Strain degradation is a common problem in many artificially-cultivated edible mushrooms. As a fungus with poor tolerance to low-temperature, Volvariella volvacea cannot delay its degradation by long-term low temperature storage like other fungi, so its degradation is particularly severe, which hinders industrial applications. Periodic mycelial subculture is a common storage method for V. volvacea, but excessive subculturing can also lead to strain degeneration. After 20 months of continuous subculturing every 3 days, V. volvacea strains S1–S20 were obtained, and their characteristics throughout the subculture process were analyzed. With increasing number of subculture, the growth rate, mycelial biomass, the number of fruiting bodies and biological efficiency gradually decreased while the production cycle and the time to primordium formation was lengthened. Strains S13–S20, obtained after 13–20 months of mycelial subculturing, also lacked the ability to produce fruiting bodies during cultivation experiments. Determination of reactive oxygen species (ROS) content as well as enzyme activity showed that decreased lignocellulase activity, along with excessive accumulation of ROS, was concomitant with the subculture-associated degeneration of V. volvacea. Reverse transcription polymerase chain reaction (RT-PCR) was eventually used to analyze the gene expression for lignocellulase and antioxidant enzymes in subcultured V. volvacea strains, with the results found to be consistent with prior observations regarding enzyme activities. These findings could form the basis of further studies on the degeneration mechanism of V. volvacea and other fungi.

Periodic mycelial subculture is a method commonly used for the storage of edible mushrooms, but excessive subculturing can lead to the degeneration of strains. In this study, the Volvariella volvacea strain V971(M0) was successively subcultured on PDA medium every 4 days, and one generation of strains was preserved every 4 months. Thus, five generations of subcultured strains (M1–M5) were obtained after 20 months of mycelial subculturing, their production traits were determined, and transcriptomic analysis was performed using RNA-seq; the differentially expressed genes were verified via RT-qPCR. The results showed that as the number of subcultures increased, the diameter of the mycelium and biological efficiency gradually decreased; in addition, the time in which the primordium formed increased and the production cycle was lengthened, while strains M4 and M5 lacked the ability to produce fruiting bodies. There were 245 differentially expressed genes between the M1–M5 and M0 strains, while the highest number of differentially expressed genes was between M3 and M0, at 1439; the smallest number of differentially expressed genes was between M2 and M0, at 959. GO enrichment analysis showed that the differentially expressed genes were mainly enriched in metabolic processes, organelle components, and catalytic activities. KEGG enrichment analysis showed that the differentially expressed genes were mainly enriched in metabolic pathways. The further annotation of differentially expressed genes showed that 39, 24, and 24 differentially expressed genes were related to substrate degradation, amino acid synthesis and metabolism, and reactive oxygen species metabolism, respectively. The downregulation of the related differentially expressed genes would lead to the excessive accumulation of reactive oxygen species, inhibit nutrient absorption and energy acquisition, and lead to the degradation of V. volvacea. These findings could form a theoretical basis for the degeneration mechanism of V. volvacea, and also provide a basis for the molecular function study of the genes related to strain degradation.

Manganese is one of the trace elements necessary for organisms to maintain normal biological activities and is also a cofactor for manganese superoxide dismutase (Mn-SOD) and manganese peroxidase (MnP). In order to find a simple and effective method to rejuvenate the degenerated V. volvacea strains, we explored the effect of the exogenous addition of MnSO₄ on the antioxidant vigour and productivity of degenerated strains of V. volvacea. The results showed that the exogenous MnSO₄ had no significant effect on the non-degenerated strain T0, but it could effectively increase the mycelial growth rate, mycelial biomass, and LBL decolouring ability of the degenerated strains T10 and T19, and reduce the production cycle and increased the biological efficiency of T10; it helped the severely degenerated T19 to regrow its fruiting body; and it also significantly increased the viability of the matrix-degrading enzymes such as EG, Lac, Xyl, etc. of T10 and T19. Meanwhile, exogenous MnSO₄ significantly increased the activity of GPX, GR, CAT, SOD, and the content of GSH, polyphenols, minerals, and polysaccharides in T10 and T19 strains, which resulted in a significant decrease in the accumulation of ROS, such as O₂⁻ and H₂O₂ in T10 and T19. The correlation analysis showed that there was a significant correlation between antioxidant activity and the production ability of V. volvacea. This study can provide theoretical reference and technical support for the rejuvenation research of degenerated strains of V. volvacea and other edible fungi.

Stipe elongation is one of the main characteristics of the postharvest quality deterioration of Flammulina velutipes fruiting bodies. In order to reduce the postharvest stipe elongation of F. velutipes and prolong its shelf life, in this study, using polyethylene (PE) packaging and unpackaged treatments as controls, the effects of a polyethylene nano-packaging on postharvest stipe elongation of F. velutipes were investigated and its regulatory mechanisms were explored from the physiological and biochemical aspects. The results showed that the CO₂ content in polyethylene nano-packaging boxes was higher than in control boxes, and the O₂ content was lower than in the control group, indicating that nano-packaging could reduce F. velutipes respiration during low-temperature storage. The stipe elongation rate, chitinase activity, and β-1,3-glucanase activities of F. velutipes were lower in the nano-packaging group than in PE-packaged and unpackaged control groups, and the nano-packaging inhibited the increase in chitin and β-glucan, the main components of the cell wall. The levels of auxin (IAA), abscisic acid (ABA), gibberellin (GA), and cytokinin (CTK) were also lower in the nano-packaged group than in controls at most timepoints. After 18 d of storage, polyethylene nano-packaging inhibited the stipe elongation of fruiting bodies, and maintained mushroom quality, with stipe elongation of only 34.7% and 76.7% of PE-packaged and unpackaged control groups, respectively. The results showed that nano-packaging could effectively inhibit the harvest stipe elongation and prolong the shelf quality of F. velutipes.