Christophe Billette

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.

Extracellular laccase production during hyphal interactions between Trichoderma sp. and Shiitake, Lentinula edodes

Lentinula edodes (Berk.) Pegler was cultivated in liquid media containing malt and yeast extract. Extracellular laccase activity, measured in the culture fluids, was 5–18 times higher in cultures incubated for 29 days than in cultures incubated for 24 days. The addition of water-soluble lignin derivatives or Trichoderma sp. in cultures of L. edodes incubated for 11 days increased laccase activity 3- to 20 fold. The higher response was obtained with live mycelium of Trichoderma sp., but cell-free culture fluids of Trichoderma sp. in pure cultures were also effective. Trichoderma sp. induced changes in the laccase isoenzyme pattern as a result of the alteration of laccases secreted by L. edodes and not the induction of new isoforms.

Interspecific interactions with Trichoderma longibrachiatum induce Pleurotus ostreatus defence reactions based on the production of laccase isozymes

Laccases are phenoloxidases involved in aromatic compound transformation but also in stress response towards antagonist species such as Trichoderma sp. In this study intracellular isoforms of laccases produced by Pleurotus ostreatus in liquid cultures with or without Trichoderma longibrachiatum showed five isoforms with various intensities depending on the culture conditions suggesting a basal expression of these enzymes, which can be induced by interspecific interactions. A first attempt to analyse the induction of P. ostreatus laccase-gene expression by a biotic factor was realized using semi-quantitative RT-PCR. We showed that the transcription of a laccase gene of P. ostreatus can be modified by a biotic stress such as T. longibrachiatum.

Characterization of the aap1 gene of Agaricus bisporus, a homolog of the yeast YAP1

The structure, homologies, polymorphism and expression profiles of a new gene, aap1, have been studied for precisely characterizing it and defining its putative involvement in thermo-tolerance of both vegetative mycelium growth and sporophore differentiation. Sequence polymorphism was analyzed in 3 homokaryons of A. bisporus and 24 strains having different abilities for mycelial growth at temperatures above 30 °C and for producing mature fruiting bodies at 25 °C. The level of gene expression was measured by real-time PCR both in vegetative mycelium after transfer from 25 to 32 °C and in primordia and fruiting bodies produced during cultures at 17 or 25 °C. The results indicated that aap1 gene belong to a new subfamily of the yeast YAP1 homologs. It is not a dominant contributor to the thermo-tolerance of A. bisporus, but the protein it encodes may be involved as an overall stress resistance transcription factor. The way Aap1 senses redox level differs from that of AP-1-like transcription factor Yap1.

Halotolerant laccases from Chaetomium sp., Xylogone sphaerospora, and Coprinopsis sp. isolated from a Mediterranean coastal area.

Laccases (EC 1.10.3.2) are phenoloxidases involved in the transformation of the recalcitrant fraction of organic matter in soil. These enzymes are also able to transform certain aromatic pollutants such as polycyclic aromatic hydrocarbons (PAHs) and are known to be inhibited by chloride ions. This study aims to test the potential of some fungal strains newly isolated from natural environments subjected to high osmotic pressure such as coastal ecosystems, to produce chloride tolerant laccases. Three strains were identified as Chaetomium sp., Xylogone sphaerospora (two Ascomycota), and Coprinopsis sp. (a Basidiomycota) and the laccases produced by these fungi were weakly inhibited by chloride ions compared with previous data from literature. Moreover, we tested their reactivity towards various PAHs which are widespread anthropic pollutants. They were able to transform anthracene to 9,10-anthraquinone and we determine 7.5xa0eV as the threshold of ionization potential for PAH oxidation by these laccases.