Viral diversity in Phytophthora cactorum population infecting strawberry.

Eighty-eight Phytophthora cactorum strains isolated from crown or leather rot of strawberry in 1971-2019 were screened for viruses using RNA-seq and RT-PCR. Remarkably, all but one isolate were virus-infected, most of them harbouring more than one virus of different genera or species. The most common virus occurring in 94% of the isolates was the Phytophthora cactorum RNA virus 1 (PcRV1) resembling members of Totiviridae. Novel viruses related to members of Endornaviridae, named Phytophthora cactorum alphaendornaviruses 1-3 (PcAEV1-3), were found in 57% of the isolates. Four isolates hosted viruses with affinities to Bunyaviridae, named Phytophthora cactorum bunyaviruses 1-3 (PcBV1-3), and a virus resembling members of the proposed genus 'Ustivirus', named Phytophthora cactorum usti-like virus (PcUV1), was found in a single isolate. Most of the virus species were represented by several distinct strains sharing ≥81.4% aa sequence identity. We found no evidence of spatial differentiation but some temporal changes in the P. cactorum virus community were observed. Some isolates harboured two or more closely related strains of the same virus (PcAEV1 or PcRV1) sharing 86.6%-96.4% nt identity in their polymerase sequence. This was surprising as viruses with such a high similarity are typically mutually exclusive.

Role and function of Chondrostereum purpureum in biocontrol of trees.

A decay fungus, Chondrostereum purpureum (Pers. Ex Fr.) Pouzar, has been investigated in Europe, Northern America and New Zealand for its ability to decay hardwood stumps and thus prevent sprouting. The aim of these investigations has been to find an alternative to mechanical (cutting only) and chemical sprout control (cutting and applying chemicals to stumps in order to prevent sprouting). Mechanical sprout control is not an efficient option due to hardwood tree species' ability to re-sprout efficiently after cutting, and therefore management costs are high. Chemicals would be efficient but due to their harmful effects on the environment, alternatives are needed. The fungal treatment, i.e., cutting accompanied with C. purpureum inoculum is an environmentally friendly and efficient option for sprout control. This mini-review comprises the role and function of C. purpureum in biocontrol of trees: the ecology of C. purpureum, its sprout control efficacy, factors affecting sprout control efficacy, devices in biological sprout control, potential risks, and the future perspectives of biological sprout control. KEY POINTS: • A fungus Chondrostereum purpureum is efficient in preventing sprouting of hardwoods • C. purpureum is not sensitive to environmental conditions • Devices should be developed for cost-efficient biological sprout control.

Rainstorm effects on the biocontrol efficacy of the decay fungus Chondrostereum purpureum against birch sprouting in boreal forests.

In forest regeneration areas, alongside roads and railways, under electric power lines and above gas pipe lines, there is a need for regular sprout control. A biocontrol method against broadleaved sprouting with formulations including the decay fungus Chondrostereum purpureum (Pers. Ex Fr.) Pouzar has been shown to be effective. Yet, heavy rain during spreading of this fungal inoculum on freshly cut stumps may affect the efficacy of the treatment, i.e., stump mortality during the following years. Thus, we performed an experiment where freshly cut birch stump surfaces (Betula pendula Roth and Betula pubescens Ehrh.) were treated with fungal inoculum under heavy irrigation and without it. Furthermore, two different adjuvants which aimed to fix the fungal inoculum to freshly cut stumps during irrigation and to protect against solar radiation were tested. Our results revealed that the artificial rainstorm treatment caused a delay in the efficacy of C. purpureum, but after three growing seasons, there was no significant difference in the mortality of birch stumps treated under irrigation or without it (stump mortalities 74 and 86%, respectively). Adjuvants did not improve the efficacy in stumps treated under irrigation nor in those treated without irrigation. KEY POINTS: • Heavy rain delayed the sprout control efficacy of a fungus Chondrostereum purpureum. • Final efficacy of formulations was the same in wet and dry conditions. • No additional adjuvants are needed to improve formulations.

Heterobasidion Partitivirus 13 Mediates Severe Growth Debilitation and Major Alterations in the Gene Expression of a Fungal Forest Pathogen.

