γ-Aminobutyric (GABA) acid is a nutrient and signaling molecule existing in many plants, participating in the regulation of metabolism and various physiological activities. Two strains of Hypsizygus marmoreus (a white variety and a brown variety) were investigated to study the impact of exogenous GABA on mycelial growth and the response to stress. Mycelial growth, microscopic morphology, antioxidant profile, and gad2 expression in H. marmoreu were investigated under salt, dehydration, or cold stress. The results indicated that 5 mM GABA stimulated mycelial growth under standard cultivation conditions, whereas GABA addition over 10 mM hindered the growth. Under salt, dehydration, or cold stress, treatment with 5 mM GABA significantly enhanced the mycelial growth rate and density of both H. marmoreus strains by promoting front hyphae branching. Meanwhile, the activities of key antioxidant enzymes such as peroxidase (POD), catalase (CAT), and ascorbate peroxidase (APX) were enhanced by GABA, thereby augmenting the defensive network against abiotic stress. Additionally, gad2 expression and GABA concentration were increased under abiotic stresses as a resistance regulation response. The exogenous addition of GABA strengthened the upregulation of gad2 expression and GABA production. These findings indicated that exogenously adding low concentrations of GABA effectively enhanced the mycelial growth and antioxidant profile of H. marmoreus, thereby improving its resistance against stresses.

A novel fungal immunomodulatory protein (FIP), identified as FIP-hma, was discovered in the genome of an edible mushroom Hypsizygus marmoreus. Bioinformatics analysis suggested FIP-hma contained the cerato-platanin (CP) conserved domain and was categorized into Cerato-type FIP. In phylogenetic analysis, FIP-hma was clustered into a new branch of the FIP family, displaying large system divergence from most of the other FIPs. The higher gene expression of FIP-hma was observed during the vegetative growth stages than that during the reproductive growth stages. In addition, the cDNA sequence of FIP-hma was cloned and successfully expressed in Escherichia coli (E. coli) BL21(DE3). The recombinant protein of FIP-hma (rFIP-hma) was neatly purified and isolated by Ni-NTA and SUMO-Protease. The iNOS, IL-6, IL-1β, and TNF-α levels of RAW 264.7 macrophages were upregulated by rFIP-hma, indicating its activation of an immune response by regulating central cytokines. No cytotoxic effects were observed in an MTT test. The findings of this work discovered a novel immunoregulatory protein from H. marmoreus, provided a systematic bioinformatic profile, suggested an effective approach for its heterologous recombinant production, and reported its potent immunoregulatory activity in macrophages. This study sheds light on the physiological function research of FIPs and their further industrial utilization.

Postharvest Flammulina filiformis is prone to quality degradation, adversely impacting its commercial value. Cordyceps militaris, rich in antioxidant and antibacterial components, shows promise as a natural biological preservative. This study aimed to explore the potential of C. militaris extract (CME) as a preservative for F. filiformis. Through analyzing indicators such as browning, stipe elongation, and cap diameter, this study confirmed the effectiveness of CME in delaying oxidation and inhibiting microbial growth during storage. Additionally, transcriptome analysis revealed that CME modulated gene expression in F. filiformis, enhancing its antioxidant defense mechanisms. The results demonstrated that CME could effectively extend the shelf life of F. filiformis, providing valuable insights into preservation strategies for this and other edible fungi.

Core germplasm, a strategically selected subset of the original germplasm, aims to maximize the representation of genetic diversity within the entire collection. Establishing a germplasm resource bank is essential for the effective management and sustainable utilization of genetic resources. This study developed a core germplasm repository for Hypsizygus marmoreus, a commercially important mushroom species, to capture the genetic diversity of the original collection with a minimal sample size. Genetic diversity and cluster analyses were conducted on 57 representative strains of H. marmoreus, including both cultivated and wild accessions from different regions, using 15 pairs of simple sequence repeat (SSR) markers. DNA molecular identity cards were generated for all germplasms, and cultivation trials with agronomic trait assessments were performed on 24 core accessions. A total of 115 distinct alleles were identified, with genetic similarity coefficients ranging from 0.70 to 1.00. Clustering at a similarity threshold of 0.76 classified the strains into five groups. The core germplasm panel, comprising 24 accessions (42.11% of the total collection), retained full allelic diversity and preserved the genetic and phenotypic variability of the original population, confirming its suitability for parental selection in breeding programs. unique molecular identity codes were developed for each H. marmoreus germplasm by integrating SSR marker profiles with data on geographical origin, fruiting body color, and cultivation traits. These were converted into DNA molecular ID codes, providing a reliable system for rapid identification and traceability of germplasm resources. The findings offer a valuable reference for breeding improvement and the protection of edible fungal varieties with independent intellectual property rights.

Mitochondria regulate nuclear genomes and their own genetic material, primarily to provide energy in eukaryotes. Currently, high-throughput sequencing technologies are being used to resolve the mitochondrial genomes of various edible fungi. However, the application of pan-genomes for the analysis of edible mushroom mitochondrial genomes remains unexplored. In this study, we conducted a comparative mitochondrial genome analysis of 31 Hypsizygus marmoreus strains (four newly sequenced monotypes and 27 public datasets), ranging from 98,284 to 111,087 bp. This variation was determined to be primarily driven by dynamic changes in non-coding regions, particularly intronic polymorphisms in the cox1 gene. Further, transfer RNA (tRNA) secondary structures exhibited atypical globular and elongated conformations alongside copy number variations. Additionally, codon usage showed a pronounced A/T bias, whereas core respiratory chain genes demonstrated an evolutionary pattern of strong purifying selection. Furthermore, the 31 mitochondrial genomes of H. marmoreus were found to harbor eight gene rearrangement patterns and five genetic clusters, and the pan-genome analysis (220,364 bp, 217 nodes) captured abundant single-nucleotide polymorphisms (SNPs), insertions/deletions (InDels), and structural variations. This study provides breeding-relevant genetic markers and a genomic framework for H. marmoreus germplasm classification, genetic improvements, and the molecular breeding of stress-resilient varieties.