Fungal endophytes are the most ubiquitous plant symbionts on earth and are phylogenetically diverse. Studies on the fungal endophytes in tobacco have shown that they are widely distributed in the leaves, stems, and roots, and play important roles in the composition of the microbial ecosystem of tobacco. Herein, we analyzed and quantified the endophytic fungi of healthy tobacco leaves at the seedling stage (SS), resettling growth stage (RGS), fast-growing stage (FGS), and maturing stage (MS) at three altitudes (600, 1000, and 1300 m). We sequenced the internal transcribed spacer (ITS) region of fungal samples to delimit operational taxonomic units (OTUs) and phylogenetically characterize the communities. The results showed that the numbers of clustering OTUs at SS, RGS, FGS, and MS were 516, 709, 469, and 428, respectively. At the phylum level, species in Ascomycota and Basidiomycota had absolute predominance, representing 97.8% and 2.0% of the total number of species, respectively. We also found the number of fungi at the RGS and FGS stages was higher than those at the other two stages. Additionally, OTU richness was determined by calculating the Observed Species, Shannon, Simpson, Chao1, abundance-based coverage estimator (ACE), Good’s coverage and phylogenetic distance (PD)_whole_tree indices based on the total number of species. Our results showed RGS samples had the highest diversity indices. Furthermore, we found that the diversity of fungal communities tended to decrease with increasing altitude. The results from this study indicated that tobacco harbors an abundant and diverse endophytic fungal community, which provides new opportunities for exploring their potential utilization.
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