The fungal genus Heterobasidion includes some of the most devastating conifer pathogens in the boreal forest region. In this study, we showed that the alphapartitivirus Heterobasidion partitivirus 13 from Heterobasidion annosum (HetPV13-an1) is the main causal agent of severe phenotypic debilitation in the host fungus. Based on RNA sequencing using isogenic virus-infected and cured fungal strains, HetPV13-an1 affected the transcription of 683 genes, of which 60% were downregulated and 40% upregulated. Alterations observed in carbohydrate and amino acid metabolism suggest that the virus causes a state of starvation, which is compensated for by alternative synthesis routes. We used dual cultures to transmit HetPV13-an1 into new strains of H. annosum and Heterobasidion parviporum The three strains of H. parviporum that acquired the virus showed noticeable growth reduction on rich culturing medium, while only two of six H. annosum isolates tested showed significant debilitation. Based on reverse transcription-quantitative PCR (RT-qPCR) analysis, the response toward HetPV13-an1 infection was somewhat different in H. annosum and H. parviporum We assessed the effects of HetPV13-an1 on the wood colonization efficacy of H. parviporum in a field experiment where 46 Norway spruce trees were inoculated with isogenic strains with or without the virus. The virus-infected H. parviporum strain showed considerably less growth within living trees than the isolate without HetPV13-an1, indicating that the virus also causes growth debilitation in natural substrates.IMPORTANCE A biocontrol method restricting the spread of Heterobasidion species would be highly beneficial to forestry, as these fungi are difficult to eradicate from diseased forest stands and cause approximate annual losses of €800 million in Europe. We used virus curing and reintroduction experiments and RNA sequencing to show that the alphapartitivirus HetPV13-an1 affects many basic cellular functions of the white rot wood decay fungus Heterobasidion annosum, which results in aberrant hyphal morphology and a low growth rate. Dual fungal cultures were used to introduce HetPV13-an1 into a new host species, Heterobasidion parviporum, and field experiments confirmed the capability of the virus to reduce the growth of H. parviporum in living spruce wood. Taken together, our results suggest that HetPV13-an1 shows potential for the development of a future biocontrol agent against Heterobasidion fungi.

Looking for relationships between the populations of Dothistroma septosporum in northern Europe and Asia.

Dothistroma septosporum, a notorious pine needle pathogen with an unknown historical geographic origin and poorly known distribution pathways, is nowadays found almost in all areas inhabited by pines (Pinus spp.). The main aim of this study was to determine the relationship between North European and East Asian populations. In total, 238 Eurasian D. septosporum isolates from 11 countries, including 211 isolates from northern Europe, 16 isolates from Russian Far East and 11 isolates from Bhutan were analysed using 11 species-specific microsatellite and mating type markers. The most diverse populations were found in northern Europe, including the Baltic countries, Finland and European Russia. Notably, D. septosporum has not caused heavy damage to P. sylvestris in northern Europe, which may suggest a long co-existence of the host and the pathogen. No indication was obtained that the Russian Far East or Bhutan could be the indigenous area of D. septosporum, as the genetic diversity of the fungus there was low and evidence suggests gene flow from northern Europe to Russian Far East. On the western coast of Norway, a unique genetic pattern was observed, which differed from haplotypes dominating other Fennoscandian populations. As an agent of dothistroma needle blight, only D. septosporum was documented in northern Europe and Asia, while D. pini was found in Ukraine and Serbia.

Distribution of Viruses Inhabiting Heterobasidion annosum in a Pine-Dominated Forest Plot in Southern Finland.

We investigated the diversity and spatial distribution of viruses infecting strains of the root rot fungus Heterobasidion annosum collected from pine stumps at a heavily infected forest site. Four different partitiviruses were detected in 14 H. annosum isolates at the study site, constituting approximately 29% of all Heterobasidion isolates investigated (N = 48). Two of the viruses detected were new partitiviruses designated here as Heterobasidion partitivirus 16 (HetPV16) and HetPV20, and two were previously known partitiviruses: HetPV7 and HetPV13. The two new partitiviruses found, HetPV16-an1 and HetPV20-an1, shared ~70% RdRp nucleotide sequence identity with the alphapartitivirus Rosellinia necatrix partitivirus 2, and less than 40% identity with known viruses of Heterobasidion spp. HetPV7-an1 was closely similar to HetPV7-pa1 isolated earlier from Heterobasidion parviporum, supporting the view of conspecific virus pools in different Heterobasidion species. Three fungal isolates were found to be co-infected with two different partitivirus strains (HetPV7-an1 and HetPV13-an2 or HetPV16-an1 and HetPV20-an1). Different isolates representing each host clone had variable virus compositions, and virus strains occurring in more than one host clone showed minor sequence variations between clones.

