dsRNA Components Research Articles

Molecular characterization of a new botybirnavirus that infects Alternaria sp. from tobacco.

The genus Alternaria comprises many important fungal pathogens that infect a wide variety of organisms. In this report, we present the discovery of a new double-stranded RNA (dsRNA) mycovirus called Alternaria botybirnavirus 2 (ABRV2) from a phytopathogenic strain, XC21-21C, of Alternaria sp. isolated from diseased tobacco leaves in China. The ABRV2 genome consists of two dsRNA components, namely dsRNA1 and dsRNA2, with lengths of 6,162 and 5,865 base pairs (bp), respectively. Each of these genomic dsRNAs is monocistronic, encoding hypothetical proteins of 201.6 kDa (P1) and 2193.3 kDa (P2). ABRV2 P1 and P2 share 50.54% and 63.13% amino acid sequence identity with the corresponding proteins encoded by dsRNA1 of Alternaria botybirnavirus 1 (ABRV1). Analysis of its genome organization and phylogenetic analysis revealed that ABRV2 is a new member of the genus Botybirnavirus.

A special satellite-like RNA of a novel hypovirus from Pestalotiopsis fici broadens the definition of fungal satellite.

Satellites associated with plant or animal viruses have been largely detected and characterized, while those from mycoviruses together with their roles remain far less determined. Three dsRNA segments (dsRNA 1 to 3 termed according to their decreasing sizes) were identified in a strain of phytopathogenic fungus Pestalotiopsis fici AH1-1 isolated from a tea leaf. The complete sequences of dsRNAs 1 to 3, with the sizes of 10316, 5511, and 631 bp, were determined by random cloning together with a RACE protocol. Sequence analyses support that dsRNA1 is a genome of a novel hypovirus belonging to genus Alphahypovirus of the family Hypoviridae, tentatively named Pestalotiopsis fici hypovirus 1 (PfHV1); dsRNA2 is a defective RNA (D-RNA) generating from dsRNA1 with septal deletions; and dsRNA3 is the satellite component of PfHV1 since it could be co-precipitated with other dsRNA components in the same sucrose fraction by ultra-centrifuge, suggesting that it is encapsulated together with PfHV1 genomic dsRNAs. Moreover, dsRNA3 shares an identical stretch (170 bp) with dsRNAs 1 and 2 at their 5' termini and the remaining are heterogenous, which is distinct from a typical satellite that generally has very little or no sequence similarity with helper viruses. More importantly, dsRNA3 lacks a substantial open reading frame (ORF) and a poly (A) tail, which is unlike the known satellite RNAs of hypoviruses, as well as unlike those in association with Totiviridae and Partitiviridae since the latters are encapsidated in coat proteins. As up-regulated expression of RNA3, dsRNA1 was significantly down-regulated, suggesting that dsRNA3 negatively regulates the expression of dsRNA1, whereas dsRNAs 1 to 3 have no obvious impact on the biological traits of the host fungus including morphologies and virulence. This study indicates that PfHV1 dsRNA3 is a special type of satellite-like nucleic acid that has substantial sequence homology with the host viral genome without encapsidation in a coat protein, which broadens the definition of fungal satellite.

Characterization of a novel botybirnavirus isolated from a phytopathogenic Alternaria fungus.

Alternaria fungi are important pathogens infecting a wide variety of organisms. Here, we report a novel double-stranded RNA (dsRNA) mycovirus named Alternaria botybirnavirus 1 (ABRV1) isolated from a phytopathogenic Alternaria sp. strain (SCFS-3) infecting a pear tree in China. ABRV1 has two dsRNA components (dsRNAs 1 and 2) with the sizes of 6,188 and 5,903bp, containing two putative open reading frames encoding two polyproteins (202 and 192kDa, respectively). The polyprotein encoded by ABRV1 dsRNA1 shares 41% amino acid (aa) sequence identity with the one encoded by dsRNA2 (instead of dsRNA1) of Sclerotinia sclerotiorum botybirnavirus 1 (SsBRV1). Conversely, the polyprotein encoded by ABRV1 dsRNA2 shares 46% aa sequence identity with the one (i.e., cap-pol fusion protein) encoded by SsBRV1 dsRNA1. ABRV1 has isometric spherical virus particles (~40nm in diameter), putatively composed of the 60-, 70- and 80-kDa structural proteins. The genomic organization and phylogenetic analyses revealed that ABRV1 belongs to a newly proposed family "Botybirnaviridae", and to our knowledge, this is the first report of a botybirnavirus infecting an Alternaria sp. strain.

