Folia Biologica et Oecologica 10: 130–138 (2014) Acta Universitatis Lodziensis

Gasteroid fungi – the morphological characteristics of selected endangered and rare noted in Poland

JANUSZ ŁUSZCZYŃSKI* & AGNIESZKA TOMASZEWSKA** Department of Botany, Institute of Biology, Jan Kochanowski University, Świętokrzyska 15, 25-406 Kielce, Poland E-mail: * [email protected]; **[email protected]

ABSTRACT

The aim of the work was to present the characteristics of selected species from Disciseda, and genera which due to the small differences in morphology of their fruit bodies may pose some identification problems. The selected species of of these genera are described based on the materials collected during the course of our studies. All materials were gathered during the research into macromycetes in xerothermic habitats located in the Nida Basin. Taxa noted by us are considered to be very rare in the mycobiota of Poland and are highly endangered.

KEY WORDS: Gasteromycetes, thermophilic fungi, endangered species of fungi, , Geastraceae, Lycoperdaceae

Introduction The gasteroid fungi (formerly pseudoparenchyma; Pilát 1958, Gasteromycetes) are polyphyletic group Rudnicka-Jezierska 1991). of fungi which currently belong to Fruit bodies of gasteroid fungi different taxa in the class develop underground (hypogeously) and (Hibbett & Thorn 2001, Binder & are spherical. As they mature they Bresinsky 2002). Biological and emerge over ground becoming epigeous. morphological properties as well as the Gasteroid fungi are mainly saprobionts development, maturation and dispersal of that grow in woodless areas, xerothermic, are characteristic features of the sandy and steppe sites, in forests, but also Gasteromycetes. Gasteroid fungi are in wet places and even on the moors usually spherical, piriform and clavate. (Pilát 1958, Rudnicka-Jezierska 1991). The hymenium is enclosed inside the Species that differ by small macro- fruit body until spores mature. Fruit and micromorphological characters of the bodies consist of three basic parts: fruit body’s structure in individual genera peridium (the wall), (the fertile were selected for morphological analysis. area) and trama (sterile hyphae that form A scrupulous submicroscopic and molecular examination of the structure of DOI: 10.2478/fobio-2014-0005 FOLIA BIOLOGICA ET OECOLOGICA similar taxa helps to identify fruit bodies from Disciseda, Geastrum and correctly and reduces the risk of Tulostoma genera, which due to the small determination errors (e.g. Tomaszewska differences in morphology of their fruit et al. 2011). bodies may pose some identification The aim of this study was to present problems. the characteristics of selected species

Material and methods Species were collected during the size and colour of the endoperidium, the research into macromycete fungi in colour of the exoperidium and the xerothermic habitats located in the Nida manner in which it flakes, and also the Basin (south of Poland) between 1991 pigmentation and the structure of the and 2013. The studies were intensified stem surface, were all noted during from 2010 until 2013. The investigations collections. were conducted in protected areas such The laboratory examinations were as nature reserves (Krzyżanowice, conducted using light microscopy (LM) Skorocice), landscape parks and Natura and scanning electron microscopy 2000 sites (Nida Landscape Park – (SEM). The structure, size and shape of PLH260003 Ostoja Nidziańska, Szaniec the capillitium and the spores were Landscape Park – PLH260034 Ostoja measured using standard reagents and Szaniecko-Solecka, Kozubów Landscape light microscope. The measurements Park – PLH260029 Ostoja Kozubowska). were performed using 400x and 1000x The examined plant communities are magnification. The episporium sculpture protected under the Habitats Directive. was investigated using SEM. The investigations conducted in these The material (gleba samples with respected areas also provided data about spores) was mounted on an aluminium their functioning and the interactions stub and coated with 24-carat gold between fungi and xerothermic (Karcz 1996, 2009). The electron vegetation. micrographs were taken at the The mycological investigations were magnifications of: 3000, 5000, 10000 conducted using permanent research and 12000x. The studies with the help of plots and were supplemented with the the scanning electron microscope (SEM) route method. A total of 30 plots (each of were carried out in the Department of them of 100 m2) were established in six Environment Protection of Jan communities of xerothermic vegetation Kochanowski University in Kielce and in from Festuco-Brometea class, such as: the Laboratory of Field Emission, Adonido-Brachypodietum pinnati, Scanning Electron Microscopy and Festucetum pallentis, Inuletum Microanalysis at the Institute of ensifoliae, Sisymbrio-Stipetum capillatae, Geological Sciences of Jagiellonian Seslerio-Scorzoneretum purpureae and University in Kraków. Thalictro-Salvietum pratensis (names The following studies were used for according to Matuszkiewicz 2012). The taxonomic identification: Pilát (1958), observations and collections of fruit Wright (1987), Rudnicka-Jezierska bodies were carried out at intervals. The (1991), Sarasini (2005) and Sunhede number of species fruit bodies, the (1990). The nomenclature of the taxa is organoleptic properties, i.e. the shape, given after (2014).