Cadophora margaritata sp. nov. and other fungi associated with the longhorn beetles Anoplophora glabripennis and Saperda carcharias in Finland.

Symbiosis with microbes is crucial for survival and development of wood-inhabiting longhorn beetles (Coleoptera: Cerambycidae). Thus, knowledge of the endemic fungal associates of insects would facilitate risk assessment in cases where a new invasive pest occupies the same ecological niche. However, the diversity of fungi associated with insects remains poorly understood. The aim of this study was to investigate fungi associated with the native large poplar longhorn beetle (Saperda carcharias) and the recently introduced Asian longhorn beetle (Anoplophora glabripennis) infesting hardwood trees in Finland. We studied the cultivable fungal associates obtained from Populus tremula colonised by S. carcharias, and Betula pendula and Salix caprea infested by A. glabripennis, and compared these to the samples collected from intact wood material. This study detected a number of plant pathogenic and saprotrophic fungi, and species with known potential for enzymatic degradation of wood components. Phylogenetic analyses of the most commonly encountered fungi isolated from the longhorn beetles revealed an association with fungi residing in the Cadophora-Mollisia species complex. A commonly encountered fungus was Cadophora spadicis, a recently described fungus associated with wood-decay. In addition, a novel species of Cadophora, for which the name Cadophora margaritata sp. nov. is provided, was isolated from the colonised wood.

The efficacy of six elite isolates of the fungus Chondrostereum purpureum against the sprouting of European aspen.

The sprouting of broad-leaved trees after cutting is problematic in forest regeneration areas, along roads and railways, under electric power and above gas pipe lines. In Finland, one of the most difficult species to control in these areas is the European aspen (Populus tremula), which produces both stump sprouts and root suckers after saplings have been cut. In this study, we investigated whether a decay fungus of broad-leaved trees, Chondrostereum purpureum, could be used as a biological control agent against aspen sprouting. The efficacy of six elite strains of C. purpureum (improved earlier in a breeding process) was investigated on aspen for three years. The most efficient C. purpureum strain, R53, tested earlier on birch (Betula pendula and B. pubescens), was efficient in causing mortality of aspen stumps and preventing the development of root suckers. With this strain, stump mortality was 78%, while significantly lower in control stumps which were cut only (47%). Aspen trees in the vicinity of the treatments (within a 10 m radius around each sapling) decreased the efficacy of C. purpureum. This study shows that the decay fungus C. purpureum can successfully be used in the sprout control of aspen saplings.

Diagnosis and discovery of fungal viruses using deep sequencing of small RNAs.

Analysis of virus-derived small RNAs with high-throughput sequencing has been successful for detecting novel viruses in plants and invertebrates. However, the applicability of this method has not been demonstrated in fungi, although fungi were among the first organisms reported to utilize RNA silencing. Here, we used virus-infected isolates of the fungal species complex Heterobasidion annosum sensu lato as a model system to test whether mycovirus genome segments can be detected with small RNA deep sequencing. Species of the genus Heterobasidion are some of the most devastating forest pathogens in boreal forests. These fungi cause wood decay and are commonly infected with species of the family Partitiviridae and the unassigned virus species Heterobasidion RNA virus 6. Small RNA deep sequencing allowed the simultaneous detection of all eight double-stranded RNA virus strains known to be present in the tested samples and one putative mitovirus species (family Narnaviridae) with a single-stranded RNA genome, designated here as Heterobasidion mitovirus 1. Prior to this study, no members of the family Narnaviridae had been described as infecting species of Heterobasidion. Quantification of viral double- and single-stranded RNA with quantitative PCR indicated that co-infecting viral species and viruses with segmented genomes can be detected with small RNA deep sequencing despite vast differences in the amount of RNA. This is the first study demonstrating the usefulness of this method for detecting fungal viruses. Moreover, the results suggest that viral genomes are processed into small RNAs by different species of Heterobasidion.

Fusarium circinatum isolates from northern Spain are commonly infected by three distinct mitoviruses.

