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PubMed2025

A positive-sense single-stranded RNA virus acquired a negative-sense open reading frame through recombination.

Although positive- and negative-sense single-stranded RNA viruses are ubiquitous in nature, there is currently no evidence of recombination or reassortment between viruses with these two major forms of genome organization. Here, we describe the discovery of brine shrimp virga-like virus 1 (BSVV1), a novel positive-sense single-stranded RNA virus with a recombinant genome structure derived from two viral phyla with differing genome organizations. The genome of BSVV1 comprises three open reading frames (ORFs). ORF1 resembles the RNA-dependent RNA polymerase of Ips virga-like virus 1 (a positive-sense RNA virus), while ORF2, transcribed in the positive orientation, is related to the glycoprotein of Hubei bunya-like virus 10 and other negative-sense RNA viruses. The predicted ORF3 was unique to BSVV1 without known homologs identified. The presence of the three protein products was verified by mass spectrometry. Notably, our analysis also revealed that BSVV1 is geographically widespread and found in brine shrimp from at least eight countries on four continents. In addition, BSVV1 was successfully cultured and proliferated to high viral loads during brine shrimp development. In sum, we provide compelling evidence of an ancient recombination event between negative- and positive-sense single-stranded RNA viruses, enriching our understanding of the evolution of genome structures in RNA viruses.

Open Reading Frames↗

PubMed2026

Structure and dynamics of the Nipah virus RNA-dependent RNA polymerase.

Nipah virus (NiV) is a highly pathogenic, nonsegmented, negative-sense RNA virus (nsNSV) from the Mononegavirales order that causes frequent outbreaks, with no approved treatment available. Replication and transcription of its genome are carried out by a viral RNA-dependent RNA polymerase (RdRp) complex composed of the large catalytic protein (L) and the tetrameric phosphoprotein (P). Recently, structural insights into the NiV RdRp complex have emerged at an unprecedented pace. In particular, snapshots of the complex in precatalytic, early-elongation, and inhibitor-bound states have been reported. In this article, we review how these data shed light on the molecular mechanisms of RNA synthesis and inhibition in NiV and explore how these insights expand our understanding of nsNSV RdRps in general.

Nipah Virus↗

PubMed2026

Development and evaluation of an ARTIC-based amplicon sequencing assay for whole-genome characterization of respiratory syncytial virus.

Respiratory syncytial virus (RSV), a ~15.2 kb negative-sense RNA virus, causes acute respiratory infections in infants and older adults. Its two subtypes, RSV-A and RSV-B, evolve rapidly, making ongoing monitoring of circulating strains essential. The Georgia Public Health Laboratory (GPHL) developed and evaluated an amplicon-based whole-genome sequencing (WGS) assay for RSV surveillance. A total of 214 de-identified remnant clinical specimens (102 RSV-A and 112 RSV-B) with RT-PCR Cq values <31 were included. RSV genomes were amplified using ARTIC-style and custom primer sets, with the ARTIC set showing superior performance. Libraries were prepared using a modified Illumina COVIDSeq protocol, sequenced on NextSeq 1000/2000 instruments, and analyzed using the GPHL-RSV-PIPE bioinformatics pipeline. Among genomes meeting validation criteria, sequencing depth was slightly higher for RSV-A (median 53,433×; mean 51,076×) than RSV-B (median 49,699×; mean 46,945×), whereas genomic coverage was slightly lower for RSV-A (median 97.5%; mean 96.6%) than RSV-B (median 98.3%; mean 97.6%). Predominant lineages were A.D.3.1 and A.D.5.2 for RSV-A and B.D.E.1 for RSV-B. For RSV-A, the assay showed 92.8% accuracy, 96.2% sensitivity, 87.2% specificity, 92.6% positive predictive value, and 93.2% negative predictive value. Intra- and inter-run precision assessed using 16 and 53-57 genomes, respectively, showed nearly 100% consensus genome identity with 0-5 nucleotide differences. Specificity testing of 31 non-RSV specimens produced no false-positive detections. Limits of detection were 4.4 TCID50/mL for RSV-A and 18.6 TCID50/mL for RSV-B. These results demonstrate that the ARTIC-based RSV WGS assay enables near real-time surveillance and strengthens data-driven public health responses to future outbreaks.IMPORTANCERSV, with two major subtypes, RSV-A and RSV-B, causes acute respiratory infections that can be severe in infants under 6 months and older adults. Current RSV surveillance at the GPHL relies on the Thermo Fisher TaqMan Gene Expression Capillary assay, which detects and subtypes RSV but lacks resolution for lineage classification and identification of emerging variants. To address this critical gap, GPHL developed and evaluated an amplicon-based WGS assay using 214 de-identified RSV clinical specimens. Genomes were amplified using ARTIC-style and custom-primer sets, with ARTIC primers showing superior performance. The assay demonstrated strong sequencing depth, genomic coverage, specificity, repeatability, reproducibility, and low limits of detection. RSV lineages were accurately determined based on genetic variation. These results establish that the ARTIC-based WGS assay enables near real-time genomic surveillance, supporting monitoring of circulating RSV strains and informing data-driven public health responses.

