Richard W. Kerrigan

Genome sequence of the button mushroom Agaricus bisporus reveals mechanisms governing adaptation to a humic-rich ecological niche

Agaricus bisporus is the model fungus for the adaptation, persistence, and growth in the humic-rich leaf-litter environment. Aside from its ecological role, A. bisporus has been an important component of the human diet for over 200 y and worldwide cultivation of the “button mushroom” forms a multibillion dollar industry. We present two A. bisporus genomes, their gene repertoires and transcript profiles on compost and during mushroom formation. The genomes encode a full repertoire of polysaccharide-degrading enzymes similar to that of wood-decayers. Comparative transcriptomics of mycelium grown on defined medium, casing-soil, and compost revealed genes encoding enzymes involved in xylan, cellulose, pectin, and protein degradation are more highly expressed in compost. The striking expansion of heme-thiolate peroxidases and β-etherases is distinctive from Agaricomycotina wood-decayers and suggests a broad attack on decaying lignin and related metabolites found in humic acid-rich environment. Similarly, up-regulation of these genes together with a lignolytic manganese peroxidase, multiple copper radical oxidases, and cytochrome P450s is consistent with challenges posed by complex humic-rich substrates. The gene repertoire and expression of hydrolytic enzymes in A. bisporus is substantially different from the taxonomically related ectomycorrhizal symbiont Laccaria bicolor. A common promoter motif was also identified in genes very highly expressed in humic-rich substrates. These observations reveal genetic and enzymatic mechanisms governing adaptation to the humic-rich ecological niche formed during plant degradation, further defining the critical role such fungi contribute to soil structure and carbon sequestration in terrestrial ecosystems. Genome sequence will expedite mushroom breeding for improved agronomic characteristics.

Morphological, genetic, and interfertility analyses reveal a novel, tetrasporic variety of Agaricus bisporus from the Sonoran Desert of California

A distinctive variety of A. bisporus has been discovered in several habitats in the low Sonoran Desert of the Coachella Valley of California at elevations below sea level. The tetrasporic character of the basidia is consistently predominant in this local population. The spores of this population tend to be shorter and narrower than those of the bisporic strains. The genotypes of two individuals from this population clearly place them in the species A. bisporus, where they form a cohesive group in specieswide cluster analyses of nuclear genotypic similarity. The relative genetic divergence of this population from others in the species is of a similar order to that observed between other geographically isolated, bisporic populations of the species. The tetrasporic individuals exhibit complete interfertility with several bisporic strains of European and Coastal Californian origin. Because of their basidial and spore morphologies, these mushrooms do not fall within the traditional circumscription of A. bisporus. Varietal status is proposed for these distinctive, tetrasporic members ofthe species, and A. bisporus var. burnettii is described. Primary habitat of these mushrooms is being destroyed rapidly near our field sites. Geographical range of the variety is unknown.

A phylogenetic reconstruction and emendation of Agaricus section Duploannulatae.

Agaricus section Duploannulatae comprises the group of species allied with A. bisporus and A. bitorquis. Disagreement exists in the literature regarding the composition of this group. We used DNA sequence data from the ITS segments of the nuclear ribosomal DNA region, in a sample of European and North American isolates, to identify characters shared by this group, to further delimit species-level taxa within the section, and to develop a phylogenetic hypothesis. Shared polymorphisms that suggest a natural limit for section Duploannulatae were found. ITS1 data were assessed using parsimony, distance and maximum likelihood methods of phylogeny. The section Duploannulatae comprised six robust clades. Five clades corresponded to well characterized species from the temperate Northern Hemisphere (A. bisporus, A. subfloccosus, A. bitorquis, A. vaporarius, A. cupressicola). The sixth clade encompassed an A. devoniensis complex. Species concepts, nomenclature, and relationships are discussed and compared with prior reports.

