Many fungi-infecting viruses, which are termed mycoviruses, have been identified, and most do not cause any visible symptoms. Some mycoviruses, however, can attenuate the virulence of the infected fungi, a phenomenon referred to as hypovirulence. To study fungus responses to virus infection, we established a model system composed of Fusarium graminearum and four mycoviruses including FgV1 (Fusarium graminearum virus 1), FgV2, FgV3, and FgV4. FgV1 and FgV2 infections caused several phenotypic alterations in F. graminearum including abnormal colony morphology, defects in perithecium development, and reductions in growth rate, conidiation, and virulence. In contrast, FgV3 and FgV4 infections did not cause any phenotypic change. An RNA-Seq-based analysis of the host transcriptome identified four unique Fusarium transcriptomes, one for each of the four mycoviruses. Unexpectedly, the fungal host transcriptome was more affected by FgV1 and FgV4 infections than by FgV2 and FgV3 infections. Gene ontology (GO) enrichment analysis revealed that FgV1 and FgV3 infections resulted in down-regulation of host genes required for cellular transport systems. FgV4 infection reduced the expression of genes involved in RNA processing and ribosome assembly. We also found 12 genes that were differentially expressed in response to all four mycovirus infections. Unfortunately, functions of most of these genes are still unknown. Taken together, our analysis provides further detailed insights into the interactions between mycoviruses and F. graminearum.

No abstract

There is increasing concern regarding the use of fungicides to control plant diseases, whereby interest has increased in the biological control of phytopathogenic fungi by the application of hypovirulent mycoviruses as a possible alternative to fungicides. Transmission of hypovirulence-associated double-stranded RNA (dsRNA) viruses between mycelia, however, is prevented by the vegetative incompatibility barrier that often exists between different species or strains of filamentous fungi. We determined whether protoplast fusion could be used to transmit FgV1-DK21 virus, which is associated with hypovirulence on F. boothii (formerly F. graminearum strain DK21), to F. graminearum, F. asiaticum, F. oxysporum f. sp. lycopersici, and Cryphonectria parasitica. Relative to virus-free strains, the FgV1-DK21 recipient strains had reduced growth rates, altered pigmentation, and reduced virulence. These results indicate that protoplast fusion can be used to introduce FgV1-DK21 dsRNA into other Fusarium species and into C. parasitica and that FgV1-DK21 can be used as a hypovirulence factor and thus as a biological control agent.

bThe accumulation of viral RNA depends on many host cellular factors. The hexagonal peroxisome (Hex1) protein is a fungal protein that is highly expressed when the DK21 strain of Fusarium graminearum virus 1 (FgV1) infects its host, and Hex1 affects the accumulation of FgV1 RNA. The Hex1 protein is the major constituent of the Woronin body (WB), which is a peroxisome-derived electron-dense core organelle that seals the septal pore in response to hyphal wounding. To clarify the role of Hex1 and the WB in the relationship between FgV1 and Fusarium graminearum, we generated targeted gene deletion and overexpression mutants. Although neither HEX1 gene deletion nor overexpression substantially affected vegetative growth, both changes reduced the production of asexual spores and reduced virulence on wheat spikelets in the absence of FgV1 infection. However, the vegetative growth of deletion and overexpression mutants was increased and decreased, respectively, upon FgV1 infection compared to that of an FgV1-infected wild-type isolate. Viral RNA accumulation was significantly decreased in deletion mutants but was significantly increased in overexpression mutants compared to the viral RNA accumulation in the virus-infected wild-type control. Overall, these data indicate that the HEX1 gene plays a direct role in the asexual reproduction and virulence of F. graminearum and facilitates viral RNA accumulation in the FgV1-infected host fungus.T he interactions between viral elements and host factors are important for maintaining the infection cycles of RNA viruses in host cells. Viruses utilize numerous host factors that play essential roles in virus infection. Therefore, understanding the role(s) of host factors can provide insight into the molecular mechanism(s) of host and virus interactions. Relative to host factors affecting RNA viruses of animals and plants, host factors affecting fungal viruses (mycoviruses) are poorly understood.Several host and viral components required for virus life in cells have been identified and characterized in the model organism Saccharomyces cerevisiae. The knowledge obtained by studying S. cerevisiae as a host for several double-stranded RNA (dsRNA) and single-stranded RNA (ssRNA) viruses has greatly extended our understanding of mycovirus-host interaction (1). With respect to filamentous fungi, the host factors required for mycovirus replication and symptom induction have been well described for the interaction between the prototypic hypovirus Cryphonectria hypovirus 1 strain EP713 (CHV1) and its host, the chestnut blight fungus (Cryphonectria parasitica). One of these host factors, NAM-1, modulates symptom induction in the fungus in response to CHV1 infection (2). The hypovirus-responsive host transcription factor gene pro1 is required for female fertility of C. parasitica, development of its asexual spores, and the maintenance of CHV1 infection (3). The host gene Cpbir1 (bir1 of C. parasitica), which encodes the IAP (inhibitor of apoptosis protein) CpBir1 and which is required for fungal...

