Journal of Fungi

Species of the ascomycetous genus Cladobotryum (Hypocreales, Hypocreaceae) are ecologically and economically important mycoparasites that cause cobweb disease in cultivated and wild mushrooms. Despite their significance as fungal pathogens and producers of bioactive metabolites, the taxonomy of Cladobotryum remains unresolved due to extensive morphological plasticity, complex teleomorph–anamorph connections, and the presence of cryptic species. This study employs an integrative approach combining micro- and macromorphological characterization, multi-locus phylogeny (ITS, rpb2, and tef-1a), and comparative genomics to clarify the taxonomic position of the Greek isolate Cladobotryum sp. ATHUM 6904, previously designated as an unclassified red-pigmented (URP) strain. Phylogenetic analyses demonstrated that URP strains form a distinct, well-supported clade closely related to C. tenue and C. rubrobrunnescens, yet genetically and morphologically distinct from both. Comparative genomic analyses of isolate ATHUM 6904 and the ex-type strains of C. tenue and C. rubrobrunnescens revealed pronounced divergence in transposable element content, mitochondrial genome architecture, gene order, orthologous gene composition, secondary metabolite biosynthetic potential, and overall genomic distance. Micro- and macromorphological comparisons further supported the differentiation of isolate ATHUM 6904 from both reference species. Based on the combined molecular, morphological, and genomic evidence, the Greek isolate ATHUM 6904 is described as a novel species, Cladobotryum rhodochroum sp. nov.

Although RNA sequencing (RNA-seq) enables rapid transcriptome profiling, functional annotation of fungal transcriptomes remains challenging. Existing tools prioritize broad taxonomic coverage, and reference genomes are scarce for non-model species. This study aimed to develop a fungal-specific functional annotation workflow to support rapid and accurate functional analyses downstream of RNA-seq, independent of reference genome availability. To evaluate the workflow, RNA-seq data from 57 samples of Lentinula edodes strain H600 (shiitake mushroom) were retrieved, along with full-length transcript sequencing (Iso-Seq) data and corresponding RNA-seq data from 20 samples of Phakopsora pachyrhizi (Asian soybean rust) from public databases. The workflow successfully annotated over 96% of protein-coding transcripts and demonstrated applicability to Iso-Seq data. Functional enrichment analyses revealed higher-resolution functional detection than existing annotation tools. Furthermore, integrating homology searches against fungal-specific databases with expression pattern-based annotations highlighted the workflow’s utility for target identification in genome editing and other applications. Overall, the results of this study highlight the potential of the developed workflow in facilitating the discovery of functionally important transcripts and their translation into biotechnological applications.

Protein phosphorylation modification plays a role in cells’ response to oxidative stress, a key factor leading to postharvest browning of Flammulina filiformis. However, the molecular mechanism by which protein phosphorylation contributes to postharvest browning of F. filiformis remains unclear. This study aimed to characterize the basal phosphoproteomic landscapes associated with variations in different browning phenotypes of F. filiformis. Using data-independent acquisition (DIA) mass spectrometry, we comprehensively profiled the phosphorylation dynamics in susceptible-to-browning (SB) and resistant-to-browning (RB) cultivars at harvest and after 24 h storage. We identified 84,244 phosphorylation sites on 4494 phosphoproteins, with the SB cultivar displaying more altered sites (21,195) than the RB (16,087). Functional enrichment analysis revealed that the differential phosphorylation was significantly implicated in kinases and energy metabolism pathways. Notably, the SB cultivar exhibited a more pronounced phosphorylation profile on key proteins involved in ATP synthesis and glycolysis. Protein–protein interaction (PPI) network analysis further indicated a kinase-mediated regulatory network targeting core energy metabolism components, including ATP synthase and 6-phosphofructokinase. This distinct phosphosignature in the SB cultivar correlated with its more severe browning phenotype and a sharper decline in ATP content during storage. Our findings suggest that divergent phosphorylation-mediated regulation of energy metabolism is strongly associated with the differential postharvest browning susceptibility between these two cultivars, providing a valuable molecular resource for future functional studies.