Pterosporomyces herrerae (Guevara, Gómez, Castellano) Guevara, Gómez & Z.W. Ge, gen. nov. & comb. nov.

MycoBank number: MB 840660; Index Fungorum number: IF 840660Facesoffungi number: FoF

= Aroramyces herrerae Guevara, Gómez & Castellano, IMA Fungus 7(2) 235-238, 2016 (basionimo)

GenBank: MZ343611 (Type), MZ343612 (paratype) & MK811032 Holotype Guevara 1211 (ITCV1211), paratype Guevara 1225 (ITCV 1225)

Macromorphology. Fruiting bodies 5-18 x 6-15 x 5-10 mm, irregular, globose or subglobose. Basidiomes surface white, pale tan to brownish, mottled dark brown with pale areas when handled and dried, smooth when fresh, much wrinkled when dried, with some white mycelial strands, soil, and organic matter adherent to surface. Peridium <0.5 mm wide, somewhat separable. Hymeneal gleba olive green when young, green-brown to dark brown when mature, nearly black when dried, locules ellipsoid to elongate, stuffed with spores, columella absent when young, thin dendroid in mature, gelatinized, grayish. Rhizomorphs few, small, white attached at base, brownish when handled or when herborized. Odor organic solvent-like. Taste not recorded. Macrochemical characters. Positive reaction with KOH (5%), brown to blackish on the surface of dried specimens.

Micromorphology. Peridium 70-400 µm wide, two- layered. Epicutis 45-175 µm wide, usually on the thinner side with some areas with wart-like protrusions, of septate hyphae, thin-walled, pale yellow-brown to yellow-brown in KOH, repent hyphae 4.5-6.5 µm broad, occasionally inflated cells up to 18 µm broad, with interspersed small crystalline particles scattered across the layer, subcutis 110-135 µm wide, of septate, thin- walled, hyaline in KOH, interwoven to subparallel or cross-weaved hyphae, 6.5-11.0 (-15.0) µm wide. Mycelial strands on peridium of dark brown, filiform, branched hyphae, 2-3 µm broad, encrusted with small crystalline particles, clamp connection present. Trama 37-112 µm wide, hyaline in KOH, thin-walled, compactly interwoven to parallel hyphae, 2-5 µm wide, in a gelatinized matrix, clamp connections present. Basidia not observed. Spores without utricle and hilar appendage 10.5-12.3 x 5.3-7.0 µm, mean = 11.3 x 6.0 µm, with utricle and hilar appendage (12.3-) 13.2-14.0 x 7.9-9.7 (-10.5) µm, walls up to 1 µm thick, oblong fusoid, ellipsoid, symmetrical, smooth when young, spinny within the utricle when mature and not encompassing hilar appendage, often not equally inflated, rarely encompassing entire spore, commonly laterally inflated up to 5 (-6) µm broad, hyaline to yellow-orange singly, pale brown in mass in KOH, inamyloid, non-dextrinoid in Meltzer reagent.

b) Distribution, habit, habitat, and ecology

México, Michoacán, in the Trans-Mexican Volcanic belt, hypogeous, solitary to groups, under Quercus castanea Muhl. Q. obtusata Bonpl., Q. magnoliifolia Née, Q. rugosa Née, Pinus leiophyla, Pinus pseudostrobus Schl. & Cham., and Pinus michoacana Mtz. at approximately 2160 m elevation, September and October.c) c) Specimens examined

State of Michoacán, MÉXICO, locality Puerto Madroño; ejido Atécuaro, Municipality of Morelia, 19° 32’ 113”, 101° 12’ 5”, 18 Oct. 2011, G Guevara 1211 (ITCV 1211 holotype), G. Guevara 1225 (ITCV 1225 paratype), G. Guevara 1218 (ITCV 1218). Ichaqueo, 20 Sept. 2014, V Gomez-Reyes 863, 877, (EBUM paratype).