bioinformatics pipeline↗

PubMed2024

Heat Inactivation of Nipah Virus for Downstream Single-Cell RNA Sequencing Does Not Interfere with Sample Quality.

Single-cell RNA sequencing (scRNA-seq) technologies are instrumental to improving our understanding of virus-host interactions in cell culture infection studies and complex biological systems because they allow separating the transcriptional signatures of infected versus non-infected bystander cells. A drawback of using biosafety level (BSL) 4 pathogens is that protocols are typically developed without consideration of virus inactivation during the procedure. To ensure complete inactivation of virus-containing samples for downstream analyses, an adaptation of the workflow is needed. Focusing on a commercially available microfluidic partitioning scRNA-seq platform to prepare samples for scRNA-seq, we tested various chemical and physical components of the platform for their ability to inactivate Nipah virus (NiV), a BSL-4 pathogen that belongs to the group of nonsegmented negative-sense RNA viruses. The only step of the standard protocol that led to NiV inactivation was a 5 min incubation at 85 °C. To comply with the more stringent biosafety requirements for BSL-4-derived samples, we included an additional heat step after cDNA synthesis. This step alone was sufficient to inactivate NiV-containing samples, adding to the necessary inactivation redundancy. Importantly, the additional heat step did not affect sample quality or downstream scRNA-seq results.

Nipah Virus↗

PubMed2025

A Respiratory Syncytial Virus trailer sequence modulates viral replication and copy-back defective viral genome generation and propagation kinetics.

Copy-back defective viral genomes (cbDVGs) are key inducers of antiviral responses during negative-sense RNA virus infection. Once considered byproducts of in vitro viral replication, cbDVGs have since been detected in clinical specimens and implicated in affecting infection outcomes. The molecular mechanism of cbDVG generation remains unclear, thereby hindering our ability to manipulate cbDVG production during infection for therapeutic gain. Previous work showed that respiratory syncytial virus (RSV) cbDVG re-initiation sites cluster in trailer-end hotspots R1, R2, and R3, and that a poly-U mutation in R1 selectively reduced cbDVG formation at the mutated region. Here, we reported that a 10U mutation in R2 drastically reduced cbDVGs in this region in both minigenome and recombinant virus systems. Furthermore, during high-MOI passaging of the R2-10U virus, we observed delayed detection of cbDVGs with re-initiation sites in R1-R3 (trailer cbDVGs) compared to WT, while no differences in virus titers were observed. Interestingly, we observed the rapid emergence and accumulation of a viral variant bearing a 2-ribonucleotide deletion (R2-8U) within the R2-10U mutation sequence as early as P0. Compared to R2-10U, the R2-8U virus was stable, displayed faster generation and accumulation of trailer cbDVGs, restored cbDVGs with R2 re-initiation sites, and exhibited enhanced genomic replication. Overall, our data identify a sequence in the RSV trailer whose mutation critically modulates both viral replication and the generation/propagation of trailer cbDVGs. Our data also suggest that cbDVG generation, particularly near the trailer, may be an evolutionary tradeoff for more rapid virus genomic replication.

defective viral genome generation and accumulation↗

PubMed2025

Small interfering RNAs generated from the terminal panhandle structure of negative-strand RNA virus promote viral infection.

