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Taxonomic status of Genabea, with two new species of Genea (Pezizales)
Myca!. Res. 95 (8): 986-994 (1991)
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Taxonomic status of Genabea, with two new species of Genea (Pezizales)
BIN-CHENG ZHANG* Department of Mycology, Institute of Microbiology, Academia Sinica, Beijing 100080, China
The taxonomic position of Genabea in the Pezizales is discussed. Due to discovery of an intermediate species between Genea and Genabea in China, the modern concepts of these two genera become confused. In this paper, Genabea is reduced to subgeneric rank in the genus Genea, Le. Genea subgen. Genabea (TuL & C. TuL) stat. nov. (basionym: Genabea TuL & C. TuL), typified by Genea fragilis (TuL & C. Tul) comb. nov. (basionym: Genabea fragilis TuL & C. TuL). The species formerly included in Genabea are revised. Three species of Genea found in China are described and illustrated in detail, including Genea sinensis, Genea variabilis which are described as new and Genea fragilis as new to the Chinese mycoflora. A key to the species is provided. In the earlier literature, different authors included different genera in the Geneaceae. Fischer (1938) suggested that the Geneaceae included Petchiomyces Fischer & Matt., Genea Vitt., Myrmecocystis Harkn. and Genabea Tul. & C Tul. This was followed by Hawker (1954) and Gilkey (1954b, 1961). Based on his studies on the type material, Trappe (1975) considered that Myrmecocystis and Genabea were congeneric, and redelimited Genea and Genabea. When the name Geneaceae Trappe was validly published by Trappe (1979), only Genea and Genabea were included. Since then, Trappe's (1979) system has been widely accepted (Hawksworth, Sutton & Ainsworth, 1983; Eriksson & Hawksworth, 1988). On taxonomic status of the Geneaceae, however, Pfister (1984) and Zhang (1991) had different opinions. Recently, I found a few collections referable to both Genea and Genabea. These collections are described as new taxa below, and they apparently challenge the current concepts of Genea and Genabea. This paper deals with this problem, and describes species of Genea found in China. Material used in this study was from K and HMAS. Under light microscopy, the measurements and colours were obtained based on material mounted in lactophenol unless otherwise mentioned. Thin sections for study of peridial structure were made by a freezing microtome. For SEM observations, part of the hymenium of a dried ascoma was rehydrated on a clean glass slide, then stuck onto an adhesive layer on a small clean coverslip, then coated with gold and observed using a Hitachi S570 SEM.
Taxonomic status of Genabea The separation of Genahea from Genea was questioned by Korf (1973). He considered Genea, Genabea and Myrmecocystis to be congeneric. Trappe (1975) synonymized Myrmecocystis with Genabea, but he retained Genabea as a distinct genus, although • Present address: CAB International, Wallingford, Oxon, U.K.
he agreed that. strictly from the standpoint of ascocarp structure, Genea and Genabea may be considered to be congeneric. Trappe (1975) redelimited two genera as follows: Genea: spores verrucose, uniseriate; asci cylindric; hymenia only occasionally interrupted by sterile zones of paraphyses. Genabea: spores echinulate, uniseriate, biseriate or randomly arranged; asci clavate to ellipsoid; hymenia regularly separated into pockets by sterile zones of paraphyses or isodiametric cells. Discovery of Genea variabilis B.-C Zhang sp. nov. described below makes this delimitation very difficult. G. variabilis has coloured ascospores ornamented with spines (a feature of Genabea); asci are mostly cylindrical and contain uniseriate spores (a feature of Genea), but are sometimes clavate to ellipsoidal with biseriate to irregularly arranged spores (a feature of Genabea); hymenium is continuous or tending to be interrupted by sterile mycelial strands (a feature of Genea) (Figs 16-24). Other characters such as ascomatal gross morphology and peridial structure are similarly referable to both genera. If ascospore ornamentation and colour are emphaSized, this species certainly belongs to Genabea sensu Trappe (1975). But it should be remembered that some species in Genea sensu stricto have ascospores bearing blunt spines, e.g. in Genea sinensis B.-C Zhang sp. nov. (Figs 14-15). If continuity of hymenium is emphasized, it should be placed in Genea, but its hymenium tending to be interrupted provides a connexion between the two genera. The morphology of asci and the arrangement of spores within showed no borderline between Genea and Genabea. I thus agree with Korf (1973) to synonymize the two genera, and the name Genea takes priority over Genabea, Genabea species (including G. variabilis), however, have coloured, spinose ascospores, which are different from the species in Genea sensu stricto. For convenience of identification and for reflecting phylogenetic progression within the genus,
Bin-Cheng Zhang I propose that Genabea is reduced to a subgeneric rank in
Genea sensu lato. Genea Vin. subgen. Genea, Monogr. Tuberac.: 27 (1831). Type species: G. verrucosa Vitt. (lectotype, fide Trappe, 1979). Genea Vitt. subgen. Genabea (Tul. & C TuL) B.-C Zhang, comb. nov.
987 other authentic material, I will not make the transfer in the present paper.
Genabea sphaerospora Matt., Malpighia 14: 256 (1900). This is a lesser mown species. Its placement in Genea needs a type study.
Genabea TuL & C TuL, Giorn. Bot. Ital. 1 (2): 60 (1845).
Genabea tasmanica Massee & Rodway, Kew Bull. 1989: 125 (1989), nom. illegit. (fide Hawksworth & Udagawa, 1976).
Type species: Genea fragilis (TuL & C TuL) B.-C Zhang. Species formerly included in Genabea are revised as follows:
Sphaerodes beatonii (D. Hawksw.) P. Cannon & D. Hawksw., Bot. ]. Linn. Soc. (London) 84: 145 (1982).
Myrmecocystis cerebriformis Harm., Proc. Calif. Acad. Sci. 3 (I):
This species was described under Genabea, but was based on the asci and ascospores of Sphaerodes beatonii parasitizing ascomata of Labyrinthomyces species. The type specimen (K) was examined.
269 (1899). Genea cerebriformis (Harm.) Gilkey, Univ. Calif. Publ. Bot 6: 304 (1916).
Species of Genea in China
Genabea cerehriformis (Harm.) Trappe, Mycotaxon 2: 118 (1975).
