torres-Torres et al. 2015). Raymundo et al. (2013) and López-Peña et al. (2016) also reported G. sessiliforme from xerophylic vegetation with Quercus sp. in Sonora, but the voucher specimens were not available for confirmation. In Martinique, the species was found in mesophylic to distinctly xerophylic forests, which could represent, locally, its preferential habitat. Several collections came from The Caravelle Peninsula, which is characterized by a seasonally dry season.

Discussion
The current morphological concept of Ganoderma weberianum dated back from Steyaert (1972) and was based on Fomes weberianus. However, on one hand, the very identification of F. weberianus remained questioned. As raised by Yombiyeni and Decock (2017), there was confusion around the modern interpretation of this taxon and its generic placement was debated; the species was either considered in Ganoderma, following Steyaert (1972), or in Phylloporia, following Ryvarden (1972). On another hand, the circumscription of G. weberianum sensu Steyaert remained questioned and, consequently, its distribution range remained uncertain.
Fomes weberianus, originating from Samoa ("in insula Samoa"), was first described by Saccardo (1891). This author did not specify a type or any reference specimens, mentioning only "Exempl. in Museo berolin" (nowadays B). The current concept of G. weberianum was developed based on a specimen held in B (#700007410) stamped as type; this specimen represents indeed a species of Ganoderma, hence G. weberianum sensu stricto (Steyaert 1972). However, the same year, Ryvarden (1972) recombined F. weberianus into Phylloporia, although without citing any reference specimen.
Nonetheless, in addition to the type cited by Steyaert (1972), two other specimens annotated as "Fomes weberianus, Weber, Samoa" exist, of which one also is stamped as type. One of these two specimens is located at B [Samoa Island, Weber "Fomes Weberi" det. P. Henn., Fomes weberianus (#700021870!)], and the second in the Bresadola herbarium in S [Samoa Island, Weber, det. P. Henn. and Bresadola as Fomes weberianus "n. sp." "Typus!" F15098!]. These two latter specimens do not represent a species of Ganoderma but a species of Phylloporia; their morphological features agree very well with the modern, morphological concept of this genus (e.g., Wagner and Ryvarden 2002). Moreover, and essentially, the morphological characters of these two specimens are in complete agreement with the original diagnosis of F. weberianus (Saccardo 1891).
This diagnosis was, partly, a copy of a handwritten description, contemporary to, or previous to Saccardo (1891), and which is still present within the folder #700007410 in B. It emphasized a duplex context ("strato duplice") made of an upper tomentose to floccose layer ("superiori tomentoso-floccoso") and a lower, corky layer ("inferiori suberosolignoso"), separated one from the other by a thin black line ("a superiore linea nigra limitato"). Saccardo (1891), following the above-cited note, related F. weberianus to Polyporus circinatus (Fr.) Fr. and P. tomentosus Fr., two Hymenochaetaceae nowadays accepted in Onnia (Ryvarden 1990). Subsequent early interpretations of F. weberianus (e.g., Bresadola 1914, 1916, 1925, Lloyd 1915, Cunningham 1950) also associated this species to taxa that are, mainly, akin to species of Phylloporia as currently accepted. As far as we had been able to ascertain, there was no pre- Steyaert (1972) interpretation of this taxon as a species of Ganoderma.
This casted doubts on the interpretation of F. weberianus; considering the original diagnosis and both specimens from B and Bresadola herbarium in S, Phylloporia weberiana sensu Ryvarden (1972), most likely, is the correct interpretation. Hence, a lectotype should be designated. This will be discussed in more detail later on.
Our results, following Moncalvo (2000), confirmed that G. weberianum sensu Steyaert was polyphyletic and encompassed several species. Multilocus phylogenetic inferences had shown distinct, well-supported clades and an overall phylogenetic structure corresponding to a geographical pattern (Fig. 1). The G. weberianum sensu Steyaert lineage was divided into two main sublineages, viz. a Neotropical and Paleotropical sublineages.
