Further notes on the morphology and distribution of Neopaxillus echinospermus ( Agaricales , Basidiomycota ) in Southern Brazil

Neopaxillus echinospermus is a common but poorly understood agaric species from South America (Argentina, Bolivia, Brazil and Paraguay). In this paper, we discuss conflicting morphological features reported in the literature and expand the distribution of the species to Seasonal Deciduous and Semideciduous Forests from Rio Grande do Sul and Paraná states, respectively, in Southern Brazil. Hyphal structure of the pileipellis, presence of cheilocystidia and caulocystidia, presence of clamp connections and oleiferous (thrombopleurous) hyphae, and the basidiospore morphology under scanning electron microscopy are illustrated and discussed in detail.

Neopaxillus echinospermus (Speg.)Singer is a mushroom species, with a limited and poorly documented distribution in South America, comprising Argentina, Brazil, Paraguay and Bolivia (Watling and de Meijer 1997).This agaric was originally described by Singer (1948) as Neopaxillus echinosporus Singer, based on specimens collected by J. Rick supposedly in Rio Grande do Sul, in Southern Brazil.This name was put into synonymy based on further examination of Naucoria echinosperma Speg.from Brazil (Spegazzini 1889), which was considered the correct basionym of the species; for a detailed nomenclatural discussion, see Watling and Aime (2013).
For many years, Neopaxillus Singer remained as a monotypic genus in the Crepidotaceae (Imai) Singer, until Singer and Lodge (1988) described N. plumbeus from Costa Rica, Central America.Based on both macroand micromorphological similarities, some researchers suggested that the genus would be related to the gilled boletes, proposing its placement in the Paxillaceae Lotsy (Singer 1986) or Serpulaceae Jarosch & Bresinsky (Binder and Hibbett 2006).Recently, Vizzini et al. (2012) described N. dominicanus from Central America and their phylogenetic resulted in the return of Neopaxillus to the Crepidotaceae.Finally, Watling and Aime (2013) analyzed morphology and 28s ribosomal DNA region of N. echinospermus and reassured the position of the type species in the latter family.
In spite of the current and relatively consistent position of the genus, supported on molecular analysis, conflicting morphological data have been noted in the literature, such as the absence or presence of cystidia and clamp connections, and the hyphal structure of the pileipellis.
Thus, the aim of this paper is to provide modern and detailed descriptions of N. echinospermus, updating the morphological concept of the species, based on collections from Atlantic Forest in Southern Brazil, and to expand the known distribution of the fungus in Seasonal Semideciduous and Deciduous Forests of that biome.
Samples were collected, during recent fieldwork in Seasonal Semideciduous Forest from the western region of Paraná state, and previous collections in Seasonal Deciduous Forests from central region of Rio Grande do Sul state.These ecosystems are two common vegetation types in the Atlantic Forest biome of Southern Brazil, in which a portion of the trees defoliates during the dry season (IBGE 2012).Moreover, these forests are characterized by the presence of tree species of Anacardiaceae, Apocynaceae, Bignoniaceae, Boraginaceae, Caricaceae, Fabaceae, Malvaceae, Meliaceae, Moraceae, Rutaceae and Sapindaceae (Roderjan et al. 2002).
Habitat: In the forest, terricolous, among litterfall, solitary to gregarious or subcaespitose, in small clusters, on bare soil.The literature presents some controversial morphological data on Neopaxillus echinospermus.In the original description, Singer (1948) described the species as without cystidia, but with cystidioles on the gill margin, clamp connections in all septa, and pileipellis composed of a trichoderm-palisade.On the other hand, Watling and de Meijer (1997) and Watling and Aime (2013) reported specimens from Paraná state without any type of cystidia, absence of clamp connections, and pileipellis formed of filamentous, somewhat incrusted, and irregularly arranged hyphae.De Meijer (2008) presented a detailed description and illustrations of N. echinospermus and, in agreement to our examined material, he described the presence of both cheilocystidia and clamp connections.Vizzini et al. (2012), on discussing N. dominicanus Angelini & Vizzini, also examined specimens of N. echinospermus from Paraná (collected by A.A.R de Meijer) and not only observed cheilocystidia but also inconspicuous pleurocystidia, which we have not found in our materials.
All examined samples from Paraná and Rio Grande do Sul exhibited conspicuous clamp connections (Figure 13) and cheilocystidia (Figures 6-8), with variable morphology, including presence of catenulate cheilocystidia (Figure 7); the pileipellis of these mushrooms also presented the same trichoderm-palisadoderm pattern described by Singer (1948), which is mainly composed of clavate or (sub-) cylindrical elements.It is probable that older specimens can be more difficult to note the presence and position of such structures, but when fresh collections are examined they are conspicuous.In addition, we also noted the presence of oleiferous (thrombopleurous) hyphae in all parts of the basidiomata (Figure 13), a feature only mentioned in the recently described N. dominicanus (Vizzini et al. 2012).The great variation on the color of pileus is also illustrated here (Figures 3 and 4), however this feature alone is of poor taxonomic value, since the basidiomata persist for several days and are subject to numerous environmental factors that change the basidiomata colors.
The probable mycorrhizal status of this mushroom is to be determined, since the genus was considered in the past close to Paxillaceae and Cortinariaceae, two families with numerous ectomycorrhizal genera.Our field observations suggest that this species would not be a mycorrhizal fungus, because when the basidiomata are carefully removed from the substrate, we have not noticed the presence of rhizomorphs or other morphological evidences of this partnership.No molecular or culture studies were performed to determine (or not) the mycorrhizal status in Neopaxillus members, but as previously pointed, the absence of typical plant partners in this part of Atlantic Forests, as well the current placement of the species in the nonmycorrhizal family Crepidotaceae are strong evidences suggesting saprotrophic condition (Aime et al. 2005;Vizzini et al. 2012;Watling and Aime 2013).
Neopaxillus echinospermus is known from South American tropics and subtropical regions (Argentina, Paraguay, Bolivia, and Southern and Southeastern Brazil); reports from Central America and México were considered by Vizzini et al. (2012) as misidentifications of N. dominicanus.In Brazil, the species has been previously reported from Ombrophilous Forests, especially the Mixed Ombrophilous Forest from Southern (de Meijer 2008, from Paraná) and Dense Ombrophilous Forest in Southeastern (Spegazzini 1889, from São Paulo).The type locality of N. echinosporus (Singer 1948) is not precise; the material was collected by J. Rick in a place called Couto which, according to Watling and de Meijer (1997), may to represent a locality near the municipality of Rio Pardo in the central region Rio Grande do Sul state.If correct, this area belongs to the Pampa biome of southern Brazil, and comprises swampy and grassland vegetation, composed mainly of native grasses, with tree vegetation found only of riparian forest (Silva et al. 2011).With the new findings, the distribution of this species is now expanded to the Seasonal Semideciduous Forests from the state of Paraná and Seasonal Deciduous Forests of Rio Grande do Sul state, both in Southern Brazil.New reports of N. echinospermus are expected from other areas of Central Brazil and South America, requiring future mycological fieldwork on these areas.