I have not examined any specimen of this species, but Trappe (1975) considered it to be an acceptable species. For other synonyms, see Trappe (1975).
Genabea spinospora (Gilkey) Trappe, Mycotaxon 2: 118 (1975).
Myrmecocystis spinospora Gilkey, Mycologia 53: 25 (1961). According to Gilkey (1961) and Trappe (1975), this species is a good species. Because I have not seen the type or any
Tao (1988) reported Genea globospora Liu & Tao (nom. ined.) and Genabea spinospora found in China. I examined the specimens deposited in the Mycological Herbarium of Shanxi University (MHSU). Because the specimen identified as Genabea spinospora is not identical to Gilkey's (1961) description and not a member of Genabea, and because G. globospora has not been validly published yet, they will not be discussed further in this paper. Based on my collections, three species are described below; among them two species are described as new.
Key to the species of Genea known to China 1. Ascomata with brown superficial hyphae; ascospores hyaline, warted 1. Ascomata without superficial hyphae; ascospores pale brown to brown, spinose
. G. sinensis 2
2. Ascospores globose; asci and paraphyses hyaline, not fragile 2. Ascospores ellipsoidal; asci and paraphyses brown, fragile.
G. variabi/is . G. fragi/is
Genea fragilis (TuL & C Tul) B.-C Zhang, comb. nov. (Figs 1-9)
tissue a textura prismatica to angularis when close to peridium and epithecium. Paraphyses brown, multi-septate, sometimes constricted at the septa, fragile, often anastomosed to form a textura prismatica surrounding asci, the apical cells differentiated to form an epithecium. Epithecium similar to the peridium in structure but slightly thinner. Asci clavate to ellipsoidal, 85-150 x 40-60 !-lm, brown, thin-walled, fragile, with 73-6 uniseriate to irregularly arranged spores (because asci are fragile, it is so difficult to keep asci intact while sectioning that it is not possible to count accurately the number of spores), irregularly arranged among paraphyses. Ascospores ellipsoidal to broadly ellipsoidal, pale brown to brown, 23-30 x 19-21!-lm excluding ornamentation, ornamented with walls about I !-lm thick, with dense spines up to 3-5 !-lm high, thin, often attached with fragments of asci and some deposits.
Genabea fragi/is TuL & C TuL, Giorn. Bot. Ital. 1 (2): 60 (1845).
Ascomata irregular, nodose, lobed, 4-6 mm diam., greyish brown, dried brown, hollow, without conspicuous apical hole, surface warted to roughened; the interior slightly paler, warted to roughened; dark brown on the cut surface, separated by pale sterile tissue visible under dissecting microscope. Peridium 150-250 !-lm thick including warts up to 70 !-lm high, of 2 layers; the outer (ectal excipulum) composed of cells globose to subglobose, pale brown to yellowish brown inward, 17-40 ( - 50) !-lm diam., thick-walled, forming a textura globulosa; some outermost cells irregularly outgrowing to multiple globose to subglobose cells simply piling up to form primitive warts; the inner (medullary excipulum) subhyaline, composed of cells more or less isodiametric, 7-17!-lm diam., with walls l!-lm thick, forming a textura angularis, sometimes forming a textura intricata when close to the hymenium. Hymenium composed of irregularly arranged asci and paraphyses, interrupted and separated by sterile tissue into many fertile pockets (discontinuous hymenia); sterile
Specimen examined: China: Sichuan, Wenchuan, Wolong Natural Reserve, nr Shanwan, alt. 2250 m, hypogeous in Larix mastersiana Hedh. & Wils. woods, B.-C. Zhang 645, 22 Sept. 1989 (HMAS 60282). Genea fragi/is is a new record to the Chinese mycoflora, and is the first report in Asia. The Chinese collection generally agrees with the' authentic' specimen in K which I examined,
Taxonomy of Genea and Genabea
Figs 1-5. Genea fragilis (B. C. Zhang 645). Fig. 1. Section of ascoma, showing interrupted hymenia, peridium and epithecium (bar = 200 I.lm). Fig. 2. Peridial structure (bar = 50 I.lm). Fig. 3. Epithecium (bar = 50 I.lm). Fig. 4. Fragile sterile tissue separating hymenia (bar = 20 I.lm). Fig. 5. Fragile, coloured asci and paraphyses (bar = 20 I.lm).
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Bin-Cheng Zhang
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Figs 6-9. Genea fragilis (B.-C. Zhang 645). Fig. 6. Light micrograph of ascospores (bar = 20 ~m). Figs 7-8. Scanning electron micrographs of ascospores, showing the ornamentation, note ascus fragments on the surface (bar = 5 ~m). Fig. 9. Detail of spore ornamentation (SEM, bar = 1 ~m).
especially in fragility of asci and paraphyses, and ascospore morphology (Figs 5-9), but somewhat different is ascomatal colour and ascospore size. The authentic specimen and collections from Switzerland and Canada are black or sooty (Tulasne & Tulasne, 1845; Gilkey, 1954a), but our material is greyish brown. The ascospores from the authentic specimen
are 27-38 x 20-27 !-lm excluding ornamentation, larger than those of our material. Because colour changes of ascomata often occur at different developmental stages and with drying conditions, and because the spore size is often variable, it is suitable to assign our collection to G. fragilis. Genea fragilis differs from other species in subgen. Genabea
990
Taxonomy of Genea and Genabea in its fragile, coloured asci and paraphyses, and ellipsoidal ascospores (Figs 5-6). Coloured asci and paraphyses are very peculiar in the hypogeous Ascomycetes. In our materiaL the colour of asci and paraphyses is obviously much darker than that of sterile tissue and peridium, and nearly the same as that of ascospores. This feature is doubtless very useful in species separation. Genea fragilis was found only in association with Larix mastersiana in China and shares the same habitat with Genea variabilis. L. mastersiana is an indigenous species in China, and Sichuan is the main distribution area (Zhou, 1989). The association of G. fragilis with L. mastersiana is worth further study. Genea sinensis B.-C. Zhang, sp. nov.
