Elucidating the effects of volatile organic compounds on the inhibition of the phytopathogen Sporisorium scitamineum

Sugarcane holds substantial agricultural and industrial importance in Brazil and worldwide, primarily due to its use in biofuel production and for a range of biomolecules of diverse biochemical applications. The drive towards sustainable agricultural practices has fostered the interest in the exploration of bioproducts to reduce the dependence on agrochemicals. In this context, microbial volatile organic compounds (VOCs) have emerged as potential biofungicides, offering several advantages over conventional chemical treatments. This study focused on evaluating their antagonistic potential against Sporisorium scitamineum, the causative agent of sugarcane smut disease, and investigated the molecular mechanisms underlying their inhibitory effects. Two Pseudomonas bacterial strains isolated from sugarcane roots exhibited remarkable antagonistic activity against S. scitamineum. Notably, ITA P2F2 completely inhibited the growth of the phytopathogen and was identified as a novel species within the Pseudomonas genus. A total of 63 VOCs were identified from both bacterial isolates, with eight showing inhibitory activity against the phytopathogen. Transcriptomic analyses revealed significant changes in gene expression of S. scitamineum when exposed to VOCs from ITA P2F2, including the downregulation of genes involved in central metabolic pathways, such as carbohydrate metabolism and fatty acid ß-oxidation, and the upregulation of genes associated with DNA repair. Potential DNA damage caused by the VOCs was further evidenced through temporal analysis using Fourier Transform Infrared (FTIR) spectroscopy. Additionally, electron microscopy analysis showed structural damage to the fungal hyphae after VOCs exposure. This multidisciplinary study is the first to investigate the inhibition of S. scitamineum using VOCs. Our findings contribute to the molecular understanding of these signaling molecules and highlight their potential as promising bioproducts to replace conventional agrochemicals in the control of sugarcane pathogens. Overall design: To investigate changes in the gene expression of S. scitamineum and to identify potential genes involved in its growth inhibition, we conducted transcriptomic analyses using RNAseq. S. scitamineum samples were pre-growth for 5 days at 28°C. The collection of cellular pellets were performed after 2 days of co-culture with ITA P2F2, E coli DH5a, and with no VOC-treatment. We used 4 replicates per treatment. RNA extraction was performed with RNeasy Plant Mini Kit (Qiagen). Twelve libraries were prepared using the TruSeq Stranded mRNA Sample Prep LS Kit (Illumina) using 1.5 micrograms of total RNA as imput and sequenced on the Hiseq 2500 instrument.