As far the Neotropics are concerned, at least two species were confirmed, G. mexicanum (previously variably reported as G. sessiliforme and G. subamboinense var. laevisporum) and G. parvulum (previously known as G. subamboinense). Furthermore, the specimen MUCL 43522 from Cuba could represent G. perzonatum. Although we are of the opinion that G. perzonatum may well represent a species on its own, more material, ideally from various localities, and DNA sequences, is necessary to draw a definitive conclusion. On the other hand, the specimen Guzmán-Dávalos 9569 (IBUG!) from Mexico, basal to the G. parvulum s.l. / G. mexicanum s.l. clade in the ITS-based phylogenetic inferences (Fig. 2) and with chlamydospores ornamented with isolated pillars, also could represent a distinct taxon. This demonstrates a likely higher than known phylogenetic and morphological diversity and, ahead, taxonomic diversity. Several additional names also remain of uncertain status and, if any, unknown affinities; it includes G. argillaceum and G. praelongum, or still G. multiplicatum and G. vivianimercedianum. Collections from their type localities and DNA sequences data are highly needed.
The Paleotropical sublineage was further divided into three clades, including an African, an Indian, and a tropical Asian / Australasian clade, representing, at the least, as many phylogenetic species or species complexes. As regard to the situation in Central Africa, at least one species could be segregated from the G. weberianum sensu Steyaert. Ganoderma carocalcareum Douanla-Meli (Douanla-Meli and Langer 2009) could apply for this taxon but three previous priority names might apply too, which will require a revision of their type specimens.
On the basis of our phylogenetic inferences, we conclude that G. weberianum in Southeast Asia and Australia (which would correspond to G. weberianum sensu Moncalvo 2000) also is a complex of species. It would include, at least, G. rivulosum (S F181158!) and G. microsporum (isotype BPI!). Moncalvo et al. (1995) suggested that there are few differences between G. microsporum and G. weberianum, and later on, Moncalvo (2000) considered both names as synonyms (Moncalvo 2000), which was also the opinion of Smith and Sivasithamparam (2003) and Wang et al. (2005). However, this synonymy needs to be ascertained.
The sister clade of G. weberianum sensu Steyaert lineage is, in our phylogenetic analyses, hitherto, the clade D, which comprised specimens from Central Africa and China, the latter referenced at GenBank as G. hoehnelianum. Ganoderma hoehnelianum was described by Bresadola (1912) from Java (Indonesia) having basidiomes with a "crusta, tenui, opaca". This "crust" was observed in the type specimen (S F181067!) and is different from the laccate pileal surface of the G. weberianum complex, made of cuticular cells organized in a dense palisade. Therefore, the identity of this clade also should be ascertained.
This study also confirmed that G. resinaceum sensu auctores from China, East Africa, Europa, and both North and South America represented a species complex. Loyd et al. (2018) showed that G. resinaceum sensu American auctores encompassed at least two distinct species, viz. G. polychromum and G. sessile. These results agreed with those of Moncalvo (2000), who distinguished European and North American "populations" of G. resinaceum, on the basis of which it was proposed that these "disjunct and genetically isolated [populations]" "may warrant recognition at the species level". Our study showed that the G. resinaceum sensu European auctores also represented a species complex, with two well-supported phylogenetic species (Fig. 1); thus G. resinaceum in Europe also could hide a larger than expected diversity.
As emphasized by Moncalvo (2000) and Richter et al. (2015), the identification of species in Ganoderma was commonly based on the microanatomy of the pileus surface, the basidiospores morphology and size, and in some cases, the host relationships. The occurrence of chlamydospores in the basidiomes or in in vitro cultures also was highlighted as a valuable feature (Moncalvo 2000, Hong and Jung 2004, Richter et al. 2015, Loyd et al. 2019, although this character, as suggested by Steyaert (1980), also could be environment dependent. Our results concerning the Neotropical species of the G. weberianum complex confirmed the presence of chlamydospores, in basidiomes and in in vitro cultures, and their ornamentation, as pertinent taxonomic features for the systematic of this group. Three ornamentation types, smooth, with pillars, or with ridges have been observed in the Neotropical species. A similar situation could occur in G. weberianum sensu Moncalvo in Southeast Asia. Steyaert (1972) described G. weberianum with both smooth and ornamented chlamydospores with pillars, "columns", or "partitions". Smith and Sivasithamparam (2003) confirmed these observations based on examination of the type specimen (cited by Steyaert 1972) and specimens from Australia and the south Pacific regions.