Wolfina aurantiopsis, a rare species in the family ()

Carlo AGNELLO Summary: A detailed morphological study of the rare ascomycete aurantiopsis (Ellis) Eckblad is pre- Matteo CARBONE sented after the thorough study of two collections from Tennessee (USA). Macroscopic and microscopic Christine BRAATEN color photographs as well as line drawings are included. Notes regarding and nomenclature of W. aurantiopsis and related species are proposed. Keywords: , Galiella, Trichaleurina, , Wolfina oblongispora, Wolfina papuana, Tennes- Ascomycete.org, 5 (1) : 39-45. see, eastern USA, taxonomy. Janvier 2013 Mise en ligne le 03/01/2013 Riassunto: Viene presentato uno studio morfologico dettagliato della rara Wolfina aurantiopsis (Ellis) Eckblad basato su due raccolte effettuate nel Tennessee (USA). Lo studio è corredato da fotocolor in habitat, foto al microscopio e disegno a tratto. Vengono inoltre fornite note tassonomico-nomenclaturali e discusso un confronto con specie simili. Parole chiave: Ascomycota, Galiella, Trichaleurina, Sarcosoma, Wolfina oblongispora, Wolfina papuana, Ten- nessee, U.S.A. orientali, tassonomia.

Résumé : une étude morphologique détaillée du rare ascomycète Wolfina aurantiopsis (Ellis) Eckblad est présentée d’après l’examen approfondi de deux récoltes du Tennessee (États-Unis). Des photographies ma- croscopiques et microscopiques ainsi que des dessins au trait sont ajoutés. Des notes concernant la taxino- mie et la nomenclature de W. aurantiopsis et des espèces associées sont proposées. Mots-clés : Ascomycota, Galiella, Trichaleurina, Sarcosoma, Wolfina oblongispora, Wolfina papuana, Tennes- see, est des États-Unis, taxinomie.

Introduction Materials and methods

The Wolfina was established by SEAVER (1937) and was Microscopical characters are based on fresh and dry specimens. based on Peziza aurantiopsis Ellis. ECKBLAD (1968) then validated the Three optical microscopes were used: Nikon Eclipse E600 com- genus due to the previous lack of a Latin diagnosis (required at that pound microscope, Olympus CX41 trinocular and Optika B353 tri- time, ex art. 36.1 I.C.B.N.): “Apothecia juvenilia clausa, matura dehis- nocular with plan-achromatic objectives 4×, 10×, 40×, 60× and 100× centia, cupulata, excipulo toto textura intricata constructo, extus to- in oil immersion. The following main reagents were used: Melzer’s mentosa, tomento e stratis cellularibus externis formato; pili tomenti reagent, cotton blue and Congo red. Water mounts were used for semplice, septati, recti vel undulati, pariete fusco, crasso instructi, laeve the observation of the pigmentation and spore size. At least 30 vel basin versus exasperati. Asci cylindrici, non amyloidei, octospori, pa- spores were measured from each apothecium and from spore de- riete crasso instructi. Sporae ellipsoideae, hyalinae, non guttulatae, posit. leaves vel longitudinaliter leviter striatae. Paraphyses filiformes, rectae, septatae”. Finally, the genus was placed into the family Chorioacti- Taxonomy daceae Pfister by FISTERP et al. (2008). Until now, only two other species have been described in this Wolfina aurantiopsis genus, Wolfina oblongispora (J.Z. Cao) W.Y. Zhuang & Zheng Wang (Ellis) Seaver ex Eckblad, Nytt. Mag. Bot., from China and Wolfina papuana Otani from Papua New Guinea (see 15(1-2): 126 (1968). discussion). However, further molecular and systematic study is nee- ded to determine their phylogenetic placement. Basionym: Peziza aurantiopsis Ellis, Bull. Torrey Bot. Club, 9: 18 After careful evaluation of the family Kobayasi, (1882). the authors of this paper were obliged to consider and explore the sister family Chorioactidaceae Pfister (PFISTER et al., 2008). Phyloge- Synonymy: ≡ netic studies conducted on Sarcosomataceae by CARBONE et al. Lachnea aurantiopsis (Ellis) Sacc., Syll. Fung., 8: 180 (1889); Scu- (2013), lead