Viral synergism suppresses R gene-mediated resistance by impairing downstream defense mechanisms in soybean.

Viral synergism occurs when mixed infection of a susceptible plant by 2 or more viruses leads to increased susceptibility to at least 1 of the viruses. However, the ability of 1 virus to suppress R gene-controlled resistance against another virus has never been reported. In soybean (Glycine max), extreme resistance (ER) against soybean mosaic virus (SMV), governed by the Rsv3 R-protein, manifests a swift asymptomatic resistance against the avirulent strain SMV-G5H. Still, the mechanism by which Rsv3 confers ER is not fully understood. Here, we show that viral synergism broke this resistance by impairing downstream defense mechanisms triggered by Rsv3 activation. We found that activation of the antiviral RNA-silencing pathway and the proimmune mitogen-activated protein kinase 3 (MAPK3), along with the suppression of the proviral MAPK6, are hallmarks of Rsv3-mediated ER against SMV-G5H. Surprisingly, infection with bean pod mottle virus (BPMV) disrupted this ER, allowing SMV-G5H to accumulate in Rsv3-containing plants. BPMV subverted downstream defenses by impairing the RNA-silencing pathway and activating MAPK6. Further, BPMV reduced the accumulation of virus-related siRNAs and increased the virus-activated siRNA that targeted several defense-related nucleotide-binding leucine-rich repeat receptor (NLR) genes through the action of the suppression of RNA-silencing activities encoded in its large and small coat protein subunits. These results illustrate that viral synergism can result from abolishing highly specific R gene resistance by impairing active mechanisms downstream of the R gene.

Identifying transcription factors associated with Fusarium graminearum virus 2 accumulation in Fusarium graminearum by phenome-based investigation.

Fusarium graminearum virus 2 (FgV2) infection induces phenotypic changes like reduction of growth rate and virulence with an alteration of the transcriptome, including various transcription factor (TFs) gene transcripts in Fusarium graminearum. Transcription factors are the primary regulator in many cellular processes and are significant in virus-host interactions. However, a detailed study about specific TFs to understand interactions between FgV2 and F. graminearum has yet to be conducted. We transferred FgV2 to a F. graminearum TF gene deletion mutant library to identify host TFs related to FgV2 infection. FgV2-infected TF mutants were classified into three groups depending on colony growth. The FgV2 accumulation level was generally higher in TF mutants showing more reduced growth. Among these FgV2-infected TF mutants, we found several possible TFs that might be involved in FgV2 accumulation, generation of defective interfering RNAs, and transcriptional regulation of FgDICER-2 and FgAGO-1 in response to virus infection. We also investigated the relation between FgV2 accumulation and production of reactive oxygen species (ROS) and DNA damage in fungal host cells by using DNA damage- or ROS-responsive TF deletion mutants. Our studies provide insights into the host factors related to FgV2 infection and bases for further investigation to understand interactions between FgV2 and F. graminearum.

Identification of viral genes involved in pepper mottle virus replication and symptom development in Nicotiana benthamiana.

Pepper mottle virus (PepMoV) infects primarily Capsicum species, including pepper and bell pepper which are important vegetable and spice crops in Korea. We have previously collected 13 PepMoV isolates from nine regions comprising five provinces, causing different symptoms on inoculated indicator host plants in Korea. To further identify the responsible symptom determinant(s) and explore viral protein functions of PepMoV, two out of 13 isolates, including 134 and 205136, were used in this study. Isolate 134 causes necrosis and yellowing, while 205136 causes severe mottle and yellowing symptoms on Nicotiana benthamiana. All chimeric and site-directed mutants contain the PepMoV 134 genome as a backbone with specific regions switched for those from counterparts of PepMoV 205136. Effects of all mutants compared with 134 after inoculation onto N. benthamiana by agroinfiltration. Results from our study provide direct evidence that the helper component-proteinase (HC-Pro) and the nuclear inclusion protein b (NIb)-coat protein (CP) regions are involved in virus accumulation and symptom determinants. In addition, we mapped to amino acid residues tyrosine, glycine, and leucine at position 360, 385, and 527, respectively, in the HC-Pro region participate in faster viral accumulation or movement in the plant. The residue valine at position 2773 of NIb plays an essential role in isolate 134 symptom development. As part of this study, we seek to gain insight into viral factors involved in the PepMoV infection cycle and a better understanding of plant-virus interactions. These findings complement the insufficiency of the gene function study of the PepMoV virus and provide a novel perspective for the protein function study of the Potyvirus.