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Results Eight selected species of gasteroid farinaceous surface of the endoperidium fungi were examined. Macro- and is a characteristic feature of the species. micromorphological characters were Peristome cristate with delimited bulge, used to describe the species. Species from 12 to 20 ridges (Fig. 2a). Gleba descriptions are based on material dark brown. Spores globose, finely collected in our investigations. A list of verrucose, 4.8–7(–8) µm. It should be similarities and differences between the noted that there is a possibility of described species is given in Tables 1, 2 mistaking this species with Geastrum and 3. berkeleyi Massee which also has Disciseda (Klotzsch) Henn. granulately surface of the endoperidium Mature fruit bodies globose, rarely and cristate peristome. flattened. Exoperidium white, whitish Schwein. yellow, mature brownish ashen. Exoperidium splits into 6 to 12 not Endoperidium is rigid, pergameneous hygroscopic segments that reach 3 to 4 and nut-like coloured. Spores globose, cm in diam. when expanded. (5–)6.5–7.8(–8.6) µm in diam. Endoperidium globose is 0.4 to 1.2 cm in (according to Lizárraga et al. 2010: 4–7 diam., grey-brown, ochraceous-brown or µm in diam.), distinctly strongly grey-white. A white layer of fine crystals verrucose, verrucae 1–1.5 µm, without of calcium oxalate on the endoperidium sterigmata (Fig. 1a). Capillitium is light is the main diagnostic trait of the species. yellow, hyaline, quite thick-walled. Peristome sericeous-fimbriate is lighter Capillitium threads are wavy, brittle and than the remainder of theendoperidium, 2.7–3.5 µm thick. with a collar delimited by a bulge (Fig. Disciseda candida (Schwein.) Lloyd 2b). Gleba dark brown. Spores globose, Mature fruit bodies are loaf-like. 3.5–5.5(–7) µm in diam., minutely Exoperidium is dirty whitish yellowish, verrucose. mature, earth brown. Endoperidium Geastrum schmidelii Vittad. strong, leathery, matt, brown-grey to Exoperidium splits into 5 to 10 ashen in colour. Spores globose, entirely not hygroscopic segments that punctate, delicately verrucose or reach 1-3 cm in diam. when expanded. glabrous, (3.8–)4.5–5 µm (according to Endoperidium globose is brown-grey to Bates et al. 2009: 4.0–5.6(–6.4) × 4.0– brown at the bottom, whitish at the top, 5.6(–6.4) µm in diam.), without especially in young fruit bodies. sterigmata (Fig. 1b). Capillitium light Peristome sulcate, from 10 to 19 ridges, yellow, hyaline and thin-walled. delimited by a furrow, covered with Capillitium threads wavy, brittle and 2.5 farinose coating in young fruit bodies µm thick. (Fig. 2c). Gleba dark brown. Spores Morgan globose, 4.7–7.5 µm (according to Exoperidium splits into 5 to 12 Sarasini 2005: (4–)4.2–4.8(–5.5) µm in triangular segments that are hygroscopic diam.), distinctly thickly verrucose. or subhygroscopic. It is beige-coloured, Tulostoma brumale Pers. grey-brown, light brown to dark brown Exoperidium whitish, membranous, and when expanded 3 to 5 cm in diam. on soon flaking away. Mature endoperidium average. Endoperidium globose is 0.5 to ochraceous-white, sometimes with rusty- 2 cm in diam. and only partially with brown stains, also yellowish brown. apophysis. The granulately rough, Peristome tubular, mouth area darker

