Complete genome sequence of a novel badnavirus infecting Fatsia japonica in China.

The complete genome sequence of a novel badnavirus, tentatively named "fatsia badnavirus 1" (FaBV1, OM540428), was identified in Fatsia japonica. The infected plant displayed virus-like symptoms on leaves, including yellowing and chlorosis. The genome of FaBV1 is 7313 bp in length and similar in size and organization to other members of the genus Badnavirus (family Caulimoviridae), containing four open reading frames (ORFs), three of which are found in all known badnaviruses, and the other of which is only present in some badnaviruses. The virus has the genome characteristics of badnaviruses, including a tRNAMet binding site (5'-TCTGAATTTATAGCGCTA-3') and two cysteine-rich domains (C-X-C-2X-C-4X-H-4X-C and C-2X-C-11X-C-2X-C-4X-C-2X-C). Pairwise sequence comparisons of the RT+RNase H region indicated that FaBV1 shares 61.4-71.2% nucleotide (nt) sequence identity with other known badnaviruses, which is below the threshold (80% nt sequence identity in the RT+RNase H region) used for species demarcation in the genus Badnavirus. Phylogenetic analysis revealed that FaBV1, ivy ringspot-associated virus (IRSaV, MN850490.1), and cacao mild mosaic virus (CMMV, KX276640.1) together form a separate clade within the genus Badnavirus, suggesting that FaBV1 is a new member of the genus Badnavirus in the family Caulimoviridae. To our knowledge, this is the first report of a badnavirus infecting F. japonica.

A novel R2R3-MYB transcription factor PbMYB1L of Pyrus bretschneideri regulates cold tolerance and anthocyanin accumulation.

KEY MESSAGE: PbMYB1L enhances the cold tolerance and anthocyanin accumulation of transgenic Arabidopsis by regulating the expression of genes related to the cold-responsive genes pathway and anthocyanin synthesis pathway. MYB transcription factors (TFs) have been demonstrated to play diverse roles in plant growth and development. In the present study, we identified a novel R2R3-MYB transcription factor, PbMYB1L, from the peel of 'Red Zaosu' pear (Pyrus bretschneideri), which was induced by cold stress and acted as a positive regulator in anthocyanin biosynthesis. Notably, the transgenic Arabidopsis lines exhibited enhanced tolerance to cold stress. Compared to the Arabidopsis wild-type plants, the transgenic lines displayed longer primary roots and reduced reactive oxygen species (ROS) levels including O2-, hydrogen peroxide (H2O2), and malondialdehyde (MDA). Furthermore, significant upregulation of key cold-responsive genes AtCBF1, AtCBF2, AtCBF3, AtCBF4, and AtKIN1 was observed in the transgenic plants under cold stress conditions compared to wild type. Arabidopsis plants overexpressing PbMYB1L had significant anthocyanin accumulation in leaves after cold treatment with quantitative results indicating higher expression of anthocyanin structural genes compared to wild type. These findings suggest that PbMYB1L not only plays a vital role in conferring cold tolerance but also acts as a crucial regulator of anthocyanin biosynthesis.

First report of Lily virus X infecting Paris polyphylla var. yunnanensis in Yunnnan, China.