Virus-derived small interfering RNAs (vsiRNAs) have been widely recognized to play an antiviral immunity role. However, it is unclear whether vsiRNAs can also play a positive role in viral infection. Here, we characterized three highly abundant vsiRNAs mapped to the genomic termini of rice stripe virus (RSV), a negative-strand RNA virus transmitted by insect vectors. The three vsiRNAs shared 11 nucleotides due to the conservative genomic termini and were likely generated from viral terminal panhandle structure, depending on both Dicer1 and Dicer2 in insects. In addition to targeting viral RNAs in a miRNA-like manner, the three vsiRNAs coordinately downregulated the expression of DOPA decarboxylase, thereby suppressing the prophenoloxidase immune reaction in insect vectors. In vsiRNA-silenced transgenic rice, the viral titer significantly decreased, indicating that these vsiRNAs promote RSV replication in rice. This study elucidates a unique function of vsiRNAs derived from the conserved panhandle structure of negative-strand RNA viruses in enhancing viral infection.

RNA, Small Interfering↗

PubMed2025

Marburg Virus Minigenome Assays.

This chapter describes minigenome systems for Marburg virus (MARV), which reconstitute the viral polymerase complex functions of gene expression and genome replication. Procedures covered herein include passage and seeding of cells, transfection, sample collection, and reporter gene assays.

Marburgvirus↗

PubMed2026

Genome characterization of two novel mitoviruses and a negative-sense single-stranded RNA mycovirus from the phytopathogenic fungus Clarireedia jacksonii.

Clarireedia jacksonii is a phytopathogenic fungus responsible for dollar spot disease in turfgrass worldwide. In this study, we characterized the complete genome sequences of three novel mycoviruses isolated from C. jacksonii isolate MBCT-836 using next-generation sequencing and the fragmented and primer-ligated dsRNA sequencing (FLDS) method. Two of these viruses, designated Clarireedia jacksonii mitovirus 1 (CjMV1) and Clarireedia jacksonii mitovirus 2 (CjMV2), possess positive-sense single-stranded RNA genomes of 2,575 bp and 2,856 bp, respectively. Both viruses contain a single open reading frame that utilizes the mitochondrial genetic code and encodes an RNA-dependent RNA polymerase (RdRp). Phylogenetic analysis placed CjMV1 and CjMV2 within the genera Unuamitovirus and Duamitovirus, respectively, in the family Mitoviridae. The third virus, Clarireedia jacksonii negative-stranded RNA virus 1 (CjNSV1), features a bisegmented negative-sense RNA genome consisting of a large segment (7,961 nt) encoding an RdRp with a conserved Bunya_RdRp domain, and a small segment (1,444 nt) encoding a protein showing homology to bunyavirus nucleocapsid proteins. Phylogenetic analysis revealed that CjNSV1 clusters with members of the proposed family Sclerobunyaviridae within the order Bunyavirales. To our knowledge, this study provides the first report of complete genome sequences of mycoviruses infecting C. jacksonii, expanding our understanding of the mycovirosphere in economically significant turfgrass pathogens.

Genome, Viral↗

PubMed2025

Molecular characterisation of a novel citrus-infecting emaravirus, citrus emaravirus 1.