to the accommodation of two additional genera into tellinia aurantiopsis Kuntze, Rev. Gen. Pl., 2: 869 (1891). Chorioactidaceae: the new genus Pseudosarcosoma M. Carbone, = Sarcosoma carolinianum Durand, J. Mycol., 9: 103 (1903), fide Agnello & P. Alvarado, and the established genus Trichaleurina SEAVER (1937). Rehm. Nevertheless, our knowledge of the four original Chorioacti- daceae genera (i.e., Kupfer ex Eckblad, Lib., Original diagnosis: “Peziza (Sarcoscypha) aurantiopsis – Sessile, Paden & Tylutki and Wolfina Seaver ex Eckblad) was limi- about one inch across, with a coarse, felt-like, black-brown myce- ted to Neournula pouchetii (Berthet & Riousset) Paden only. lium at the base, matted together on hardwood branches, etc., on Wolfina aurantiopsis has a presumably restricted geographic which it grows; the outside of the cup also coated with coarse range limited to an area comprising several states in the eastern Uni- (.00025’ diam.) brown, smooth sparingly-branched, continuous ted States. Recently, WANG (2011) reported this species from Taiwan, hairs; disk clear pale yellow, nearly sulphur yellow, becoming dull but we believe that it should be investigated genetically to figure orange when dry; margin thin, pale, narrow, erect; flesh white, thick out if it is conspecific with the American collections. Anyway, after (1/8’-1/4’), firm, elastic, dry; asci cylindrical, subtruncate above, thorough examination of collections made from Fall Creek Falls .0013-.0014 × .0006’; paraphyses stout, septate, slightly thickened State Park and the Great Smoky Mountains National Park (GSMNP) above; sporidia oblong-elliptical, granular, .0009’-.0010’ × .0006’, in Tennessee, we are now able to present a detailed study of this un- epispore smooth. On the bare soil and on decaying wood and leaves common species. in low, sandy, oak and pine woods, Sept. 1881.” 39 Description of studied collections clavate-cylindrical, narrowed below to a long, slender pedicel, apex Macroscopical characters rounded, not blue with iodine, 400-450 × 18 μ, opening by a lid. Apothecium shallow cup-shaped, sessile, 2.5–4.5 cm wide. Hy- Spores 8, uniseriate, hyaline, smooth, continuous, elliptical, 25-30 × menium shallow, pale yellow to pale orangish-ochraceous in age, 15 μ (the majority 28 × 15 μ); Paraphyses cylindrical, hyaline, sep- smooth. External surface rough, black, convoluted. Flesh thick, tate, very little thickened at the free tips, 3 μ thick”. white, firm, corky in dried specimen; odor and taste not distinctive. Notes on similar species Microscopical characters Morphological characters reported here are clearly identical with Asci cylindrical, 340–430 × 14.5–20 μm, operculate, inamyloid, 8- those made by many authors (DURAND, 1903, as Sarcosoma caroli- spored, with walls up to 2 μm thick; base tapering and flexuous.Pa- nianum; SEAVER, 1911, 1937, 1942; ECKBLAD, 1968; ROODY, 2003; BES- raphyses filiform, septate, anastomosing, 3–4.5 μm wide, as long as SETTE et al., 2007; PFISTER et al., 2008). The only exception is in ELLIS the asci, not or only slightly enlarged at the tips up to 5 (–5.5) μm. (1882) where we find slightly smaller spores of ca. 25 × 15 μm. Al- Spores elliptical to subcylindrical, 25.4–32 × 10.4–15 μm (on ave- though collections of this species have been recorded from a wide rage 28.3 × 12.5 μm), Q = 2.03–2.52 (average 2.27), hyaline, with gra- but confined area in the eastern United States (PFISTER et al., 2008), nular content, sometimes embedded in a gelatinous sheath; the we decided to discuss some similar species not present in this area. surface is marked by thin longitudinal, sometimes anastomosing Wolfina oblongispora originally described as Galiella oblongispora striations. Subhymenium of a thick textura intricata made up of cy- J.Z. Cao (CAO et al., 1992), was then amended and transferred to