Plants interfere with non-self recognition of a phytopathogenic fungus via proline accumulation to facilitate mycovirus transmission.

Non-self recognition is a fundamental aspect of life, serving as a crucial mechanism for mitigating proliferation of molecular parasites within fungal populations. However, studies investigating the potential interference of plants with fungal non-self recognition mechanisms are limited. Here, we demonstrate a pronounced increase in the efficiency of horizontal mycovirus transmission between vegetatively incompatible Sclerotinia sclerotiorum strains in planta as compared to in vitro. This increased efficiency is associated with elevated proline concentration in plants following S. sclerotiorum infection. This surge in proline levels attenuates the non-self recognition reaction among fungi by inhibition of cell death, thereby facilitating mycovirus transmission. Furthermore, our field experiments reveal that the combined deployment of hypovirulent S. sclerotiorum strains harboring hypovirulence-associated mycoviruses (HAVs) together with exogenous proline confers substantial protection to oilseed rape plants against virulent S. sclerotiorum. This unprecedented discovery illuminates a novel pathway by which plants can counteract S. sclerotiorum infection, leveraging the weakening of fungal non-self recognition and promotion of HAVs spread. These promising insights provide an avenue to explore for developing innovative biological control strategies aimed at mitigating fungal diseases in plants by enhancing the efficacy of horizontal HAV transmission.

Complete genome sequence of a novel alternavirus isolated from the phytopathogenic fungus Colletotrichum fioriniae.

A novel double-strand RNA (dsRNA) mycovirus, named "Colletotrichum fioriniae alternavirus1" (CfAV1), was isolated from the strain CX7 of Colletotrichum fioriniae, the causal agent of walnut anthracnose. The complete genome of CfAV1 is composed of three dsRNA segments: dsRNA1 (3528 bp), dsRNA2 (2485 bp), and dsRNA3 (2481 bp). The RNA-dependent RNA polymerase (RdRp) is encoded by dsRNA1, while both dsRNA2 and dsRNA3 encode hypothetical proteins. Based on multiple sequence alignments and phylogenetic analysis, CfAV1 is identified as a new member of the family Alternaviridae. This is the first report of an alternavirus that infects the phytopathogenic fungus C. fioriniae.

Characterization of a novel hypovirus isolated from the phytopathogenic fungus Colletotrichum camelliae.

Some members of genus Colletotrichum are important plant pathogens. Here, we report a novel positive single-stranded RNA virus, Colletotrichum camelliae hypovirus 1 (CcHV1), from strain GXNN11-2 of Colletotrichum camelliae. The complete genome of CcHV1 is 9907 nucleotides (nt) in length and contains a single large open reading frame (ORF) from nt 352 to 9006. This ORF encodes a polyprotein with four conserved domains, namely UDP-glycosyltransferase, RNA-dependent RNA polymerase (RdRp), peptidase, and DEAD-like helicase. The CcHV1 polyprotein shares the highest similarity with Fusarium concentricum hypovirus 1. Phylogenetic analysis indicated that CcHV1 clustered with members of the genus Betahypovirus within the family Hypoviridae. This is the first report of a hypovirus in a member of the genus Colletotrichum.

Molecular characterization of a novel fungal alphaflexivirus reveals potential inter-species horizontal gene transfer.

Sclerotinia sclerotiorum is a notorious phytopathogenic fungus that harbors diverse mycoviruses. A novel positive-sense single-stranded RNA virus, Sclerotinia sclerotiorum alphaflexivirus 2 (SsAFV2), was isolated from the hypovirulent strain 32-9 of S. sclerotiorum, and its complete genome was determined. The SsAFV2 genome contains 7,162 nucleotides (nt), excluding the poly (A) structure, and is composed of four open reading frames (ORF1-4). ORF1 encodes a polyprotein that contains three conserved domains: methyltransferase, helicase, and RNA-dependent RNA polymerase (RdRp). The ORF3 putative encodes coat proteins (CP), with ORF2 and ORF4 encoding hypothetical proteins of unknown functions. Phylogenetic analysis revealed that SsAFV2 clustered with Botrytis virus X (BVX) based on multiple alignments of helicase, RdRp, and CP, but the methyltransferase of SsAFV2 was most closely related to Sclerotinia sclerotiorum alphaflexivirus 1, suggesting that SsAFV2 is a new member of the Botrexvirus genus within the Alphaflexiviridae family, and also revealed the occurrence of potential inter-species horizontal gene transfer events within the Botrexvirus genus during the evolutionary process. Our results contribute to the current knowledge regarding the evolution and divergence of Botrexviruses.

