Genus Laccaria Research Articles

Endophytic Bacterial Community, Core Taxa, and Functional Variations Within the Fruiting Bodies of Laccaria.

Macrofungi do not exist in isolation but establish symbiotic relationships with microorganisms, particularly bacteria, within their fruiting bodies. Herein, we examined the fruiting bodies' bacteriome of seven species of the genus Laccaria collected from four locations in Yunnan, China. By analyzing bacterial diversity, community structure, and function through 16S rRNA sequencing, we observed the following: (1) In total, 4,840,291 high-quality bacterial sequences obtained from the fruiting bodies were grouped into 16,577 amplicon sequence variants (ASVs), and all samples comprised 23 shared bacterial ASVs. (2) The Allorhizobium-Neorhizobium-Pararhizobium-Rhizobium complex was found to be the most abundant and presumably coexisting bacterium. (3) A network analysis revealed that endophytic bacteria formed functional groups, which were dominated by the genera Allorhizobium-Neorhizobium-Pararhizobium-Rhizobium, Novosphingobium, and Variovorax. (4) The diversity, community structure, and dominance of ecological functions (chemoheterotrophy and nitrogen cycling) among endophytic bacteria were significantly shaped by geographic location, habitat, and fungal genotype, rather than fruiting body type. (5) A large number of the endophytic bacteria within Laccaria are bacteria that promote plant growth; however, some pathogenic bacteria that pose a threat to human health might also be present. This research advances our understanding of the microbial ecology of Laccaria and the factors shaping its endophytic bacterial communities.

Three new species of Laccaria (Agaricales, Basidiomycota) from Southwest China (Yunnan) based on morphological and multi-gene sequence data.

The genus Laccaria has been reported from temperate and tropical areas and is an important constituent in forest ecosystems. However, the species diversity of Laccaria in Southwest China (Yunnan) has been underestimated. In this paper, descriptions based on morphological and multi-gene sequence data from internal transcribed spacer (ITS) region, large subunit ribosomal RNA gene (nrLSU), translation elongation factor 1-α (TEF1α) and the polymerase II second largest subunit (RPB2) of three new Laccaria species from Southwest China (Yunnan) are reported. Two of these were characterized by orange pileus and globose to subglobose basidiospores: L. cinnabarina and L. spinulosa. While L. cinnabarina has orange red colored basidiocarps with conspicuously pellucid-striate pattern, and a fibrillose stipe with longitudinally striations, L. spinulosa has a brownish orange to brown fruiting body with light white pruinae and 2-spored basidia. Laccaria longistriata is characterized by brown to flesh-colored basidioma, prominently striate to sulcate pileus and globose to subglobose basidiospores. The three new species were described, illustrated and compared with closely related species in morphology and phylogeny.

Laccaria guizhouensis sp. nov. (Agaricales, Basidiomycota) from Southwest China

ABSTRACT The genus Laccaria (Hydnangiaceae, Agaricales) is an important ectomycorrhizal symbiont of a broad range of host plants. Laccaria guizhouensis was collected from a subtropical mixed forest dominated by Fagaceae in Southwest China and described based on morphological characteristics and molecular data. Laccaria guizhouensis is characterised by its medium-sized basidiocarps and strong striate or sulcatus, flesh-coloured to brown pileus, flesh-coloured to brown lamellae, 2- to 4-spored basidia, globose to obellipsoid, hyaline, moderately echinulate basidiospores, and 1–1.5 μm-long echinulate. Phylogenetic analyses based on ITS sequences (HMAS352265 and HMAS352266) indicated that L. guizhouensis represented a new species separated from all other Laccaria species.

Morphology and Molecular Phylogeny Reveal Five New Species of Laccaria (Hydnangiaceae, Agaricales) from Southern China.

The genus Laccaria is a type of cosmopolitan and ecologically important fungal group. Members can form ectomycorrhizal associations with numerous trees, and some species are common edible fungi in local markets. Although some new species from China are recently published, the species diversity of Laccaria is still unclear in China. In this study, some samples of Laccaria were collected from southern China, and morphological characteristics and phylogenetic analyses based on the multilocus dataset of ITS-LSU-tef1-rpb2 confirmed five new species. Laccaria miniata, L. nanlingensis and L. neovinaceoavellanea were collected from subtropical broad-leaved forests, and L. rufobrunnea and L. umbilicata were collected from subtropical mixed forests of southwest China. Full descriptions, illustrations, comparisons with similar species and phylogenetic analysis are provided.

