# Genomic and Metabolomic Analysis of the Endophytic Fungus Alternaria alstroemeriae S6 Isolated from Veronica acinifolia: Identification of Anti-Bacterial Properties and Production of Succinic Acid

## TL;DR

This study explores a fungus found in a plant, showing it can produce antibacterial compounds and succinic acid, which could be useful in biotechnology.

## Contribution

The discovery of A. alstroemeriae S6's antibacterial properties and its potential as a succinic acid producer is novel.

## Key findings

- The fungal extract showed strong antibacterial activity against both Gram-positive and Gram-negative bacteria.
- Genome analysis revealed 58 biosynthetic gene clusters, including pathways for peptides, terpenes, and polyketides.
- Succinic acid was identified as the major metabolite, with 20 minor bioactive compounds also detected.

## Abstract

Background: Endophytic fungi are prolific sources of bioactive metabolites with potential in pharmaceutical and biotechnological applications. Methods: Here, the endophytic fungus, Alternaria alstroemeriae S6, was isolated from Veronica acinifolia (speedwell), and conducted its anti-microbial activities, whole-genome sequencing and metabolome analysis. Results: The ethyl acetate extract of this fungus exhibited strong anti-bacterial activity and the inhibition zones, induced by the fungal extract at 20 mg/mL, reached 16.25 ± 0.5 mm and 26.5 ± 0.5 mm against Gram-positive and Gram-negative bacteria. To unravel the biosynthetic potential for anti-bacterial compounds, whole-genome sequencing was conducted on A. alstroemeriae S6, resulting in a high-quality assembly of 42.93 Mb encoding 13,885 protein-coding genes. Comprehensive functional genome annotation analyses, including gene ontology (GO) terms, clusters of orthologous groups (COGs), Kyoto encyclopedia of genes and genomes (KEGG), carbohydrate-active enzymes (CAZymes), and antibiotics and secondary metabolites analysis shell (antiSMASH) analyses, were performed. According to the antiSMASH analysis, 58 biosynthetic gene clusters (BGCs), including 16 non-ribosomal peptide synthetases (NRPSs), 21 terpene synthases, 12 polyketide synthetases (PKSs), and 9 hybrids, were identified. In addition, succinic acid was identified as the major metabolite within the fungal extract, while 20 minor bioactive compounds were identified through LC-MS/MS-based molecular networking on a GNPS database. Conclusions: These findings support the biotechnological potential of A. alstroemeriae S6 as an alternative producer of succinic acid, as well as novel anti-bacterial agents.