Molecular analysis of six segments of Tobacco leaf enation virus, a novel phytoreovirus from tobacco

Tobacco leaf enation virus (TLEV) is a putative member of the genus Phytoreovirus within the family Reoviridae. Previous western blot analysis of structural viral proteins (apparent molecular weights of 93 kDa; 58 kDa; 48 kDa; 39 kDa and 36 kDa) associated with TLEV, isolated from infected tobacco in South Africa, suggested that these proteins may correspond to structural Wound tumor virus (WTV) proteins. To further establish the nature of this novel virus disease phenotype in tobacco, molecular characterization of six dsRNA components was undertaken. Full-length cDNA clones were obtained by an optimized modified single-primer amplification sequence-independent dsRNA cloning method. Results of this study revealed the conserved terminal sequence: 5'GG(U/C)...UGAU 3' of segments S6-S12, while adjacent to these conserved terminal sequences are imperfect inverted repeats (7-15 bp in length), both features being common to reoviruses. The complete nucleotide sequences of segments S5 (2,610 bp), S7 (1,740 bp), S8 (1,439 bp), S10 (1,252 bp), S11 (1,187 bp) and S12 (836 bp) were determined. Comparison of full-length nucleotide sequences with corresponding segments of other phytoreoviruses, Rice gall dwarf virus (RGDV), Rice dwarf virus (RDV) and WTV has shown nucleotide and predicted amino acid identities within the range of 30-60%. TLEV consistently shows a higher identity to WTV than to other phytoreovirus species where sequence data is available. Each segment had a single predicted open reading frame encoding proteins with calculated molecular weights of S5 (90.6 kDa); S7 (58.1 kDa); S8 (47.7 kDa); S10 (39.8 kDa); S11 (35 kDa) and S12 (19.5 kDa). The relatively low nucleotide and amino acid identity to other members of the genus demonstrates that TLEV is a novel phytoreovirus, distinct from the only other reported dicotyledenous-infecting WTV and is the first phytoreovirus reported to emerge in Africa.

97. Type I Interferon Response Against Virus Vectors and Non-Viral Gene Transfer

The first line of defense that gene delivery vectors, viral or non-viral, confront in cells is the type I interferon (IFN) response. Cells produce type I IFNs as a response to dsRNA (intermediates of viral replication) and other viral components. IFNs induce the expression of IFN stimulated genes that can be used as markers for IFN response. One of these IFN stimulated genes is the Myxovirus resistance protein A (MxA).

Double‐stranded RNA: distribution and analysis among isolates of Rhizoctonia solani AG‐2 to ‐13

The occurrence, distribution, and genetic relatedness of double‐stranded RNA (dsRNA) components from 36 isolates of Rhizoctonia solani belonging to nine anastomosis groups (AGs) were studied using electrophoretic analysis and RNA–RNA blot hybridization. DsRNA was consistently detected in all 36 isolates. The size of the dsRNA components varied considerably, ranging from 0·74 to 23 kb. Two thirds of isolates possessed different size dsRNA components. Only two of the isolates had small size dsRNA between 0·5 and 1·0 kb. The biotin‐labelled dsRNA probes provided the sensitivity and specificity required to study genetic relationships of dsRNA. As little as 10 pg of ‘hybridizing’ dsRNA could be detected using biotin‐labelled total dsRNA with no detectable nonspecific‐hybridization. Results from several dot‐spot as well as RNA–RNA gel hybridization experiments revealed considerable sequence heterogeneity among dsRNA components within each isolate or isolates from the same AG.

Characterization of mycoviruses and analyses of chitinase secretion in the biocontrol fungus Metarhizium anisopliae.