Laccaria murina and L. pumila from Pakistan, a study based on SEM and nrITS molecular marker.

Two species of the genus Laccaria, are described as new reports for Pakistan. Laccaria murina has been collected from a Himalayan moist temperate forest in Khanspur, KP, while L. pumila was found in the moist temperate forests of Kumrat Valley in Dir Upper, KP, and at higher altitudes of the Deosai plains of Gilgit-Baltistan, Pakistan. Phylogenetic analysis based on the nrITS region clustered the Pakistani species with L. murina sequences with other Asian collections and L. pumila from USA and Netherlands. SEM of basidiospores along with detailed micro-morphological data are provided. General distribution, habitat, ecology, and diagnostic features are also discussed. RESEARCH HIGHLIGHTS: It has new reports from Pakistan, studies based on light, scanning electron microscopy, and nrITS molecular markers. These species have been described with detailed micro-morphological and molecular phylogenetic analyses. General distribution, ecology, diagnostic features, and comparisons with closely related specimens have been provided. Graphical representation of DNA extraction and geographical locations of sampling sites are also illustrated (Figures1 and 2). Very few members of this genus are already described from Pakistan.

Determination of Diversity, Distribution and Host Specificity of Korean Laccaria Using Four Approaches

The genus Laccaria (Hydnangiaceae, Agaricales) plays an important role in forest ecosystems as an ectomycorrhizal fungus, contributing to nutrient cycles through symbiosis with many types of trees. Though understanding Laccaria diversity and distribution patterns, as well as its association with host plants, is fundamental to constructing a balanced plant diversity and conducting effective forest management, previous studies have not been effective in accurately investigating, as they relied heavily on specimen collection alone. To investigate the true diversity and distribution pattern of Laccaria species and determine their host types, we used four different approaches: specimen-based analysis, open database search (ODS), NGS analysis, and species-specific PCR (SSP). As a result, 14 Laccaria species have been confirmed in Korea. Results regarding the species distribution pattern were different between specimen-based analysis and SSP. However, when both were integrated, the exact distribution pattern of each Laccaria species was determined. In addition, the SSP revealed that many Laccaria species have a wide range of host types. This study shows that using these four different approaches is useful in determining the diversity, distribution, and host of ECM fungi. Furthermore, results obtained for Laccaria will serve as a baseline to help understand the role of ECM fungi in forest management in response to climate change.

Nematicidal anthranilic acid derivatives from Laccaria species

Three undescribed natural products, the anthranilic acid derivatives laccanthrilic acids A, B, and C, as well as the known (3S)-1,2,3,4-tetrahydro-3-β-carboline-3-carboxylic acid were isolated from fruiting bodies of Laccaria laccata. The structures were established by 1D and 2D NMR spectroscopy, HR-(+)-ESIMS and chemical synthesis. The absolute configuration of laccanthrilic acids A and B was determined by GC-MS after hydrolytic cleavage and derivatisation of the resulting glutamic acid with methanol and Mosher's reagent and subsequent comparison with authentic synthetic samples of known absolute configuration. The absolute configuration of laccanthrilic acid C was determined by comparison of the CD spectra of laccanthrilic acids B and C with each other. Metabolic profiling of related species showed that the compounds are common in the genus Laccaria. Laccanthrilic acid B exhibited moderate nematicidal effects against Caenorhabditis elegans, which might explain to some degree the beneficial role of these fungi for the growth and survival of their host plants.

Laccaria violaceotincta: a new species from tropical India based on morphology and molecular phylogeny

Laccaria violaceotincta sp. nov. is described from Kerala State, India based on morphology and molecular phylogeny. All the collections of the species were made from a freshwater swamp forest predominantly composed of species of Myristica. The Maximum Likelihood (ML) analysis of sequence of the internal transcribed spacer (nrITS) of the ribosomal RNA gene clearly revealed its novelty as well as its position within the genus Laccaria. A taxonomic description, photographs of the basidiocarps and the micro-structures, comparisons with phenetically similar and phylogenetically related species and a phylogram depicting the placement of the new species are provided.