Lily virus X (LVX) is a positive-sense ssRNA virus belonging to the genus Potexvirus in the family Alphaflexiviridae. LVX is known to infect plants of the genera Lilium and Tricyrtis in the family Liliacea. LVX was first reported in an asymptomatic lily (Lilium formosanum) from England (Stone, 1980), but has been shown to infect plants in the Netherlands (Chen et al. 2005), the United States (Jordan et al. 2008) and Japan (Nijo et al. 2018). To date, the complete genomes of two LVX isolates from the Netherlands and Japan have been reported. Paris polyphylla var. yunnanensis, known as Dianchonglou in China, is a perennial plant of the family Melanthiaceae (formerly belonging to the family Trillium). In China, its rhizome is commonly used as an antispasmodic agent for stroke and cancer treatment (Chang et al. 2017). From 2019 to 2022, leaf mottle and shrinkage which are typical symptoms of viral infections were observed on the leaves of P. polyphylla var. yunnanensis plants in Dianchonglou fields in Qujing, Yunnan. Disease incidence ranged from 19% to 45% across 5 fields (90 plants per field) in Qujing. To identify the possible viral pathogen(s) associated with the disease, the mirVanaTM miRNA isolation Kit was used to extract total RNA was from a mixed sample pool of 5 symptomatic leaf samples collected from the 5 fields. RNA sequencing library was constructed using TruSeqTM RNA sample preparation kit. Sequencing on the Illumina HiSeqTM 2500 platform (Illumina, USA) with 125-bp paired-end reads yielded 23,077,786 raw reads. 22,534,100 clean reads were obtained by removing reads of low quality and poly-N using Trimmomatic software (Bolger et al. 2014). By utilizing the paired-end splicing method in Trinity software (Grabherr et al. 2011) the the raw reads were De novo assembled into 184,596 contigs, of which 303 were related to viruses, including Paris mosaic necrosis virus (PMNV), Pear alphapartitivirus (PAPV), Dahlia mosaic virus (DMV), and Lily virus X (LVX). BLASTn analysis revealed that 12 contigs (lengths ranging from 344 nt to 5,981 nt, query cover 6% to 99%) were most similar (57.32% to 91.67% nt identities) to the genome sequences of LVX, suggesting a possible infection of LVX in the plants. To confirm the result, a full-length genomic sequence of LVX was obtained by reverse transcription polymerase chain reaction (RT-PCR) using specific primers designed based on the sequence of the assembled contigs. The PCR products were cloned into pGEM-T vector (Promega Corporation, USA) and sequenced using the Sanger method (Sangon Biotech, Shanghai, China). The obtained full-length genomic sequence of the LVX isolate (LVX-PP, accession number OM100017) was 5,981 nt in length. BLASTp analysis demonstrated that the putative Rep and CP of LVX-PP shared 76.27% to 81.05% and 80.81% to 81.82% aa sequence similarities with that of other LVX isolates, respectively. Maximum-likelihood phylogenetic trees inferred from the Rep and CP aa sequences showed that LVX-PP clustered closely with LVX isolates. The leaf samples were further analyzed using a lily virus X (LVX) ELISA kit (DEIAPV181, Creative Diagnostics, U.S.A.). Healthy P. polyphylla var. yunnanensis leaves were taken as a negative control and buffer solution as a blank control. The results showed a positive reaction for all five symptomatic plants (OD = 1.259 ± 0.007) relative to the negative (OD = 0.099) and blank (OD = 0.073) controls. These results indicate that LVX can infect P. polyphylla var. yunnanensis. To our knowledge, this is the first report that LVX has been detected in P. polyphylla var. yunnannensis. This study will serve as an important reference for the study of the host range of LVX. Further studies will be required to determine how LVX spreads between P. polyphylla var. yunnannensis and other host plants.

Complete genome sequence analysis of Valeriana jatamansi tymovirus 1: a novel member of the genus Tymovirus infecting Valeriana jatamansi Jones.

A new positive-sense, single-stranded RNA virus, tentatively named "Valeriana jatamansi tymovirus 1" (VaJV1, OQ730267), was isolated from Valeriana jatamansi Jones displaying symptoms of vein-clearing in Yunnan Province, China. The complete genome of VaJV1 consists of 6,215 nucleotides and contains three open reading frames (ORFs). The genome structure of VaJV1 is typical of members of the genus Tymovirus. BLASTn analysis and multiple sequence alignments showed that the complete genome and coat protein of VaJV1 shared the most sequence similarity (65.5% nucleotides and 50.5% amino acid sequence identity) with an isolate of the tymovirus okra mosaic virus (NC_009532). Phylogenetic analysis confirmed that VaJV1 clustered most closely with other tymoviruses. We propose that Valeriana jatamansi tymovirus 1 represents a new species within the genus Tymovirus.

Complete genome sequence of a novel fusarivirus from the phytopathogenic fungus Fusarium sp.

Fusarium diseases include wilts, blights, rots, and cankers of many horticultural, field, ornamental, and forest crops in both agricultural and natural ecosystems, and they significantly hinder food plant production. Here, we describe a novel mycovirus, tentatively designated as "Fusarium fusarivirus 1" (FuFV1), which was discovered in an isolate of the phytopathogenic fungus Fusarium sp. FuFV1 has a positive-sense single-stranded RNA (+ssRNA) genome of 6,391 nucleotides (nt) containing three open reading frames (ORFs). ORF1 encodes a large polypeptide of 1,501 amino acids (aa) with conserved RNA-dependent RNA polymerase (RdRp) and helicase (Hel) domains. ORF2, overlapping ORF1 by 122 nucleotides, encodes a polypeptide with a conserved Smc domain. The third and smaller ORF (ORF3) encodes a polypeptide with an unknown function. BLASTp analysis of the ORF1-encoded polypeptide revealed that FuFV1 shares the highest aa sequence similarity (68.5% identity, E-value 0.0) with Fusarium poae fusarivirus 1 (FpFV1, genus Alphafusarivirus). Phylogenetic analysis of the RdRp and helicase (Hel) sequences indicated that FuFV1 clustered closely with FpFV1 in a separate branch within the clade containing members of the genus Alphafusarivirus. Based on these results, we propose that FuFV1 should be considered a novel mycovirus belonging to the genus Alphafusarivirus of the family Fusariviridae.

Complete genome sequence of a novel closterovirus isolated from Dregea volubilis.