Metarhizium anisopliae is the best-characterized entomopathogen and is used to control insect pests in sugar cane plantations in Brazil on a commercial scale. We have previously reported the infection of some M. anisopliae strains by dsRNA mycoviruses. Here we describe the purification and characterization of the viruses (MaV-A1, MaV-M5, MaV-RJ) in terms of dsRNA content, capsid proteins, electron microscopy, Western blot, and hybridization patterns. One spontaneous mutant lost some of the high molecular weight dsRNA components and showed significant alterations in colony morphology and spore production, suggesting that viral genes interfere with fungal phenotype. A comparison between dsRNA mycovirus-free and infected M. anisopliae isolates showed that virus-free isolates have increased endochitinase secretion. By comparing the following parameters: the buoyant density in CsCl of the presumed virions; the number and estimated molecular weight of the dsRNA components and the molecular mass of the capsid proteins to other mycoviruses previously described, we suggest the inclusion of MaV-A1 and MaV-M5 in the family Totiviridae and MaV-RJ in the family Partitiviridae.

Selective occurrence of endogenous double-stranded RNAs in insects

Noninfectious endogenous double-stranded RNAs (dsRNAs) of 1–15 kb in length are widely distributed in both the plant and fungal kingdoms. Here we report the first evidence of the selective occurrence of such plasmid-like dsRNAs in the animal kingdom, a variety of symptomless insects. A considerable portion of larval, pupal and adult individuals of butterflies were found to harbor characteristic sets of dsRNA components at defined levels. Some cultured cell lines from the cabbage armyworm, Mamestra brassicae, also harbored indigenous dsRNAs, which had no lytic effects on the host cells and did not interfere with their normal growth. The moth dsRNAs were incapable of naturally infecting noncarrier cells during culture, a result that suggests efficient transmission of the dsRNAs to progeny cells through cell division. The total copy number of the dsRNAs in the cultured moth cells was estimated to be about 3000. The moth dsRNAs were found broadly around the mitochondrial fraction after subcellular fractionation, and they did not appear to form distinct virus-like particles.

Detection of grapevine leafroll‐associated closterovirus III by molecular hybridization

Highly purified double‐stranded (ds) RNA was obtained from cortical scrapings of mature canes of vines infected by grapevine leafroll‐associated closterovirus III (GLRaV III) using phenol‐chloroform extraction, chromatography on CF‐11 cellulose minicolumns and enzymatic digestion. Complementary (c) DNA fragments of various lengths, obtained by random priming denatured dsRNA templates, were cloned into the plasmid pUC‐18 at the Smal site in Escherichia coli strain DH5α. Two 32P‐labelled cDNA clones denoted p16ds (c. 1100 bp) and p23ds (c. 1500 bp) were successfully used as probes for detecting GLRaV III sequences in grapevine extracts from leaves and petioles, or cortical tissues. Probe p23ds was virus‐specific and did not hybridize with total RNA from healthy controls, or from vines infected by grapevine leafroll‐associated closterovirus I (GLRaV I), or with genomic RNA from purified grapevine closterovirus A (GVA) and B (GVB). A riboprobe (pGEM23ds) transcribed from p23ds in transcription vector pGEM3zf specifically recognized GLRaV III sequences, but not GLRaV I or GVA sequences, in extracts from differently infected vines. Moreover, in Northern blots, the same probe hybridized also with smaller dsRNA components, which may be replicative forms of subgenomic RNAs.

A third cryptic virus in beet (Beta vulgaris)

Japanese leaf beet Beta vulgaris var. cicla cv. Fudanso plants were found to contain four double‐stranded RNA (dsRNA) components in apparently healthy beet plants. Two were identified as from beet cryptic virus 1 (BCV1), but the other two showed different mobilities on gel electrophoresis and were transcribed into complementary DNA (cDNA) and cloned. Hybridization analysis showed no significant sequence homology between these two dsRNAs and the dsRNA components of BCV1 or the other known cryptic virus of beet, BCV2. Slot‐ and dot‐blot hybridization were used with cDNA clones as probes to identify plants containing these two dsRNA components. Virus particles were purified from these plants and were shown to contain the two new dsRNA components, thus demonstrating the existence of a new beet cryptic virus, which we have called BCV3.

