Cryphonectria nitschkei chrysovirus 1 with unique molecular features and a very narrow host range

Cryphonectria nitschkei chrysovirus 1 (CnCV1), was described earlier from an ascomycetous fungus, Cryphonectria nitschkei strain OB5/11, collected in Japan; its partial sequence was reported a decade ago. Complete sequencing of the four genomic dsRNA segments revealed molecular features similar to but distinct from previously reported members of the family Chrysoviridae. Unique features include the presence of a mini-cistron preceding the major large open reading frame in each genomic segment. Common features include the presence of CAA repeats in the 5′-untranslated regions and conserved terminal sequences. CnCV1-OB5/11 could be laterally transferred to C. nitschkei and its relatives C. radicalis and C. naterciae via coculturing, virion transfection and protoplast fusion, but not to fungal species other than the three species mentioned above, even within the genus Cryphonectria, suggesting a very narrow host range. Phenotypic comparison of a few sets of CnCV1-infected and -free isogenic strains showed symptomless infection in new hosts

Establishment of Neurospora crassa as a model organism for fungal virology

The filamentous fungus Neurospora crassa is used as a model organism for genetics, developmental biology and molecular biology. Remarkably, it is not known to host or to be susceptible to infection with any viruses. Here, we identify diverse RNA viruses in N. crassa and other Neurospora species, and show that N. crassa supports the replication of these viruses as well as some viruses from other fungi. Several encapsidated double-stranded RNA viruses and capsid-less positive-sense single-stranded RNA viruses can be experimentally introduced into N. crassa protoplasts or spheroplasts. This allowed us to examine viral replication and RNAi-mediated antiviral responses in this organism. We show that viral infection upregulates the transcription of RNAi components, and that Dicer proteins (DCL-1, DCL-2) and an Argonaute (QDE-2) participate in suppression of viral replication. Our study thus establishes N. crassa as a model system for the study of host-virus interactions. The fungus Neurospora crassa is a model organism for the study of various biological processes, but it is not known to be infected by any viruses. Here, Honda et al. identify RNA viruses that infect N. crassa and examine viral replication and RNAi-mediated antiviral responses, thus establishing this fungus as a model for the study of host-virus interactions

A novel deltapartitivirus from red clover

The family Partitiviridae has five genera, among which is the genus Deltapartitivirus. We report here the complete genome sequence of a deltapartitivirus from red clover, termed “red clover cryptic virus 3” (RCCV3). RCCV3 has a bisegmented double-stranded (ds) RNA genome. dsRNA1 and dsRNA2 are 1580 and 1589 nucleotides (nt) in length and are predicted to encode an RNA-directed RNA polymerase (RdRP) and a capsid protein (CP), respectively. The RCCV3 RdRP shares the highest sequence identity with the RdRP of a previously reported deltapartitivirus, Medicago sativa deltapartitivirus 1 (MsDPV1) (76.5%), while the RCCV3 CP shows 50% sequence identity to the CP of MsDPV1. RdRP- and CP-based phylogenetic trees place RCCV3 into a clade of deltapartitiviruses. The sequence and phylogenetic analyses clearly indicate that RCCV3 represents a new species in the genus Deltapartitivirus. RCCV3 was detectable in all three tested cultivars of red clover

Identification of novel totiviruses from the ascomycetous fungus Geotrichum candidum

Mycoviruses are widely distributed across the kingdom Fungi, including ascomycetous yeast strains of the class Saccharomycetes. Geotrichum candidum is an important fungal pathogen belonging to Saccharomycetes and has a diverse host range. Here, we report the characterization of four new classical totiviruses from two distinct Geotrichum candidum strains from Pakistan. The four identified viruses were tentatively named “Geotrichum candidum totivirus 1, 2, 3a, and 3b” (GcTV1-3b). The complete dsRNA genomes of the identified totiviruses are 4621, 4592, 4576, and 4576 bp in length, respectively. All totivirus genomes have two open reading frames, encoding a capsid protein (CP) and an RNA-dependent RNA polymerase (RdRP), respectively. The downstream RdRP domain is assumed to be expressed as a CP-RdRP fusion product via -1 frameshifting mediated by a heptameric slippery site. Sequence comparisons and phylogenetic analysis showed that each of the discovered viruses belongs to a new species of the genus Totivirus in the family Totiviridae, with GcTV1 and GcTV3 (a and b strains) clustering in one subgroup and GcTV2 in another subgroup

A new tetra-segmented splipalmivirus with divided RdRP domains from Cryphonectria naterciae, a fungus found on chestnut and cork oak trees in Europe

Positive-sense (+), single-stranded (ss) RNA viruses with divided RNA-dependent RNA polymerase (RdRP) domains have been reported from diverse filamentous ascomycetes since 2020. These viruses are termed splipalmiviruses or polynarnaviruses and have been characterized largely at the sequence level, but ill-defined biologically. Cryphonectria naterciae, from which only one virus has been reported, is an ascomycetous fungus potentially plant-pathogenic to chestnut and oak trees. We molecularly characterized multiple viruses in a single Portuguese isolate (C0614) of C. naterciae, taking a metatranscriptomic and conventional double-stranded RNA approach. Among them are a novel splipalmivirus (Cryphonectria naterciae splipalmivirus 1, CnSpV1) and a novel fusagravirus (Cryphonectria naterciae fusagravirus 1, CnFGV1). This study focused on the former virus. CnSpV1 has a tetra-segmented, (+)ssRNA genome (RNA1 to RNA4). As observed for other splipalmiviruses reported in 2020 and 2021, the RdRP domain is separately encoded by RNA1 (motifs F, A and B) and RNA2 (motifs C and D). A hypothetical protein encoded by the 5′-proximal open reading frame of RNA3 shows similarity to a counterpart conserved in some splipalmiviruses. The other RNA3-encoded protein and RNA4-encoded protein show no similarity with known proteins in a blastp search. The tetra-segment nature was confirmed by the conserved terminal sequences of the four CnSpV1 segments (RNA1 to RNA4) and their 100% coexistence in over 100 single conidial isolates tested. The experimental introduction of CnSpV1 along with CnFGV1 into a virus free strain C0754 of C. naterciae vegetatively incompatible with C0614 resulted in no phenotypic alteration, suggesting asymptomatic infection. The protoplast fusion assay indicates a considerably narrow host range of CnSpV1, restricted to the species C. naterciae and C. carpinicola. This study contributes to better understanding of the molecular and biological properties of this unique group of viruses