The endophyte Fusarium oxysporum 3RR genome analysis identifies bioactive compounds targeting biofilm formation

Abstract

Introduction The rising global challenge of antimicrobial resistance and biofilm-associated infections necessitates the discovery of novel bioactive compounds. Endophytic fungi, recognized for their metabolic versatility and ecological adaptability, represent a promising, yet underexplored, source of such molecules. This study aimed to comprehensively characterize the biosynthetic potential of Fusarium oxysporum strain 3RR and establish mechanistic links between its genomic architecture, enzymatic capacity, and observed antibiofilm and antibacterial activities. Methods An integrated genomics-metabolomics approach was employed. Whole-genome sequencing combined Illumina paired-end and Oxford Nanopore long-read technologies, with hybrid assembly via SPAdes. Gene prediction was performed using BRAKER3 followed by functional annotation via InterProScan, eggNOG-mapper, while biosynthetic gene clusters (BGCs) were predicted using antiSMASH v8.0.4. Antibacterial activity was assessed by agar disk diffusion and microdilution, and antibiofilm activity by crystal violet quantification and microscopy. The extract composition was analyzed using Gas Chromatography–Mass Spectrometry (GC-MS). Results The assembled genome (41.14 Mb, N50 = 3.52 Mb) was annotated, and predicted to contain 11,738 genes. 36 BGCs were identified, predominantly terpenes, NRPS, and PKS clusters. CAZy analysis revealed a notable CE10 esterase expansion. GC-MS identified 14 major compounds in the extracellular ethyl acetate extract, dominated by 2,4-di-tert-butylphenol (2,4-DTBP, 16.72%), oleic acid (10.14%), and 8-hydroxyisotrichodermin (7.79%). The extract exhibited antibacterial activity against Staphylococcus epidermidis (up to 34 mm inhibition zone) and Escherichia coli, and remarkable antibiofilm effects, inhibiting S. epidermidis biofilm formation by 96.11% and eradicating Staphylococcus aureus biofilms by up to 54.85%, confirmed microscopically. Discussion The study reveals a sophisticated metabolic architecture in F. oxysporum 3RR, driven by CAZy enzymes and secondary metabolite BGCs. CE10 may act as a metabolic interface by facilitating the liberation or transformation of phenolic and lipid-derived compounds, thereby increasing their potential bioavailability. Rather than attributing the observed bioactivity to 2,4-DTBP or oleic acid alone, the present work places these known metabolites within their genomic and ecological context, linking them to the strain’s biosynthetic capacity and its plant-endophytic lifestyle. Conclusion F. oxysporum 3RR possesses a powerful chemical defense and offense system, positioning this endophytic fungus as a promising source for novel therapeutic agents against microbial biofilms.