From ascomatal gross morphology and ascospore morphology, G. sinensis apparently belongs to the G. hispidula Berk. group, including G. hispidula, G. arenaria Harkn., G. brachytheca Gilkey and G. echinospora Gilkey. G. sinensis is closely related to G. hispidula, but the former differs from the latter in its superficial hairs much shorter and sparser, epithecium thinner and without hairs, and ascospores smaller and with conical warts to blunt spines (Figs 14-15; see Hawker, 1954; Minter & Zhang, 1989). G. sinensis is also similar to G. brachytheca in gross morphology, but the latter has much shorter asci with clustered ascospores ornamented with papillae (Gilkey, 1954). All these features are different from our new species. G. arenaria and G. echinospora are obviously different from G. sinensis (Gilkey, 1939, 1954a).
(Figs 10-15)
Ascomala applanato globosa, cava, lobata, 5-9 mm diam., brunneola, basi caespitibus mycelialibus; aspeetus exteriores sparse hirsuti, verrucosi, hyphae superficiales usque ad 750 ~m longae, brunneolae vel brunneae; aspeetus interiores luteoli, verrucosi. Peridium exterius 100-- ISO ~m crassum, lexlura angulari; peridium interius 60- IIO ~m crassum, lexlura inlricala. Epilhecium 50--100 ~m crassum, lexlura angulari vel globulosa. Asci cylindrici, 220--270 x 24-32 ~m, 8sporigeri. Ascosporae ellipsoideae vel late ellipsoideae, raro subglobosae, hyalinae, (IS -) 20-32 x (12 -) 16-23 ~m exclusae omamentis, conice verrucosae vel obtuse spinosae, uninucleatae. In terra sub Quercus uariabili. China: Beijing, Tanzhesi, Y. C Huang (B.-C Zhang 529), 16 Nov. 1988 (HMAS, holotypus). Ascomata flattened globose, lobed, 5-9 mm diam, pale brown, hollow, with or without an apical opening, surface with numerous superficial hyphae, verrucose, with basal mycelial tuft; the interior yellowish brown, verrucose. Peridium of two layers, the outer (ectal excipulum) 1OD-150 IJm thick, with cells more or less isodiametric, 10-37'5 IJm diam, walls slightly thickened, forming a textura angularis; the outermost cells outgrowing (simply piling up) to form minute warts, bearing superficial hyphae up to 750 IJm high, 13 IJm diam. near the base, 5 IJm diam. near apex, with walls up to 3 IJm thick, pale brown to brown, smooth; the inner (medullary excipulum) 60-110 IJm thick, yellowish brown, composed of hyphae 2'5-5 IJm diam, nearly arranged in paralleL forming a textura intricata. Hymenium 300 IJm thick excluding epithecium, composed of cylindrical asci and paraphyses arranged in a palisade. Paraphyses hyaline, 300 IJm long, 3-5 IJ diam., the apical cells differentiated to form an epithecium. Epithecium 50-100 IJm thick, composed of cells filamentous but often inflated, 8-15 x 18-30 IJm, hyaline, nearly isodiametric when close to the interior, (24 - ) 28-50 ( - 65) IJm diam., yellowish brown, forming a textura angularis or globulosa. Asci cylindricaL 220-270- X 24-32 IJm, thin-walled with 8 uniseriate spores. Ascospores ellipsoidal to broadly ellipsoidaL occasionally subglobose, hyaline, (15 -) 20-32 x (12 -) 16-22'5 IJm excluding ornamentation, with walls up to 2'5 IJm thick, ornamented with conical warts or blunt spines 1'5-3 IJm high, truncose or obtuse at the apices, sometimes slightly branched, uninucleate. Specimens examined: China: Beijing, Tanzhesi, alt. 350m, hypogeous to subhypogeous under Quercus uariabilis Bl. in a mixed forest, Y. C Huang (B.-C Zhang 529), 16 Nov. 1988 (HMAS 60278, holotype); loc. cit., B.-C Zhang 591, 21 Nov. 1989 (HMAS 60279).
Genea variabilis B.-c. Zhang, sp. nov. Figs 16-24 Etym.: variabilis (L., variable), indicating the great variability in ascomatal morphology, continuity of hymenium, shape of asci and spore size
Ascomata cupulata, applanato globosa vel irregularia, cava, saepe lobata, 4-13 mm diam., luteo-brunneola vel brunneola, aspeetus exteriores et interiores verrucosi. Peridium (excipulum) 220--500 ~m crassum cum verruca, textura angulari; verrucae 100--350 ~m alt., lexlura globulosa. Hymenium continuum, raro interruptum. Epithecium a peridio simile. Asci plerumque cylindrici vel clavati, raro elliptici vel late elliptici, 75-200 (- 240) x 40--{)0 ~m, (3 -) 8-sporigeri. Ascosporae uni- vel biseriatae, raro irregulare dispositae, globosae, brunneolae, 25-34 (- 46) ~m diam exclusae omamentis, spinosae, saepe conjunetae, aspeetu similis brevi crista, uninucleatae. In terra sub Laricis mastersianae. China, Sichuan, Wolong Natural Reserve, B.-C Zhang 621, 2 Sept. 1989 (HMAS, holotypus). Ascomata flattened globose to irregular, simply Peziza-like discoid to globose and hollow with, occasionally without, an apical opening, often lobed, yellowish brown to pale brown, dried brown to dark brown, 4-13 mrn diam., surface verrucose, without basal mycelial tuft; the interior concolorous and verrucose. Peridium (excipulum) 220-500 IJm thick including warts 100-350 IJm high, a single layer, composed of cells globose to polygonaL yellowish brown, (12 -) 20-45 (- 55) IJm diam., with walls 1 IJm thick, forming a textura angularis; the outermost cells outgrowing to multiple globose to subglobose cells, simply piling up to form warts, cells loosely arranged, forming a textura globulosa. Subhymenium conspicuous, about 25 IJm thick, with hyphae 4-12 IJm diam., arranged in parallel or subparallel. Hymenium continuous, sometimes tending to be interrupted by sterile tissue, occasionally interrupted. Paraphyses thin, septate, often anastomosed, the apical cells differentiated to form a thick epithecium. Epithecium nearly the same structure as that of peridium. Asci mostly cylindrical to clavate, occasionally ellipsoidal to broadly ellipsoidaL thin-walled, 75-200 ( - 240) x 40-60 IJm, stalked, with (3 - ) 8 uni- to biseriate or irregularly arranged spores. Ascospores globose, pale brown in lactophenoL reddish brown in Melzer's reagent, 25-34 (- 46) IJm diam. excluding ornamentation, with walls up to 5 IJ thick, ornamented with spines or bracket-shaped ridges, spines up to 7'5 IJm high, apices often contacted to each other or anastomosed to form very short ridges, appearing spinose-reticulate under light microscope, but when viewed
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Bin-Cheng Zhang
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10
Figs 10-15. Genea sinensis (type). Fig. 10. Section of ascoma, showing verrucose peridium, continuous hymenium and epithecium (bar = 100 j..lm). Fig. 11. Superficial hyphae (bar = 100 j..lm). Figs 12, 13. Light micrographs of ascospores with different foci (bar = 20 j..lm). Fig. 14. Scanning electron micrograph of ascospore, showing the ornamentation (bar = 5 j..lm). Fig. 15. Detail of spore ornamentation (SEM, bar = 2 j..lm).