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FOLIA BIOLOGICA ET OECOLOGICA coloured, yellowish or dirty brown. Stem 1997: 4.5–5 µm, without the fibrillose, ochraceous-fawn, minutely and ornamentation). Capillitium subhyaline, very delicately squamulose, straight (Fig. thick-walled, well visible lumen, 3a). Spores (4.2–)4.7–5.64 µm in diam. colourless and not thickened at septa, (according to Sarasini 2005: (3.5–)4.2– moderately branched, without crystals. 4.7(–5.5) µm in diam.), globose, light Tulostoma squamosum Gmelin yellow, minutely verrucose (Fig. 3b). Exoperidium dark, sometimes Capillitium hyaline, thick-walled, lumen whitish, thin-walled, persisting longer. small, coloured at septa and swollen in Endoperidium pergameneously rigid, septa, branched, external surface covered light yellow, white-ochraceous, with fine crystals. becoming chestnut-coloured. Peristome Tulostoma melanocyclum Bres. tubular, mouth area concolourous with Exoperidium whitish or light the head, stem cinnamon-coloured or ochraceous, hyphal, persisting quite long. brown-red, unevenly covered with Endoperidium ashen-ochraceous or protruding, sharp squamulae (Fig. 3e). ochraceous-rusty. Peristome tubular, Spores globose and subglobose, yellow- dark, dirty brown. Stem ochraceous- brown, small spiny, (4.5–)5.64–7.52 µm brown to dark brown, sulcate, covered (Fig. 3f). Capillitium hyaline, thick- with fine, adherent squamulae (Fig. 3c). walled, lumen visible, slightly yellowish Spores are globose and subglobose (Fig. and often broader at septa, branched, 3d), brown, spinulose, (6.58–)7.52–8.46 without crystals. µm (according to Hansen & Knudsen

Table 1. The comparison of selected morphological and anatomical characteristics in similar Disciseda species.

Feature Disciseda candida Dirty whitish yellowish, mature White, whitish yellow, mature Exoperidium from brownish to earthy brown pale grey coloured Rigid, pergameneous, hazel nut Strong, leathery, Endoperidium like coloured pergameneous, brown-grey Peristome Frayed Frayed, frimbrillate Gleba Red brown Brightly brown, rusty brown (5–)6.5–7.8(–8.6) µm in diam. (3.8–)4.5–5 µm in diam. Spores Distinctly strongly verrucose Smooth or puncticulate, very (verrucae 1-1.5 µm) fine verrucose Sterigmata Absent Absent 2.7–3.5 µm thick, 2.5 µm thick, Capillitium wavy, fragile wavy, fragile

Discussion The new information regarding many and for this reason it can be considered rare and endangered species of gasteroid for more frequent. Other species are very fungi in Poland was collected in our rare and are considered as endangered study. Only one species, Tulostoma (E) in Polish Red List (Wojewoda & brumale has a larger number of localities, Ławrynowicz 2006).

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The aim of the study was to draw the our research has shown the need to attention to the selected features of fungi conduct parallel complementary studies fruit bodies from the Disciseda, with the use of LM and SEM to carry out Geastrum and Tulostoma genera which the correct determination of these fungi. due to their variability are difficult to The climatic and habitat factors can also identify. Full descriptions of the taxa are impact phenotypic characters based on macro- and micromorphological (Tomaszewska et al. 2012, 2014). The characters recorded in field studies and identification of taxa based only on laboratory examinations. The important observations of features by using the LM differences between fruit bodies within may contribute to erroneous genera and species may be noticeable but determinations. Further complex are strongly influenced by the sample investigations into gasteroid fungi are size being collected and how well the needed in to verify fully the morphology is being preserved. They morphological structure and also to also become less distinct as fruit bodies identify taxa correctly. mature and become old. Consequently,

Table 2. The comparison of selected morphological and anatomical characteristics in similar Geastrum species.

Geastrum Feature Geastrum minimum Geastrum schmidelii campestre Splits into 5–12 Splits into 6–12 Splits into 5–10 triangular segments; segments; segments; not hygroscopic; not hygroscopic; hygroscopic or Exoperidium subhygroscopic; Grey brown, Grey white to brown Beige-coloured, ochraceous brown or grey-brown, light grey white brown to dark brown Globose, 0.5–2 cm Globose, 0.4–1.2 cm Globose, 0.3–2.5 cm in diam; in diam; in diam; Endoperidium Presence of crystals of Smooth surface Granulately rough, calcium oxalate on the farinaceous surface surface Cristate, with Sericeous-fimbriate, Sulcate, delimited by delimited bulge, lighter than the a furrow, covered 12–20 ridges remainder of the with farinose coating Peristome endoperidium, with a in young fruit bodies, collar delimited by a 10–19 ridges bulge Gleba Dark brown Dark brown Dark brown 4.8–7(–8) µm in 3.5–5.5(–7) µm in 4.7–7.5 µm diam, diam, diam, globose, distinctly Spores globose, finely globose, minutely thickly verrucose verrucose verrucose

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Table 3. The comparison of selected morphological and anatomical characteristics in similar Tulostoma species.