Characterization of a Fungal Virus Representing a Novel Genus in the Family Alphaflexiviridae.

Sclerotinia sclerotiorum is an ascomycetous fungus and hosts various mycoviruses. In this study, a novel fungal alphaflexivirus with a special genomic structure, named Sclerotinia sclerotiorum alphaflexivirus 1 (SsAFV1), was cloned from a hypovirulent strain, AHS31. Strain AHS31 was also co-infected with two botourmiaviruses and two mitoviruses. The complete genome of SsAFV1 comprised 6939 bases with four open reading frames (ORFs), a conserved 5'-untranslated region (UTR), and a poly(A) tail in the 3' terminal; the ORF1 and ORF3 encoded a replicase and a coat protein (CP), respectively, while the function of the proteins encoded by ORF2 and ORF4 was unknown. The virion of SsAFV1 was flexuous filamentous 480-510 nm in length and 9-10 nm in diameter. The results of the alignment and the phylogenetic analysis showed that SsAFV1 is related to allexivirus and botrexvirus, such as Garlic virus X of the genus Allexivirus and Botrytis virus X of the genus Botrevirus, both with 44% amino-acid (aa) identity of replicase. Thus, SsAFV1 is a novel virus and a new genus, Sclerotexvirus, is proposed to accommodate this novel alphaflexivirus.

Discovery and Evolution of Six Positive-Sense RNA Viruses Co-infecting the Hypovirulent Strain SCH733 of Sclerotinia sclerotiorum.

Sclerotinia sclerotiorum is a well-known phytopathogenic fungus with a wide host range. Identifying novel mycoviruses in phytopathogenic fungi is necessary to develop novel strategies for plant health protection and contribute to understanding the origin of viruses. Six new mycoviruses with positive single-stranded RNA genomes co-infecting the hypovirulent strain SCH733 of S. sclerotiorum were identified using a metatranscriptomic approach, and their complete genome sequences were molecularly determined. These mycoviruses belong to the following five families: Narnaviridae, Mitoviridae, Deltaflexviridae, Botourmiaviridae, and Ambiguiviridae. Three of these mycoviruses belong to existing International Committee on Taxonomy of Viruses (ICTV)-recognized species. Two of these newly identified mycoviruses have unique genomic features that are significantly different from those of all known mycoviruses. Phylogenetic analysis revealed that these six mycoviruses included close as well as distant relatives of known mycoviruses, thereby providing new insight into virus evolution and classification. Mycovirus horizontal transmission and elimination experiments revealed that Sclerotinia sclerotiorum narnavirus 5 is associated with hypovirulence of S. sclerotiorum, although we have not shown that it is independently responsible for the hypovirulence phenotype. This study broadens the diversity of known mycoviruses infecting S. sclerotiorum and provides a clue toward limiting hypovirulence in S. sclerotiorum.

Mycoviromic Analysis Unveils Complex Virus Composition in a Hypovirulent Strain of Sclerotinia sclerotiorum.

Mycoviruses are ubiquitous in pathogenic fungi including Sclerotinia sclerotiorum. Using RNA sequencing, more mycoviruses have been identified in individual strains, which were previously reported to be infected by a single mycovirus. A hypovirulent strain of S. sclerotiorum, HC025, was previously thought to harbor a single mitovirus, Sclerotinia sclerotiorum mitovirus 1 (SsMV1), based on the analysis of the conventional dsRNA extraction method. We found HC025 to be co-infected by five mycoviruses. In addition to SsMV1, four mycoviruses were identified: Sclerotinia sclerotiorum narnavirus 4 (SsNV4), Sclerotinia sclerotiorum negative-stranded RNA virus 1 (SsNSRV1), Sclerotinia sclerotiorum ourmia-like virus 14 (SsOLV14), and SsOLV22. Three mycoviruses including SsNV4, SsNSRV1, and SsOLV14 share high replicase identities (more than 95%) with the previously reported corresponding mycoviruses, and SsOLV22 shows lower identity to the known viruses. The complete genome of SsOLV22 is 3987 nt long and contains a single ORF-encoded RdRp, which shares 24.84% identity with the RNA-dependent RNA polymerase (RdRp) of Hubei narna-like virus 10 (query coverage: 26%; e-value: 8 × 10-19). The phylogenetic tree of RdRp suggests that SsOLV22 is a new member within the family Botourmiaviridae. All of the mycoviruses except for SsNSRV1 could horizontally co-transfer from HC025 to the virulent strain Ep-1PNA367 with hypovirulent phenotypes, and converted a later strain into a hypovirulent strain. In summary, we molecularly characterized the hypovirulent strain HC025 and identified five RNA mycoviruses including a new member within Botourmiaviridae.