The complete genome sequence of a putative novel closterovirus, tentatively named "Dregea volubilis closterovirus 1" (DvCV1, GenBank accession no. MZ779122), infecting Dregea volubilis in China was determined using high-throughput sequencing (HTS). The complete genome sequence of DvCV1 consists of 16,165 nucleotides (nt) and contains nine ORFs. The genome structure of DvCV1 is typical of members of the genus Closterovirus. Complete genome sequence analysis showed that DvCV1 shares 41.4-48.4% nucleotide sequence identity with other known closteroviruses. The putative RNA-dependent RNA polymerase (RdRp), heat shock protein 70-like protein (HSP70h), and coat protein (CP) of DvCV1 share 46.80-62.65%, 31.06-51.80%, and 28.34-37.37% amino acid sequence identity, respectively, with the RdRp, HSP70h and CP of other closteroviruses. Phylogenetic analysis based on HSP70h aa sequences placed DvCV1 alongside other members of the genus Closterovirus in the family Closteroviridae. These results suggest that DvCV1 is a new member of the genus Closterovirus. This is the first report of a closterovirus infecting D. volubilis.

Complete genome sequence analysis of a novel citlodavirus isolated from the leaves of Myrica rubra in Yunnan.

Through high-throughput sequencing, a novel citlodavirus, tentatively named "Myrica rubra citlodavirus 1" (MRV1, accession no. OP374189), was isolated from the leaves of Myrica rubra in Yunnan exhibiting narrow deformity of leaf tips, shrinkage, and chlorosis along the veins. The complete genome sequence was determined and analyzed using cloning and Sanger sequencing. MRV1 is a single-stranded circular non-enveloped DNA virus with a genome size of 3775 nucleotides and contains six open reading frames (ORFs). The virion-sense genome strand encodes a coat protein (CP, nt 750-1,493, 247 aa), two hypothetical movement proteins (V3, nt 382-666, 94 aa; and V2, nt 461-895, 144 aa), and one movement protein (MP, nt 1,527-2,438, 303 aa). The complementary strand of the genome encodes two replication proteins (RepA, nt 3,712-2,834, 292 aa; Rep, nt 2,867-2,553, 104 aa). The MRV1 genome contains the stem-loop motif 5'-TAATATTAC-3', which is a highly conserved nonanucleotide motif found in the origin of virion-strand replication in geminiviruses. Genome sequence alignment analysis showed that citrus chlorotic dwarf associated virus (CCDaV, accession no. JQ920490) shared the highest nucleotide sequence similarity (66.10% identity) with MRV1. Phylogenetic analysis showed that CCDaV is the closest known relative of MRV1, and that these viruses clustered in a single branch within a clade consisting of citlodaviruses. These results indicate that MRV1 should be regarded as a new species of the genus Citlodavirus in the family Geminiviridae.

Complete genome sequence analysis of Paris alphapartitivirus 1: a novel member of the genus Alphapartitivirus infecting Paris polyphylla var. yunnanensis.

A novel double-stranded RNA (dsRNA) virus, tentatively named "Paris alphapartitivirus 1" (ParAPV1, OL960006-OL960007), was detected in Paris polyphylla var. yunnanensis plants exhibiting leaf chlorosis and shrinkage symptoms in Yunnan. Its complete genome sequence was determined using Illumina and Sanger sequencing. ParAPV1 has a bipartite genome that consists of dsRNA1 (1,917 bp) encoding the viral RNA-dependent RNA polymerase (RdRp), and dsRNA2 (1,818 bp) encoding the putative coat protein (CP). Sequence comparisons showed that the RdRp and CP of ParAPV1 are most similar to those of pear alphapartitivirus (PpPV2), with 69.97% and 54.21% amino acid sequence identities respectively. Phylogenetic analysis of the RdRp amino acid sequences of ParAPV1 and other partitiviruses showed that ParAPV1 cluster with viruses in a clade containing alphapartitiviruses, and that its closest known relatives are PpPV2 (BBA66577) and rose partitivirus (RoPV, ANQ45203S). Taken together, these results suggest that ParAPV1 should be regarded as a new member of genus Alphapartitivirus in the family Partitiviridae. This is the first report of a partitivirus infecting P. polyphylla var. yunnanensis.

Identification and analysis of sucrose synthase gene family associated with polysaccharide biosynthesis in Dendrobium catenatum by transcriptomic analysis.

