The natural product citral has previously been demonstrated to possess antifungal activity against Magnaporthe oryzae. The purpose of this study was to screen and annotate genes that were differentially expressed (DEGs) in M. oryzae after treatment with citral using RNA sequencing (RNA-seq). Thereafter, samples were reprepared for quantitative real-time PCR (RT-qPCR) analysis verification of RNA-seq data. The results showed that 649 DEGs in M. oryzae were significantly affected after treatment with citral (100 μg/mL) for 24 h. Kyoto Encyclopedia of Genes and Genomes (KEGG) and a gene ontology (GO) analysis showed that DEGs were mainly enriched in amino sugar and nucleotide sugar metabolic pathways, including the chitin synthesis pathway and UDP sugar synthesis pathway. The results of the RT-qPCR analysis also showed that the chitin present in M. oryzae might be degraded to chitosan, chitobiose, N-acetyl-D-glucosamine, and β-D-fructose-6-phosphate following treatment with citral. Chitin degradation was indicated by damaged cell-wall integrity. Moreover, the UDP glucose synthesis pathway was involved in glycolysis and gluconeogenesis, providing precursors for the synthesis of polysaccharides. Galactose-1-phosphate uridylyltransferase, which is involved in the regulation of UDP-α-D-galactose and α-D-galactose-1-phosphate, was downregulated. This would result in the inhibition of UDP glucose (UDP-Glc) synthesis, a reduction in cell-wall glucan content, and the destruction of cell-wall integrity.
Gray blight disease is a serious disease of tea (Camellia sinensis (L.) Kuntz), for which there is currently no effective control or preventative measure apart from fungicides. Screening for effectiveness of a natural anti-microbial against this pathogen and identifying its mode of action could contribute to the management of this disease. Antifungal activity of the anti-microbial Ningnanmycin (NNM) from Streptomyces noursei var. xichangensis against the pathogen causing gray blight disease, Pseudopestalotiopsis camelliae-sinensis (F. Liu & L. Cai) strain GZHS-2017-010 was confirmed in vitro by the mycelial growth rate method. Optical microscopy, scanning electron microscopy and transmission electron microscopy were used to observe morphological change of hyphae of Ps. camelliae-sinensis treated with NNM. RNA sequencing, bioinformatics and quantitative real-time PCR were used to identify genes in the hyphae which were differentially expressed in response to treatment with NNM. Thirty-eight genes from sixteen pathways, known as targets of antifungal agents, were used to investigate gene expression in hyphae at the half-maximal effective concentration (EC50) dosage, EC30, and EC70 for 1, 7, or 14 h. The results indicated that NNM can inhibit the growth of hyphae in vitro, with an EC50 of 75.92 U/mL, inducing morphological changes in organelles, septa, and extracellular polysaccharides, targeting ribosomes to disturb translation in protein synthesis, and influencing some biosynthetic functions of the hyphae.
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