Wol- lindrical, septate, 5–8 (–9) μm wide hyphae, arranged perpendicular fina by ZHUANG & WANG (1998). The most emphasized difference is a to the asci; brownish at low magnification.Medullary excipulum larger spore size, 36–45 × (15–) 17.5–22 μm, however, as reported by of textura intricata made up of hyaline, closely septate, 5–8 (–10) μm ZHUANG & WANG (1998) this species is characterized by spores orna- wide hyphae. Ectal excipulum: not a real ectal excipulum but a sim- mentated by longitudinal striations mixed with some warts. The ori- ple extension of the medullary excipulum where darker hyphae are ginal description: “Apothecia sessilia vel breviter stipitata, obconica, arranged in palisade, mixed with basal part of hairs. External hairs 3.5-4.5 cm alta, 3-5 cm lata. Hymenium flavidum vel alboluteum. Fa- cylindrical, 5–9 (–12) μm wide, septate, with walls up to 1 μm thick, cies exterior pilei nigro-fusca. Hypothecium distinctum, denso-intrica- brown, entirely covered by warts or prickles when young or not well tum. Excipuli medullaris textura intricata in matrica gelatinosa sita. developed, then smooth in almost their length except in the basal Excipuli exterior 80-100 μm crassa, ad texturam angularem vel globu- part where they remain ornamented. losam, cellulis brunneo-tunicatis. Capillae brunneo-tunicata, flexuo- sae, 80-600 × 7-12.5 μm. Asci longo-cylindrici, I-, 8-spori, 400-450 × Ecology and studied collections 18-21 μm. Ascosporae uniseriatae, oblongae, 33-42 × 16-18 μm, ver- Solitary or in small clusters, broadly attached to fallen twigs and ruculosus. Paraphyses filiformis, 2.5-3 μm diam”. branches of hardwood in a section of a trail dominated by Rhodo- Wolfina papuana Otani (OTANI, 1975) was tentatively included in dendron maximum. Wolfina, with the following Latin diagnosis: “Apothecia sparsa, sessi- UNITED STATES OF AMERICA. Tennessee, Van Buren County, Fall Creek lia, disco tenuiter concavo vel fere plano margine humili, circa 1.5- Falls State Park, 518 m a.s.l., 15 July 2012, legit and det. C. Braaten, 2.0 cm in diametro, flavo ubi novo salmoneo ubi exsiccato, receptaculo herbarium number TENN 67714. Tennessee, Cocke County, Great hemisphaerico carne gelatinosa crassa, circa 1 cm crasso, extus rugoso, Smoky Mountains National Park, Low Gap Trail, 610 m a.s.l., 6 Au- castaneo, tomentoso. Excipulum externum tenue 17.5-25.0 μ crassum, gust 2012, legit and det. C. Braaten, herbarium number TENN 67128, strato 2- vel 3-cellularum, cellula extima brunneitunicata oblongata Genbank numbers (LSU - KC306743; ITS - KC306744). 5.0-10.0 × 5.0-7.5 μ, pilis longis simplici ab cellulis extremis projectis, cylindraceis, obtusis, tenuitunicatis, laevibus, septatis, undatis, laxe Discussion conglomeratis, brunneis circa basim, superne pallidioribus; excipulum medullosum maxime crassum, circa 3-5 mm vel plus etiam, textura in- Brief historical and nomenclatural notes tricata , hyphis tenuibus, 2.5-3.0 μ in diametro, laxe intertextis, in gela- Originally described from New Jersey (U.S.A.) as Peziza aurantiop- tina inclusis; hymenium circa 800-1000 μ crassum, massis sis (ELLIS, 1882), Wolfina aurantiopsis was then transferred first in the columelliformibus crystallorum flavorum cubicorum vel oblongorum genus Lachnea by SACCARDO (1889) and subsequently in Scutellinia interspersum. Asci cylindracei, longi, apud basim angustati, crassi tu- by KUNTZE(1891). nicati, suboperculati, liquori jodi non caerulescentes, octospori, 600- SEAVER (1911) applied the name Lachnea aurantiopsis, however he 650 × 15.0-17.5 μ; ascosporae longifusiformes, non-septatae, laeves, (SEAVER, 1937) made the decision to include this species into his new flavidae, modice crassitunicatae, guttulatae, 42.5-52.5 × 13.5-15.0 μ, genus Wolfina due to its apparent lack of affinity with genera that paraphyses anguste cylindraceae, septatae, rectae, semplice, 