Taxonomy of Genea and Genabea
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Figs 16-20. Genea variabilis (type). Fig. 16. Section of ascoma, showing venucose peridium and epithecium, and continuous hymenium (bar = 200 ~m). Fig. 17. Structure of wart, showing textura globulosa (bar = 20 ~m). Fig. 18. Part of hymenium, showing arrangement of asci (bar = 40 ~m). Fig. 19. Hymenium tending to be intenupted (bar = 40 ~m). Fig. 20. Intenupted hymenia (bar = 40 ~m).
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Bin-Cheng Zhang
Figs 21-24. Genabea variabilis (type) Figs 21-23 Scanning electron micrographs of ascospores, showing spore ornamentation (21, bar = 20 ~m; 22, bar = 10 ~m; 23, bar = 3 ~m). Fig. 24. Uninucleate protoplasts of ascospores (bar = 10 ~m).
under SEM without well-developed reticulations, spines connected to each other, uninucleate.
Specimens examined: China: Sichuan, Wenchuan, Wolong Natural Reserve, near Shanwan, alt. 2200 m, hypogeous in pure Larix mastersiana woods, B.-C. Zhang 621, 22 Sept. 1989 (HMAS 60280, holotype); lac. cit., Heroes' Valley, alt. 2300 m, hypogeous under Betula sp. in a mixed forest with Betula sp. and L. mastersiana, B.-C. Zhang 626, 23 Sept. 1989 (HMAS 60281). As indicated by the specific epithet, this Chinese fungus greatly varies in ascomatal shape, hymenial continuity, ascus 63
shape and ascospore size. G. variabilis possesses many features of both subgenera, and provides a connexion between subgen. Genea and subgen. Genabea. Based on its ascospore colour and ornamentation, it apparently belongs to subgen. Genabea, and is the closest species in the subgenus to the subgen. Genea. In subgen. Genabea, G. variabilis differs from the other species in its pale brown, hollow and verrucose ascomata and ascospores ornamented with spines and ridges. Genabea sphaerospora MaHirolo, an Il:aHan species, is similar to our new species in ascospore morphology. Attempts to borrow Mal:tirolo's material yielded no response. According to MYC 95
Taxonomy of Genea and Genabea Saccardo (1902) and Ceruti (1960) (the latter author had examined Mattirolo's specimens), however, G. sphaerospora has ascomata convoluted, umbrinous; gleba dark brown, separated by sterile tissue into many fertile pockets; asci about 300 Ilm long. All these features are very different from G. variabilis. This study was supported by the National Science Foundation, China. I warmly thank Professor Y. N. Yu, who served as my supervisor and reviewed the manuscript. Mr Dong Guang-Jun and Ms Yun Nan-Cui are acknowledged for their assistance in SEM observation and printing the photographs. I would like to express my appreciation to Dr D. Minter for reviewing the manuscript. REFERENCES Ceruti, A. (1960). Elaphomycetales et Tuberales. In G. Bresadola's lconographia Mycologica 28, 1-48. Eriksson, O. E. & Hawksworth, D. L. (1988). Outline of the ascomycetes1988. Systema Ascomycetum 7, 119-315. Fischer, E. (1938). Tuberineae. In Die Naturlichen Pflanzenfamilien (ed H. G. A. Engler & K. A. E. Prantl), 5b (8), 1-42. Gilkey, H. M. (1939). Tuberales of America. Oregon State College Studies in Botany I, 1-63. Gilkey, H. M. (1954a). Tuberales. North American Flora 2 (1), 1-36.
(Received for publication 4 September 1990)
994 Gilkey, H. M. (1954 b). Taxonomic notes on Tuberales. Mycologia 46, 783-793. Gilkey, H. M. (1961). New species and revisions in the order Tuberales. Mycologia 53, 215-220. Hawker, L. E. (1954). British hypogeous fungi. Philosophical Transactions of the Royal Society of London, series B, Biological Sciences no. 650, 237, 429-546. Hawksworth, D. L., SuHon, B. C. & Ainsworth, G. C (1983). Ainsworth & Bisby's Dictionary of the Fungi, 7th edn. Kew, England: International Mycological Institute. KorL R. P. (1973). Discomycetes and Tuberales. in The Fungi IVA (ed. G. C Ainsworth, F. K. Sparrow & A. S. Sussman), pp. 249-319. Academic Press: London. Minter, D. W. & Zhang, B.-C (1989). An illustrated key to British underground Ascomycetes. The Mycologist 3, 61-67. Pfister, D. H. (1984). Genea-]afneadelphus - a tuberalean-pezizalean connection. Mycologia 76, 170-172. Saccardo, P. (1902). Sylloge Fungorum 16, 808--816. Padova. Tao, K. (1988). Taxonomic study of hypogeous fungi from China. M.5c. Thesis, Shanxi University, Taiyuan. Trappe, J. M. (1975). Generic synonyms in the Tuberales. Mycotaron 2, 109-122. Trappe, J. M. (1979). Orders, families and genera of hypogeous Ascomycotina (truffles and their relatives). Mycotaron 9, 297-340. Tulasne, L. R. & Tulasne, C (1845). Fungi, nonnull, hypogaei, novi v. minus cogniti. Giornale Botanici Halici ser. I, 2, 55-63. Zhang, B.-C (1991). A revised classification of hypogeous families and genera in the Pezizales based on ascospore nuclear number. Mycotaron (in press). Zhou, S. Q. (1989). Larir mastersiana, a rare tree species in area along Ming River, Sichuan. Plant Magazine 5, 10-11.