The application of high-throughput sequencing (HTS) has accelerated the discovery of novel viruses and the genome sequencing of poorly characterised viruses in fruit crops, revealing a greater complexity of plant viromes than previously understood. Here, we report the identification and genomic characterisation of a novel emaravirus, tentatively named "citrus emaravirus 1" (CiEV1), from Citrus limon leaves collected in the North West Province of South Africa. HTS analysis identified contigs corresponding to the four conserved emaravirus genomic segments (RNA1-RNA4), each encoding characteristic proteins: RNA-dependent RNA polymerase, glycoprotein, nucleocapsid, and movement protein. Comparison of terminal sequences and phylogenetic analysis suggest placing CiEV1 within clade D of the genus Emaravirus (family Fimoviridae). To date, no association with symptoms has been identified. This is the first report of an emaravirus detected in citrus, expanding the known host range of the genus and contributing to the growing evidence of viral diversity in citrus. Further investigation is required to determine the epidemiological significance of CiEV1 in citrus orchards.

Citrus↗

PubMed2025

Metagenomics reveals cryptic circulation of zoonotic viruses in Nigeria.

Zoonotic spillover events pose an ongoing threat to global health, with historic and recent viral diseases of international concern emerging from animal reservoirs 1-6. In Nigeria, limited surveillance of animal hosts at the human and animal interface continues to hinder our understanding of viruses that are cryptically circulating in animals near human dwellings with potential for consequential spillover events. We performed unbiased metagenomic next-generation sequencing (mNGS) on tissue and swab samples collected from 240 individual animals across 11 taxa (rodents, shrews, bats, goats, sheep, pigs, dogs, cats, chickens, cattle egrets, and lizards) in two Lassa-affected Nigerian states (Ondo and Ebonyi). Host-depleted sequencing reads were assembled into contigs, taxonomically classified, and subjected to phylogenetic analyses to characterize viral diversity, host associations, and evidence of cross-species transmission. Across all samples, we identified 214 distinct viral taxa spanning 33 families, of which 41% (n = 83) represent novel species by ICTV criteria. Positive-sense RNA viruses dominated (Coronaviridae, Picornaviridae, Astroviridae), followed by negative-sense RNA, single- and double-stranded DNA, and double-stranded RNA viruses. Notably, human-associated enteroviruses-including Hepatitis A virus (genotype 1b), echoviruses, coxsackieviruses, and noroviruses-were detected in goats, pigs, dogs, and chickens, indicating cryptic circulation of human pathogens in peridomestic and domesticated animals. Phylogenetic reconstructions revealed multiple cross-species viral sharing events, particularly among rodents, goats, sheep, and pigs, and extensive recombination within Nigerian Betacoronavirus 1 lineages. Interestingly we found a putative novel avian like coronavirus in rodents, goats and sheep. Ecological modelling demonstrated that host species identity, sample type, and sampling effort were primary drivers of viral richness and abundance, and that higher overall viral diversity strongly predicted cross-species transmission potential. Our integrated mNGS approach uncovered a rich and dynamic virome within animals inhabiting human-dominated environments in Nigeria, including undetected circulation of human enteric viruses. These findings underscore the importance of broad-taxonomic, real-time surveillance at human-animal interfaces to inform early-warning systems and pandemic preparedness, particularly in low-resource settings.

Journal Article↗

PubMed2026

Direct interaction between RSV polymerase L and active Rab11a mediates viral ribonucleoprotein transport to assembly sites.