Tulostoma Tulostoma Feature Tulostoma brumale melanocyclum squamosum Dark, Whitish or pale sometimes Whitish, membranous, Exoperidium ochraceous, relatively whitish, thinly early flaking long-term membranous, more durable White ochraceous, Grey and ochraceous, White Endoperidium yellowish brown cinereous ochraceous ochraceous Tubular, around the Tubular, around mouth dark coloured – Tubular, dark, dirty the mouth Peristome yellowish or dirty brown colored as the brown peridium Pale Pale ochraceous, Gleba Pale ochraceous ochraceous, ferrugineous ferrugineous 23–33 × 2–3 mm, 18–24(–28) × 16–29 × 2–2.8 mm; ochraceous brown to 2.5–4.5 mm; fibrous, pale dark brown, furrowed, cinereous or Stem ochraceous, covered covered with brown red, with tiny and very uniformly delicate scales covered with coarse (4.2–)4.7–5.64 µm in (6.58–)7.52-–8.46 µm in (4,5–)5.64–7.52 diam; diam; µm in diam; globose, pale globose, brown, delicate globose or Spores ochraceous, with small spiny subglobose, verrucae yellow brown, small, spiny Width of capillitium: Width of (4.7–)5.64–6.58 µm, capillitium: Width of capillitium: width of septa: 5.64 µm. 4.7–6.58 µm, 3.7–5.64 µm, width of septa: width of septa: (6.11–) Almost hyaline, thick- 4.7–5.64 µm. 8.64–9.4 µm. walled with a visible Hyaline, thick- Hyaline, thick-walled lumen, not swollen at walled with a Capillitium with small lumen, the uncoloured septa, visible lumen, swollen at the coloured branched, without slightly swollen septa, branched, crystalline plaques at the yellowish external surface septa, branched, covered with without crystalline plaques crystalline plaques

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Figure 1. Spores of Disciseda species in SEM: A – D. bovista, B – D. candida.

Figure 2. The morphology of the peristome and the structure of the endoperidium surface in Geastrum species (photo by A. Tomaszewska): A–G. campestre, B–G. minimum, C – G. schmidelii. Scale bars = 5 mm.

Figure 3. The morphology of the fruit body stem and the texture (in SEM) of Tulostoma species: A, B – T. brumale; C, D – T. melanocyclum; E, F – T. squamosum. Scale bars: A, C, E = 3 mm, B = 3 µm, D = 5 µm, F = 1µm, (photos A, C, E by G. Wołczyk).

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Acknowledgements The authors would like to thank the Laboratory of Field Emission Scanning Electron Microscopy and Microanalysis at the Institute of Geological Sciences of Jagiellonian University in Kraków (Poland) for making the scanning electron microscope (JEOL JSM 5410, JEOL, Tokio, Japan; purchased with the help of the Foundation of Polish Science, Subin 94 programme) available and thus making microphotography possible.