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Taxonomic status of Genabea, with two new species of Genea (Pezizales)
BIN-CHENG ZHANG* Department of Mycology, Institute of Microbiology, Academia Sinica, Beijing 100080, China
The taxonomic position of Genabea in the Pezizales is discussed. Due to discovery of an intermediate species between Genea and Genabea in China, the modern concepts of these two genera become confused. In this paper, Genabea is reduced to subgeneric rank in the genus Genea, Le. Genea subgen. Genabea (TuL & C. TuL) stat. nov. (basionym: Genabea TuL & C. TuL), typified by Genea fragilis (TuL & C. Tul) comb. nov. (basionym: Genabea fragilis TuL & C. TuL). The species formerly included in Genabea are revised. Three species of Genea found in China are described and illustrated in detail, including Genea sinensis, Genea variabilis which are described as new and Genea fragilis as new to the Chinese mycoflora. A key to the species is provided. In the earlier literature, different authors included different genera in the Geneaceae. Fischer (1938) suggested that the Geneaceae included Petchiomyces Fischer & Matt., Genea Vitt., Myrmecocystis Harkn. and Genabea Tul. & C Tul. This was followed by Hawker (1954) and Gilkey (1954b, 1961). Based on his studies on the type material, Trappe (1975) considered that Myrmecocystis and Genabea were congeneric, and redelimited Genea and Genabea. When the name Geneaceae Trappe was validly published by Trappe (1979), only Genea and Genabea were included. Since then, Trappe's (1979) system has been widely accepted (Hawksworth, Sutton & Ainsworth, 1983; Eriksson & Hawksworth, 1988). On taxonomic status of the Geneaceae, however, Pfister (1984) and Zhang (1991) had different opinions. Recently, I found a few collections referable to both Genea and Genabea. These collections are described as new taxa below, and they apparently challenge the current concepts of Genea and Genabea. This paper deals with this problem, and describes species of Genea found in China. Material used in this study was from K and HMAS. Under light microscopy, the measurements and colours were obtained based on material mounted in lactophenol unless otherwise mentioned. Thin sections for study of peridial structure were made by a freezing microtome. For SEM observations, part of the hymenium of a dried ascoma was rehydrated on a clean glass slide, then stuck onto an adhesive layer on a small clean coverslip, then coated with gold and observed using a Hitachi S570 SEM.
Taxonomic status of Genabea The separation of Genahea from Genea was questioned by Korf (1973). He considered Genea, Genabea and Myrmecocystis to be congeneric. Trappe (1975) synonymized Myrmecocystis with Genabea, but he retained Genabea as a distinct genus, although • Present address: CAB International, Wallingford, Oxon, U.K.
he agreed that. strictly from the standpoint of ascocarp structure, Genea and Genabea may be considered to be congeneric. Trappe (1975) redelimited two genera as follows: Genea: spores verrucose, uniseriate; asci cylindric; hymenia only occasionally interrupted by sterile zones of paraphyses. Genabea: spores echinulate, uniseriate, biseriate or randomly arranged; asci clavate to ellipsoid; hymenia regularly separated into pockets by sterile zones of paraphyses or isodiametric cells. Discovery of Genea variabilis B.-C Zhang sp. nov. described below makes this delimitation very difficult. G. variabilis has coloured ascospores ornamented with spines (a feature of Genabea); asci are mostly cylindrical and contain uniseriate spores (a feature of Genea), but are sometimes clavate to ellipsoidal with biseriate to irregularly arranged spores (a feature of Genabea); hymenium is continuous or tending to be interrupted by sterile mycelial strands (a feature of Genea) (Figs 16-24). Other characters such as ascomatal gross morphology and peridial structure are similarly referable to both genera. If ascospore ornamentation and colour are emphaSized, this species certainly belongs to Genabea sensu Trappe (1975). But it should be remembered that some species in Genea sensu stricto have ascospores bearing blunt spines, e.g. in Genea sinensis B.-C Zhang sp. nov. (Figs 14-15). If continuity of hymenium is emphasized, it should be placed in Genea, but its hymenium tending to be interrupted provides a connexion between the two genera. The morphology of asci and the arrangement of spores within showed no borderline between Genea and Genabea. I thus agree with Korf (1973) to synonymize the two genera, and the name Genea takes priority over Genabea, Genabea species (including G. variabilis), however, have coloured, spinose ascospores, which are different from the species in Genea sensu stricto. For convenience of identification and for reflecting phylogenetic progression within the genus,
Bin-Cheng Zhang I propose that Genabea is reduced to a subgeneric rank in
Genea sensu lato. Genea Vin. subgen. Genea, Monogr. Tuberac.: 27 (1831). Type species: G. verrucosa Vitt. (lectotype, fide Trappe, 1979). Genea Vitt. subgen. Genabea (Tul. & C TuL) B.-C Zhang, comb. nov.
987 other authentic material, I will not make the transfer in the present paper.
Genabea sphaerospora Matt., Malpighia 14: 256 (1900). This is a lesser mown species. Its placement in Genea needs a type study.
Genabea TuL & C TuL, Giorn. Bot. Ital. 1 (2): 60 (1845).
Genabea tasmanica Massee & Rodway, Kew Bull. 1989: 125 (1989), nom. illegit. (fide Hawksworth & Udagawa, 1976).
Type species: Genea fragilis (TuL & C TuL) B.-C Zhang. Species formerly included in Genabea are revised as follows:
Sphaerodes beatonii (D. Hawksw.) P. Cannon & D. Hawksw., Bot. ]. Linn. Soc. (London) 84: 145 (1982).
Myrmecocystis cerebriformis Harm., Proc. Calif. Acad. Sci. 3 (I):
This species was described under Genabea, but was based on the asci and ascospores of Sphaerodes beatonii parasitizing ascomata of Labyrinthomyces species. The type specimen (K) was examined.
269 (1899). Genea cerebriformis (Harm.) Gilkey, Univ. Calif. Publ. Bot 6: 304 (1916).
Species of Genea in China
Genabea cerehriformis (Harm.) Trappe, Mycotaxon 2: 118 (1975).
I have not examined any specimen of this species, but Trappe (1975) considered it to be an acceptable species. For other synonyms, see Trappe (1975).