2.5-3.0 μ had been described at that time. In addition, SEAVER (1937) revised in diametro, contentis flavidis”. We are not currently aware of any fur- the type specimen of Sarcosoma carolinianum Durand (DURAND, ther collections or descriptions of this species and so we can only 1903) and synonymized it with Wolfina aurantiopsis because no use the protologue to summarize the following important charac- morphological differences were observed. We have not yet exami- ters: gelatinized context, fusiform spores, smooth external hairs, pre- ned Durand’s type material, however, given the convincing charac- sence of crystals in the hymenium, thin ectal excipulum consisting terization from Durand’s original description, at present we do not of 2–3 layers of elements. Our opinion is consistent with that of have reason to doubt the synonymy. The original description of Sar- ZHUANG & WANG (1998) in concluding that this species should be ex- cosoma carolinianum is as follows: cluded from the genus Wolfina. “Plants solitary, sessile, attached by a dark brown tomentum; at Macroscopically, biogeographically, and ecologically, Galiella rufa first closed, then opening by a pore at the apex, expanding and en- (Schwein.) Nannf. & Korf appears akin to W. aurantiopsis. However, larging finally becoming saucer-shaped, up to 4 cm. diam. Disk G. rufa is well distinguished by globose to shallow cupulate (sub)ses- tawny-ochraceous, externally brown, covered with a thick, appres- sile apothecia with a irregularly toothed margin, reddish-brown to sed, brown tomentum, threads very long 7-8 μ thick, septate, rather orange-brown hymenium, blackish-brown external surface covered shining, but little wrinkled; substance tough-gelatinous, not at all with hairs that give it a tomentose appearance; the flesh is gelati- watery, so that the plant nearly retains its shape, size and color when nous and rubbery; microscopically it is distinguished by the subfu- dry. Consistency of the dry plant corky, not horny and brittle, and soid ornamented spores (SEAVER, 1928, 1942; BESSETTE et al., 1997; exterior nearly even. Flesh white, excipulum composed entirely of PHILLIPS, 2005). The genera Galiella Nannf. & Korf and Trichaleurina interwoven hyphae which are thick and septate, 5-6 μ diam. Asci Rehm are in need of revision. In any case, all species (tropical or tem- 40 perate) retained in these two genera possess subfusoid to fusoid, glossum Boud. However, from the time of their discovery in Sarco- warted spores. somataceae, the term “hymenial hairs” has been used for those ele- Sarcosoma globosum (Schmidel: Fr.) Casp. is recorded from eas- ments distinguished from the true paraphyses by the lack of septa tern North America (BESSETTE et al., 1997) and is easily distinguished and a larger width. It must be noted that sometimes the hymenial macroscopically by a larger and more fleshy apothecium with a dark hairs are not easily detected, in fact, in some species they can be hymenium. Microscopical features are also particularly distinct due disguised among bundles of paraphyses cohered by an amorphous to the moniliod external hairs (present in immature specimens too). exudate. The absence of hymenial hairs in W. aurantiopsis is noteworthy Notes on hymenial hairs due to our observation that other species in the family Chorioacti- As reported in CARBONE et al. (2013), the term “hymenial hairs” — daceae (i.e. Neournula pouchetii, Pseudosarcosoma latahense, Tri- or setae in the sense of PADEN (1983) — refers to structures that are chaleurina javanica and T. celebica) also lack this microscopic not true hymenial hairs, or setae, because these designations are character. Moreover we currently can say that this feature is present better applied, e.g. to the hymenial elements of the genus Tricho- in all studied Sarcosomataceae species.