Background: Dendrobium catenatum is a valuable traditional medicinal herb with high commercial value. D. catenatum stems contain abundant polysaccharides which are one of the main bioactive components. However, although some genes related to the synthesis of the polysaccharides have been reported, more key genes need to be further elucidated. Results: In this study, the contents of polysaccharides and mannose in D. catenatum stems at four developmental stages were compared, and the stems' transcriptomes were analyzed to explore the synthesis mechanism of the polysaccharides. Many genes involved in starch and sucrose metabolisms were identified by KEGG pathway analysis. Further analysis found that sucrose synthase (SUS; EC 2.4.1.13) gene maybe participated in the polysaccharide synthesis. Hence, we further investigated the genomic characteristics and evolution relationships of the SUS family in plants. The result suggested that the SUS gene of D. catenatum (DcSUS) had undergone the expansion characterized by tandem duplication which might be related to the enrichment of the polysaccharides in D. catenatum stems. Moreover, expression analyses of the DcSUS displayed significant divergent patterns in different tissues and could be divided into two main groups in the stems with four developmental stages. Conclusion: In general, our results revealed that DcSUS is likely involved in the metabolic process of the stem polysaccharides, providing crucial clues for exploiting the key genes associated with the polysaccharide synthesis.

Complete genome sequence of a novel mitovirus detected in Paris polyphylla var. yunnanensis.

Paris mitovirus 1 (ParMV1) is a positive-sense RNA virus that was detected in diseased Paris polyphylla var. yunnanensis plants in Wenshan, Yunnan. The complete genome sequence of ParMV1 is 2,751 nucleotides in length, and the genome structure is typical of mitoviruses. The ParMV1 genome has a single open reading frame (ORF; nt 358-2,637) that encodes an RNA-dependent RNA polymerase (RdRp) with a predicted molecular mass of 86.42 kDa. ParMV1 contains six conserved motifs (Ι-VΙ) that are unique to mitoviruses. The 5' and 3' termini of the genome are predicted to have a stable secondary structure, with the reverse complementary sequence forming a panhandle structure. Comparative genome analysis revealed that the RdRp of ParMV1 shares 23.1-40.6% amino acid (aa) and 32.3-45.7% nucleotide (nt) sequence identity with those of other mitoviruses. Phylogenetic analysis based on RdRp aa sequences showed that ParMV1 clusters with mitoviruses and hence should be considered a new member of the genus Mitovirus in the family Mitoviridae. This is the first report of a novel mitovirus infecting Paris polyphylla var. yunnanensis.

Complete genome sequence analysis of a novel coguvirus isolated from Paris polyphylla var. yunnanensis.

A novel negative-stranded (ns) RNA virus tentatively named "Yunnan paris negative-stranded virus" (YPNSV), was isolated from Paris polyphylla var. yunnanensis plants exhibiting leaf chlorosis and mosaic symptoms in Yunnan. Its complete genome sequence was determined using Illumina and Sanger sequencing. YPNSV has a bipartite genome that consists of a negative-stranded (ns) RNA1 encoding the viral RNA-dependent RNA polymerase (RdRp, p251), an ambisense RNA2 coding for the putative movement protein (MP, p46) and nucleocapsid protein (NP, p39), with the two open reading frames separated by a long intergenic region that is rich in A and U. Sequence comparisons showed that the RdRp, MP, and NP of YPNSV are most similar to those of watermelon crinkle leaf-associated virus 2 (WCLaV-2), with 69.1%, 50.4%, and 60.9% amino acid sequence identity, respectively. Phylogenetic analysis based on deduced amino acid sequences of RdRp and NP showed that YPNSV clustered in a clade with coguviruses and that its closest known relative is WCLaV-2. Based on the above results, YPNSV should be regarded as a new member of genus Coguvirus, family Phenuiviridae.

Demethylation alters transcriptome profiling of buds and leaves in 'Kyoho' grape.

BACKGROUND: Grape buds and leaves are directly associated with the physiology and metabolic activities of the plant, which is monitored by epigenetic modifications induced by environment and endogenous factors. Methylation is one of the epigenetic regulators that could be involved in DNA levels and affect gene expression in response to stimuli. Therefore, changes of gene expression profile in leaves and bud through inhibitors of DNA methylation provide a deep understanding of epigenetic effects in regulatory networks. RESULTS: In this study, we carried out a transcriptome analysis of 'Kyoho' buds and leaves under 5-azacytidine (5-azaC) exposure and screened a large number of differentially expressed genes (DEGs). GO and KEGG annotations showed that they are mainly involved in photosynthesis, flavonoid synthesis, glutathione metabolism, and other metabolic processes. Functional enrichment analysis also provided a holistic perspective on the transcriptome profile when 5-azaC bound to methyltransferase and induced demethylation. Enrichment analysis of transcription factors (TFs) also showed that the MYB, C2H2, and bHLH families are involved in the regulation of responsive genes under epigenetic changes. Furthermore, hormone-related genes have also undergone significant changes, especially gibberellin (GA) and abscisic acid (ABA)-related genes that responded to bud germination. We also used protein-protein interaction network to determine hub proteins in response to demethylation. CONCLUSIONS: These findings provide new insights into the establishment of molecular regulatory networks according to how methylation as an epigenetic modification alters transcriptome patterns in bud and leaves of grape.