The Growth and Conidiation of Purpureocillium lavendulum Are Co-Regulated by Nitrogen Sources and Histone H3K14 Acetylation

**Background:** Plant-parasitic nematodes cause hundreds of billions of dollars in annual crop losses worldwide. Nematophagous fungi like Purpureocillium lavendulum are natural biocontrol agents that infect nematodes, with conidia (asexual spores) serving as key infectious structures. While nitrogen starvation is known to induce conidiation in fungi, the molecular mechanisms—particularly epigenetic regulation—linking nitrogen availability to growth versus reproduction in carnivorous fungi remain poorly understood. This study investigates how nitrogen sources and histone H3K14 acetylation co-regulate vegetative growth and conidiation in P. lavendulum.

**Methods:** The whole genome of P. lavendulum YMF1.00683 was sequenced using Illumina (3.2 Gb, 70× coverage) and PacBio (2.89 Gb, 60× coverage) platforms, assembled into 21 contigs (46 Mb total, N50=4.2 Mb), with 8,862 predicted genes. The ku80 strain (high-efficiency homologous recombination background) was used as the parent strain. Phenotypic comparisons were performed on PDA (high C/N ratio) and MM (minimal medium with Vogel's salts) media, with systematic addition of individual Vogel's components to identify key nutrients. Site-directed mutagenesis created histone H3K14R (non-acetylatable) and H3K14Q (acetylation-mimicking) mutants. The histone acetyltransferase PlGCN5 was identified by local BLAST and knocked out via homologous recombination (chlorimuron-ethyl resistance marker). H3K14ac levels were assessed by western blot. ChIP-seq using anti-H3K14ac antibody was performed on mycelia cultured in PDB (nitrogen-poor) and PDB+1% (NH₄)₂SO₄ (nitrogen-supplemented) liquid media for 5 days. The TOR pathway was investigated using rapamycin (150 ng/mL) inhibition.

**Key Results:** (1) On PDA medium, the ku80 strain showed limited colony extension but abundant conidia production, with satellite colonies forming from dispersed conidia. On MM medium, colonies were significantly larger with reduced conidiation. (2) NH₄NO₃ was identified as the key component in Vogel's salts promoting growth. Both (NH₄)₂SO₄ and KNO₃ independently increased colony size in a concentration-dependent manner. Carbon sources had minimal effect on conidiation. (3) The H3K14R mutant showed significantly increased colony size and decreased conidiation on PDA compared to ku80, resembling ku80 grown on MM. H3K14ac was detected in ku80 but not in H3K14R or H3K14Q strains by western blot. H3K14R showed no significant phenotypic difference across the four media tested, indicating that nitrogen regulation depends on H3K14 modification. (4) ΔPlgcn5 showed reduced H3K14ac levels, slower growth, smaller colonies, and sharply reduced conidiation—phenocopying H3K14Q. (5) ChIP-seq identified 4,856 peaks in PDB samples (67.52% in promoter-TSS regions) and 2,331 peaks in PDB+N samples (62.27% in intergenic regions), indicating nitrogen-dependent redistribution of H3K14ac. A total of 4,994 genes were associated with H3K14ac peaks (4,146 in PDB, 1,256 in PDB+N, 426 shared). KEGG enrichment showed MAPK signaling pathway genes (33 genes in PDB, 8 in PDB+N) and conidiation core regulators (brlA, abaA, wetA, fluG) were associated with H3K14ac, with higher acetylation levels under nitrogen starvation. (6) Rapamycin inhibited growth of all strains but did not alter the differential growth pattern between H3K14R and ku80 under nitrogen-poor versus nitrogen-rich conditions, suggesting TOR signaling acts upstream of the nitrogen-epigenetic regulation axis.

**Clinical Implications:** While this is a basic science study rather than a clinical investigation, the findings have direct relevance to agricultural biotechnology. Understanding how nitrogen availability and histone acetylation regulate conidiation in P. lavendulum could enable optimization of mass production of fungal spores for biocontrol formulations. The identification of PlGCN5 and H3K14ac as key regulators of conidiation provides molecular targets for genetic engineering to enhance spore yield and virulence of this nematophagous fungus against plant-parasitic nematodes, potentially reducing reliance on chemical nematicides.