Plate 1 – Wolfina aurantiopsis. Pictures: C. Braaten. 41 Plate 2 – Wolfina aurantiopsis. Microscopic characters. A: Spores in ascus in Congo red. B: Empty ascus with operculum in Congo red. C: Upper part of paraphyses in Congo red. D-N: Spores in water. Scale bars: all = 10 μm except C = 5 μm. Pictures: C. Agnello & M. Carbone. 42 Plate 3 – Wolfina aurantiopsis. A: Section of a fresh apothecium, with highlighted parts used for pictures B and E. B: Section. C: Medullary excipulum. D: External part of the excipulum. E: External part of the excipulum and external hairs. F: Young external warted hairs. G: Smooth and warty external hairs. H: Young warty external hairs. I: External hair with tip becoming smooth with growth. Scale bars: all = 10 μm except E = 50 μm. Pictures: C. Agnello & M. Carbone. 43 Plate 4 – Wolfina aurantiopsis. Microscopic characters. A: Section. B: Spores. C: Ascus tip. D: Paraphyses. E: External hairs and lower part of the excipulum. Drawing: C. Agnello. 44 Acknowledgements OTANI Y. 1975. — Some discomycetes collected in Papua New Gui- nea. In: OTANI Y. (ed.). Reports on the Cryptogams in Papua New Gui- nea. National Science Museum, Tokyo: 5-41. We want to thank Dr. Pablo Alvarado (Alvalab) for his service of PADEN J.W. 1983. — Sarcosomataceae (Pezizales, Sarcoscyphineae). DNA sequencing and for the assistance in interpreting the results. Flora Neotropica 37. New York Botanical Garden, 17 p. PHILLIPS R. 2005. — Mushrooms and other Fungi of North America. Fi- References refly Books Ltd. PFISTER D.H., SLATER C. & HANSEN K. 2008. — Chorioactidaceae: a new fa-

BESSETTE A.E, BESSETTE A.R. & FISCHER D.W. 1997. — Mushrooms of Nor- mily in the Pezizales (Ascomycota) with four genera. Mycological theastern North America. Syracuse University Press, 582 p. Research, 112 (5): 513-527. ROODY W.C. 2003. — Mushrooms of West Virginia and the Central Ap- BESSETTE A.E., ROODY W.C., BESSETTE A.R. & DUNAWAY D.L. 2007. — Mush- palachians. Lexington, The University Press of Kentucky. rooms of the Southeastern United States. Syracuse, Syracuse Uni- SACCARDO P.A. 1889. — Sylloge Fungorum. Vol. VIII. Patavia. versity Press. SEAVER F.J. 1911. — Iowa Discomycetes. Bulletin from the Laboratories CAO J.Z., FAN L. & LIU B. 1992. — Notes on the genus Galiella in China. of Natural History of the State University of Iowa, 6 (2): 41-219, plates Mycologia, 84: 261-263. 1-41. CARBONE M., AGNELLO C. & ALVARADO P. 2013. — Phylogenetic studies in SEAVER F.J. 1928. — North American Cup-Fungi (Operculates). New the family Sarcosomataceae (Ascomycota, Pezizales). Ascomy- York. cete.org, 5 (1): 1-12. SEAVER F.J. 1937. — Photographs and descriptions of cup-fungi DURAND E.J. 1903. — The Genus Sarcosoma in North America. Journal XXVIII. A proposed genus. Mycologia, 29: 678-680. of Mycology, 9: 102-104. SEAVER F.J. 1942. — The North American Cup-fungi (Operculates). Sup- ECKBLAD F.-E. 1968. — The genera of operculate discomycetes: a ree- plementary Edn. New York. valuation of their taxonomy, phylogeny and nomenclature. Nytt WANG Y.Z. 2011. — The sarcosomataceous discomycetes in Taiwan. Magasin for Botanikk, 15: 1-191. Fungal Science, 26 (1): 49-56. KUNTZE O. 1891. — Revisio Generum Plantarum. Vol. 2. Paris, C. Klinck- ZHUANG W.-Y. & WANG Z. 1998. — Sarcosomataceous discomycetes in sieck. China. Mycotaxon, 67: 355-364. ef

Carlo Agnello Via Antonio Gramsci 11 72023 Mesagne Italy [email protected] Matteo Carbone Via Don Luigi Sturzo 173 16148 Genova Italy [email protected] Christine Braaten University of Tennessee, Knoxville TN 37996-1610 USA [email protected]

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