Pitch canker is a serious disease of pines caused by the ascomycete fungus Gibberella circinata (anamorph = Fusarium circinatum). Three distinct mitovirus strains have been described in this fungus: Fusarium circinatum mitovirus 1 (FcMV1), FcMV2-1 and FcMV2-2. Here, we investigated the frequency and population variation of these viruses and closely related sequence variants in northern Spain using RT-PCR and sequencing. Each virus strain and similar sequence variants shared >95 % sequence identity and were collectively designated as virus types. All virus types were relatively common in Spain, with estimated prevalence of 18.5 %, 8.9 % and 16.3 % for FcMV1, FcMV2-1 and FcMV2-2, respectively.

Description and prevalence of a putative novel mycovirus within the conifer pathogen Gremmeniella abietina.

The European race of Gremmeniella abietina (Lagerberg) Morelet is the causal agent of stem canker and shoot blight on numerous conifers in Europe and North America. It comprises different species and biotypes in which the presence of mycoviruses has been determined. In this report, we describe the full-length sequence of the RNA-dependent RNA polymerase (RdRp) of a putative novel virus, Gremmeniella abietina RNA virus 6 (GaRV6), with 2165 nt and a GC content of 54.7 %. A BLASTp search using the deduced RdRp amino acid sequence confirmed GaRV6 to be related to members of a still unassigned virus taxon, which includes, e.g., Fusarium graminearum dsRNA mycovirus 4 (FgV-4) and the mutualistic Curvularia thermal tolerance virus (CThTV). The prevalence and genetic diversity of GaRV6 was also studied within the European race of G. abietina. We examined 162 isolates originating from Canada, the Czech Republic, Finland, Italy, Montenegro, Serbia, Spain, Switzerland, Turkey and the United States. According to direct specific reverse transcription (RT) PCR screening based on the RdRp sequence, the virus appears to be present only in Spain, where it is relatively abundant but genetically highly uniform.

Three mitovirus strains infecting a single isolate of Fusarium circinatum are the first putative members of the family Narnaviridae detected in a fungus of the genus Fusarium.

Fusarium circinatum Nirenberg & O'Donnell (teleomorph = Gibberella circinata) is the causal agent of pitch canker disease of pines. Since 2004 it has been present in Europe, particularly in northern Spain, affecting P. radiata and P. pinaster in plantations and nurseries. The disease has now also spread to other European countries, including France, Italy and Portugal. In this report, we describe three novel members of the genus Mitovirus from a Spanish isolate of F. circinatum: Fusarium circinatum mitovirus 1 (FcMV1), FcMV2-1 and FcMV2-2. Using a mitochondrial translation table, the complete 2419-bp genome of FcMV1 encodes an RNA-dependent RNA polymerase of 731 amino acids (GC-content ca 30 %). The partial genomes of FcMV2-1 and FcMV2-2 (2193 and 1973 bp, respectively) share ca 48 % RdRp sequence similarity at the aa level and might be regarded as conspecific, while FcMV1 is clearly distinct, showing 32-35 % polymerase similarity to the other strains. However, FcMV1 shared 46 % protein-level similarity with Thielaviopsis basicola mitovirus. This is the first study to report viruses in F. circinatum, as well as the first time that mitovirus genome sequences are described from Fusarium spp.

Complex transmission of partiti-, ambi- and ourmiaviruses in the forest pathogen Heterobasidion parviporum.

Utilizing Heterobasidion partitivirus 13 strain an1 (HetPV13-an1) and 15 strain pa1 (HetPV15-pa1) in co-infection is considered a potential biocontrol approach against Heterobasidion root and butt rot. Both partitiviruses mediate debilitating effects in most Heterobasidion host isolates and are generally transmitted efficiently between host strains. In this investigation, we conducted transmission experiments in the laboratory (in vitro) using several H. parviporum isolates to test whether using dual partitivirus infections is a more efficient way of transmitting viruses to new hosts compared to using single partitivirus infections, and whether co-occurring single-stranded RNA (ssRNA) viruses are co-transmitted during the process. The results showed that H. parviporum donors carrying both partitiviruses, HetPV13-an1 and HetPV15-pa1, transmitted HetPV15-pa1 more efficiently to recipients than the same donors infected with only HetPV15-pa1. In contrast, the transmission of HetPV13-an1 did not differ significantly between donors infected with both or only one partitivirus. Altogether, the transmission rates of HetPV13-an1 and HetPV15-pa1 were high on artificial media. Moreover, the transmission of the ssRNA viruses Heterobasidion ourmia-like virus 1(HetOlV1-pa7) and 4 (HetOlV4-an1) as well as Heterobasidion ambi-like virus 3 (HetAlV3-pa4) across different recipients were found to be variable. This study demonstrated for the first time the transmission of ambi- and ourmiaviruses between H. parviporum isolates in dual cultures and showed that H. parviporum mycelia can be cured of these ssRNA viruses using heat treatment.