Respiratory syncytial virus (RSV) is an enveloped, negative-sense, single-stranded RNA virus whose ribonucleoproteins (vRNPs) must be transported from cytoplasmic viral factories to the plasma membrane for efficient virion assembly. Viral vRNPs comprise genomic RNA encapsidated by nucleoprotein N and associated with the polymerase complex (L, P, and M2-1). It was previously demonstrated that newly synthesized vRNPs are transported along microtubules by hijacking Rab11a, a small GTPase involved in the regulation of recycling endosomes. In our previous study, we showed an interaction between Rab11a and vRNPs in infected cells by immunoprecipitation assays, nevertheless the molecular mechanisms underlying Rab11a viral hijacking remained unknown. Here, we provide the first comprehensive characterization of the interaction between RSV vRNPs and Rab11a using immunoprecipitation, immunofluorescence colocalization, GST pull-down assays, and biolayer interferometry. We demonstrate that the viral polymerase L is the sole vRNPs component responsible for Rab11a recognition: immunoprecipitation of L specifically co-precipitates HA-tagged Rab11a, whereas other vRNPs proteins show no interaction. In vitro binding studies confirm that L interacts directly and specifically with the active, GTP-bound form of Rab11a with sub-micromolar affinity. Domain mapping using truncated constructs reveals that this interaction requires the C-terminal methyltransferase and CTD domains of L (residues 1756-2165) and depends on Rab11a's Switch I region, known to mediate interactions with cellular Rab11a partners. Mutagenesis further highlights leucine 1860 in the L polymerase as critical for Rab11a binding. Competitive inhibition of the interaction between Rab11a and L using the minimal Rab11a-binding domain significantly impairs vRNP dynamics during infection, indicating that Rab11a-L binding is involved in the transport of vRNPs. Together, these findings establish RSV polymerase L as the key mediator of Rab11a engagement, define the molecular interface of their interaction, and reveal a potentially conserved viral strategy for genome transport. Targeting the L-Rab11a interaction could therefore be a promising strategy for the development of RSV-specific or broad-spectrum antiviral therapies.

rab GTP-Binding Proteins↗

PubMed2025

Ebola virus VP35 NNLNS motif modulates viral RNA synthesis and MIB2-mediated signaling.

Ebola virus (EBOV) is a nonsegmented, negative-sense virus (NNSV) with a single-stranded RNA genome. EBOV encodes for a limited number of proteins and thus depends on host factors to facilitate viral replication and pathogenesis. Of the virus-encoded proteins, multifunctional EBOV VP35 (eVP35) is necessary for host immune evasion and viral RNA synthesis. Previous proteomics studies identified an interaction between eVP35 and the host E3 ubiquitin ligase Mindbomb 2 (MIB2). Here, we show how an NNLNS (Asn-Asn-Leu-Asn-Ser) motif (residues 201 to 205) within eVP35 serves as a binding site for MIB2. This motif is critical for eVP35-dependent inhibition of MIB2-mediated interferon induction. It is also important for EBOV RNA synthesis as MIB2 binding to eVP35 inhibited EBOV minigenome activity. Altogether, these findings highlight the importance of the eVP35 protein and the role of host factors in EBOV infection.

Ebolavirus↗

PubMed2001

Bunyamwera bunyavirus nonstructural protein NSs is a nonessential gene product that contributes to viral pathogenesis.

Bunyamwera virus (family Bunyaviridae, genus Bunyavirus) contains a tripartite negative-sense RNA genome. The smallest RNA segment, S, encodes the nucleocapsid protein N and a nonstructural protein, NSs, in overlapping reading frames. We have generated a mutant virus lacking NSs, called BUNdelNSs, by reverse genetics. Compared with the wild-type (wt) virus, BUNdelNSs exhibited a smaller plaque size and generated titers of virus approximately 1 log lower. In mammalian cells, the mutant expressed greatly increased levels of N protein; significantly, the marked inhibition of host cell protein synthesis shown by wt virus was considerably impaired by BUNdelNSs. When inoculated by the intracerebral route BUNdelNSs killed BALB/c mice with a slower time course than wt and exhibited a reduced cell-to-cell spread, and titers of virus in the brain were lower. In addition, the abrogation of NSs expression changed Bunyamwera virus from a noninducer to an inducer of an interferon-beta promoter. These results suggest that, although not essential for growth in tissue culture or in mice, the bunyavirus NSs protein has several functions in the virus life cycle and contributes to viral pathogenesis.

Aedes↗

PubMed2005

Attenuation of bunyavirus replication by rearrangement of viral coding and noncoding sequences.