­­­Laccaria rubroalba sp. nov. (Hydnangiaceae, Agaricales) from Southwestern China

The novel species Laccaria rubroalba is described from Southwestern China by using both morphological characteristics and molecular data. It is characterized by small basidiomata; reddish white pileus when moist or young, white to pale when dry; 4-spored basidia and globose to broadly ellipsoid, hyaline, moderately echinulate basidiospores. Phylogenetic relationships inferred from ITS sequence data confirmed the separation of this new species from other closely related species in the genus Laccaria. The new species is compared with similar taxa. A description, line drawings and colour photographs of the new species, and phylogenetic tree to show the placement of the new species are provided.

Evolution of ectomycorrhizas as a driver of diversification and biogeographic patterns in the model mycorrhizal mushroom genus Laccaria.

Summary A systematic and evolutionary ecology study of the model ectomycorrhizal (ECM) genus Laccaria was performed using herbarium material and field collections from over 30 countries covering its known geographic range.A four‐gene (nrITS, 28S, RPB2, EF1α) nucleotide sequence dataset consisting of 232 Laccaria specimens was analyzed phylogenetically. The resulting Global Laccaria dataset was used for molecular dating and estimating diversification rates in the genus. Stable isotope analysis of carbon and nitrogen was used to evaluate the origin of Laccaria's ECM ecology.In all, 116 Laccaria molecular species were identified, resulting in a near 50% increase in its known diversity, including the new species described herein: Laccaria ambigua. Molecular dating indicates that the most recent common ancestor to Laccaria existed in the early Paleocene (56–66 million yr ago), probably in Australasia. At this time, Laccaria split into two lineages: one represented by the new species L. ambigua, and the other reflecting a large shift in diversification that resulted in the remainder of Laccaria. L. ambigua shows a different isotopic profile than all other Laccaria species.Isotopes and diversification results suggest that the evolution of the ECM ecology was a key innovation in the evolution of Laccaria. Diversification shifts associated with Laccaria's dispersal to the northern hemisphere are attributed to adaptations to new ecological niches.

A preliminary study of the ectomycorrhizal fungi associated with introduced Eucalyptus in Kenya

The assemblage of ectomycorrhizal fungi associated with introduced Eucalyptus in Kenya was characterized using molecular methods. DNA extracted from root-tips collected at 47 sites in the country yielded sequences of at least 21 taxa, with eight of these known to form ectomycorrhizal relationships with trees. Hydnangium carneum, a hypogenous fungus, and species of the agaric genus Laccaria appear to be the most common ectomycorrhizal associates of Eucalyptus. Other taxa recorded from the rhizosphere of Eucalyptus included two species of entomopathogenic fungi and a number of other species considered to be saprotrophic. As a general observation, based on the data obtained in the present study, the level of diversity of fungi associated with Eucalyptus appears to be relatively low.

Soil contamination with silver nanoparticles reduces Bishop pine growth and ectomycorrhizal diversity on pine roots.

Soil contamination by silver nanoparticles (AgNP) is of potential environmental concern but little work has been carried out on the effect of such contamination on ectomycorrhizal fungi (EMF). EMF are essential to forest ecosystem functions as they are known to enhance growth of trees by nutrient transfer. In this study, soil was experimentally contaminated with AgNP (0, 350 and 790 mg Ag/kg) and planted with Bishop pine seedlings. The effect of AgNP was subsequently measured, assessing variation in pine growth and ectomycorrhizal diversity associated with the root system. After only 1 month, the highest AgNP level had significantly reduced the root length of pine seedlings, which in turn had a small effect on above ground plant biomass. However, after 4 months growth, both AgNP levels utilised had significantly reduced both pine root and shoot biomass. For example, even the lower levels of AgNP (350 mg Ag/kg) soil, reduced fresh root biomass by approximately 57 %. The root systems of the plants grown in AgNP-contaminated soils lacked the lateral and fine root development seen in the control plants (no AgNP). Although, only five different genera of EMF were found on roots of the control plants, only one genus Laccaria was found on roots of plants grown in soil containing 350 mg AgNP/kg. At the higher levels of AgNP contamination, no EMF were observed. Furthermore, extractable silver was found in soils containing AgNP, indicating potential dissolution of silver ions (Ag+) from the solid AgNP.Electronic supplementary materialThe online version of this article (doi:10.1007/s11051-015-3246-4) contains supplementary material, which is available to authorised users.

