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Effects of AthCV1 infection on the colony morphology of Aspergillus species. (A) polymerase chain reaction (PCR) assay showing successful AthCV1 transfection to different virus-free Aspergillus species; reverse transcription PCR, lane L = 1 kb plus DNA ladder; -Ve= PCR negative control (RNA sample was replaced with ultrapure water), +Ath1 = AthCV1 +ve control; +fu = AthCV1 transfected A. fumigatus (Afu-13); +ng = AthCV1 transfected A. niger (Ang-9); +nd = AthCV1 transfected A. nidulans (And-1); –fu, –ng and –nd = original virus-free isolates of A. fumigatus (Afu-13), A. niger (Ang-9) and A. nidulans (And-1), respectively. (B) Afu-13 grown at 37 °C; normal growth in the virus-free isolate, sectors formed in isogenic AthCV1-transfected line (arrow). (C) Afu-13 grown at 20 °C; normal growth in the virus-free isolate, sectors formed in isogenic AthCV1-transfected line (arrow). (D) And-1 grown at 37 °C; normal growth in the virus-free isolate, ascospore-rich sectors formed in isogenic AthCV1-transfected line (arrow). (E) And-1 grown at 37 °C; normal growth in both virus-free and AthCV1-transfected lines. (F) Ang-9 grown at 37 °C; normal growth in the virus-free isolate, conidial-free sectors with sclerotia formed (arrow) in AthCV1-transfected line. (G) Ang-9 grown at 20 °C; normal growth in the virus-free isolate, conidial- and sclerotia-free sectors in AthCV1-transfected line (arrow).
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This study determined the effects of Aspergillus thermomutatus chrysovirus 1 (AthCV1), isolated from Aspergillus thermomutatus, on A. fumigatus, A. nidulans and A. niger. Protoplasts of virus-free isolates of A. fumigatus, A. nidulans and A. niger were transfected with purified AthCV1 particles and the phenotype, growth and sporulation of the isoge...
Citations
... Hyphal fusion is a naturally-occurring process. An artificial method for introducing viruses into cells using the protoplast PEG method has been established (Castro et al., 2003;Chiba et al., 2013;Ejmal et al., 2018;Hillman et al., 2004;Kanematsu et al., 2010;Niu et al., 2016;Suharto et al., 2022;Zheng et al., 2014). Specifically, protoplasts of the recipient fungus are prepared, after which purified virus particles from the donor are introduced directly into the protoplasts. ...
... This approach was used to introduce a partitivirus derived from Rosellinia necatrix (Sordariomycetidae) into Glomerella cingulata (Hypocreomycetidae) (Kanematsu et al., 2010). In addition, Aspergillus fumigatus, A. nidulans, and A. oryzae, which are closely related species, were previously infected with a chrysovirus derived from A. thermomutatus (Ejmal et al., 2018). Furthermore, Rhizoctonia solani can be artificially infected with the cucumber mosaic virus derived from plants (Andika et al., 2017). ...
Viruses are genetic elements that parasitize self-replicating cells.Therefore, organisms parasitized by viruses are not limited to animals andplants but also include microorganisms. Among these, viruses that parasitizefungi are known as mycoviruses. Mycoviruses with an RNA genome persistentlyreplicate inside fungal cells and coevolve with their host cells, similar to acellular organelle. Within host cells, mycoviruses can modulate various fungalcharacteristics and activities, including pathogenicity and the production ofenzymes and secondary metabolites. In this review, we provide an overview ofthe mycovirus research field as introduction to fungal researchers.Recognition of all genetic elements in fungi aids towards better understandingand control of fungi, and makes fungi a significant model system for studyingmicroorganisms containing multiple genetic elements.