Bunyamwera virus (BUN) is the prototype virus of the family Bunyaviridae. BUN has a tripartite negative-sense RNA genome comprising small (S), medium (M), and large (L) segments. Partially complementary untranslated regions (UTRs) flank the coding region of each segment. The terminal 11 nucleotides of these UTRs are conserved between the three segments, while the internal regions are unique. The UTRs direct replication and transcription of viral RNA and are sufficient to allow encapsidation of viral RNA into ribonucleoprotein complexes. To investigate the segment-specific functions of the UTRs, we have used reverse genetics to recover a recombinant virus (called BUN MLM) in which the L segment open reading frame (ORF) is flanked by the M segment UTRs. Compared to wild-type virus, BUN MLM virus shows growth attenuation in cultured mammalian cells and a slower disease progression in mice, produces small plaques, expresses reduced levels of L mRNA and L (RNA polymerase) protein, synthesizes less L genomic and antigenomic RNA, and has an increased particle-to-PFU ratio. Our data suggest that the packaging of BUN RNAs is not segment specific. In addition, the phenotype of BUN MLM virus supports the finding that BUN UTRs differ in their regulation of RNA synthesis but suggests that the interplay between each segment UTR and its cognate ORF may contribute to that regulation. Since BUN MLM virus is attenuated due to an essentially irreversible mutation, the rearrangement of UTRs is a feasible strategy for vaccine design for the more pathogenic members of the Bunyaviridae.

Animals↗

PubMed2025

Purification and Ultramicroscopic Observation of the Influenza A Virus Ribonucleoprotein Complex.

Influenza A virus (IAV) has an eight-segmented, single-stranded, negative-sense viral genomic RNA (vRNA). Each vRNA strand associates with nucleoproteins and an RNA-dependent RNA polymerase complex to form a viral ribonucleoprotein (vRNP) complex. IAV vRNPs adopt a flexible double-helical configuration that varies in length. Although the transcription and replication of vRNA take place in the context of vRNPs, the precise structural conformation of vRNPs during RNA synthesis remains partially elucidated. To unravel the intricate ultrastructure of the vRNP, it is necessary to purify it while preserving its native functionality. Herein, we introduce a comprehensive protocol for the purification of IAV vRNPs using glycerol gradient ultracentrifugation. Furthermore, we provide a method for the high-speed atomic force microscopy observation of vRNPs during viral RNA synthesis.

Ribonucleoproteins↗

PubMed2025

Efficient Expression of Oropouche Virus Nonstructural Proteins NSs and NSm.

Oropouche fever, a mosquito- or midge-borne emerging zoonotic disease endemic to South and Central America, manifests as a dengue-like acute febrile illness with occasional occurrences of meningitis or meningoencephalitis. The causative agent, Oropouche virus (OROV), belongs to the genus Orthobunyavirus within the family Peribunyaviridae. Its tripartite negative-sense RNA genome comprises small (S), medium (M), and large (L) segments, encoding structural N, Gn/Gc, and L proteins, respectively. Additionally, the S- and M-segments encode nonstructural proteins: NSs and NSm, which may act as virulence factors. OROV NSs functions as an interferon antagonist with an unknown mechanism, while the roles of OROV NSm remain elusive. This chapter introduces efficient expression systems for OROV NSm and NSs proteins. Validating the presence of a signal peptide at the N-terminus of NSm protein is essential for its expression. Furthermore, expressing OROV NSs protein independently of an RNA polymerase II promoter is crucial to prevent restricted gene expression, potentially caused by NSs inhibiting cellular RNA polymerase II, as observed in closely related bunyavirus NSs proteins. These protein expression strategies offer insights into the molecular characterization of OROV NSm and NSs proteins, facilitating a deeper understanding of their virulence mechanisms.

Viral Nonstructural Proteins↗

PubMed2026

The cellular protein TIAR mediates rapid initiation of West Nile virus genome RNA synthesis.