Structural Studies of the Enveloped dsRNA Bacteriophage θ6 of Pseudomonas syringae by Raman Spectroscopy: II. Nucleocapsid Structure and Thermostability of the Virion, Nucleocapsid and Polymerase Complex

Structures and thermostabilities of the double-stranded (ds) RNA bacteriophage θ6 and of its isolated nucleocapsid-polymerase complex (nucleocapsid core) and dsRNA components have been investigated by Raman spectroscopy. The spectra show that proteins of the θ6 virion are collectively deficient in β-sheet secondary structure. In particular, the major protein (P8) of the outer spherical shell of the θ6 nucleocapsid exhibits a secondary structure dominated largely by α-helix and irregular conformations. The absence of appreciable β-structure in the P8 subunit suggests a tertiary conformation lacking the β-barrel motif common to subunits of most other spherical viral capsids. In addition, the Raman spectra show that subunits of the dodecahedral nucleocapsid core are also predominantly α-helical. The results thus indicate a largely α-helical secondary structure for the major subunit (P1) of the θ6 nucleocapsid core, as well as for the P8 subunit of the outer spherical shell.Using Raman difference spectroscopy, we demonstrate that proteins of the nucleocapsid core (P1, P2, P4 and P7) interact extensively with the packaged θ6 RNA genome, and further, that conformational stability of the packaged RNA is reduced upon removal from the core. Also, we find that proteins of the θ6 nucleocapsid are significantly more thermostable than proteins of the viral membrane envelope, which are reported in the accompanying paper (Li et al., 1993). The present results suggest that both the architectural principles and modes of protein-RNA interaction in the θ6 virion differ fundamentally from those of icosahedral single-stranded RNA viruses.Both Raman and circular dichroism spectra indicate that the dsRNA genome of θ6 is an A-form structure. The Raman marker bands signify the presence only of C3′-endo/anti nucleoside conformers. The Raman signature of dsRNA, revealed in the spectrum of the θ6 genome, is discussed here as a model for assessing base-pairing and base-stacking interactions in other ribonucleoprotein assemblies.

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<h3>Abstract</h3> Satellites associated with plant or animal viruses have been largely detected and characterized, while those from mycoviruses together with their roles remain far less determined. Three dsRNA segments (dsRNA 1 to 3 termed according to their decreasing sizes) were identified in a strain of phytopathogenic fungus <i>Pestalotiopsis fici</i> AH1-1 isolated from a tea leaf. The complete sequences of dsRNAs 1 to 3, with the sizes of 10316, 5511, and 631 bp, were determined by random cloning together with a RACE protocol. Sequence analyses support that dsRNA1 is a genome of a novel hypovirus belonging to a newly proposed genus “Alphahypovirus” of the family <i>Hypoviridae</i>, tentatively named Pestalotiopsis fici hypovirus 1 (PfHV1); dsRNA2 is a defective RNA (D-RNA) generating from dsRNA1 with septal deletions; and dsRNA3 is the satellite component of PfHV1 since it could be co-precipitated with other dsRNA components in the same sucrose fraction by ultra-centrifuge, suggesting that it is encapsulated together with PfHV1 genomic dsRNAs. Moreover, dsRNA3 shares an identical stretch (170 bp) with dsRNAs 1 and 2 at their 5′ termini and the remaining is heterogenous, which is distinct from a typical satellite that generally has very little or no sequence similarity with helper viruses. More importantly, dsRNA3 lacks a substantial open reading frame (ORF) and a poly (A) tail, which is unlike the known satellite dsRNAs of hypoviruses, as well as unlike those in association with <i>Totiviridae</i> and <i>Partitiviridae</i> since the latters are encapsidated in coat proteins. As up-regulated expression of dsRNA3, dsRNA1 was significantly down-regulated, suggesting that dsRNA3 negatively regulates the expression of dsRNA1, whereas dsRNAs 1 to 3 have no obvious impact on the biological traits of the host fungus including morphologies and virulence. This study indicates that PfHV1 dsRNA3 is a special type of satellite-like nucleic acid that has substantial sequence homology with the host viral genome without encapsidation in a coat protein, which broadens the definition of satellite. <h3>IMPORTANCE</h3> Satellites in association with plant or animal viruses have been largely detected and characterized, while those from mycoviruses together with their roles remain far less determined. Here, a special satellite-like dsRNA (SatL-dsRNA) together with its helper virus, a novel hypovirus from <i>Pestalotiopsis fici</i>, was identified and characterized. This SatL-dsRNA lacks a substantial open reading frame and a poly (A) tail, which is unlike the known satellite dsRNAs of hypoviruses. It is also unlike those in association with <i>Totiviridae</i> and <i>Partitiviridae</i> since the latters are encapsidated in coat proteins. As up-regulated expression of the SatL-dsRNA, the helper virus genome was significantly down-regulated, suggesting that it negatively regulates the genomic expression of the helper virus. This special SatL-dsRNA has substantial sequence homology with the host viral genome and is not encapsidated in the coat protein of its helper virus, which represents a novel class of satellite-like nucleic acids, and it broadens the definition of satellite.

