Fusarium head blight (FHB) caused by Fusarium graminearum is a devastating disease of cereal crops worldwide. Recently, a novel fungicide JS399-19 has been launched into the marketplace to manage FHB. It is compelling that JS399-19 shows highly inhibitory activity towards some Fusarium species, but not to other fungi, indicating that it is an environmentally compatible fungicide. To explore the mode of action of this species-specific compound, we conducted a whole-genome transcript profiling together with genetic and biochemical assays, and discovered that JS399-19 targets the myosin I of F. graminearum (FgMyo1). FgMyo1 is essential for F. graminearum growth. A point mutation S217L or E420K in FgMyo1 is responsible for F. graminearum resistance to JS399-19. In addition, transformation of F. graminearum with the myosin I gene of Magnaporthe grisea, the causal agent of rice blast, also led to JS399-19 resistance. JS399-19 strongly inhibits the ATPase activity of the wild-type FgMyo1, but not the mutated FgMyo1(S217L/E420K) . These results provide us a new insight into the design of species-specific antifungal compounds. Furthermore, our strategy can be applied to identify novel drug targets in various pathogenic organisms.

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MBC(R) isolates have high fitness and competition in field, conferred by a novel molecular mechanism.

Spray-induced gene silencing (SIGS) is an innovative strategy for crop protection. However, the mechanism of SIGS is not known. Here, we first demonstrate that secondary small interfering RNA (siRNA) amplification limits the application of SIGS. A myosin5 gene (Myo5) was chosen as the target of SIGS in an agronomically important pathogen-Fusarium asiaticum. Five segments corresponding to the different regions of the Myo5 gene were found to efficiently silence Myo5, resulting in cell wall defects, life cycle disruption and virulence reduction. Myo5-8 (one of the Myo5 segments) induced sequence-specific RNA interference (RNAi) activity in F. asiaticum, F. graminearum, F. tricinctum and F. oxysporum, but not in other fungi, in vitro. Remarkably, the silencing of Myo5 lasted for only 9 h unless the double-stranded RNA (dsRNA) was continuously supplied, because F. asiaticum is unable to maintain siRNA amplification. After spraying on plants, dsRNAs were more efficiently taken up via the wounded surface. The antifungal activity of dsRNAs taken up by plant cells was higher and longer lasting than that dried onto the plant surface. In contrast with dsRNAs in fungi, dsRNAs in plant cells could efficiently turn into substantial siRNAs via secondary amplification machinery. Our findings provide new implications to develop SIGS as a mainstream disease control strategy against Fusarium and other fungi.

To determine the mechanism of resistance to the fungicide phenamacril (JS399-19) in Fusarium graminearum, the causal agent of Fusarium head blight, we sequenced and annotated the genome of the resistant strain YP-1 (generated by treating the F. graminearum reference strain PH-1 with phenamacril). Of 1.4 million total reads from an Illumina-based paired-end sequencing assay, 92.80% were aligned to the F. graminearum reference genome. Compared with strain PH-1, strain YP-1 contained 1,989 single-nucleotide polymorphisms that led to amino acid mutations in 132 genes. We sequenced 22 functional annotated genes of another F. graminearum sensitive strain (strain 2021) and corresponding resistant strains. The only mutation common to all of the resistant mutants occurred in the gene encoding myosin-5 (point mutations at codon 216, 217, 418, 420, or 786). To confirm whether the mutations in myosin-5 confer resistance to phenamacril, we exchanged the myosin-5 locus between the sensitive strain 2021 and the resistant strain Y2021A by homologous double exchange. The transformed mutants with a copy of the resistant fragment exhibited resistance to phenamacril, and the transformed mutant with a copy of the sensitive fragment exhibited sensitivity to phenamacril. These results indicate that mutations in myosin-5 confers resistance to phenamacril in F. graminearum.