References Bates, S.T., Roberson, R.W. & Desjardin, D.E. Rudnicka-Jezierska, W. 1991. Purchawkowate 2009. Arizona gasteroid fungi I: (Lycoperdales), Tęgoskórowe Lycoperdaceae (, ). (Sclerodermatales), Pałeczkowate Fungal Diversity, 37: 153-207. (Tulostomatales), Gniazdnicowate Binder, M. & Bresinsky, A. 2002. Derivation of a (Nidulariales), Sromotnikowate (Phallales), polymorphic lineage of gasteromycetes from Osiakowe (Podaxales). In: Skirgiełło A., (ed.), boletoid ancestors. Mycologia, 94:85–98. Grzyby (Mycota) 23. Wydawnictwo Naukowe, Hansen, L. & Knudsen, H. 1997. Nordic PWN, Kraków. Macromycetes vol. 3. Heterobasidioid, Sarasini, M. 2005. Gasteromiceti epigei, Aphyllophoroid and Gasteromycetoid Associazione Micologica Bresadola, Trento. Basidiomycetes. Nordsvamp, Copenhagen. Sunhede, S. 1990. Geastraceae (Basidiomycotina). Hibbett, D.S., Thorn, R.G. 2001. Morphology, ecology and systematics with Homobasidiomycetes. The Mycota 7 Part B.. special emphasis on the north European In: McLaughlin D. J., McLaughlin E. G., species. Syn. Fung. 1. Fungiflora, Oslo. Lemke P. A. (eds.), Systematics and evolution. Tomaszewska, A., Łuszczyński, J. & Lechowicz, Ł. Springer-Verlag, Berlin, pp. 121–170. 2014. Macrofungi (Basidiomycetes) as Index Fungorum 2014. bioindicators of communities of xerothermic [http://www.indexfungorum.org/names/names. vegetation. Acta Mycol. (submitted). asp – accessed 17.06.2014]. Tomaszewska, A., Łuszczyński, J., Łuszczyńska, B. Karcz, J. 1996. Skaningowy mikroskop & Jaworska, J. 2011. Taxonomic notes on the elektronowy w badaniach karpologicznych. Polish Tulostoma species. Acta Mycologica, Wiadomości Botaniczne 40(3/4): 55–65. 46(2): 179–186. Karcz, J. 2009. Mikroskopia elektronowa Tomaszewska, A., Łuszczyński, J., Łuszczyńska, B. skaningowa w biologii. Pracownia Mikroskopii & Jaworska, J. 2012. The contribution and Elektronowej Skaningowej. Uniwersytet Śląski importance of rare macrofungi in selected plots Wydział Biologii i Ochrony Środowiska. of endangered xerothermic grasslands in the Katowice. Nida Basin. Acta Agrobotanica, 65(2): 147– Lizárraga M., Esqueda M., Gutiérrez A., Piña C. & 154. Barredo-Pool F. 2010. El género Disciseda Wojewoda, W. & Ławrynowicz M. 2006. Red List (Agaricales, Agaricaceae) en la Planicie of the macrofungi in Poland. In: Mirek Z., Central del Desierto Chihuahuense, México. Zarzycki K., Wojewoda W. & Szeląg Z. (eds), Revista Mexicana de Micologia, 32: 41–47. Red list of plants and fungi in Poland. W. Matuszkiewicz W. 2012. Przewodnik do Szafer Institute of Botany, Polish Academy of oznaczania zbiorowisk roślinnych Polski. Sciences, Kraków, pp. 53–70. Wydawnictwo Naukowe PWN, Warszawa. Wright, J.E. 1987. The Tulostoma Pilát, A. 1958. Gasteromycetes houby – břichatky. (Gasteromycetes) – a world monograph. Bibl. In: Novak F. A. (ed.), Flora ČSR, B. 1. Nakl. Mycol. 113, J. Cramer, Berlin, Stuttgart. Československé Akademie Věd, Praha.

Streszczenie W pracy opisano osiem wybranych gatunków grzybów gasteroidalnych należących do rodzajów: Disciseda, Geastrum oraz Tulostoma. Były to następujące gatunki: Disciseda bovista, D. candida, Geastrum campestre, G. minimum, G. schmidelii, Tulostoma brumale, T. melanocyclum oraz T. squamosum. Wszystkie gatunki zebrano podczas badań nad grzybami wielkoowocnikowymi na siedliskach kserotermicznych Niecki Nidziańskiej głównie w latach 2010-2013. Badania prowadzono w sześciu

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FOLIA BIOLOGICA ET OECOLOGICA zbiorowiskach roślinności kserotermicznej – Adonido-Brachypodietum pinnati, Festucetum pallentis, Inuletum ensifoliae, Sisymbrio-Stipetum capillatae, Seslerio- Scorzoneretum purpureae and Thalictro-Salvietum pratensis (nazewnictwo za Matuszkiewicz 2012). W fitocenozach tych zespołów wyznaczono trzydzieści powierzchni badawczych, na których prowadzono obserwacje w regularnych dwutygodniowych odstępach czasu. Zanotowane grzyby należą do gatunków silnie zagrożonych w naszym kraju. Tylko jeden z nich – Tulostoma brumale – posiada większą liczbę stanowisk i z tego powodu można uznać go za częstszy. Pozostałe należą do grzybów bardzo rzadkich, a także do gatunków wymierających w Polsce, kategoria E (Wojewoda & Ławrynowicz 2006). Do analizy morfologicznej wybrano gatunki, które w obrębie rodzajów wykazują niewielkie różnice w budowie owocników. Na podstawie cech makro- i mikromorfologicznych, które zaobserwowano podczas badań terenowych i laboratoryjnych, sporządzono pełne opisy zanotowanych taksonów. Istotne różnice w budowie owocników poszczególnych taksonów wynikają ze stopnia rozwoju i zachowania cech poszczególnych owocników (Tomaszewska et al. 2011). Cechy te zacierają się w miarę dojrzewania i starzenia się tych struktur. Na wykształcenie cech fenotypowych mogą także wpływać między innymi czynniki pogodowe i siedliskowe (Tomaszewska et al. 2012). Otrzymane wyniki wskazują na potrzebę dalszych badań tej grupy grzybów, w celu poznania pełnej zmienności cech budowy morfologicznej i poprawnej identyfikacji taksonów.

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