Identification of a new strain of ligustrum virus A causing leaf necrosis and chlorosis symptoms in Syringa oblata var. dilatata (Nakai) Rehder.

In 2020, lilac trees showing virus-like symptoms such as leaf necrosis and chlorosis were observed in Korea. After RT-PCR detection with specific primer sets designed based on previously reported nucleotide sequences of viruses in lilac, the agent was identified as ligustrum virus A (LVA). The complete genome of the virus was sequenced and used for phylogenetic analysis. The genome of this novel strain of LVA, LVA-SNU, is 8524 nucleotides long, excluding the poly(A) tail, and shares the highest nucleotide sequence identity (77.28%) with LVA-Sob, which was detected in a plant of the same species, Syringa oblata, in China, whereas LVA-Sob shares higher sequence identity (97.89%) with LVA-SK, which has been detected in host plants of various species.

Pepper Mottle Virus and Its Host Interactions: Current State of Knowledge.

Pepper mottle virus (PepMoV) is a destructive pathogen that infects various solanaceous plants, including pepper, bell pepper, potato, and tomato. In this review, we summarize what is known about the molecular characteristics of PepMoV and its interactions with host plants. Comparisons of symptom variations caused by PepMoV isolates in plant hosts indicates a possible relationship between symptom development and genetic variation. Researchers have investigated the PepMoV-plant pathosystem to identify effective and durable genes that confer resistance to the pathogen. As a result, several recessive pvr or dominant Pvr resistance genes that confer resistance to PepMoV in pepper have been characterized. On the other hand, the molecular mechanisms underlying the interaction between these resistance genes and PepMoV-encoded genes remain largely unknown. Our understanding of the molecular interactions between PepMoV and host plants should be increased by reverse genetic approaches and comprehensive transcriptomic analyses of both the virus and the host genes.

A Phenome-Wide Association Study of the Effects of Fusarium graminearum Transcription Factors on Fusarium Graminearum Virus 1 Infection.

The Fusarium graminearum virus 1 (FgV1) causes noticeable phenotypic changes such as reduced mycelial growth, increase pigmentation, and reduced pathogenicity in its host fungi, Fusarium graminearum. Previous study showed that the numerous F. graminearum genes including regulatory factors were differentially expressed upon FgV1 infection, however, we have limited knowledge on the effect(s) of specific transcription factor (TF) during FgV1 infection in host fungus. Using gene-deletion mutant library of 657 putative TFs in F. graminearum, we transferred FgV1 by hyphal anastomosis to screen transcription factors that might be associated with viral replication or symptom induction. FgV1-infected TF deletion mutants were divided into three groups according to the mycelial growth phenotype compare to the FgV1-infected wild-type strain (WT-VI). The FgV1-infected TF deletion mutants in Group 1 exhibited slow or weak mycelial growth compare to that of WT-VI on complete medium at 5 dpi. In contrast, Group 3 consists of virus-infected TF deletion mutants showing faster mycelial growth and mild symptom compared to that of WT-VI. The hyphal growth of FgV1-infected TF deletion mutants in Group 2 was not significantly different from that of WT-VI. We speculated that differences of mycelial growth among the FgV1-infected TF deletion mutant groups might be related with the level of FgV1 RNA accumulations in infected host fungi. By conducting real-time quantitative reverse transcription polymerase chain reaction, we observed close association between FgV1 RNA accumulation and phenotypic differences of FgV1-infected TF deletion mutants in each group, i.e., increased and decreased dsRNA accumulation in Group 1 and Group 3, respectively. Taken together, our analysis provides an opportunity to identify host's regulator(s) of FgV1-triggered signaling and antiviral responses and helps to understand complex regulatory networks between FgV1 and F. graminearum interaction.

The Signatures of Natural Selection and Molecular Evolution in Fusarium graminearum Virus 1.