Genabea spinospora (Gilkey) Trappe, Mycotaxon 2: 118 (1975).
Myrmecocystis spinospora Gilkey, Mycologia 53: 25 (1961). According to Gilkey (1961) and Trappe (1975), this species is a good species. Because I have not seen the type or any
Tao (1988) reported Genea globospora Liu & Tao (nom. ined.) and Genabea spinospora found in China. I examined the specimens deposited in the Mycological Herbarium of Shanxi University (MHSU). Because the specimen identified as Genabea spinospora is not identical to Gilkey's (1961) description and not a member of Genabea, and because G. globospora has not been validly published yet, they will not be discussed further in this paper. Based on my collections, three species are described below; among them two species are described as new.
Key to the species of Genea known to China 1. Ascomata with brown superficial hyphae; ascospores hyaline, warted 1. Ascomata without superficial hyphae; ascospores pale brown to brown, spinose
. G. sinensis 2
2. Ascospores globose; asci and paraphyses hyaline, not fragile 2. Ascospores ellipsoidal; asci and paraphyses brown, fragile.
G. variabi/is . G. fragi/is
Genea fragilis (TuL & C Tul) B.-C Zhang, comb. nov. (Figs 1-9)
tissue a textura prismatica to angularis when close to peridium and epithecium. Paraphyses brown, multi-septate, sometimes constricted at the septa, fragile, often anastomosed to form a textura prismatica surrounding asci, the apical cells differentiated to form an epithecium. Epithecium similar to the peridium in structure but slightly thinner. Asci clavate to ellipsoidal, 85-150 x 40-60 !-lm, brown, thin-walled, fragile, with 73-6 uniseriate to irregularly arranged spores (because asci are fragile, it is so difficult to keep asci intact while sectioning that it is not possible to count accurately the number of spores), irregularly arranged among paraphyses. Ascospores ellipsoidal to broadly ellipsoidal, pale brown to brown, 23-30 x 19-21!-lm excluding ornamentation, ornamented with walls about I !-lm thick, with dense spines up to 3-5 !-lm high, thin, often attached with fragments of asci and some deposits.
Genabea fragi/is TuL & C TuL, Giorn. Bot. Ital. 1 (2): 60 (1845).
Ascomata irregular, nodose, lobed, 4-6 mm diam., greyish brown, dried brown, hollow, without conspicuous apical hole, surface warted to roughened; the interior slightly paler, warted to roughened; dark brown on the cut surface, separated by pale sterile tissue visible under dissecting microscope. Peridium 150-250 !-lm thick including warts up to 70 !-lm high, of 2 layers; the outer (ectal excipulum) composed of cells globose to subglobose, pale brown to yellowish brown inward, 17-40 ( - 50) !-lm diam., thick-walled, forming a textura globulosa; some outermost cells irregularly outgrowing to multiple globose to subglobose cells simply piling up to form primitive warts; the inner (medullary excipulum) subhyaline, composed of cells more or less isodiametric, 7-17!-lm diam., with walls l!-lm thick, forming a textura angularis, sometimes forming a textura intricata when close to the hymenium. Hymenium composed of irregularly arranged asci and paraphyses, interrupted and separated by sterile tissue into many fertile pockets (discontinuous hymenia); sterile
Specimen examined: China: Sichuan, Wenchuan, Wolong Natural Reserve, nr Shanwan, alt. 2250 m, hypogeous in Larix mastersiana Hedh. & Wils. woods, B.-C. Zhang 645, 22 Sept. 1989 (HMAS 60282). Genea fragi/is is a new record to the Chinese mycoflora, and is the first report in Asia. The Chinese collection generally agrees with the' authentic' specimen in K which I examined,
Taxonomy of Genea and Genabea
Figs 1-5. Genea fragilis (B. C. Zhang 645). Fig. 1. Section of ascoma, showing interrupted hymenia, peridium and epithecium (bar = 200 I.lm). Fig. 2. Peridial structure (bar = 50 I.lm). Fig. 3. Epithecium (bar = 50 I.lm). Fig. 4. Fragile sterile tissue separating hymenia (bar = 20 I.lm). Fig. 5. Fragile, coloured asci and paraphyses (bar = 20 I.lm).
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Figs 6-9. Genea fragilis (B.-C. Zhang 645). Fig. 6. Light micrograph of ascospores (bar = 20 ~m). Figs 7-8. Scanning electron micrographs of ascospores, showing the ornamentation, note ascus fragments on the surface (bar = 5 ~m). Fig. 9. Detail of spore ornamentation (SEM, bar = 1 ~m).
especially in fragility of asci and paraphyses, and ascospore morphology (Figs 5-9), but somewhat different is ascomatal colour and ascospore size. The authentic specimen and collections from Switzerland and Canada are black or sooty (Tulasne & Tulasne, 1845; Gilkey, 1954a), but our material is greyish brown. The ascospores from the authentic specimen
are 27-38 x 20-27 !-lm excluding ornamentation, larger than those of our material. Because colour changes of ascomata often occur at different developmental stages and with drying conditions, and because the spore size is often variable, it is suitable to assign our collection to G. fragilis. Genea fragilis differs from other species in subgen. Genabea
990
Taxonomy of Genea and Genabea in its fragile, coloured asci and paraphyses, and ellipsoidal ascospores (Figs 5-6). Coloured asci and paraphyses are very peculiar in the hypogeous Ascomycetes. In our materiaL the colour of asci and paraphyses is obviously much darker than that of sterile tissue and peridium, and nearly the same as that of ascospores. This feature is doubtless very useful in species separation. Genea fragilis was found only in association with Larix mastersiana in China and shares the same habitat with Genea variabilis. L. mastersiana is an indigenous species in China, and Sichuan is the main distribution area (Zhou, 1989). The association of G. fragilis with L. mastersiana is worth further study. Genea sinensis B.-C. Zhang, sp. nov.