... Among these 16, total of five genes were down-regulated and 11 were up-regulated in the virus-infected isolate that showed mainly the regulation of metabolism associated with lipids, polysaccharides and amino acids (Lau et al., 2018). Other interesting examples of phenotypic changes were demonstrated for Aspergillus thermomutatus chrysovirus 1 (AthCV-1) hosted by Aspergillus thermomutatus, which led to the production of sexual spores (ascospores) in the host strain that normally reproduced by asexual spores (conidiospores) without infection (Ejmal et al., 2018). \Infection of Beauveria bassiana polymycovirus (BbPmV) 1 and 3 enhanced its host growth, pigmentation, sporulation and pathogenicity (Filippou et al., 2021;Kotta-Loizou and Coutts, 2017). ...
Mycoviruses are natural inhabitants of fungi and have been identified in almost all fungal taxonomic groups. Mycoviruses that infect phytopathogenic fungi are now becoming a hot research area due to their potential for the biocontrol of important plant pathogens. But, before considering a mycovirus for biocontrol, we should be fully aware of the effects it induces in a fungal host and its interactions with other viruses, fungal strains and even the host plants. Mycoviral infections are generally associated with different effects, ranging from hypovirulence to hypervirulence, but they can often be cryptic (latent infections). The cryptic lifestyle has been associated to many mycoviruses, but thanks to growing knowledge we are now aware that it is often associated to axenic conditions while the real effects can be observed only in nature. Other mycoviruses either promote (hypervirulence) or (hypovirulence) fungal pathogenicity by a strong impact on the fungal physiology or by blocking the production of toxins or effectors. Finally, indirect effects of mycoviral infections can also be provided to the plant that hosts the fungal isolate, highlighting not only their potential as direct biocontrol agents but also as priming agents for plant resilience to biotic and abiotic stresses. This review provides a broad overview of mycoviral interactions both with their hosts and with other mycoviruses, highlighting the most interesting examples. In contrast to what has been observed to date, we believe that the collective availability of these data will not only improve our understanding of mycoviruses, but also increase our confidence in considering them as alternative measures against fungal diseases to improve the sustainable production of food and feed commodities.
... However, careful examination of the uncategorized dsRNA profiles in Fig. 1A revealed that strains 261 and 161 may be representative of strains of the same mycovirus and that the same may be true for strains 282 and 255, while the dsRNA profiles of strains 39 and 321 differ from one another and show no obvious similarities to any known mycovirus dsRNA profiles. It should be noted that the dsRNA profiles of strains 261 and 161 are reminiscent of the genomes of members of the family Polymycoviridae [13], Hadakaviridae [22], and Chrysoviridae [7] and that at least four polymycoviruses have been isolated from Beauveria spp. [8,9]. ...
... Also, a natural environment with its large abundance of potential hosts would appear to be ideal for horizontal transmission of mycoviruses, but until next-generation sequencing is used to study the virome of entomopathogenic fungi in Korea, no reliable conclusions can be drawn. There have been no reports of extracellular transmission of RNA mycoviruses [15], and host immunity, through vegetative incompatibility, often precludes transmission [7]. These features also require further investigation for all viruses, including Korean strains. ...
Entomopathogenic fungi have potential as biocontrol agents against insect pests, and mycovirus-mediated hypervirulence may enhance their efficacy. Before initiating research on hypervirulence, the presence or absence of double-stranded (ds) RNA elements was determined in 94 Korean entomopathogenic fungi. dsRNA elements varying in size from ca. 0.8 to 7 kbp were found in 14.9% (14/94) of the strains examined, including Beauveria bassiana, Metarhizium pemphigi, M. pinghaense, M. rileyi, and Cordyceps fumosorosea. This study provides information on the incidence and electrophoretic banding patterns of dsRNA elements and is the first report of mycoviruses entomopathogenic fungi in Korea.
... Also, the natural environment with its large host abundance would appear to be ideal for horizontal transmission of mycoviruses but until next generation sequencing is used to study the virome of Korean entomopathogenic fungi no reliable conclusions can be drawn. There are no reports of extracellular transmission of RNA mycoviruses [14] and host immunity though vegetative incompatibility often precludes transmission [7] and these features also require further investigation for all viruses including Korean strains. Further studies are needed to investigate the mycovirus infection rate of entomopathogenic fungi in natural environments and agricultural elds. ...