Fate of ectomycorrhizal fungal biomass in a soil bioreactor system and its contribution to soil organic matter formation

Forest soils constitute a major carbon pool in the context of the global carbon cycle. Although ectomycorrhizal fungi (EMF) are responsible for a large part of the input of carbon into this pool, information on the transformation of EMF biomass into soil organic matter (SOM) is scarce. In order to quantitatively analyze the contribution of EMF biomass to SOM formation, we incubated a typical forest arenosol with 13C-labeled fungal biomass from the ectomycorrhizal basidiomycete Laccaria bicolor in a closed bioreactor system for 231 d, enabling the establishment of a full mass balance for the turnover. rDNA fragments specific for the genus Laccaria disappeared after 28 d, indicating death and decay of the organism. The amount of 13C PLFA as a proxy for living biomass decreased to roughly 10% of the initial value in the course of the experiment, again suggesting efficient disintegration of the fungal organism. After 231 d, however, 63.5% of the initially added 13C had been converted to SOM and 52% belonged to the non-living fraction. The analysis of 13CO2 formation (36.5% of the initially added label) suggested two-pool exponential kinetics with k1 = 0.0615 d−1 for the easily degradable and k2 = 0.0005 d−1 for the stable fraction (pool sizes 74% and 26% of the initially added label, respectively). When compared to the degradation of bacterial biomass in agricultural soils, the pool of readily degradable fungal biomass was considerably smaller and the half-lives of both pools (readily degradable and persistent C) were markedly higher for fungal biomass. In summary, our data allow the conclusion that EMF biomass is a significant source of SOM in forest ecosystems.

Three new Laccaria species from Southwest China (Yunnan)

In this paper descriptions of three new Laccaria species from Southwest China (Yunnan) are reported. Macromorphological, micromorphological, and molecular data have been studied to describe the new species and delineate them within the genus Laccaria. The first species Laccaria fulvogrisea is characterized by a grey to brownish coloured fruiting body and large echinulate spores. Laccaria aurantia is characterized by the deeply orange colour and globose balloon-like spores with a fine echinulate ornament. The third species Laccaria yunnanensis has large barrel-shaped pleurocystidia and a brownish to flesh-coloured fruiting body. All three species have 4-spored basidia and were found near or within Quercus and Lithocarpus mixed broad leaved forests at an altitude of above 2,000 m.

Multigene sequence data reveal morphologically cryptic phylogenetic species within the genus Laccaria in southern Australia

Laccaria (Hydnangiaceae, Agaricales, Basidiomycota) is one of the more intensively studied ectomycorrhizal genera; however, species boundaries within Laccaria and the closely related Hydnangium and Podohydnangium in Australia have not yet been examined with molecular sequence data. Based on morphological characters, eight native species of Laccaria are currently recognized in Australia, as well as three Hydnangium species and the monotypic Podohydnangium australe. Sequences of the internal transcribed spacer region of nuclear rDNA (ITS), RNA polymerase beta subunit II (rpb2) and translation elongation factor 1 alpha (tef-1α) were generated from 77 collections of Laccaria, Hydnangium and Podohydnangium from Australia. Ten phylogenetic species and a further 11 potential species (represented by singletons) of Laccaria in Australia are delimited from sequence analyses. Most of the morphological species contained cryptic phylogenetic species, but these species were always nested entirely within a given morphological species, although not always as sister taxa. The rpb2 locus performed best as a species barcode with pairwise and patristic distance measures. The ITS sequence region returned the least resolved gene tree of the three regions examined and was the least useful as a barcode region. Based on the phylogenetic topology, there appears to have been multiple gains and/or losses of the ectomycorrhizal association of some species with the myrtle beech, Nothofagus cunninghamii as well as of sequestrate basidiocarps and two-spored basidia.

The ability of ectomycorrhizal fungi to utilize fatty acids and a lipid as a carbon source for mycelial growth

We have investigated growth of ectomycorrhizal (ECM) fungi (i.e., 55 strains of 32 species in 15 genera) on saturated (palmitate), monounsaturated (oleate), diunsaturated (linoleate), triunsaturated (linolenate) fatty acids, and the triacylglyceride of oleate (triolein) lipid to elucidate an ability to utilize the fatty acids and lipid as a carbon source for growth. Relative utilization ratios (URs, %) based on mycelial growth on glucose suggest that ECM fungi belonging to the family Thelephoraceae have an ability to utilize palmitate. On the other hand, ECM fungi in the genus Laccaria can utilize at least either palmitate or oleate. Furthermore, Hygropharus russula grows on palmitate, oleate, and slightly on triolein. Lactarius chrysorrheus grows only on palmitate. These fatty-acid- and lipid-utilizing fungi may be promising as model fungi for further elucidation of the metabolic ability to utilize the fatty acids and lipid as a carbon source. On the contrary, the fungi in the genus Suillus were shown to scarcely utilize the fatty acids and lipid. Furthermore, most ECM fungi did not grow on either linoleate or linolenate.Key words: carbon source, ectomycorrhizal fungi, fatty acid, lipid, mycelial growth.

