Exploring the diversity, bioactivity of endophytes, and metabolome in Synsepalum dulcificum

Abstract

Synsepalum dulcificum exhibits high edible and medicinal value; however, there have been no reports on the exploration of its endophyte resources. Here, we conducted analyses encompassing plant metabolomics, microbial diversity, and the biological activities of endophytic metabolites in S. dulcificum. High-throughput sequencing identified 4,913 endophytic fungal amplicon sequence variants (ASVs) and 1,703 endophytic bacterial ASVs from the roots, stems, leaves, flowers, and fruits of S. dulcificum. Fungi were classified into 5 phyla, 24 classes, 75 orders, 170 families, and 313 genera, while bacteria belonged to 21 phyla, 47 classes, 93 orders, 145 families, and 232 genera. Furthermore, there were significant differences in the composition and content of metabolites in different tissues of S. dulcificum. Spearman's correlation analysis of the differential metabolites and endophytes revealed that the community composition of the endophytes correlated with plant-rich metabolites. The internal transcribed spacer sequences of 105 isolates were determined, and phylogenetic analyses revealed that these fungi were distributed into three phyla (Ascomycota, Basidiomycota, and Mucoromycota) and 20 genera. Moreover, 16S rDNA sequencing of 46 bacteria revealed they were distributed in 16 genera in three phyla: Actinobacteria, Proteobacteria, and Firmicutes. The antimicrobial activities (filter paper method) and antioxidant activity (DPPH and ABTS assays) of crude extracts obtained from 68 fungal and 20 bacterial strains cultured in different media were evaluated. Additionally, the α-glucosidase inhibitory activity of the fungal extracts was examined. The results showed that 88.6% of the strains exhibited antimicrobial activity, 55.7% exhibited antioxidant activity, and 85% of the fungi exhibited α-glucosidase inhibitory activity. The research suggested that the endophytes of S. dulcificum are highly diverse and have the potential to produce bioactive metabolites, providing abundant species resources for developing antibiotics, antioxidants and hypoglycemic drugs.

Keywords: Synsepalum dulcificum; biological activities; diversity; endophytes; metabolomics; microbiome.

FIGURE 1

Pictures of selected plants from Xishuangbanna.

FIGURE 2

PLS-DA (A), PCA of metabolome data (B) and KEGG pathway enriched dotplot (C).

FIGURE 3

PLS-DA and PCA analysis of uncultured fungi (A,B) and bacteria (C,D) (the same group is displayed in ellipses with a 95% confidence interval).

FIGURE 4

Venn diagram of uncultured endophytes identical and unique ASVs numbers statistics (A): fungi, (B): bacteria.

FIGURE 5

Mean relative abundance of the abundant phylum, class, family, and genus in uncultured fungi (A,C,E,G) and bacteria (B,D,F,H).

FIGURE 6

Species composition of endophytes from S. dulcificum (the number represents the count of isolated strains).

FIGURE 7

Heatmap of antimicrobial activity of strain extracts against pathogenic microorganisms (A): fungi; (B): bacteria (different colors represent varying sizes of inhibition circles, with a deeper red hue indicating a larger diameter of the inhibition circle).

FIGURE 8

Violin plot of antioxidant activity of endophyte extracts (A): fungi; (B): bacteria (the horizontal line inside the box represents the median of free radical scavenging rate; the upper and lower edges of the box represent the upper and lower quartiles; the top and bottom of the vertical line represent the maximum and minimum values).

FIGURE 9

Violin plot of α-glucosidase inhibitory activity of fungal extracts (the black horizontal line inside the box represents the median of α-glucosidase inhibition rate; the upper and lower edges of the white box indicate the upper and lower quartiles; the top and bottom ends of the vertical line represent the maximum and minimum values).