Fusarium graminearum virus 1 (FgV1) is a positive-sense ssRNA virus that confers hypovirulence in its fungal host, Fusarium graminearum. Like most mycoviruses, FgV1 exists in fungal cells, lacks an extracellular life cycle, and is therefore transmitted during sporulation or hyphal anastomosis. To understand FgV1 evolution and/or adaptation, we conducted mutation accumulation (MA) experiments by serial passage of FgV1 alone or with FgV2, 3, or 4 in F. graminearum. We expected that the effects of positive selection would be highly limited because of repeated bottleneck events. To determine whether selection on the virus was positive, negative, or neutral, we assessed both the phenotypic traits of the host fungus and the RNA sequences of FgV1. We inferred that there was positive selection on beneficial mutations in FgV1 based on the ratio of non-synonymous to synonymous substitutions (d N /d S ), on the ratio of radical to conservation amino acid replacements (p NR /p NC ), and by changes in the predicted protein structures. In support of this inference, we found evidence of positive selection only in the open reading frame 4 (ORF4) protein of DK21/FgV1 (MA line 1); mutations at amino acids 163A and 289H in the ORF4 of MA line 1 affected the entire structure of the protein predicted to be under positive selection. We also found, however, that deleterious mutations were a major driving force in viral evolution during serial passages. Linear relationships between changes in viral fitness and the number of mutations in each MA line demonstrated that some deleterious mutations resulted in fitness decline. Several mutations in MA line 1 were not shared with any of the other four MA lines (PH-1/FgV1, PH-1/FgV1 + 2, PH-1/FgV1 + 3, and PH-1/FgV1 + 4). This suggests that evolutionary pathways of the virus could differ with respect to hosts and also with respect to co-infecting viruses. The data also suggested that the differences among MA lines might also be explained by mutational robustness and other unidentified factors. Additional research is needed to clarify the effects of virus co-infection on the adaptation or evolution of FgV1 to its environments.

Identification of Viruses and Viroids Infecting Tomato and Pepper Plants in Vietnam by Metatranscriptomics.

Tomato (Lycopersicum esculentum L.) and pepper (Capsicum annuum L.) plants belonging to the family Solanaceae are cultivated worldwide. The rapid development of next-generation sequencing (NGS) technology facilitates the identification of viruses and viroids infecting plants. In this study, we carried out metatranscriptomics using RNA sequencing followed by bioinformatics analyses to identify viruses and viroids infecting tomato and pepper plants in Vietnam. We prepared a total of 16 libraries, including eight tomato and eight pepper libraries derived from different geographical regions in Vietnam. We identified a total of 602 virus-associated contigs, which were assigned to 18 different virus species belonging to nine different viral genera. We identified 13 different viruses and two viroids infecting tomato plants and 12 viruses and two viroids infecting pepper plants with viruses as dominantly observed pathogens. Our results showed that multiple infection of different viral pathogens was common in both plants. Moreover, geographical region and host plant were two major factors to determine viral populations. Taken together, our results provide the comprehensive overview of viral pathogens infecting two important plants in the family Solanaceae grown in Vietnam.

Exploration of the interactions between mycoviruses and Fusarium graminearum.

In this review, we discuss recent studies of the interaction between Fusarium graminearum viruses (FgVs) and the fungal host, Fusarium graminearum. Comprehensive transcriptome and proteome analyses have shown changes in the expression of host genes in response to infection by diverse FgVs. Using omics data and reverse genetics, researchers have determined the effects of some fungal host proteins (including FgHex1, FgHal2, FgSwi6, and vr1) on virus accumulation, virus transmission, and host symptom development. Recent reports have revealed the functions of the RNAi component in F. graminearum and the functional redundancy of FgDICERs and FgAGOs in the antiviral defense response against different FgV infections. Studies have also documented a unique mechanism used by FgV1 to overcome the antiviral response of the fungal host.

The ORF2 protein of Fusarium graminearum virus 1 suppresses the transcription of FgDICER2 and FgAGO1 to limit host antiviral defences.