During the intracellular replication cycle of West Nile virus (WNV), genome RNA synthesis is initially inefficient but increases exponentially as viral replication complexes are sequestered in invaginations in the endoplasmic reticulum. In this study, we investigated the functional role of the cellular protein TIAR (T-cell intracellular antigen-related protein) in the transcription of WNV genome RNA. Close colocalization of cytoplasmic TIAR with viral double-stranded RNA was detected by a proximity ligation assay in WNV-infected cells. TIAR binds specifically to the WNV 3'(-) SL but not to the complementary WNV 5'(+) SL in in vitro RNA binding assays. Only the 3' end of the WNV minus-strand RNA was enriched by immunoprecipitation of infected cell lysates with anti-TIAR antibody. Stable overexpression of TIAR in clonal A549 cells increased the ratio of intracellular viral plus-strand to minus-strand RNA in a dose-dependent manner. TIAR contains three RNA recognition motifs (RRMs). Biophysical data indicated that only RRM2 directly contacts RNA and that up to three TIAR molecules can bind cooperatively to the WNV 3'(-) SL RNA. These data provide additional evidence that TIAR functions as a proviral host factor facilitating exponential amplification of WNV genome production in infected cells.IMPORTANCEWest Nile virus (WNV) is a mosquito-borne orthoflavivirus associated with increasing global human disease incidence. The molecular mechanisms underlying viral replication are not fully understood. In early stages of infection, viral genome transcription is inefficient; however, in late stages, viral genome transcription increases exponentially. T-cell intracellular antigen-related (TIAR) protein is a cellular protein that has been shown to interact with the 3' end of the WNV negative-sense antigenomic RNA. We obtained data showing colocalization of cellular TIAR with viral replication complexes in infected cells and an increased ratio of intracellular genomic to antigenomic viral RNA in TIAR-overexpressing cells, and confirmed preferential binding of TIAR to the 3' end of the WNV antigenome both in vitro and in infected cell extracts. We also demonstrated that multiple TIAR proteins can bind cooperatively to the WNV 3'(-) stem-loop RNA. These data provide supporting evidence for a model of TIAR-mediated rapid initiation of nascent genome RNA synthesis in infected cells.

TIAR↗

PubMed2026

Molecular characterization and genome sequence analysis of Dichroa emaravirus, a putative novel member of the genus Emaravirus.

Hydrangea febrifuga (syn. Dichroa febrifuga) is a traditional medicinal plant distributed in China and Southeast Asia, and febrifugine, one of its principal bioactive constituents, has served as an important lead compound for antimalarial drug development. Viral infections may adversely affect the quality of medicinal plants; however, no emaravirus has previously been reported from H. febrifuga. Here, high-throughput sequencing was performed on H. febrifuga leaves exhibiting mosaic symptoms collected in Yunnan Province, China. Combined with RT-PCR, Sanger sequencing, and 5'/3' rapid amplification of cDNA ends (RACE), five full-length genomic RNA segments of a putative novel emaravirus, tentatively designated Dichroa emaravirus (DEV), were identified and characterized. The five negative-sense single-stranded RNA (-ssRNA) segments have a combined length of 12,971 nt and encode an RNA-dependent RNA polymerase (RdRp), glycoprotein precursor (GP), nucleocapsid protein (NP), movement protein (MP), and an uncharacterized accessory protein, P5. The maximum amino acid sequence identities of DEV P1-P4 with recognized emaraviruses were 73.90%, 51.82%, 65.60%, and 81.30%, respectively, whereas P5 showed a maximum identity of 49.16% with its closest homolog. Thus, three of the four core proteins had maximum identities below 80%, consistent with the current ICTV species demarcation criterion for the genus Emaravirus. Maximum-likelihood phylogenetic analyses based on the four core proteins further supported the placement of DEV within the genus Emaravirus (family Fimoviridae). These results support DEV as a putative novel emaravirus and represent the first report of an emaravirus associated with H. febrifuga.

Genome, Viral↗