... However, careful examination of the uncategorized dsRNA pro les in Fig 1A reveals that strains 261 and 161 may be representative of strains of the same mycovirus and that the same may be true for strains 282 and 255 while the dsRNA pro les of strains 39 and 321 are dissimilar from one another with no obvious similarities to any known mycovirus dsRNA pro les. It should be noted that the dsRNA pro les of strains 261 and 161 are reminiscent of the genomes of members of the family Polymycoviridae [12], Hadakaviridae [22] and Chrysoviridae [7] and that at least four polymycoviruses have been isolated from Beauveria spp. [8,9]. ...
Entomopathogenic fungi have potential as biocontrol agents against insect pests and mycovirus-mediated hypervirulence may enhance their efficacy. Before initiating research on hypervirulence, the presence or absence of double-stranded (ds) RNA elements has been determined in 94 Korean entomopathogenic fungi. DsRNA elements varying in size from ca. 0.8 to 7 kbp were found in 14.9% (14/94) of the strains examined, including Beauveria bassiana , Metarhizium pemphigi , Metarhizium pinghaense , Metarhizium rileyi and Cordyceps fumosorosea . This study provides information on the incidence and electrophoretic banding pattern of dsRNA elements and is the first report of mycoviruses in Korean entomopathogenic fungi.
... Fungi, plants and possibly insects have been reported as hosts of chrysoviruses (Kotta--Loizou et al., 2020). Like many mycoviruses, most chrysoviruses appear symptomless in their fungal hosts (Ghabrial and Suzuki, 2009), but some chrysoviruses may cause hypovirulence and debilitation in their fungal hosts: Aspergillus thermomutatus chrysovirus 1 (AthCV1) reduced sporulation of a range of Aspergillus species, associated with invasive fungal diseases in immunocompromised patients (Ejmal et al., 2018); Magnaporthe oryzae chrysovirus 1-A (MoCV1-A) and Magnaporthe oryzae chrysovirus 1-B (MoCV1-B) caused hypovirulence features in the rice blast fungus, such as impaired growth, altered colony morphology, and reduced pigmentation (Urayama et al., 2010(Urayama et al., , 2012(Urayama et al., , 2014Higashiura et al., 2019); Alternaria alternata chrysovirus 1 strain QY2 (AaCV1-QY2) can independently induce slow growth and reduce virulence of Alternaria alternata, an important fungus pathogenic to fruit trees (Okada et al.,2018;Li et al., 2022). These attenuating effects of mycovirus infection might be a potential tool for the treatment of fungal infections in human health as well as agriculture (Fuke et al., 2011;Son et al., 2015). ...
A new double-stranded RNA (dsRNA) virus has been identified in the filamentous fungus Setosphaeria turcica f.sp. sorghi, whose genome consists of four segments (dsRNA1-4). Each dsRNA carries single open reading frame (ORF) flanked by 5′ and 3′ untranslated regions (UTRs) containing strictly conserved termini. The putative protein encoded by dsRNA1 showed 80.50% identity to the RNA-dependent RNA polymerase (RdRp) of the most closely related virus, Alternaria alternata chrysovirus 1 (AaCV1), belonging to the Chrysoviridae. dsRNA2 encodes the putative coat protein, while dsRNA3 and dsRNA4 respectively encode the hypothetical proteins of unknown functions. Phylogenetic analysis based on the RdRp protein indicated the virus clustered with members of the genus Betachrysovirus in the family Chrysoviridae. Based on the dsRNA profile, amino acid sequence comparisons, and phylogenetic analyses, the mycovirus is thought to be a new member of the family Chrysoviridae and designated as Setosphaeria turcica chrysovirus 1 (StCV1). Moreover, obvious differences were observed in the colony, mycelial and spore morphology between StCV1-infected and virus-cured strains of S. turcica f.sp. sorghi. StCV1 infection strongly reduced colony growth rate, spore production ability and virulence on host fungus. To our knowledge, this is the first report about mycovirus infecting S. turcica f.sp. sorghi and also the first chrysovirus infecting S. turcica.
