Geasters in the Western Ghats and west coast of India

Inventory in different locations of the Western Ghats and west coast of India during 2011–2013 yielded six species of geasters (Geastrum fimbriatum, G. lageniforme, G. pseudostriatum, G. saccatum, G. schweinitzii and G. triplex). Based on fresh basidiomata, illustrations of geasters are presented with macroscopic and microscopic observations. Fruit bodies of G. lageniforme were gregarious and abundant followed by G. triplex, while G. pseudostriatum were rare and solitary. Among the geasters, G. triplex was ectomycorrhizal with native tree Terminalia paniculata of the west coast. Geastrum fimbriatum, G. pseudostriatum and G. schweinitzii constitute the first record for the Western Ghats of India. Distribution, substrate preference, ectomycorrhizal features, economic value and conservation of geasters are discussed.


Introduction
Geastrum (phylum: Basidiomycota; order: Geasterales; family: Geastraceae) is a cosmopolitan genus among the gasteroid fungi possessing enclosed hymenophore and commonly known as earthstars as exoperidium dehisce like star [1,2].The genus Geastrum was erected by Persoon [3] with G. coronatum as type species.This genus has been generally characterized by pointed globose or sub-globose puffball with four-layered peridium buried in litter or soil or humus strata without stipe and pileus consisting of brown globose or sub-globose spores with verrucose or echinulate surface.Traditional taxonomy of geasters mainly relay on morphological traits like peristome type (fibrillose or sulcate), its delimitation with rest of the body, presence or absence of connecting stalk between exoperidium and endoperidium, hygroscopic nature of exoperidial rays and structure of mycelial layer [1,2,[4][5][6].Recently, Zamora et al. [7] considered two important characteristics (macrochamical tests to detect phenoloxidase and crystalline deposits from rhizomorphs), which were not previously used for identification of Geastrum species.Most recently, the systematics of the genus Geastrum has been revisited using morphological, chemical and molecular phylogenetic approaches [8][9][10][11].
According to the dictionary of fungi by Kirk et al. [12] and Perez [13], 50 species of Geastrum have been described worldwide.However, based on herbarium specimens and field work, Zamora et al. [10,11] suggest the existence up to 100-120 species under the genus Geastrum.Hosaka et al. [14] proposed a new subclass Phallomycetidae in the order Geastrales.Phylogenetic relationship has been established between Gasteromycete and Geastrum spp.by sequencing large subunit of rDNA [15,16].Kasuya et al. [8] carried out molecular studies for the phylogenetic placement of Geastrum melanocephalum and Geastrum triplex, while Zamora et al. [17] redescribed a forgotten species of Geastrum argentinum based on molecular traits.Jeppson et al. [9] established phylogenetic relationships of European earthstars based on molecular sequence data (nuclear rDNA ITS1 and ITS2; LSU; Tef-α), morphological and ecological characteristics.However, substantial variation in sequence has been reported between morphologically similar taxa in Geastraceae [15,[18][19][20].Tedersoo et al. [21] studied the diversity, distribution and evolution of phylogenetic lineages among the ectomycorrhizal geasters.
Geasters in tropical region (e.g.Argentina, Bolivia, Brazil, Costa Rica, eastern Central Africa and West Africa) have been studied by various investigators [16,22,23].Many geasters occurring in the European region have been categorized under threatened, critically endangered and red listed (e.g.Bulgaria, Czech Republic, Montenegro, Poland, Slovakia, Sweden and Switzerland) [24][25][26].Western Ghats and west coast of India encompass a wide variety of habitats (e.g.forest reserves, sacred groves, agroforests, mangroves and sand dunes) supporting diverse macrofungal population.Several studies have been carried out in the Western Ghats and west coast region to document the distribution, richness and diversity of macrofungi [27][28][29][30][31][32][33].Literature search reveals a few reports on the occurrence of geasters in the southern India (including the Western Ghats) [29,31,32,[34][35][36][37] and northern India [38][39][40].Several interesting geasters were recovered during the current inventory on macrofungi in selected habitats of the Western Ghats and west coast of India.Thus, this paper presents the occurrence, distribution, habitats, substrate and morphological description of six species of geasters.

Material and methods
Survey was undertaken during southwest monsoon and post-monsoon seasons in 2011-2013 by opportunistic and transect methods.Inventory of geasters was carried out in forest reserves, Shola forests, coffee agroforests and sacred groves in the Western Ghats (Kodagu, Karnataka State).In the west coast (Mangalore, Karnataka State), locations surveyed for geasters include arboretum, Anacardium plantation, Areca agroforest, Pongamia plantation and Terminalia dominant mixed forest.
Measurement of basidiomata, mature fruit bodies and spore sac was the average of 10 specimens except for G. pseudostriatum (5 specimens), while measurement of basidiospores (mounted in phosphate-buffered-saline) inclusive of ornamentation was based on the average of 25 spores using 100× objective of Olympus System Microscope (CX41RF, USA).Selected specimens were cleaned and fixed in a mixture of water-ethanol-formaldehyde (14:5:1) and deposited in the mycological herbarium of the Department of Biosciences, Mangalore University (MUBSNCKKRSMF#034-039).