The filamentous fungus Fusarium graminearum possesses an RNA-interference (RNAi) pathway that acts as a defence response against virus infections and exogenous double-stranded (ds) RNA. Fusarium graminearum virus 1 (FgV1), which infects F. graminearum, confers hypovirulence-associated traits such as reduced mycelial growth, increased pigmentation and reduced pathogenicity. In this study, we found that FgV1 can suppress RNA silencing by interfering with the induction of FgDICER2 and FgAGO1, which are involved in RNAi antiviral defence and the hairpin RNA/RNAi pathway in F. graminearum. In an FgAGO1- or FgDICER2-promoter/GFP-reporter expression assay the green fluorescent protein (GFP) transcript levels were reduced in FgV1-infected transformed mutant strains. By comparing transcription levels of FgDICER2 and FgAGO1 in fungal transformed mutants expressing each open reading frame (ORF) of FgV1 with or without a hairpin RNA construct, we determined that reduction of FgDICER2 and FgAGO1 transcript levels requires only the FgV1 ORF2-encoded protein (pORF2). Moreover, we confirmed that the pORF2 binds to the upstream region of FgDICERs and FgAGOs in vitro. These combined results indicate that the pORF2 of FgV1 counteracts the RNAi defence response of F. graminearum by interfering with the induction of FgDICER2 and FgAGO1 in a promoter-dependent manner.

Differential Contribution of RNA Interference Components in Response to Distinct Fusarium graminearum Virus Infections.

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 Fusarium graminearum by FgV1, -2, or -3 differentially induces the gene expression of RNAi components in F. graminearum Transcripts of the DICER-2 and AGO-1 genes of F. graminearum (FgDICER-2 and FgAGO-1) 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 FgAGO-1 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 FgAGO-1 overexpression mutant. Furthermore, the levels of induction of FgAGO-1, FgDICER-2, and some of the FgRdRP 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 FgAGO-1 is required for RNAi in F. graminearum, that FgAGO-1 induction differs in response to FgV1, -2, and -3, and that FgAGO-1 might contribute to the accumulation of vsiRNAs in FgV1-infected F. graminearumIMPORTANCE To increase our understanding of how RNAi components in Fusarium graminearum 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 FgDICER-2 and FgAGO-1, in response to infection by three different FgVs. FgAGO-1 can efficiently induce a robust RNAi response against FgV1 infection, but FgDICER genes might be relatively redundant to FgAGO-1 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 Cryphonectria parasitica, which showed dicer-like protein 2 and Argonaute-like protein 2 to be important in antiviral RNA silencing, our results showed that F. graminearum 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.

Effects of the deletion and over-expression of Fusarium graminearum gene FgHal2 on host response to mycovirus Fusarium graminearum virus 1.

The mycovirus Fusarium graminearum virus 1 (FgV1) is associated with reduced virulence (hypovirulence) of Fusarium graminearum. Transcriptomic and proteomic expression profiling have shown that many F. graminearum genes are differentially expressed as a consequence of FgV1 infection. Several of these genes may be related to the maintenance of the virus life cycle. The host gene, FgHal2, which has a highly conserved 3'-phosphoadenosine 5'-phosphatase (PAP phosphatase-like) domain or inositol monophosphatase (IMPase) superfamily domain, shows reduced expression in response to FgV1 infection. We generated targeted gene deletion and over-expression mutants to clarify the possible function(s) of FgHal2 and its relationship to FgV1. The gene deletion mutant showed retarded growth, reduced aerial mycelia formation and reduced pigmentation, whereas over-expression mutants were morphologically similar to the wild-type (WT). Furthermore, compared with the WT, the gene deletion mutant produced fewer conidia and these showed abnormal morphology. The FgHal2 expression level was decreased by FgV1 infection at 120 h post-inoculation (hpi), whereas the levels were nine-fold greater for both the virus-free and virus-infected over-expression mutant than for the WT. FgV1 RNA accumulation was decreased in the deletion mutant at 48, 72 and 120 hpi. FgV1 RNA accumulation in the over-expression mutant was reduced relative to that of the WT at 48 and 120 hpi, but was similar to that of the WT at 72 hpi. The vertical transmission rate of FgV1 in the gene deletion mutant was low, suggesting that FgHal2 may be required for the maintenance of FgV1 in the host cell. Together, these results indicate that the putative 3'(2'),5'-bisphosphate nucleotidase gene, FgHal2, has diverse biological functions in the host fungus and may affect the viral RNA accumulation and transmission of FgV1.

A comparison of transcriptional patterns and mycological phenotypes following infection of Fusarium graminearum by four mycoviruses.

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.

The HEX1 gene of Fusarium graminearum is required for fungal asexual reproduction and pathogenesis and for efficient viral RNA accumulation of Fusarium graminearum virus 1.

The 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.