Two Novel Rhabdoviruses Related to Hypervirulence in a Phytopathogenic Fungus.

Rhabdoviruses are ubiquitous and diverse viruses that propagate owing to bidirectional interactions with their vertebrate, arthropod, and plant hosts, and some of them could pose global health or agricultural threats. However, rhabdoviruses have rarely been reported in fungi. Here, two newly identified fungal rhabdoviruses, Rhizoctonia solani rhabdovirus 1 (RsRhV1) and RsRhV2, were discovered and molecularly characterized from the phytopathogenic fungus Rhizoctonia solani. The genomic organizations of RsRhV1 and RsRhV2 are 11,716 and 11,496 nucleotides (nt) in length, respectively, and consist of five open reading frames (ORFs) (ORFs I to V). ORF I, ORF IV, and ORF V encode the viral nucleocapsid (N), glycoprotein (G), and RNA polymerase (L), respectively. The putative protein encoded by ORF III has a lower level of identity with the matrix protein of rhabdoviruses. ORF II encodes a hypothetical protein with unknown function. Phylogenetic trees based on multiple alignments of N, L, and G proteins revealed that RsRhV1 and RsRhV2 are new members of the family Rhabdoviridae, but they form an independent evolutionary branch significantly distinct from other known nonfungal rhabdoviruses, suggesting that they represent a novel viral evolutionary lineage within Rhabdoviridae. Compared to strains lacking rhabdoviruses, strains harboring RsRhV2 and RsRhV1 showed hypervirulence, suggesting that RsRhV1 and RsRhV2 might be associated with the virulence of R. solani. Taken together, this study enriches our understanding of the diversity and host range of rhabdoviruses. IMPORTANCE Mycoviruses have been attracting an increasing amount of attention due to their impact on important medical, agricultural, and industrial fungi. Rhabdoviruses are prevalent across a wide spectrum of hosts, from plants to invertebrates and vertebrates. This study molecularly characterized two novel rhabdoviruses from four Rhizoctonia solani strains, based on their genomic structures, transcription strategy, phylogenetic relationships, and biological impact on their host. Our study makes a significant contribution to the literature because it not only enriches the mycovirus database but also expands the known host range of rhabdoviruses. It also offers insight into the evolutionary linkage between animal viruses and mycoviruses and the transmission of viruses from one host to another. Our study will also help expand the contemporary knowledge of the classification of rhabdoviruses, as well as providing a new model to study rhabdovirus-host interactions, which will benefit the agriculture and medical areas of human welfare.

Fusarivirus accessory helicases present an evolutionary link for viruses infecting plants and fungi.

A significant number of mycoviruses have been identified that are related to plant viruses, but their evolutionary relationships are largely unexplored. A fusarivirus, Rhizoctonia solani fusarivirus 4 (RsFV4), was identified in phytopathogenic fungus Rhizoctonia solani (R. solani) strain XY74 co-infected by an alphaendornavirus. RsFV4 had a genome of 10,833 â€‹nt (excluding the poly-A tail), and consisted of four non-overlapping open reading frames (ORFs). ORF1 encodes an 825 aa protein containing a conserved helicase domain (Hel1). ORF3 encodes 1550 aa protein with two conserved domains, namely an RNA-dependent RNA polymerase (RdRp) and another helicase (Hel2). The ORF2 and ORF4 likely encode two hypothetical proteins (520 and 542 aa) with unknown functions. The phylogenetic analysis based on Hel2 and RdRp suggest that RsFV4 was positioned within the fusarivirus group, but formed an independent branch with three previously reported fusariviruses of R. solani. Notably, the Hel1 and its relatives were phylogenetically closer to helicases of potyviruses and hypoviruses than fusariviruses, suggesting fusarivirus Hel1 formed an evolutionary link between these three virus groups. This finding provides evidence of the occurrence of a horizontal gene transfer or recombination event between mycoviruses and plant viruses or between mycoviruses. Our findings are likely to enhance the understanding of virus evolution and diversity.