Results and description
A total six species of geasters (Geastrum fimbriatum, G. lageniforme, G. pseudostriatum, G. saccatum, G. schweinitzii and G. triplex) were recorded in different habitats of the Western Ghats and west coast.Among the locations surveyed, specific habitats endowed with geasters in the Western Ghats were coffee agroforests of B'Shettigeri (12°12'N, 75°88'E), sacred groves of Betoli (12°18'N, 75°78'E) and Acacia forests in Madikeri (12°42'N, 75°74'E).Four habitats in the west coast (12°48'N, 74°55'E) consists of geasters were Terminalia dominant mixed forest, Pongamia dominant mixed forest, arboretum (endemic and endangered tree species of the Wedstern Ghats) and plantations of Anacardium.Fruit bodies of G. lageniforme were the most abundant followed by G. triplex and G. schweinitzii.Geastrum fimbriatum and G. saccatum occur rarely either solitary or in pairs, while G. pseudostriatum was very rare and solitary.Among the geasters, G. triplex was ectomycorrhizal with native tree species (Terminalia paniculata) of the west coast.
Based on morphological, ecological and molecular sequence data, Jeppson et al. [9] reported that G. pseudostriatum is closely related to Geastrum berkeleyi.But, the latter requires nutrient-rich wooded/bushy habitats and produce large fruit bodies with small spores.
Immature basidiomata hypogeous, dull orange brown, onion-or pear-shaped, coarsely fibrous to squamulose and measures 2.6 (2.3-2.9)cm diam × 2.9 (2.7-3.2) cm tall.At maturity, epigeous, exoperidium splits into 5-6 non-hygroscopic starfish-like rays; involute and later curl upwards and inwards under the endoperidial body.Pseudoparenchymatous layer initially pinkish brown fading to brown with age, concentrically fissured to form a saucer-like base under the endoperidium.Mycelial layer not encrusted with soil, radially fissured and there is a basal attachment point to the belowground mycelium.Endoperidial body spherical to sub-spherical, 2.7 (2.1-2.9)cm diam, sessile and surmounting a saucer-like base.Endoperidium greyish brown, thin, papery and smooth.Peristome delimited, fibrillate, creamish to brown, conically elevated and surrounded by a distinct pallid to brownish black halo.Gleba at first pallid and firm, with age becoming brown and powdery.Spores brownish, spherical, spiny and measure 4.5 (4-4.7)µm diam (inclusive of ornamentations).SUBSTRATE AND DISTRIBUTION.On soil along with litter in Terminalia dominant mixed forest in Mangalore University campus (Mangalore; August 12, 2011; September 12, 2012, July 17, 2013; N.C.Karun; MUBSNCKKRSMF#039).Marasmius sp., Termitomyces umkowaan and Xylaria sp. were found in the vicinity of G. triplex.Geastrum triplex is ectomycorrhizal with Terminalia paniculata, and was also found in the vicinity of the tree bases of Artocarpus heterophyllus, Canarium strictum and Mangifera indica in arboretum of the west coast.Geastrum triplex occurred in grasslands and moist-deciduous to Shola forests in Sasthanada, Kerala [32].It was also recorded from the forest locations dominated by T. paniculata in Kuvempu University campus (Karnataka) [37].This geaster was ectomycorrhizal with Shorea robusta in tropical moist deciduous forests of central India [38].
According to Kasuya et al. [8], G. triplex is highly polyphyletic as the collar-forming Geastrum spp.are usually recognized under G. triplex-complex.But, the European and North American holotypes of G. triplex are not exactly similar with those from the Indonesia [8].Interestingly, the European G. triplex has been recently renamed as Geastrum michelianum by Zamora et al. [10].Further molecular studies are necessary to obtain more precise insight on this complex.