Observations. Pterosporomyces herrerae is recognized morphologically by its olive-green to the brown-green color of the gleba and spores with a distinctive inflated, wing-like appearance to the utricle (inflated up to 6 µm), associated with Quercus spp. and ITS, ATP6 gene variation. Restingomyces reticulatus is similar to P. herrerae, but the former has reticulated basidiospores. This novel taxon also resembles to Hysterangium inflatum Roadway, a member in the family Hysterangiaceae with olive-green, brown to black gleba but differs by presenting smaller spores of 9-12 x 8-9 µm including ornamentation with utricle up to 2.5 µm on the side, ectomycorrhizal associated with Eucalyptus spp. and very distant genetically in ITS and ATP genes. Similarly, Hysterangium stoloniferum Tul. & C. Tul. resembles Pterosporomyces herrerae, but the former presents bigger spores of 17-21 x 6-8 µm and a narrow adnate rugose utricle.

Discussion. The study of hypogeous fungi has been very limited in Mexico (Cázares et al. 1992; Trappe & Guzman 1971). However, in 2016 Aroramyces balanosporus and A. herrerae were proposed as new taxa based only in macro (brown to blackish gleba) and microscopic (fine spines within the utricle) morphology without molecular support (Guevara et al. 2016; Castellano et al. 2000). Posteriorly, an ATP6 and ITS gene analysis were performed on these species to confirmed their novel status. The analysis confirmed the novel status of A. balanosporus (Genbank MK811031) as expected (Peña-Ramirez et al. 2019). However, the phylogenetic study surprisingly revealed that A. herrerae belongs to the evolutive basal family Trappeaceae and Trappea darkeri, Restingomyces reticulatus, and Phallobata alba. Thus, Aroramyces herrerae was transferred from the Hysterangiaceae to Pterosporomyces within the family Trappeaceae, an ancestral taxon in Phallales. The basidiomata of Pterosporomyces herrerae in cross-section are similar to those in Restingomyces reticulatus having both brown jelly dendroid gleba as seen in Aroramyces, but differs in the reticulate spore ornamentation present in Restingomyces; in contrast, Pterosporomyces herrerae shows utricle (wing) spores up to 5(6) µm tall. It seems to be that the utricle is a convergent evolutive feature similarly observed in other hypogeous species such as Austrogautieria in the family Gallaceaceae, which possesses longitudinally ridged spores alike to those in Gautieria (Hosaka et al. 2006). Similarly, the ITS and ATP6 genes analysis showed that Pterosporomyces herrerae along with Restingomyces reticulatus is ancestral of epigeous related genera such as Clathrus, Phallus, Dictyophora, Mutinus, among other stinkhorn taxa that evolved from hypogeous gasteroid forms in the Phallales clade in agreement with Hosaka et al. (2006) and Sulzbacher et al. (2016). Fig. 1 & 2.

With these results, the members of the family Trappeaceae increase to five, Trappea darkeri, T. phillipsii, T. pinyonensis, Restingomyces reticulatus, and Pterosporomyces herrerae, hoping to answer the question, “how many more taxa await to be described from the Neotropical Forest ?”

Figure 1: Pterosporomyces herrerae, phylogenetic tree inferred under the maximum-likelihood (ML) criterium for ITS gene with Mr. Bayes . The posterior probabilities for each clade are shown on the branches. The accession numbers in the sequence labels indicate the GenBank accession numbers

Figure 2: Pterosporomyces herrerae, phylogenetic tree inferred under the maximum-likelihood (ML) criterium for ATP6 gene with Mr. Bayes . The posterior probabilities for each clade are shown on the branches. The accession numbers in the sequence labels indicate the GenBank accession numbers

Figure 3: a-i, Pterosporomyces herrerae comb. nov.(ITCV 1211Typus), a, b, & c: cross-sections of basidiome (bar = 1 cm), d: basidiomata (bar = 1 cm), e: close up of gleba, f: epicutis with pseudoparenchyma cells, (bar = 10 µm ), g: Trama (bar = 10 µm), h & i: basidiospores with inflated utricle (bar 10 = µm)