The indigenous coastal Californian population of the mushroom Agaricus bisporus, a cultivated species, may be at risk of extinction

This study employed nuclear and mitochondrial markers to assess the present‐day composition of the population of Agaricus bisporus in coastal California. Favourable weather in the fall, winter and spring of 1990–91 furnished an uncommon opportunity to collect and study field material of the ‘button mushroom’A. bisporus, a cultivated species, from the region. The previous such season occurred 13 years earlier. Ninety‐five nonredundant cultures from field material were prepared and genotypically characterized. These data were combined with data from earlier studies. Multilocus nuclear and mitochondrial genotypes were determined for 123 individuals. Genotypes were compared in pairwise fashion both within the sample and between this sample and others of diverse geographical origin or commercial provenance. Using parametric analysis and cluster analysis of nuclear similarities, and also mitochondrial data, two elements – indigenous and European – were apparent within the sample. This was consistent with our earlier results on a much smaller sample. At least 10 mitochondrial haplotypes (MTs) were present; based on genotypic similarities of associated nuclei, five (or six) MTs were Californian, four were European, and one was ambiguous. Based on MT origins, 54% of the 121 classifiable individuals in California were of European ancestry; natives constituted a minority at 46%. Even in the indigenous Monterey cypress habitat, where 84% of all individuals from California were sampled, non‐native A. bisporus appeared to have achieved parity (at 48–49%) with the native population. In all other habitats, which are far more extensive, European individuals outnumbered Californian natives by 4:1. Some evidence of hybridization between the two ancestral groups was found. European strains appear to have been resident in California for approximately one century. The extensive occupancy of the native habitat by non‐native germ plasm, the apparent inability of native strains to occupy or compete in non‐native habitats, and the disproportionately large inoculum reservoirs represented by non‐native habitat and agronomic activities all suggest that the native population is under considerable competitive pressure in what may be a very dynamic situation. If this surmise is correct, the native population may be at serious risk of further contraction, irreversible dilution through interbreeding, and possibly even extinction.

Mitochondrial DNA variation in natural populations of the mushroom Agaricus bisporus

We investigated the patterns of mitochondrial DNA variation in the global population of the commercial mushroom Agaricus bisporus. Through the analysis of RFLPs among 441 isolates from nine countries in North America and Eurasia, we found a total of 140 mtDNA haplotypes. Based on population genetic analysis, there are four genetically distinct natural populations in this species, found in coastal California, desert California, France and Alberta (Canada). While 134 of the 140 mtDNA haplotypes were unique to single geographical regions, two mtDNA haplotypes, mt001 and mt002, were found in almost every population surveyed. These two mtDNA haplotypes also predominate among cultivars used throughout the world for at least the last two decades. These two mtDNA haplotypes are more similar to the cosmopolitan groups of mtDNA haplotypes than to the indigeneous clusters of mtDNA haplotypes from the two Californian regions.

Agaricus section Xanthodermatei: a phylogenetic reconstruction with commentary on taxa

Agaricus section Xanthodermatei comprises a group of species allied to A. xanthodermus and generally characterized by basidiomata having phenolic odors, transiently yellowing discolorations in some parts of the basidiome, Schaeffer’s reaction negative, and mild to substantial toxicity. The section has a global distribution, while most included species have distributions restricted to regions of single continents. Using specimens and cultures from Europe, North America, and Hawaii, we analyzed DNA sequences from the ITS1+2 region of the nuclear rDNA to identify and characterize phylogenetically distinct entities and to construct a hypothesis of relationships, both among members of the section and with representative taxa from other sections of the genus. 61 sequences from affiliated taxa, plus 20 from six (or seven) other sections of Agaricus, and one Micropsalliota sequence, were evaluated under distance, maximum parsimony and maximum likelihood methods. We recognized 21 discrete entities in Xanthodermatei, including 14 established species and 7 new ones, three of which are described elsewhere. Four species from California, New Mexico, and France deserve further study before they are described. Type studies of American taxa are particularly emphasized, and a lectotype is designated for A. californicus. Section Xanthodermatei formed a single clade in most analyses, indicating that the traditional sectional characters noted above are good unifying characters that appear to have arisen only once within Agaricus. Deep divisions within the sequence-derived structure of the section could be interpreted as subsections in Xanthodermatei; however, various considerations led us to refrain from proposing new supraspecific taxa. The nearest neighbors of section Xanthodermatei are putatively in section Duploannulati.

New and rare taxa in Agaricus section Bivelares (Duploannulati).