Phylogenetic and recombination analysis of tomato spotted wilt virus.

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.

Insight into mycoviruses infecting Fusarium species.

Most of the major fungal families including plant-pathogenic fungi, yeasts, and mushrooms are infected by mycoviruses, and many double-stranded RNA (dsRNA) mycoviruses have been recently identified from diverse plant-pathogenic Fusarium species. The frequency of occurrence of dsRNAs is high in Fusarium poae but low in other Fusarium species. Most Fusarium mycoviruses do not cause any morphological changes in the host but some mycoviruses like Fusarium graminearum virus 1 (FgV1) cause hypovirulence. Available genomic data for seven of the dsRNA mycoviruses infecting Fusarium species indicate that these mycoviruses exist as complexes of one to five dsRNAs. According to phylogenetic analysis, the Fusarium mycoviruses identified to date belong to four families: Chrysoviridae, Hypoviridae, Partitiviridae, and Totiviridae. Proteome and transcriptome analysis have revealed that FgV1 infection of Fusarium causes changes in host transcriptional and translational machineries. Successful transmission of FgV1 via protoplast fusion suggests the possibility that, as biological control agents, mycoviruses could be introduced into diverse species of fungal plant pathogens. Research is now needed on the molecular biology of mycovirus life cycles and mycovirus-host interactions. This research will be facilitated by the further development of omics technologies.

Highly Specific Detection of Five Exotic Quarantine Plant Viruses using RT-PCR.

To detect five plant viruses (Beet black scorch virus, Beet necrotic yellow vein virus, Eggplant mottled dwarf virus, Pelargonium zonate spot virus, and Rice yellow mottle virus) for quarantine purposes, we designed 15 RT-PCR primer sets. Primer design was based on the nucleotide sequence of the coat protein gene, which is highly conserved within species. All but one primer set successfully amplified the targets, and gradient PCRs indicated that the optimal temperature for the 14 useful primer sets was 51.9°C. Some primer sets worked well regardless of annealing temperature while others required a very specific annealing temperature. A primer specificity test using plant total RNAs and cDNAs of other plant virus-infected samples demonstrated that the designed primer sets were highly specific and generated reproducible results. The newly developed RT-PCR primer sets would be useful for quarantine inspections aimed at preventing the entry of exotic plant viruses into Korea.

Genome-wide expression profiling shows transcriptional reprogramming in Fusarium graminearum by Fusarium graminearum virus 1-DK21 infection.

BACKGROUND: Fusarium graminearum virus 1 strain-DK21 (FgV1-DK21) is a mycovirus that confers hypovirulence to F. graminearum, which is the primary phytopathogenic fungus that causes Fusarium head blight (FHB) disease in many cereals. Understanding the interaction between mycoviruses and plant pathogenic fungi is necessary for preventing damage caused by F. graminearum. Therefore, we investigated important cellular regulatory processes in a host containing FgV1-DK21 as compared to an uninfected parent using a transcriptional approach. RESULTS: Using a 3'-tiling microarray covering all known F. graminearum genes, we carried out genome-wide expression analyses of F. graminearum at two different time points. At the early point of growth of an infected strain as compared to an uninfected strain, genes associated with protein synthesis, including ribosome assembly, nucleolus, and ribosomal RNA processing, were significantly up-regulated. In addition, genes required for transcription and signal transduction, including fungal-specific transcription factors and cAMP signaling, respectively, were actively up-regulated. In contrast, genes involved in various metabolic pathways, particularly in producing carboxylic acids, aromatic amino acids, nitrogen compounds, and polyamines, showed dramatic down-regulation at the early time point. Moreover, genes associated with transport systems localizing to transmembranes were down-regulated at both time points. CONCLUSION: This is the first report of global change in the prominent cellular pathways in the Fusarium host containing FgV1-DK21. The significant increase in transcripts for transcription and translation machinery in fungal host cells seems to be related to virus replication. In addition, significant down-regulation of genes required for metabolism and transporting systems in a fungal host containing the virus appears to be related to the host defense mechanism and fungal virulence. Taken together, our data aid in the understanding of how FgV1-DK21 regulates the transcriptional reprogramming of F. graminearum.

Transmission of Fusarium boothii mycovirus via protoplast fusion causes hypovirulence in other phytopathogenic fungi.

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.