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List 12(1): 1834, 28 January 2016 doi: http://dx.doi.org/10.15560/11.1.1834ISSN 1809-127X © 2016 Check List and Authors Notes oN GeoGraphic DistributioN Silva-Filho et al. | Further notes on Neopaxillus echinospermus in Southern Brazil (1978).Micrographs were made from a Motic BA310 light microscope with a Moticam 2500 digital camera, and measurements were taken through software Motic Image Plus 2.0.In the basidiospores description Q is the quotient between the length and width, Qm is the medium value of Q, and n is the number of measured basidiospores/number of analyzed basidiomata/number of collections.Scanning electron micrographs (SEM) were performed at the Center of Electron Microscopy of the Federal University of Paraná at Curitiba (CME/ UFPR), under a Jeol JSM-6360LV scanning electron microscope.Examined specimens are preserved at the herbaria HCP (Universidade Federal do Paraná, Campus Palotina) and HCB (Universidade de Santa Cruz do Sul, Departamento de Biologia e Farmácia).
Known distribution: In tropical (Amazon Rain Forest and Yungas, Bolivia) and subtropical (Bosque Chaqueño and Yungas in Argentina, Bosque Chaqueño in Paraguay, Atlantic Forest in Brazil) regions of South America(Singer 1964).In Brazil, it has been reported only for Dense Ombrophilous Forests and Mixed Ombrophilous Forest in São Paulo and Paraná states (de Meijer 2008;Watling and de Meijer 1997).The probable type locality of N. echinosporus, as discussed later, is within the Pampa