Virus transmission frequencies in the pine root rot pathogen Heterobasidion annosum.

The combined use of Heterobasidion partitiviruses 13 and 15 (HetPV13-an1 and HetPV15-pa1) is considered a promising biocontrol approach against Heterobasidion root and butt rot. In a previous study, the transmission frequency of HetPV15-pa1 was found to be higher from a double partitivirus-infected donor than from a single partitivirus-infected donor. In this study, we included a wider array of recipient isolates to assess whether the phenomenon is widespread across different host strains and conducted transmission experiments on artificial media (in vitro) using a total of 45 different H. annosum donor-recipient pairs. In addition to investigating whether double partitivirus infection improves the transmission of HetPV13-an1 and HetPV15-pa1, we examined for the first time how efficiently co-infecting ssRNA viruses are concomitantly transmitted with the partitiviruses, and whether pre-existing ssRNA viruses in the recipients affect virus transmission. Generally, the transmission rates of HetPV13-an1 and HetPV15-pa1 were high from both single partitivirus-infected and double partitivirus-infected donors to most of the H. annosum recipient strains, with few exceptions. However, in contrast to previous experiments, the transmission frequency was not higher from the double partitivirus-infected donors. Also, ourmiavirus was transmitted between H. annosum strains, but the presence of another ourmiavirus in the recipient might affect the efficacy.

Diversity and impact of single-stranded RNA viruses in Czech Heterobasidion populations.

Heterobasidion annosum sensu lato comprises some of the most devastating pathogens of conifers. Exploring virocontrol as a potential strategy to mitigate economic losses caused by these fungi holds promise for the future. In this study, we conducted a comprehensive screening for viruses in 98 H. annosum s.l. specimens from different regions of Czechia aiming to identify viruses inducing hypovirulence. Initial examination for dsRNA presence was followed by RNA-seq analyses using pooled RNA libraries constructed from H. annosum and Heterobasidion parviporum, with diverse bioinformatic pipelines employed for virus discovery. Our study uncovered 25 distinct ssRNA viruses, including two ourmia-like viruses, one mitovirus, one fusarivirus, one tobamo-like virus, one cogu-like virus, one bisegmented narna-like virus and one segment of another narna-like virus, and 17 ambi-like viruses, for which hairpin and hammerhead ribozymes were detected. Coinfections of up to 10 viruses were observed in six Heterobasidion isolates, whereas another six harbored a single virus. Seventy-three percent of the isolates analyzed by RNA-seq were virus-free. These findings show that the virome of Heterobasidion populations in Czechia is highly diverse and differs from that in the boreal region. We further investigated the host effects of certain identified viruses through comparisons of the mycelial growth rate and proteomic analyses and found that certain tested viruses caused growth reductions of up to 22% and significant alterations in the host proteome profile. Their intraspecific transmission rates ranged from 0% to 33%. Further studies are needed to fully understand the biocontrol potential of these viruses in planta.IMPORTANCEHeterobasidion annosum sensu lato is a major pathogen causing significant damage to conifer forests, resulting in substantial economic losses. This study is significant as it explores the potential of using viruses (virocontrol) to combat these fungal pathogens. By identifying and characterizing a diverse array of viruses in H. annosum populations from Czechia, the research opens new avenues for biocontrol strategies. The discovery of 25 distinct ssRNA viruses, some of which reduce fungal growth and alter proteome profiles, suggests that these viruses could be harnessed to mitigate the impact of Heterobasidion. Understanding the interactions between these viruses and their fungal hosts is crucial for developing effective, environmentally friendly methods to protect conifer forests and maintain ecosystem health. This study lays the groundwork for future research on the application of mycoviruses in forest disease management.

Virus community dynamics in the conifer pathogenic fungus Heterobasidion parviporum following an artificial introduction of a partitivirus.