... When subisolate LI41-1T1 was subjected to TEM analyses, isometric VLPs were visible in sucrose fractions (30-50%) after virus purification (Fig. 3C, right panel). To date, only dsRNA mycoviruses belonging to the family Polymycoviridae have been reported to be infectious as naked genomic dsRNAs [23,37], whereas chrysoviruses have only been transfected successfully with virions [38] but not dsRNA, in agreement with the results of the present study. ...
Fungi are generally thought to live in host plants with a single lifestyle, being parasitism, commensalism, or mutualism. The former, known as phytopathogenic fungi, cause various plant diseases that result in significant losses every year; while the latter, such as endophytic fungi, can confer fitness to the host plants. It is unclear whether biological factors can modulate the parasitic and mutualistic traits of a fungus. In this study, we isolated and characterized a mycovirus from an endophytic strain of the fungus Pestalotiopsis theae, a pathogen of tea (Camellia sinensis). Based on molecular analysis, we tentatively designated the mycovirus as Pestalotiopsis theae chrysovirus-1 (PtCV1), a novel member of the family Chrysoviridae, genus Alphachrysovirus. PtCV1 has four double-stranded (ds) RNAs as its genome, ranging from 0.9 to 3.4 kbp in size, encapsidated in isometric particles. PtCV1 significantly reduced the growth rates of its host fungus in vitro (ANOVA; P-value < 0.001) and abolished its virulence in planta (ANOVA; P-value < 0.001), converting its host fungus to a non-pathogenic endophyte on tea leaves, while PtCV1-free isolates were highly virulent. Moreover, the presence of PtCV1 conferred high resistance to the host plants against the virulent P. theae strains. Here we report a mycovirus that modulates endophytic and phytopathogenic fungal traits and provides an alternative approach to biological control of plant diseases caused by fungi.
... HTS has been successfully used to detect known and unknown viruses from fungal samples [81] as well as detect mycovirus genome segments through small RNA deep sequencing [25,82]. In addition, host transcriptome and small RNA profiling data have provided insight into the molecular mechanisms underpinning the observed changes in phenotype [83,84]. Genetic engineering research has started to concentrate on mycovirus-mediated hypo-or hypervirulence [84,85] due to the clear link between antiviral RNA silencing and mycoviral infection [86]. ...
Mycoviruses, just as the fungal endophytes they infect, are ubiquitous biological entities on Earth. Mycoviruses constitute a diverse group of viruses, and metagenomic approaches have—through recent discoveries of been mycoviruses—only recently began to provide evidence of this astonishing diversity. The current review presents (1) various mycoviruses which infect fungal endophytes and forest pathogens, (2) their presumed origins and interactions with fungi, plants and the environment, (3) high-throughput sequencing techniques that can be used to explore the horizontal gene transfer of mycoviruses, and (4) how the hypo- and hypervirulence induced by mycoviral infection is relevant to the biological control of pathogenic fungi.
... Protoplast preparation and virus transfection were undertaken following a method described previously (54) with minor modifications. Briefly, ~1 × 10 7 conidia were germinated to produce hyphae overnight in liquid yeast extract peptone dextrose broth [yeast extract (5 g/liter), microbiological peptone (3 g/liter), and dextrose (10 g/liter)]. ...
Here, we describe a tripartite circular single-stranded (ss) DNA mycovirus, named Fusarium graminearum gemytripvirus 1 (FgGMTV1). The genome of FgGMTV1 comprises three circular ssDNA segments (DNA-A, DNA-B, and DNA-C). Sequence alignments and phylogenetic analyses showed that FgGMTV1 is nested within the family Genomoviridae. We also constructed the first infectious DNA clones of a DNA mycovirus. Our results show that DNA-A and DNA-B are mutually interdependent for their replication and are associated with severely reduced colony growth and hypovirulence. DNA-C relies on DNA-A and DNA-B for replication and is necessary for the recovery of abnormal fungal phenotypes. DNA-C also enhances the accumulation of viral DNA in infected fungi and permits stable colonization and easy transmission via conidia. This is the first multipartite DNA virus isolated from a fungus. Our phylogenetic analyses also suggest that the multipartite genome of FgGMTV1 may have evolved from a monopartite genome of an ancient genomovirus.