From ascomatal gross morphology and ascospore morphology, G. sinensis apparently belongs to the G. hispidula Berk. group, including G. hispidula, G. arenaria Harkn., G. brachytheca Gilkey and G. echinospora Gilkey. G. sinensis is closely related to G. hispidula, but the former differs from the latter in its superficial hairs much shorter and sparser, epithecium thinner and without hairs, and ascospores smaller and with conical warts to blunt spines (Figs 14-15; see Hawker, 1954; Minter & Zhang, 1989). G. sinensis is also similar to G. brachytheca in gross morphology, but the latter has much shorter asci with clustered ascospores ornamented with papillae (Gilkey, 1954). All these features are different from our new species. G. arenaria and G. echinospora are obviously different from G. sinensis (Gilkey, 1939, 1954a).
(Figs 10-15)
Ascomala applanato globosa, cava, lobata, 5-9 mm diam., brunneola, basi caespitibus mycelialibus; aspeetus exteriores sparse hirsuti, verrucosi, hyphae superficiales usque ad 750 ~m longae, brunneolae vel brunneae; aspeetus interiores luteoli, verrucosi. Peridium exterius 100-- ISO ~m crassum, lexlura angulari; peridium interius 60- IIO ~m crassum, lexlura inlricala. Epilhecium 50--100 ~m crassum, lexlura angulari vel globulosa. Asci cylindrici, 220--270 x 24-32 ~m, 8sporigeri. Ascosporae ellipsoideae vel late ellipsoideae, raro subglobosae, hyalinae, (IS -) 20-32 x (12 -) 16-23 ~m exclusae omamentis, conice verrucosae vel obtuse spinosae, uninucleatae. In terra sub Quercus uariabili. China: Beijing, Tanzhesi, Y. C Huang (B.-C Zhang 529), 16 Nov. 1988 (HMAS, holotypus). Ascomata flattened globose, lobed, 5-9 mm diam, pale brown, hollow, with or without an apical opening, surface with numerous superficial hyphae, verrucose, with basal mycelial tuft; the interior yellowish brown, verrucose. Peridium of two layers, the outer (ectal excipulum) 1OD-150 IJm thick, with cells more or less isodiametric, 10-37'5 IJm diam, walls slightly thickened, forming a textura angularis; the outermost cells outgrowing (simply piling up) to form minute warts, bearing superficial hyphae up to 750 IJm high, 13 IJm diam. near the base, 5 IJm diam. near apex, with walls up to 3 IJm thick, pale brown to brown, smooth; the inner (medullary excipulum) 60-110 IJm thick, yellowish brown, composed of hyphae 2'5-5 IJm diam, nearly arranged in paralleL forming a textura intricata. Hymenium 300 IJm thick excluding epithecium, composed of cylindrical asci and paraphyses arranged in a palisade. Paraphyses hyaline, 300 IJm long, 3-5 IJ diam., the apical cells differentiated to form an epithecium. Epithecium 50-100 IJm thick, composed of cells filamentous but often inflated, 8-15 x 18-30 IJm, hyaline, nearly isodiametric when close to the interior, (24 - ) 28-50 ( - 65) IJm diam., yellowish brown, forming a textura angularis or globulosa. Asci cylindricaL 220-270- X 24-32 IJm, thin-walled with 8 uniseriate spores. Ascospores ellipsoidal to broadly ellipsoidaL occasionally subglobose, hyaline, (15 -) 20-32 x (12 -) 16-22'5 IJm excluding ornamentation, with walls up to 2'5 IJm thick, ornamented with conical warts or blunt spines 1'5-3 IJm high, truncose or obtuse at the apices, sometimes slightly branched, uninucleate. Specimens examined: China: Beijing, Tanzhesi, alt. 350m, hypogeous to subhypogeous under Quercus uariabilis Bl. in a mixed forest, Y. C Huang (B.-C Zhang 529), 16 Nov. 1988 (HMAS 60278, holotype); loc. cit., B.-C Zhang 591, 21 Nov. 1989 (HMAS 60279).
Genea variabilis B.-c. Zhang, sp. nov. Figs 16-24 Etym.: variabilis (L., variable), indicating the great variability in ascomatal morphology, continuity of hymenium, shape of asci and spore size
Ascomata cupulata, applanato globosa vel irregularia, cava, saepe lobata, 4-13 mm diam., luteo-brunneola vel brunneola, aspeetus exteriores et interiores verrucosi. Peridium (excipulum) 220--500 ~m crassum cum verruca, textura angulari; verrucae 100--350 ~m alt., lexlura globulosa. Hymenium continuum, raro interruptum. Epithecium a peridio simile. Asci plerumque cylindrici vel clavati, raro elliptici vel late elliptici, 75-200 (- 240) x 40--{)0 ~m, (3 -) 8-sporigeri. Ascosporae uni- vel biseriatae, raro irregulare dispositae, globosae, brunneolae, 25-34 (- 46) ~m diam exclusae omamentis, spinosae, saepe conjunetae, aspeetu similis brevi crista, uninucleatae. In terra sub Laricis mastersianae. China, Sichuan, Wolong Natural Reserve, B.-C Zhang 621, 2 Sept. 1989 (HMAS, holotypus). Ascomata flattened globose to irregular, simply Peziza-like discoid to globose and hollow with, occasionally without, an apical opening, often lobed, yellowish brown to pale brown, dried brown to dark brown, 4-13 mrn diam., surface verrucose, without basal mycelial tuft; the interior concolorous and verrucose. Peridium (excipulum) 220-500 IJm thick including warts 100-350 IJm high, a single layer, composed of cells globose to polygonaL yellowish brown, (12 -) 20-45 (- 55) IJm diam., with walls 1 IJm thick, forming a textura angularis; the outermost cells outgrowing to multiple globose to subglobose cells, simply piling up to form warts, cells loosely arranged, forming a textura globulosa. Subhymenium conspicuous, about 25 IJm thick, with hyphae 4-12 IJm diam., arranged in parallel or subparallel. Hymenium continuous, sometimes tending to be interrupted by sterile tissue, occasionally interrupted. Paraphyses thin, septate, often anastomosed, the apical cells differentiated to form a thick epithecium. Epithecium nearly the same structure as that of peridium. Asci mostly cylindrical to clavate, occasionally ellipsoidal to broadly ellipsoidaL thin-walled, 75-200 ( - 240) x 40-60 IJm, stalked, with (3 - ) 8 uni- to biseriate or irregularly arranged spores. Ascospores globose, pale brown in lactophenoL reddish brown in Melzer's reagent, 25-34 (- 46) IJm diam. excluding ornamentation, with walls up to 5 IJ thick, ornamented with spines or bracket-shaped ridges, spines up to 7'5 IJm high, apices often contacted to each other or anastomosed to form very short ridges, appearing spinose-reticulate under light microscope, but when viewed
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Bin-Cheng Zhang
12
10
Figs 10-15. Genea sinensis (type). Fig. 10. Section of ascoma, showing verrucose peridium, continuous hymenium and epithecium (bar = 100 j..lm). Fig. 11. Superficial hyphae (bar = 100 j..lm). Figs 12, 13. Light micrographs of ascospores with different foci (bar = 20 j..lm). Fig. 14. Scanning electron micrograph of ascospore, showing the ornamentation (bar = 5 j..lm). Fig. 15. Detail of spore ornamentation (SEM, bar = 2 j..lm).
