To identify the viruses in tree peony plants associated with the symptoms of yellowing, leaf rolling, stunted growth, and decline, high-throughput sequencing of small RNA and mRNA was conducted from a single symptomatic plant. Bioinformatic analyses and reconstruction of viral genomes indicated mixed viral infections involving cycas necrotic stunt virus, apple stem grooving virus, lychnis mottle virus, grapevine line pattern virus, and three new viruses designated as peony yellowing-associated citrivirus (PYaCV, Citrivirus in Betaflexiviridae), peony betaflexivirus 1 (PeV1, unclassified in Betaflexiviridae), and peony leafroll-associated virus (PLRaV, Ampelovirus in Closteroviridae). PYaCV was 8,666 nucleotides (nt) in length, comprising three open reading frames (ORFs), and shared 63.8 to 75.9% nt sequence identity with citrus leaf blotch virus (CLBV) isolates. However, the ORF encoding the replication-associated protein (REP) shared 57 and 52% sequence identities at the nt and amino acid (aa) level, respectively, with other reported CLBV isolates, which were below the criterion for species classification within the family Betaflexiviridae. Recombination analysis identified putative recombination sites in PYaCV, which originated from CLBV. PeV1, only identified from the transcriptome data, was 8,124 nt in length, with five ORFs encoding the REP (ORF1), triple gene block (ORF2 to 4) and coat protein (CP, ORF5). Phylogenetic analysis and sequence comparison showed that PeV1 clustered with an unassigned member, the garlic yellow mosaic-associated virus within the Betaflexiviridae family, into a separate clade. Partial genome sequence analysis of PLRaV (12,545 nt) showed it contained seven ORFs encoding the partial polyprotein 1a, the RNA-dependent RNA polymerase (RdRp), two small hydrophobic proteins p11 and p6, HSP70h, p55, and a CP duplicate, which shared low aa sequence identity with Closteroviridae family members. Phylogenetic analysis based on the aa sequences of RdRp or HSP70h indicated that PLRaV clustered with grapevine leafroll-associated virus 1 (GLRaV-1) and GLRaV-13 in the Ampelovirus genus. Field investigation confirmed the wide distribution of these viruses, causing mixed infections of peony plants in Beijing.

Background: In plants, RNA silencing is an important conserved mechanism to regulate gene expression and combat against abiotic and biotic stresses. Dicer-like (DCL) and Argonaute (AGO) proteins and RNA-dependent RNA polymerase (RDR) are the core elements involved in gene silencing and their gene families have been explored in many plants. However, these genes and their responses to stresses have not yet been well characterized in adzuki bean. Results: A total of 11 AGO, 7 DCL and 6 RDR proteins were identified, and phylogenetic analyses of these proteins showed that they clustered into six, four and four clades respectively. The expression patterns of these genes in susceptible or resistant adzuki bean cultivars challenged with drought, bean common mosaic virus and Podosphaera xanthii infections were further validated by quantitative RT-PCR. The different responses of these proteins under abiotic and biotic stresses indicated their specialized regulatory mechanisms. Conclusions: In this study, 24 genes of the DCL, AGO and RDR gene families in adzuki bean were identified, and the sequence characterization, structure of the encoded proteins, evolutionary relationship with orthologues in other legumes and gene expression patterns under drought and biotic stresses were primarily explored, which enriched our understanding of these genes in adzuki bean. Our findings provide a foundation for the comparative genomic analyses of RNA silencing elements in legume plants and further new insights into the functional complexity of RNA silencing in the response to various stresses in adzuki bean.

Lily (Lilium) is an important bulbous perennial herb that is frequently infected by one or more viruses. To investigate the diversity of lily viruses, lilies with virus-like symptoms in Beijing were collected to perform small RNA deep sequencing. Then, the 12 complete and six nearly full-length viral genomes, including six known viruses and two novel viruses were determined. Based on sequence and phylogenetic analyses, two novel viruses were considered to be members of the genera Alphaendornavirus (Endornaviridae) and Polerovirus (Solemoviridae). These two novel viruses were provisionally named lily-associated alphaendornavirus 1 (LaEV-1) and lily-associated polerovirus 1 (LaPV-1). Based on sequence, phylogenetic and recombination analyses, strawberry latent ringspot virus (SLRSV) in the genus Stralarivirus (Secoviridae) was identified for the first time in China, and shown to exhibit the highest nucleotide (nt) diversity among the available full-length SLRSV genome sequences, with the highest identities of 79.5% for RNA1 and 80.9% for RNA2. Interestingly, the protease cofactor region in RNA1 was 752 aa in length, whereas those of the other 27 characterized isolates ranged from 700–719 aa in length. The genome sequences of lily virus A (Potyvirus), lily virus X (Potexvirus), and plantago asiatica mosaic virus (Potexvirus) exhibited varying degrees of sequence diversity at the nucleotide level compared with their corresponding characterized isolates. In addition, plantago asiatica mosaic virus (PlAMV) tended to cluster on a host species-basis. One identified lily mottle virus (Potyvirus) isolate was detected as a recombinant, and which clustered in a different group with four other isolates. Seven identified lily symptomless virus (Carlavirus) isolates, including one recombinant, were clustered into three clades. Our results revealed the genetic diversity of lily-infecting viruses, and sequence insertion, host species and recombination are factors that likely contribute to this diversity. Collectively, our results provide useful information regarding the control of viral disease in lily.