Basidiospore Wall Ultrastructure of the False-Truffle Hydnangium and Its Phylogenetic Significance

in basidiomycete systematics. Arguments for and against including false truffles in the Agaricales hinge on the interpretation ofthe character suites and the degree of importance given to them in forming phylogenetic hypotheses (e.g., Kuhner, 1980, 1984; Julich, 1981; Singer, 1986). A number of mycologists have postulated a close relationship between the false-truffle genera Hydnangium Wallr. and Podohydnangium Beaton, Pegler, & Young and the mushroom genus Laccaria Berk. & Br. (e.g., Pegler and Young, 1979; Kuhner, 1980, 1984; Julich, 1981; Beaton et al., 1984; Mueller, 1992; Mueller and Ammirati, 1993) based on similar basidiospore or? namentation. There are numerous publications illustrating basidiospore ornamentation as viewed under the scanning electron microscope for one or more of these three genera (e.g., Pegler and Young, 1969, 1971, 1979; Beaton et al., 1984; Castellano et al., 1989; Mueller, 1991, 1992). Kuhner (1980, 1984) reported that the discrete, conical echinulae observed on basidiospores of both Laccaria and Hydnangium have microfibrils that run nearly parallel to the axis of the spine. He used these wall ultrastructure features, along with cytological and ecological characters, to justify placing Hydnangium and Laccaria in the same family Hydnangiaceae in his Tricholomatales (Kuhner, 1980, 1984). Based on unpublished micrographs of Hydnangium, Mueller (1992) and Mueller and Ammirati (1993) argued that this type of basidiospore wall ultrastructure is the one shared derived morphological feature that links these genera. However, while illustra? tions of the basidiospore wall ultrastructure of Laccaria were presented in Besson and Kiihner (1971) and Kiihner (1980), to our knowledge, no transmission electron micrographs showing the basidiospore wall ultrastructure of Hydnangium or Podohydnangium have been published. In this paper, we present micrographs documenting the similarity of wall ultrastructure of Hydnangium and Laccaria and discuss the phylogenetic implications of these data. Scanning electron micrographs were taken at the Field Museum of Natural History following the procedures outlined in Mueller (1991,1992). For examination on the Uppsala University Jeol 100 B transmission electron microscope, small pieces of lamellar tissue from air-dried herbari? um specimens were rehydrated in 3% KOH, fixed in 2.5% glutaraldehyde in 0.1 M phosphate buffer (pH 7.5), and washed in buffer three times. The samples were then postfixed in 2% Os04 in 0.1 M phosphate buffer for 2 h at 4 C, washed in buffer three times, dehydrated in a graded eth? anol series and propylene oxide infiltrated with Epon, and embedded in Epon that was polymerized for 48 h at 60 C. Sections of 50 nm thickness were cut with a Dupont diamond knife

Basidiospore Wall Ultrastructure of the False-Truffle Hydnangium and Its Phylogenetic Significance