Taxonomy of Genea and Genabea
992
Figs 16-20. Genea variabilis (type). Fig. 16. Section of ascoma, showing venucose peridium and epithecium, and continuous hymenium (bar = 200 ~m). Fig. 17. Structure of wart, showing textura globulosa (bar = 20 ~m). Fig. 18. Part of hymenium, showing arrangement of asci (bar = 40 ~m). Fig. 19. Hymenium tending to be intenupted (bar = 40 ~m). Fig. 20. Intenupted hymenia (bar = 40 ~m).
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Bin-Cheng Zhang
Figs 21-24. Genabea variabilis (type) Figs 21-23 Scanning electron micrographs of ascospores, showing spore ornamentation (21, bar = 20 ~m; 22, bar = 10 ~m; 23, bar = 3 ~m). Fig. 24. Uninucleate protoplasts of ascospores (bar = 10 ~m).
under SEM without well-developed reticulations, spines connected to each other, uninucleate.
Specimens examined: China: Sichuan, Wenchuan, Wolong Natural Reserve, near Shanwan, alt. 2200 m, hypogeous in pure Larix mastersiana woods, B.-C. Zhang 621, 22 Sept. 1989 (HMAS 60280, holotype); lac. cit., Heroes' Valley, alt. 2300 m, hypogeous under Betula sp. in a mixed forest with Betula sp. and L. mastersiana, B.-C. Zhang 626, 23 Sept. 1989 (HMAS 60281). As indicated by the specific epithet, this Chinese fungus greatly varies in ascomatal shape, hymenial continuity, ascus 63
shape and ascospore size. G. variabilis possesses many features of both subgenera, and provides a connexion between subgen. Genea and subgen. Genabea. Based on its ascospore colour and ornamentation, it apparently belongs to subgen. Genabea, and is the closest species in the subgenus to the subgen. Genea. In subgen. Genabea, G. variabilis differs from the other species in its pale brown, hollow and verrucose ascomata and ascospores ornamented with spines and ridges. Genabea sphaerospora MaHirolo, an Il:aHan species, is similar to our new species in ascospore morphology. Attempts to borrow Mal:tirolo's material yielded no response. According to MYC 95
Taxonomy of Genea and Genabea Saccardo (1902) and Ceruti (1960) (the latter author had examined Mattirolo's specimens), however, G. sphaerospora has ascomata convoluted, umbrinous; gleba dark brown, separated by sterile tissue into many fertile pockets; asci about 300 Ilm long. All these features are very different from G. variabilis. This study was supported by the National Science Foundation, China. I warmly thank Professor Y. N. Yu, who served as my supervisor and reviewed the manuscript. Mr Dong Guang-Jun and Ms Yun Nan-Cui are acknowledged for their assistance in SEM observation and printing the photographs. I would like to express my appreciation to Dr D. Minter for reviewing the manuscript. REFERENCES Ceruti, A. (1960). Elaphomycetales et Tuberales. In G. Bresadola's lconographia Mycologica 28, 1-48. Eriksson, O. E. & Hawksworth, D. L. (1988). Outline of the ascomycetes1988. Systema Ascomycetum 7, 119-315. Fischer, E. (1938). Tuberineae. In Die Naturlichen Pflanzenfamilien (ed H. G. A. Engler & K. A. E. Prantl), 5b (8), 1-42. Gilkey, H. M. (1939). Tuberales of America. Oregon State College Studies in Botany I, 1-63. Gilkey, H. M. (1954a). Tuberales. North American Flora 2 (1), 1-36.
(Received for publication 4 September 1990)
994 Gilkey, H. M. (1954 b). Taxonomic notes on Tuberales. Mycologia 46, 783-793. Gilkey, H. M. (1961). New species and revisions in the order Tuberales. Mycologia 53, 215-220. Hawker, L. E. (1954). British hypogeous fungi. Philosophical Transactions of the Royal Society of London, series B, Biological Sciences no. 650, 237, 429-546. Hawksworth, D. L., SuHon, B. C. & Ainsworth, G. C (1983). Ainsworth & Bisby's Dictionary of the Fungi, 7th edn. Kew, England: International Mycological Institute. KorL R. P. (1973). Discomycetes and Tuberales. in The Fungi IVA (ed. G. C Ainsworth, F. K. Sparrow & A. S. Sussman), pp. 249-319. Academic Press: London. Minter, D. W. & Zhang, B.-C (1989). An illustrated key to British underground Ascomycetes. The Mycologist 3, 61-67. Pfister, D. H. (1984). Genea-]afneadelphus - a tuberalean-pezizalean connection. Mycologia 76, 170-172. Saccardo, P. (1902). Sylloge Fungorum 16, 808--816. Padova. Tao, K. (1988). Taxonomic study of hypogeous fungi from China. M.5c. Thesis, Shanxi University, Taiyuan. Trappe, J. M. (1975). Generic synonyms in the Tuberales. Mycotaron 2, 109-122. Trappe, J. M. (1979). Orders, families and genera of hypogeous Ascomycotina (truffles and their relatives). Mycotaron 9, 297-340. Tulasne, L. R. & Tulasne, C (1845). Fungi, nonnull, hypogaei, novi v. minus cogniti. Giornale Botanici Halici ser. I, 2, 55-63. Zhang, B.-C (1991). A revised classification of hypogeous families and genera in the Pezizales based on ascospore nuclear number. Mycotaron (in press). Zhou, S. Q. (1989). Larir mastersiana, a rare tree species in area along Ming River, Sichuan. Plant Magazine 5, 10-11.