Molecular characterization of a novel penoulivirus from the phytopathogenic fungus Colletotrichum camelliae.

Colletotrichum camelliae is a widespread filamentous phytopathogenic fungus. In this study, a novel mycovirus designated as "Colletotrichum camelliae botourmiavirus 1" (CcBV1) was isolated from strain ZJQT11 of C. camelliae, and its complete genome sequence was determined. CcBV1 has a genome of 2,506 nucleotides and contains a large open reading frame (ORF) that encodes an RNA-dependent RNA polymerase (RdRp) with 672 amino acids and a predicted molecular mass of 75.23 kDa. A BLASTp search showed that RdRp encoded by CcBV1 is closely related to that of Pyricularia oryzae ourmia-like virus 1 with 73.22% identity. Phylogenetic analysis indicated that CcBV1 clustered in the penoulivirus clade within the family Botourmiaviridae. To the best of our knowledge, this is the first report of a penoulivirus in C. camelliae.

The complete genome sequence of a novel chrysovirus from the entomopathogenic fungus Beauveria bassiana Vuillemin.

Beauveria bassiana, an entomopathogenic fungus, is used for arthropod pest control worldwide. Here, we report the discovery and characterization of a novel double-stranded RNA (dsRNA) mycovirus, Beauveria bassiana chrysovirus 2 (BbCV-2), isolated from a Chinese B. bassiana strain. The genome sequence of the virus was determined by metagenomic sequencing, RT-PCR, and RACE cloning and was found to consist of four dsRNA segments that are 3441 bp, 2779 bp, 2925 bp, and 2688 bp long, respectively. Each dsRNA segment contains a single ORF. The ORF of dsRNA1 encodes a 1114-amino-acid (aa) protein (123.4 kDa) with a conserved RNA-dependent RNA polymerase (RdRp) motif, the sequence of which showed the highest identity of only 16.13% to that of Beauveria bassiana chrysovirus-1 (BbCV-1). The ORF of dsRNA2 encodes an 805-aa coat protein (CP) (84.7 kDa). The ORFs of dsRNAs 3 and 4 encodes proteins of undetermined function. The virus is a new member of the family Chrysoviridae from B. bassiana.

Complete genome sequence of a novel mitovirus associated with Lagenaria siceraria.

Here, we report the discovery and molecular characterization of a novel mitovirus isolated from tissues of Lagenaria siceraria, named "Lagenaria siceraria associated mitovirus 1" (LsaMV1). Next-generation sequencing and adapter-ligation-mediated amplification were used to obtain the full-length genome sequence of LsaMV1. The genome of LsaMV1 is 3,098 nucleotides (nt) long and contains an opening reading frame (ORF) encoding a putative RNA-dependent RNA polymerase (RdRP). Homology searches and phylogenetic analysis of the 855-aa RdRP suggested that LsaMV1 is a member of a new species in the family Mitoviridae. This is the first report of a member of the family Mitoviridae associated with the important summer vegetable bottle gourd.

Two distant helicases in one mycovirus: evidence of horizontal gene transfer between mycoviruses, coronaviruses and other nidoviruses.

Nidovirales, which accommodates viruses with the largest RNA genomes, includes the notorious coronaviruses; however, the evolutionary route for nidoviruses is not well understood. We have characterized a positive-sense (+) single-stranded (ss) RNA mycovirus, Rhizoctonia solani hypovirus 2 (RsHV2), from the phytopathogenic fungus Rhizoctonia solani. RsHV2 has the largest RNA genome size of 22,219 nucleotides, excluding the poly(A) tail, in all known mycoviruses, and contains two open reading frames (ORF1 and ORF2). ORF1 encodes a protein of 2,009 amino acid (aa) that includes a conserved helicase domain belonging to helicase superfamily I (SFI). In contrast, ORF2 encodes a polyprotein of 4459 aa containing the hallmark genes of hypoviruses. The latter includes a helicase belonging to SFII. Following phylogenetic analysis, the ORF1-encoded helicase (Hel1) unexpectedly clustered in an independent evolutionary branch together with nidovirus helicases, including coronaviruses, and bacteria helicases. Thus, Hel1 presence indicates the occurrence of horizontal gene transfer between viruses and bacteria. These findings also suggest that RsHV2 is most likely a recombinant arising between hypoviruses and nidoviruses.