The complete genomes two different dsRNA mycoviruses, Fusarium graminearum virus 3 (FgV3) and Fusarium graminearum virus 4 (FgV4), was sequenced and analyzed. The viral genome of FgV3 is 9,098 base pairs (bp) long and contains two open reading frames (ORF) encoding a putative RNA-dependent RNA polymerase (RdRp) and a protein of unknown function. The FgV4 genome is composed of two dsRNA genome segments of 2,383 bp and 1,739 bp. FgV4 dsRNA-1 contains a single ORF, which has a conserved RdRp motif, while FgV4 dsRNA-2 contains two putative ORFs coding for products of unknown function. Both the genome organization and phylogenetic analysis indicated that FgV3 was closely related to members of the families Totiviriridae and Chrysoviridae, but it was placed outside of their main clusters, whereas FgV4 formed a distinct clade with the family Partitiviridae. This is the first report of the full-length nucleotide sequences of FgV3 and FgV4 infecting Fusarium graminearum.

The mechanisms of RNA interference (RNAi) as a defense response against viruses remain unclear in many plant-pathogenic fungi. In this study, we used reverse genetics and virus-derived small RNA profiling to investigate the contributions of RNAi components to the antiviral response against Fusarium graminearum viruses 1 to 3 (FgV1, -2, and -3). Real-time reverse transcription-quantitative PCR (qRT-PCR) indicated that infection of by FgV1, -2, or -3 differentially induces the gene expression of RNAi components in Transcripts of the and genes of ( and ) accumulated at lower levels following FgV1 infection than following FgV2 or FgV3 infection. We constructed gene disruption and overexpression mutants for each of the Argonaute and dicer genes and for two RNA-dependent RNA polymerase (RdRP) genes and generated virus-infected strains of each mutant. Interestingly, mycelial growth was significantly faster for the FgV1-infected overexpression mutant than for the FgV1-infected wild type, while neither FgV2 nor FgV3 infection altered the colony morphology of the gene deletion and overexpression mutants. FgV1 RNA accumulation was significantly decreased in the overexpression mutant. Furthermore, the levels of induction of, , and some of the genes caused by FgV2 and FgV3 infection were similar to those caused by hairpin RNA-induced gene silencing. Using small RNA sequencing analysis, we documented different patterns of virus-derived small interfering RNA (vsiRNA) production in strains infected with FgV1, -2, and -3. Our results suggest that the Argonaute protein encoded by is required for RNAi in, that induction differs in response to FgV1, -2, and -3, and that might contribute to the accumulation of vsiRNAs in FgV1-infected To increase our understanding of how RNAi components in react to mycovirus infections, we characterized the role(s) of RNAi components involved in the antiviral defense response against Fusarium graminearum viruses (FgVs). We observed differences in the levels of induction of RNA silencing-related genes, including and, in response to infection by three different FgVs. can efficiently induce a robust RNAi response against FgV1 infection, but genes might be relatively redundant to with respect to antiviral defense. However, the contribution of this gene in the response to the other FgV infections might be small. Compared to previous studies of, which showed dicer-like protein 2 and Argonaute-like protein 2 to be important in antiviral RNA silencing, our results showed that developed a more complex and robust RNA silencing system against mycoviruses and that FgDICER-1 and FgDICER-2 and FgAGO-1 and FgAGO-2 had redundant roles in antiviral RNA silencing.

No abstract

Tomato spotted wilt virus (TSWV) severely damages and reduces the yield of many economically important plants worldwide. In this study, we determined the whole-genome sequences of 10 TSWV isolates recently identified from various regions and hosts in Korea. Phylogenetic analysis of these 10 isolates as well as the three previously sequenced isolates indicated that the 13 Korean TSWV isolates could be divided into two groups reflecting either two different origins or divergences of Korean TSWV isolates. In addition, the complete nucleotide sequences for the 13 Korean TSWV isolates along with previously sequenced TSWV RNA segments from Korea and other countries were subjected to phylogenetic and recombination analysis. The phylogenetic analysis indicated that both the RNA L and RNA M segments of most Korean isolates might have originated in Western Europe and North America but that the RNA S segments for all Korean isolates might have originated in China and Japan. Recombination analysis identified a total of 12 recombination events among all isolates and segments and five recombination events among the 13 Korea isolates; among the five recombinants from Korea, three contained the whole RNA L segment, suggesting reassortment rather than recombination. Our analyses provide evidence that both recombination and reassortment have contributed to the molecular diversity of TSWV.