Viruses infecting the conifer pathogenic fungus Heterobasidion annosum sensu lato are intracellular and spread via anastomosis contacts. In the laboratory, these viruses transmit readily even between somatically incompatible isolates, but their dispersal capacity in natural conditions has not been previously studied. We introduced a mycovirus to a heavily diseased forest site by inoculating Norway spruce stumps with heartrot decay using a mycelial suspension of Heterobasidion parviporum strain RT3.49C hosting the partitivirus strain HetRV4-pa1. The Heterobasidion population at the sample plot was screened for mycoviruses prior to and after the inoculation. Based on sequence analysis, the resident H. parviporum strains harbored six different strains of the virus species Heterobasidion RNA virus 6 (HetRV6) and one strain of HetRV4 prior to the inoculation. After three growth seasons, the inoculated H. parviporum host strain was not detected, but the introduced virus had infected two resident H. parviporum genets. The presence of a preexisting HetRV6 infection did not hinder spread of the introduced partitivirus but resulted in coinfections instead. The resident HetRV6 virus population seemed to be highly stable during the incubation period, while the single indigenous HetRV4 infection was not detected after the inoculation. In laboratory infection experiments, the introduced virus could be transmitted successfully into all of the resident H. parviporum genets. This study shows for the first time transmission of a Heterobasidion virus between somatically incompatible hosts in natural conditions.

Molecular characterization of HetRV8-ir1, a partitivirus of the invasive conifer pathogenic fungus Heterobasidion irregulare.

The Heterobasidion annosum (Fr.) Bref. complex includes some of the most destructive conifer pathogenic fungi in the Boreal forest region. H. irregulare, formerly known as the North American pine type of H. annosum, was introduced from North America into Italy during the Second World War and occurs as an invasive pathogen in Pinus pinea stands together with the native European species H. annosum sensu stricto. We describe the complete nucleotide sequence of a new putative partitivirus from an Italian strain of H. irregulare. The bisegmented genome of HetRV8-ir1 encodes an RNA-dependent RNA polymerase of 704 aa and a capsid protein of 638 aa. The polymerase and capsid aa sequences are relatively similar (59-78 %) to those of Fusarium poae virus 1, Pleurotus ostreatus virus 1, and grapevine-associated partitivirus 1. HetRV8-ir1 is the first virus described from H. irregulare, and it is distantly related to previously known partitiviruses of Heterobasidion species.

Efficacy of Biological and Chemical Control Agents against Heterobasidion Spore Infections of Norway Spruce and Scots Pine Stumps on Drained Peatland.

Treatment of conifer stumps with a control agent effectively prevents Heterobasidion spore infections in summer cuttings and protects the residual stand and the next tree generation from damage caused by Heterobasidion root rot. Thus far, stump treatment experiments have been carried out in mineral soils, and no information is available on the efficacy of stump treatment agents in boreal peatland conditions. In the present study, biological and chemical control agents (Phlebiopsis gigantea and urea, respectively) were tested in Scots pine and Norway spruce stands subjected to thinning, cap cutting, and clearcutting on drained peatland in Central Finland. The control efficacy of urea was high in both spruce and pine stumps (on average 99.5 and 85.3%, respectively), while the efficacy of P. gigantea was highly variable on both tree species and ranged from full protection down to negative control effect, i.e., there were more Heterobasidion infections on the treated than untreated half of the stumps. The moisture content of the stump wood or the thickness of the peat layer did not affect the control efficacy of either control agent. These results emphasize a need for further studies to determine the reasons for the unsteadiness of the biological control in peatland conditions.

Bunyaviruses Affect Growth, Sporulation, and Elicitin Production in Phytophthora cactorum.

Phytophthora cactorum is an important oomycetous plant pathogen with numerous host plant species, including garden strawberry (Fragaria × ananassa) and silver birch (Betula pendula). P. cactorum also hosts mycoviruses, but their phenotypic effects on the host oomycete have not been studied earlier. In the present study, we tested polyethylene glycol (PEG)-induced water stress for virus curing and created an isogenic virus-free isolate for testing viral effects in pair with the original isolate. Phytophthora cactorum bunya-like viruses 1 and 2 (PcBV1 & 2) significantly reduced hyphal growth of the P. cactorum host isolate, as well as sporangia production and size. Transcriptomic and proteomic analyses revealed an increase in the production of elicitins due to bunyavirus infection. However, the presence of bunyaviruses did not seem to alter the pathogenicity of P. cactorum. Virus transmission through anastomosis was unsuccessful in vitro.