Discussion and outlook
Unidentified species of geasters were recorded in moist-deciduous forests, semievergreen forests, sacred groves and coffee plantations of the Western Ghats of Karnataka [29,31].In the Western Ghat region of Kerala, G. saccatum was recorded on the decaying logs [32].Recently it was also found in the urban locations of southern India [35].In the present study several locations in the Western Ghats and west coast of India has been surveyed for geasters and revealed that they were confined to specific habitats.They were restricted to the coffee agroforests, sacred groves, the forest reserves and Shola forests in the Western Ghats.Geastrum saccatum were found in Terminalia dominant mixed forest, Anacardium or Pongamia plantations and in Areca dominant plantations in the west coast.However, in the Western Ghats of Kerala, geasters were recovered in moist deciduous, semi-evergreen, evergreen and Shola forests and grasslands [32].According to Mohanan [32], geasters in Western Ghats are usually terrestrial, rarely lignicolous or coprophilous.In our study, fruit bodies of geasters were mainly observed at the soil surrounded with leaf litter and humus except for G. lageniforme and G. schweinitzii.Geastrum lageniforme preferred soil mixed with decomposing twigs or bark of Pongamia, while G. schweinitzii was found directly on the phyllodes of Acacia auriculiformis and decaying twigs of latex-yielding plant, Sapium insigne (see Fig. 2e).Besides of our studies, the occurrence of G. schweinitzii has also been noted in Brazil [13,48].
Although Natarajan et al. [28] have reported several ectomycorrhizal fungi in dipterocarp forest of the Western Ghats, none of them belongs to the genus Geastrum.Generally the genus Geastrum is believed to be saprotrophic with an exception of one published record from Germany indicating a mycorrhizal connection between G. fimbriatum and Fagus sylvatica [57].In our study, G. fimbriatum which occurred in the vicinity of Artocarpus heterophyllus, Coffea robusta and Mangifera indica in coffee plantations in the Western Ghats needs further survey to reveal its ectomycorrhizal nature.Bakshi [55] has reported association of roots of Abies pindrow, Cedrus deodara, Pinus wallichiana and P. roxburghii with G. fimbriatum in Dehra Dun forests of northern India.Geastrum triplex was gregarious, partially growing on soil and connected underneath to the feeder roots of timber yielding tree, Terminalia paniculata in the west coast region.Preliminary microscopic examinations revealed the presence of mycelial mantle around the roots and direct connection of mycelia with roots.Geastrum triplex is common in humus accumulated moist forest floor in the vicinity of T. paniculata with thick canopy cover especially during mid rainy season (August).This geaster has been consistently discovered in such habitats over a decade.Geastrum triplex was found close to the tree bases of Artocarpus heterophyllus (fruit and timber yielding), Canarium strictum (resin and canoe wood yielding) and Mangifera indica (fruit and fuel wood yielding) in 20 years old arboretum consisting of endemic and endangered trees of the Western Ghats.Geastrum triplex similarly to G. fimbriatum needs further studies to confirm its ectomycorrhizal association with different host tree species.Occasionally G. triplex has been collected from the forest locations dominated by T. paniculata in Kuvempu University campus (Karnataka) [37].Geastrum triplex was found to be ectomycorrhizal in Sal trees (Shorea robusta) in tropical moist deciduous forest in Madhya Pradesh, central India [38].Puffballs are known for many traditional uses (e.g.aesthetic, decorative, dietary and medicinal) [58][59][60].Geasters are used for prophylactic and therapeutic applications in newborn infants [60] and to clear the discharges from ear [61].Among the geasters recorded in the present study, G. fimbriatum is edible in India and Madagascar, G. saccatum is medicinal fungus in Mexico and G. triplex is edible and medicinal in India and Mexico, and G. triplex is also medicinal in Guyana [59].According to Soosairaj et al. [36], wild G. fimbriatum growing on the wood debris is edible in the southeastern India.Dore et al. [62] has reported anti-inflammatory, antioxidant and cytotoxic actions of extract from G. saccatum.Solvent extracts of G. triplex were also active against several plant and human pathogenic bacteria [37].Squeezing the spore sac of G. saccatum and G. triplex releases spores in to bee hives and anesthetize bees for about 30 min facilitating harvest of honey [63].
Several geasters occurring in Europe have been considered as threatened, critically endangered and red listed (e.g.Bulgaria, the Czech Republic, Montenegro, Poland, Slovakia, Sweden and Switzerland) [24][25][26]64].Esqueda et al. [66] have proposed priority regions for conservation of biodiversity of geasters in Mexico.However, Nitare [66] and Benkert [67] opined that geasters serve as indicators of specific habitats in need of conservation.There is a general opinion worldwide that geasters are under threat due to anthropogenic pressures and thus those locations endowed with geasters needs special attention for restoration and conservation.

Fig. 3
Fig. 3 Different stages of immature fruit bodies (a), onion-shape immature fruit body (b; a,b: scooped out basidiomata with rhizoidal connection with tree roots), partially dehisced fruit body (c), different stages of matured fruit bodies (d), unopened hairy peristome of mature fruit body (e), opened hairy peristome of mature fruit body (f) and spiny spores (g) of Geastrum triplex (note the connections of immature fruit bodies at the basal region with roots of Terminalia paniculata).Scale bars: 1 cm (a-f); 5 μm (g).