Colletotrichum gloeosporioides isolates causing different anthracnose diseases on Stylosanthes in Australia carry distinct double-stranded RNAs

The electrophoretic patterns of conidial protein and mycelial double-stranded (ds) RNA were investigated in 14 Australian isolates of Colletotrichum gloeosporioides . All isolates showed virtually identical patterns of conidial proteins but there was considerable variation in their dsRNA components. Each of seven isolates causing Type A anthracnose contained only two major dsRNA bands which were either absent or present in only minor amounts in seven Type B isolates. Considerable variation was observed between different Type B isolates but all isolates contained three major bands which were absent in Type A isolates, thus providing biochemical evidence to confirm the existence of two distinct pathogen populations in Australia.

Structural properties of double-stranded RNAs associated with biological control of chestnut blight fungus

Double-stranded RNAs (ds RNAs) are thought to be the cytoplasmic determinants responsible for the phenomenon of transmissible hypovirulence in the chestnut blight fungus Endothia parasitica [Murr.] Anderson. The three major ds RNA components associated with the North American hypovirulent strain, Grand Haven 2, were characterized with respect to molecular-hybridization specificity and RNase T1-digestion patterns. The large (L-RNA; approximately 9 kilobase pairs) and middle-sized (M-RNA; approximately 3.5 kilobase pairs) ds RNA components cross-hybridized under stringent conditions and exhibited indistinguishable partial and complete RNase T1 digestion patterns relative to their 5' and 3' termini. These results suggest that M-RNA was derived from L-RNA by an internal deletion event. The small (S-RNA; approximately 1 kilobase pair) RNA was unrelated to L- and M-RNA by these criteria. However, all three ds RNA components contained RNase T1-resistant oligonucleotides at one 5' terminus and at the corresponding 3' terminus of the complementary strand. These RNase T1-resistant species exhibited properties consistent with stretches of poly(uridylic acid) and poly(adenylic acid), respectively. The combined results are discussed in terms of the structural organization of hypovirulence-associated ds RNA molecules and their similarities to "double-stranded" RNA molecules observed in plant and animal cells infected with single-stranded RNA viruses.

Molecular Cloning of the Double-stranded RNA of Beet Cryptic Viruses

Summary Three of the four dsRNA components of purified beet cryptic virus (BCV) were copied into cDNA and cloned into pUC9. Clones corresponding to RNAs 1, 3 and 4 did not hybridize to each other or to RNA 2, suggesting that there is no significant sequence homology between the four dsRNA components. RNA extracted from 15 BCV-infected beet plants was analysed by Northern blotting using the cDNA clones as probes. Nine plants were found to contain RNAs 1, 3 and 4 whereas in six plants only RNAs 3 and 4 were detectable. The results are compatible with the occurrence of two different viruses. The sensitivity and specificity of the cDNA hybridization assay was greater than that of immunosorbent electron microscopy in the detection of BCVs.

Naked dsRNA Associated with Hypovirulence of Endothia parasitica Is Packaged in Fungal Vesicles

SUMMARY Hypovirulence in Endothia parasitica is known to be associated with dsRNA. The characteristics of this cytoplasmically transmissible element suggest it might be viral in nature. Attempts to isolate viral particles indicate the presence of two particulate fractions in hypovirulent (HV) strains and a similar one in virulent (V) strains. Composition analyses of the fractions show these to be similar except that those from HV strains include dsRNA. Also present in these particles are carbohydrate, protein and chloroform-methanol-extractable substances. The protein composition is too low to indicate the presence of a capsid. The neutral sugars that are present (arabinose, mannose, galactose, glucose) in these particles are also present in the fungal cell wall. Silver- and ethidium bromide-stained polyacrylamide gels resolve several minor as well as the known major dsRNA components. These components are arbitrarily divided into four mol. wt. groups: 4.5 × 106 to 5.0 × 106, 1.2 × 106 to 1.5 × 106, 5.8 × 105 to 6.2 × 105 and &lt; 4.6 × 105. Not all segments are present at all times, although the phenotype of the culture is not affected. Proteins are not detectable in vesicles of HV strains, but six proteins are detectable from V strains by gel electrophoresis. Assays for [32P]UMP incorporation indicate the presence of a RNA polymerase closely associated with the dsRNA. Our results suggest that the transmissible element responsible for hypovirulence is a naked dsRNA genome packaged within vesicles formed by the host.