Molecular characterization of a novel fusarivirus infecting the edible fungus Auricularia heimuer.

Here, we describe a novel mycovirus, Auricularia heimuer fusarivirus 1 (AhFV1), isolated from the edible fungus Auricularia heimuer strain CCMJ1296. The virus has a single-stranded positive-sense [+ssRNA] genome of 7,127 nucleotides containing two overlapping open reading frames (ORFs) and a poly(A) tail. The large ORF1 encodes a polyprotein of 1,637 amino acids (aa) with conserved RNA-dependent RNA polymerase (RdRp) and DEAD-like helicase superfamily (DEXDc) domains. ORF2 encodes a putative 633-aa protein with unknown function. A BLAST search showed that the nucleotide sequence of the AhFV1 genome is 41.28% identical to that of Sclerotium rolfsii fusarivirus 2 and 40.49% identical to that of Sclerotium rolfsii fusarivirus 1. Phylogenetic analysis based on RdRp and helicase (Hel) sequences indicated that AhFV1 is related to unclassified mycoviruses and other fusariviruses. Our data suggest that AhFV1 should be classified as a member of the newly proposed family "Fusariviridae". This is the second virus and the first full genome sequence of a fusarivirus from A. heimuer.

Molecular characteristics of a novel ssRNA virus isolated from Auricularia heimuer in China.

A novel positive-sense single-stranded RNA virus was isolated from strain CCMJ1271 of the fungus Auricularia heimuer, and the complete genome sequence of the virus was determined. Database searching, sequence alignment, and phylogenetic analysis revealed that this fungal virus and some viruses of family Virgaviridae clustered into a single branch of a phylogenetic tree, and we thus tentatively named the virus "Auricularia heimuer mycovirgavirus 1" (AhMV1). The AhMV1 genome consists of 9,934 nucleotides and contains a short poly(A) tail and three open reading frames (ORFs). ORF1 encodes an RNA-dependent RNA polymerase (RdRp), ORF2 encodes a protein that is homologous to movement proteins of plant virgaviruses, and ORF3 encodes a coat protein (CP). AhMV1 is the first virus to be discovered in A. heimuer.

The complete genome sequence of a novel hypovirus infecting Bipolaris oryzae.

Hypoviruses are positive-sense single-stranded RNA mycovirus that infect filamentous fungi. However, hypoviruses have not been reported in Bipolaris oryzae, an important phytopathogenic fungus in water bamboo and rice. Here, we report the characterization of a novel hypovirus, tentatively named "Bipolaris oryzae hypovirus 1" (BoHV1), isolated from strain ES35 of B. oryzae infecting water bamboo. The complete genome of BoHV1 consists of 13,596 nucleotides and a poly(A) tail at the 3' end. BoHV1 has single open reading frame (ORF) and encodes a putative polyprotein (4,218 amino acids) containing four potential conserved domains for a papain-like protease, a protein of unknown function (DUF3525), RNA-dependent RNA polymerase (RdRp), and helicase. Phylogenetic analysis of the polyprotein, RdRp, and helicase domains suggested that BoHV1 belongs to the genus Hypovirus within the family Hypoviridae. This is the first report of the presence of a hypovirus in the phytopathogenic fungus B. oryzae.

Molecular characterization of a novel fusarivirus infecting the plant-pathogenic fungus Botryosphaeria dothidea.

A novel virus, Botryosphaeria dothidea fusarivirus 1 (BdFV1), was isolated from a fungal strain, SDAU11-86 of Botryosphaeria dothidea, and its complete genome sequence was determined. BdFV1 has a single-stranded positive-sense (+ssRNA) genome with 6,179 nucleotides, excluding the poly(A) tail. The genome of BdFV1 contains two putative open reading frames (ORFs). The first ORF encodes a large polyprotein of 1,544 amino acids (aa) with conserved RNA-dependent RNA polymerase and viral helicase domains. The second ORF encodes a putative 481-aa protein with unknown function. Sequence comparisons and phylogenetic analysis suggested that BdFV1 is a novel mycovirus belonging to the newly proposed family "Fusariviridae". This is the first report of a +ssRNA mycovirus in B. dothidea.

