Pleurotus ostreatus is one of the widely cultivated edible fungi across the world. Mycelial subculture is an indispensable part in the process of cultivation and production for all kinds of edible fungi. However, successive subcultures usually lead to strain degeneration. The degenerated strains usually have a decrease in stress resistance, yield, and an alteration in fruiting time, which will subsequently result in tremendous economic loss. Through proteomic analysis, we identified the differentially expressed proteins (DEPs) in the mycelium of Pleurotus ostreatus from different subcultured generations. We found that the DNA damage repair system, especially the double-strand breaks (DSBs), repairs via homologous recombination, was impaired in the subcultured mycelium, and gradual accumulation of the DSBs would lead to the strain degeneration after successive subculture. The TUNEL assay further confirmed our finding about the DNA breaks in the subcultured mycelium. Interestingly, the enzyme activity of laccase, carboxylic ester hydrolase, α-galactosidase, and catalase directly related to passage number could be used as the characteristic index for strain degeneration determination. Our results not only reveal for the first time at the molecular level that genomic instability is the cause of degeneration, but also provide an applicable approach for monitoring strain degeneration in process of edible fungi cultivation and production.

Pleurotus ostreatus is a widely cultivated edible fungus around the world. At present, studies on the developmental process of the fruiting body are limited. In our study, we compared the differentially expressed proteins (DEPs) in the stipe and cap of the fruiting body by high-throughput proteomics. GO and pathway analysis revealed the great differences in the metabolic levels, including sucrose and starch metabolism, and sphingolipid signaling and metabolism, and the differences of 16 important DEPs were validated further by qPCR analysis in expression level. In order to control the cap and stipe development, several chemical inducers were applied to the primordium of the fruiting body according to the pathway enrichment results. We found that CaCl₂ can affect the primordium differentiation through inhibiting the stipe development. EGTA (ethyleneglycol bis (β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid) treatment confirmed the inhibitory role of Ca²⁺ in the stipe development. Our study not only shows great metabolic differences during the cap and stipe development but also reveals the underlying mechanism directing the primordium differentiation in the early development of the fruiting body for the first time. Most importantly, we provide a reliable application strategy for the cultivation and improvement of the Pleurotus ostreatus, which can be an example and reference for a more edible fungus.

Background: Strains XG04 and XGT2 of Lentinus edodes (Berk.) Singer demonstrate a high degree of genomic similarity, with XGT2 representing a systematic selection of XG04 and exhibiting enhanced phenotypic traits. Methods: An investigation into the differences between these strains was conducted using untargeted metabolomics to identify potential causal factors. Five exogenous inducers were assessed for their relationship with the observed phenotypes, and their impacts on fruiting body characteristics were analyzed. Results: Notably, the exogenous inducer rolipram, at a concentration of 0.4%, was found to increase cAMP expression levels in L. edodes primordia, which subsequently affected gill development, leading to the formation of gill-free fruiting bodies. Morphological differences between the two strains were evident; XG04 exhibited a spherical morphology with absent gills, rendering it commercially unviable, whereas XGT2 displayed a thicker cap and a more robust stipe, maintaining its characteristic umbrella shape. Conclusions: As the concentration of rolipram increased, both cap retraction and gill reduction in XGT2 occurred in a dose-dependent manner. The endogenous cAMP levels in the fruiting bodies were measured before and after rolipram treatment, revealing that the cap retraction and gill reduction in XGT2 progressed in a dose-dependent manner alongside increasing cAMP expression levels. Furthermore, a positive correlation was observed between cAMP levels and rolipram concentration. This study provides a foundation for improving the quality and productivity of mushroom cultivation by manipulating fruiting body characteristics through external stimuli.