Ongoing field and laboratory studies have led to our recognition of new taxa in Agaricus section Bivelares, a recent combination and now the earliest synonym and correct name of section Duploannulati. Agaricus cupressophilus and A. tlaxcalensis, in the new Agaricus subsection Cupressorum, and A. agrinferus, A. devoniensis subsp. bridghamii, and A. subsubensis in Agaricus subsection Hortenses, are described. Agaricus subfloccosus is lectotypified. Phylogeny reconstruction methods with ITS1+2 DNA sequences were used to determine appropriate placements of the new taxa. Collectively these new taxa and phylogenetic associations represent a substantial augmentation and clarification of our knowledge of section Bivelares; described, sequenced species-level taxa in the northern hemisphere are increased from six to 10, a distinct subsectional lineage is revealed and infraspecific resolution within A. devoniensis is improved. An anomalous ITS1+2 sequence is documented in one collection of A. subsubensis. Preliminary data on another novel member of Bivelares from France also are provided. Several of these taxa are rare, highlighting opportunities and challenges for documenting biodiversity in this group. Additional comments on related taxa treated in recentPUBLICations are also provided.

Population and phylogenetic structure within the Agaricus subfloccosus complex

The name Agaricus subfloccosus (J. E. Lange) Pilat, based originally on Danish specimens, has been applied to mushrooms growing in coastal northwestern Europe as well as to mushrooms associated with Picea (and Abies ) at higher elevations in western Europe. Corresponding populations are also found in these two habitat zones in western North America. Material of both lowland and highland forms from Europe and North America was studied using morphological, cultural, and genotypic approaches. Homothallism was consistently observed in those isolates studied. Individual and population level genetic data are also consistent with homothallic reproduction. Dissimilarity analysis of multilocus nuclear and mitochondrial genotypes based on allozyme and DNA RFLP markers provided strong evidence that, within either ecologically distinct group, the amphiatlantic populations were genotypically very similar. Conversely, there were large genotypic differences between the two ecologically distinct groups, whether within or between continents. Supported by cultural and morphological evidence, these data indicate that two ancient, phylogenetically distinct entities exist within current concepts of A. subfloccosus . A formal taxonomic resolution is complicated by the lack of a holotype for Langes species.

Evidence for amphithallism and broad geographical hybridization potential among Agaricus subrufescens isolates from Brazil, France, and Thailand

Agaricus subrufescens is a cultivated edible and medicinal mushroom. Its known geographical distribution encompasses the Americas, Europe, Oceania, and Asia. The objective of this study was to assess mating compatibility and interfertility of strains originating from Brazil, France, and Thailand. Progeny of each strain were analyed with codominant molecular markers. Multilocus genotype tests revealed that the three strains were amphithallic with percentages of heterokaryotic single spore progenies of 75% for the Thai strain and around 40% for the Brazilian and French strains. In mating tests A. subrufescens had a multiallelic unifactorial system of sexual incompatibility. The three parent strains were interfertile based on experimental pairings of single-spore isolates, the recovery of hybrid heterokaryons from compatible matings, and the ability of hybrids to produce mushrooms and fertile spores. This biological approach supports the inclusion of the European strains within the species and the extension of the geographical distribution range to Asia. Our data should help to develop breeding strategies and to better manage and exploit the diversity existing in A. subrufescens.

Studies in Agaricus I. Agaricus Pattersonae

Agaricus pattersonae Peck is one of several poorly known species of Agaricus described from California. It has a history of confusion that predates its publication in 1907. This paper is intended to resolve the conflicts surrounding that species. For several years I collected a robust, red-staining mushroom in old groves of Cupressus macrocarpa Hartw. ex Gord. (Monterey cypress), a California endemic. Although this fungus bore a striking resemblance to the photograph of the type of A. pattersonae published by Smith in 1940 (7), discrepancies in the type description precluded such a determination. An examination of the type material was therefore imperative. In addition to the holotype at Albany (NYS), portions of the original collection (A. M. Patterson 18) were located in Bronx (NY) and in Beltsville (BPI). Also located at Beltsville were Pattersons field notes, along with notes and watercolors by S. Nohara, and correspondence from Peck. Patterson and Nohara, botany students at the Leland Stanford Junior University, began to make numbered collections of fleshy fungi during the winter of 1906-1907. On 31 January 1907 Patterson described, sketched and photographed two separate collections of Agaricus. Patterson 17 had a whitish pileus streaked with light brownish fibrils, an entire annulus, and was found under pines. Patterson 18 was characterized by a pileus covered with large, appressed brown scales, a strikingly two-layered annulus, and a habitat under cypress. The following day Nohara described and painted an Agaricus collection (Nohara 25) corresponding to Patterson 18, taken under cypress from the same area: the Stanford racetrack/stock farm.