Age-dependent variations in leaf-endophytic microbiota and biocontrol potential in Camellia oleifera

Endophytic microorganisms play important roles in plant health, but their diversity and functions can vary with host developmental stage. Camellia oleifera, a major oil-producing crop in China, offers a valuable system for exploring these dynamics. We compared endophytic microbial communities in young (5-year) and old (15-year) C. oleifera leaves using high-throughput 16 S rRNA and ITS sequencing, supported by culture-dependent isolation and inhibition assays. Diversity indices, taxonomic composition, and predicted bacterial functions were analyzed with QIIME2 and PICRUSt2. Ten fungal and nine bacterial strains were isolated and tested for antagonism against key leaf pathogens in confrontation assays (n = 3 replicates). A total of 607 bacterial and 778 fungal ASVs were identified. Young leaves contained greater bacterial richness and diversity (Observed ASVs, Shannon, Simpson indices, P < 0.05), while old leaves hosted higher fungal richness. Beta diversity (Bray-Curtis PCoA) showed clear separation of bacterial communities by leaf age, with weaker separation for fungi. Bacterial functional prediction revealed enrichment of carbohydrate metabolism and amino acid biosynthesis in young leaves, while secondary metabolism and stress response pathways were more prominent in old leaves. Among isolated strains, Coniochaeta velutina inhibited Colletotrichum gloeosporioides, Alternaria alternata, and Botryosphaeria dothidea by 67.4%, 54.8%, and 65.2%, respectively. The bacterium Burkholderia ambifaria achieved the highest inhibition rate of 87.2% against B. dothidea. These findings suggest that leaf age shapes endophytic microbial diversity and bacterial functional potential in C. oleifera. Moreover, C. velutina and B. ambifaria represent promising candidates for biocontrol applications. While limited by small sample size (n = 3 replicates), this exploratory study provides foundational insights into age-associated shifts in endophyte communities and their biocontrol potential.