... AfuPV-1 does not cause any obvious symptoms in the fungal host (Bhatti et al., 2011), while AfuCV affects host colony morphology, but not virulence of the fungal host (Jamal et al., 2010). AthCV1 reduces conidia formation (Ejmal et al., 2018). AfuTmV-1 modestly suppresses A. fumigatus virulence in an insect model, but does not affect the host animal's fungal burden (Kanhayuwa et al., 2015). ...
Aspergillus fumigatus is an airborne fungal pathogen that causes severe infections with invasive growth in immunocompromised patients. Several mycoviruses have recently been isolated from A. fumigatus strains, but there are presently no reports of mycoviral-mediated reduction or elimination of fungal pathogenicity in vertebrate models. Here, we report the biological features of a novel mycovirus, A. fumigatus chrysovirus 41362 (AfuCV41362), isolated from the hypovirulent A. fumigatus strain IFM 41362. The AfuCV41362 genome is comprised of four dsRNAs, each of which contains a single ORF (ORF1-4). ORF1 encodes a protein with sequence similarity to RNA-dependent RNA polymerases of viruses in the family Chrysoviridae, while ORF3 encodes a putative capsid protein. Viral RNAs are expressed primarily during the germination stage, and RNA-seq analysis of virus-infected A. fumigatus at the germination stage suggested that the virus suppressed expression of several pathogenicity-associated host genes, including hypoxia adaptation and nitric oxide detoxification genes. In vitro functional analysis revealed that the virus-infected strain had reduced tolerance to environmental stressors. Virus-infected A. fumigatus strain IFM 41362 had reduced virulence in vivo compared to the virus-free strain in a mouse infection model. Furthermore, introduction of the mycovirus to a natively virus-free KU A. fumigatus strain induced virus-infected phenotypes. To identify mycovirus genes responsible for the reduced virulence of A. fumigatus, each viral ORF was ectopically expressed in the virus-free KU strain. Ectopic expression of the individual ORFs only nominally reduced virulence of the host fungus in a mouse infection model. However, we found that ORF3 and ORF4 reduced tolerance to environmental stresses in in vitro analysis. Based on these results, we suggest that the AfuCV41362 mycovirus ORF3 and ORF4 reduce fungal virulence by suppressing stress tolerance together with other viral genes, rather than alone.
... Similarly, mycovirus infections have been shown to affect the transcription levels of genes associated with basic cellular functions in different fungal hosts (e.g. Li et al. 2008;Lee et al. 2014;Ejmal et al. 2018;Vainio et al. 2018). Some mycoviruses are also able to suppress host RNA silencing (Segers et al. 2007;Yaegashi et al. 2013a;Chiba et al. 2016). ...
Soils support a myriad of organisms hosting highly diverse viromes. In this minireview, we focus on viruses hosted by true fungi and oomycetes (members of Stamenopila, Chromalveolata) inhabiting bulk soil, rhizosphere and litter layer, and representing different ecological guilds including fungal saprotrophs, mycorrhizal fungi, mutualistic endophytes and pathogens. Viruses infecting fungi and oomycetes are characterized by persistent intracellular non-lytic lifestyles and transmission via spores and/or hyphal contacts. Almost all fungal and oomycete viruses have genomes composed of single stranded or double stranded RNA, and recent studies have revealed numerous novel viruses representing yet unclassified family-level groups. Depending on the virus-host combination, infections can be asymptomatic, beneficial or detrimental to the host. Thus, mycovirus infections may contribute to the multiplex interactions of hosts, therefore likely affecting the dynamics of fungal communities required for the functioning of soil ecosystems. However, the effects of fungal and oomycete viruses on soil ecological processes are still mostly unknown. Interestingly, new metagenomics data suggest an extensive level of horizontal virus transfer between plants, fungi and insects.