UvCom1 Is an Important Regulator Required for Development and Infection in the Rice False Smut Fungus Ustilaginoidea virens.

Ustilaginoidea virens is an economically important biotrophic fungal pathogen that causes rice false smut worldwide. However, the regulatory mechanisms of smut ball formation under U. virens infection remain unclear. Here, we identified an important transcription factor, UvCom1, from this pathogen, which regulates the formation of smut balls on rice spikelets. UvCom1 contains two conserved internal repeat 1 (RPT) domains and is found only in fungi, with specific conservation in species of Basidiomycetes and Ascomycetes. UvCom1 protein N- or C-terminal fusion vectors both showed transactivation activity in yeast. Deletion of UvCom1 significantly affected the vegetative growth and conidiation of U. virens. UvCom1 negatively regulated the responses to oxidative, osmotic, and cell wall stresses. Remarkably, UvCom1 was found to be essential for the formation of rice smut balls, and UvCom1 deletion mutants lost the ability to stably utilize nutrients from the rice host. UvCom1 was also highly expressed at the mycelial expansion stage. Transcriptomic analysis and quantitative real-time PCR revealed that UvCom1 could affect the expression of genes significantly enriched in transmembrane transport. This study demonstrates that UvCom1 is a key transcription factor governing smut ball formation of this biotrophic fungus.

Investigation of East Asian clouds with polarization light detection and ranging.

In this paper we present results of investigation of the main optical properties of East Asian clouds with a ground-based polarization lidar placed in Daejeon, Republic of Korea. Asian dust is located in elevated layers of the atmosphere in spring, travels long distances, and causes significant damage to ecology. We present backscattering matrices of clouds obtained from polarimetric remote measurements which comprise information on the scattering and absorption properties of cloud particles, their morphology, and spatial orientation. Theory of our applied lidar polarization experiment is presented in terms of the instrumental vectors of a transmitter and a receiver. Methods of solving linear and nonlinear systems of equations comprising echo signals are considered. Some numerical and measurement results are presented to illustrate the efficiency and versatility of the method of estimating the cloud parameters.

Blood lipid peroxidation, antioxidant enzyme activities and hemorheological changes in autistic children.

OBJECTIVES: Early infantile autism is a severe form of childhood psychiatric disease with characteristic symptoms. Hyperserotoninaemia in 43.5%, lactic acidosis 43% and hyperpyruvataemia in 30% were biochemically demonstrated in autistic children. Our earlier results led to the postulation that a dissequilibrium in the blood redox is involved in infantile autism; the oxidative loading and the antioxidant defending enzyme system were investigated together with the hemorheological parameters in infantile autism. METHODS: Malonyl-dialdehyde (MDA) endproduct of lipid peroxidation and activities of the antioxidant enzymes: superoxide dismutase (SOD), catalase (C-ase), glutathione peroxidase (GP-ase) and reduced glutathione (GSH) were biochemically determined from plasma and red blood cells. PATIENTS: The antioxidant specificities were investigated in plasma and red blood cell haemolysate from 25 infantile autistic children. RESULTS: Significantly increased superoxide dismutase (SOD) (2.89 vs. 1.32 U/mg protein, p < 0.01) and decreased glutathione peroxidase (0.620 vs. 0.910 U/mg protein, p < 0.01) levels as well as catalase (0.463 vs. 4.948 BU/mg protein, p < 0.001) activities were detected; while the plasma and erythrocyte lipid peroxidation and the reduced glutathione (GSH) levels did not change. The results of the investigated prooxidant and the antioxidant status provide evidence that there exists an oxidative stress in children with infantile autism. While investigating the hemorheological parameters of 25 infantile autistic patients, some characteristic pathological parameters were detected: the initial filtration rate (Fi) (0.72 vs. 0.75 p < 0.01) and the clogging rate (CR) (1.926 vs. 2.912, p < 0.01) values of red blood cells (RBC) decreased while the mean transit time (Tc) (8.93 vs. 7.39, p < 0.001) increased suggesting reduced RBC deformability.