in basidiomycete systematics. Arguments for and against including false truffles in the Agaricales hinge on the interpretation ofthe character suites and the degree of importance given to them in forming phylogenetic hypotheses (e.g., Kuhner, 1980, 1984; Julich, 1981; Singer, 1986). A number of mycologists have postulated a close relationship between the false-truffle genera Hydnangium Wallr. and Podohydnangium Beaton, Pegler, & Young and the mushroom genus Laccaria Berk. & Br. (e.g., Pegler and Young, 1979; Kuhner, 1980, 1984; Julich, 1981; Beaton et al., 1984; Mueller, 1992; Mueller and Ammirati, 1993) based on similar basidiospore or? namentation. There are numerous publications illustrating basidiospore ornamentation as viewed under the scanning electron microscope for one or more of these three genera (e.g., Pegler and Young, 1969, 1971, 1979; Beaton et al., 1984; Castellano et al., 1989; Mueller, 1991, 1992). Kuhner (1980, 1984) reported that the discrete, conical echinulae observed on basidiospores of both Laccaria and Hydnangium have microfibrils that run nearly parallel to the axis of the spine. He used these wall ultrastructure features, along with cytological and ecological characters, to justify placing Hydnangium and Laccaria in the same family Hydnangiaceae in his Tricholomatales (Kuhner, 1980, 1984). Based on unpublished micrographs of Hydnangium, Mueller (1992) and Mueller and Ammirati (1993) argued that this type of basidiospore wall ultrastructure is the one shared derived morphological feature that links these genera. However, while illustra? tions of the basidiospore wall ultrastructure of Laccaria were presented in Besson and Kiihner (1971) and Kiihner (1980), to our knowledge, no transmission electron micrographs showing the basidiospore wall ultrastructure of Hydnangium or Podohydnangium have been published. In this paper, we present micrographs documenting the similarity of wall ultrastructure of Hydnangium and Laccaria and discuss the phylogenetic implications of these data. Scanning electron micrographs were taken at the Field Museum of Natural History following the procedures outlined in Mueller (1991,1992). For examination on the Uppsala University Jeol 100 B transmission electron microscope, small pieces of lamellar tissue from air-dried herbari? um specimens were rehydrated in 3% KOH, fixed in 2.5% glutaraldehyde in 0.1 M phosphate buffer (pH 7.5), and washed in buffer three times. The samples were then postfixed in 2% Os04 in 0.1 M phosphate buffer for 2 h at 4 C, washed in buffer three times, dehydrated in a graded eth? anol series and propylene oxide infiltrated with Epon, and embedded in Epon that was polymerized for 48 h at 60 C. Sections of 50 nm thickness were cut with a Dupont diamond knife

CYTOLOGICAL STUDIES IN LACCARIA (AGARICALES). I. MEIOSIS AND POSTMEIOTIC MITOSIS

In the genus Laccaria, basidial formation, dikaryotic basidia, karyogamy, and meiosis were generally similar to structures and phenomena reported for other Agaricales. The haploid nuclei of dikaryotic basidia resided side by side in the basidium prior to karyogamy. Following karyogamy a single nucleolus was observed in L. montana (four-spored species); several nucleoli remained in the nucleus of L. tortilis (two-spored species). The haploid number of chromosomes for L. montana appeared to be n = 9. Postmeiotic mitosis typically occurred in the basidiospores resulting in binucleate basidiospores for four-spored species and quadrinucleate basidiospores for two-spored species. Postmeiotic mitosis sometimes occurred in the sterigmata and basidia proper. In instances where postmeiotic mitosis occurred in basidia, mature basidiospores were not formed and the basidia were collapsed, and contained up to eight nuclei.

Numerical Taxonomic Analyses on Laccaria (Agaricales)

Numerical taxonomic analyses (UPGMA and PCOR) of data obtained through detailed examinations of basidiocarps and somatic cultures were conducted to develop an objective classification scheme for Laccaria in North America. Analyses were also undertaken to investigate the effect which different character sets had on the classification scheme. Classi? fication schemes based on incomplete data sets differed significantly from the scheme obtained when all ofthe available data were utilized. A complete circumscription of Laccaria species, therefore, required data on cultural features as well as basidiocarp characters. A major problem in delimiting stable and natural taxa within the agaric genus Laccaria Berkeley & Broome (Tricholomataceae) has been the paucity of stable, diagnostic characters (Mueller, 1982). Consequently, a critical examination was made of cultural and basidiocarp macro- and micromorphological characters during a taxonomic study of the genus for the continental United States and Canada (Mueller, 1982). Numerical taxonomic techniques were used to analyze objectively and efficiently the large amount of accumulated data. As mentioned by Wolfe (1984), numerical taxonomic methods have been used infrequently in basidiomycete taxonomy. The goal of this part of the study was two-fold: 1) to develop a phenetic classification scheme for the genus in the study region based on all available characters, and 2) to evaluate the use of restricted character sets in delimiting taxa. The hypothesis was that it was necessary to have information on basidiocarp macro- and micromorphology along with data acquired through somatic culture mat analyses to circumscribe accurately taxa of Laccaria.