Evaluation of the presence of double-stranded RNA in Ceratocystis ulmi

Nineteen cultures of Ceratocystis ulmi were classified by mating type and growth rate in vitro. Eleven of the cultures contained double-stranded RNA (dsRNA) with 52% of the segments having molecular weights (MWs) ranging from 1.5 × 106 to 2.0 × 106. Eight of 11 fast-growing (aggressive) isolates contained one to five dsRNA segments with MWs ranging from 0.6 × 106 to 3.1 × 106, whereas 3 of 8 slower growing (nonaggressive) cultures had one or two dsRNA species with MWs ranging from 1.8 × 106 to 2.7 × 106. In addition, cultures with both A and B mating types were found to either contain or lack dsRNA. Thus, there was no clear association between the presence of dsRNA or specific components of dsRNA and aggressiveness or nuclear mating type in the cultures of C. ulmi examined in this study.

Infrequent Transmission of Double-stranded RNA Virus Particles but Absence of DNA Proviruses in Single Ascospore Cultures of Gaeumannomyces graminis

SUMMARY A field isolate, 3b1a, of the wheat take-all fungus Gaeumannomyces graminis var. tritici, was previously shown to be infected with three serologically unrelated viruses, A, B, and C. It is now shown that virus B can be separated into two distinct strains, designated B1 and B2. All four viruses, A, B1, B2 and C, were faithfully transmitted into conidia. However, six out of eight single ascospore cultures derived from 3b1a single conidial cultures were shown to be virus-free. The remaining two single ascospore cultures each contained only one virus, which appeared to be the same in each culture. This virus was serologically indistinguishable from virus B1, but had dsRNA components of mol. wt. lower than those of virus B1 and had only the smaller of the two capsid polypeptide species of virus B1. No DNA provirus molecules homologous to viruses B1, B2 or C could be detected in two of the virus-free ascospore cultures or in one of the virus-infected ascospore cultures. Very low concentrations of virus particles were detected in hyphal tip isolates of G. graminis. After prolonged storage and subculturing of these isolates, the concentration of virus particles had increased to the level of the parent culture from which the hyphal tip isolates were derived.

A Satellite Double-stranded RNA in a Virus from Gaeumannomyces graminis

SUMMARY The double-stranded (ds)RNA components of two serologically related viruses (019/6-A and 38-4-A), obtained from Gaeumannomyces graminis, have been compared. 019/6-A virus has two RNA components, ds1 and ds2, whereas 38-4-A virus has three RNA components, ds1, ds2 and ds3. T1 oligonucleotide fingerprint analysis and saturation hybridization experiments indicated that 019/6-A ds1 and 38-4-A ds1 were very similar, whereas 019/6-A ds2 and 38-4-A ds2 were about 50% homologous. 38-4-A ds3 had little homology with ds1 or ds2 of either virus. The single-stranded (ss)RNA transcripts of 38-4-A virus ds1, ds2 and ds3 each directed the synthesis in vitro of a single polypeptide, with mol. wt. 62000, 55000 and 52000 respectively. Peptide mapping showed that these three polypeptides were unrelated in sequence and that ds2 encoded the virus capsid polypeptide. The ds3 is probably not required for the replication of 38-4-A virus and may be regarded as a satellite RNA.

Stability of double-stranded RNA components of Endothia parasitica through transfer and subculture

Strains of Endothia parasitica with cytoplasmic hypovirulence contain double-stranded (ds) RNA. Virulent (V) strains can be converted to hypovirulent (H) strains by hyphal anastomosis and dsRNA transfer. The dsRNA components of a French and an Italian H strain were transferred into separate cultures of the same V strain. The resulting two new H strains were used to convert six V strains in three vegetative compatibility (v-c) groups. The major dsRNA components were faithfully transferred in all conversions and maintained during subsequent subculturing. Morphological variation was noted and representative pathogenicities were determined. The similarity of the two dsRNA populations in their effects on the fungus suggests that either can be used therapeutically for control of chestnut blight.