Ectropis obliqua picorna-like virus (EoPV) is an insect RNA virus that causes a lethal granulosis infection of larvae of the tea looper (Ectropis obliqua). An internal ribosome entry site (IRES) mediates translation initiation of EoPV RNA. Here, bicistronic constructs were used to examine the 5' untranslated region (UTR) of EoPV for IRES activity. The capacities of the EoPV 5' UTR IRES and another insect virus IRES, the cricket paralysis virus intergenic region IRES, to mediate internal translation initiation in a variety of translation systems were also compared. The results demonstrated that the EoPV IRES functioned efficiently not only in mammalian cell-derived systems, but also in an insect cell-derived translation system. However, it functioned inefficiently in a plant cell-derived translation system. This study reveals the host preferences of the EoPV IRES and important differences in IRES function between the EoPV IRES and other characterized picorna-like insect viral IRESs.

The smoky-brown cockroach (Periplaneta fuliginosa) densovirus (PfDNV) has previously shown potential in urban pest control. To improve its efficacy as a biopesticide, the genome of PfDNV was engineered by inserting the insect-specific toxin gene BmKIT1 in the open reading frame encoding the major structural proteins. A green fluorescent protein (GFP) marker was tagged to the BmKIT1 at its C-terminus for in vivo imaging using Confocal laser scanning microscopy (CFSM). Using a virion rescue strategy, the genomes of recombinant and wild-type (wt) PfDNV were then cotransfected in P. fuliginosa nymphs. Reverse transcription PCR (RT-PCR) showed that the inserted BmkIT1 genes were expressed in the P. fuliginosa nymphs 48 h after cotransfection. CFSM and transmission electron microscopy also confirmed the generation of virus particles and expression of BmKIT1-GFP fusion protein in the cotransfected nymphs. The recombinant viruses remained infective to P. fuliginosa nymphs in feeding tests. Using the LT(50) bioassay method, the coninfection of the recombinant and wt PfDNV killed the P. fuliginosa nymphs approximate 32% faster than wt PfDNV only. This is the first report showing the improvement of engineered densovirus for the potential application of biopesticide.

Wuhan nodavirus (WhNV) particles are isometric, non-enveloped, and about 29 nm in diameter. In the previous study, we determine its physiochemical characterization and the nucleotide sequence of the larger genomic segment, RNA1 and identify it a nodavirus. WhNV RNA1 is 3149 nt in length, encoding protein A, catalytic subunit of RNA-dependent RNA polymerase (RdRp). In this report, we complete the sequence determination of the smaller genomic segment, RNA2 of WhNV. WhNV RNA2 is determined to be 1562 nt long, containing a 430-amino-acid open reading frame (ORF) encoding the coat protein of WhNV with a calculated molecular mass of 47,856 Da. The homology of the coat protein of WhNV and the homologous proteins of other nodaviruses either alphanodaviruses or betanodaviruses is very low. WhNV coat protein shares the highest identity (24%) with that of Lates calcarifer encephalitis virus (LCEV), a betanodavirus, and shares less than 16% identical amino acids with each of the alphanodaviruses. Furthermore, the prediction of WhNV capsid structure by 3D-PSSM shows that the capsid structure of WhNV resembles that of tomato bushy stunt virus (TBSV), a tombusvirus, which contains two domains, rather than the expected single-domain capsid protein of insect nodaviruses. The phylogenetic analysis indicates that WhNV is the most distantly related of both the alphanodaviruses and betanodaviruses, which provides significant new data for understanding the evolution of the nodavirus family.

A new insect nodavirus is isolated from Pieris rapae larvae in Wuhan city, China and tentatively designated Wuhan nodavirus (WhNV). We here report the physicochemical characterization of WhNV and determine the nucleotide sequences of its larger segment of genome, RNA1. The results show that WhNV particles are isometric, non-enveloped, with a diameter of about 29nm. The virus has a major capsid protein and a minor capsid protein with estimated molecular mass of 40 and 44kDa, respectively. WhNV RNA1 is determined to be 3149nt long, containing a 1014-amino-acid open reading frame (ORF) encoding protein A with a calculated molecular mass of 114,608Da. The protein A shows 39 and 27% identity to its homologues in Pariacoto virus (PaV) and Striped jack necrosis nervous virus (SJNNV), respectively, but shows only 24% or less identity to its homologues in other insect Nodaviruses such as Nodamura virus (NoV), Black beetle virus (BBV), Boolarra virus (BoV) and Flock house virus (FHV). Predicted domains for six RNA-dependent RNA polymerase motifs and putative ORFs (protein B) are confirmed by sequence analysis of WhNV RNA1.

The nucleotide sequence of a novel icosahedral DNA virus infecting Dendrolimus punctatus has been determined. The genome is 5039 nt long and includes inverted terminal repeats of 200 nt containing 131 nt long J-shaped terminal hairpins. The 'plus' strand of the genome contains three large open reading frames (ORFs), the left and the mid-ORFs (within the left ORF) in the left-half encoding the non-structural proteins and the right ORF in the right-half encoding viral capsid proteins. NS1 protein contains conserved replication initiation and DNA-dependent ATPase/helicase domains. VP1 protein contains a conserved PGY and phospholipase A2 motifs and shows high identities with VPs of Casphalia extranea densovirus and Bombyx mori densovirus-1 belonging to the genus Iteravirus. Phylogenetic analysis also revealed that this virus is most closely related to Casphalia extranea densovirus and Bombyx mori densovirus-1. Consequently, this virus was considered as a new third member of the genus Iteravirus of the subfamily Densovirinae, and designated Dendrolimus punctatus densovirus.

In this study, Dendrolimus punctatus tetravirus (DpTV) has been identified as a new member of the genus Omegatetravirus of the family Tetraviridae that may be related serologically to Nudaurelia capensis omega virus (NomegaV). DpTV particles are isometric, with a diameter of about 40 nm and a buoyant density of 1.281 g cm(-3) in CsCl. The virus has two capsid proteins (of 62 500 and 6800 Da) and two single-stranded RNA molecules (RNA1 and RNA2), which are 5492 and 2490 nt long, respectively. RNA1 has a large open reading frame (ORF) encoding a polypeptide of 180 kDa; RNA2 contains two partially overlapping ORFs encoding polypeptides of 17 and 70 kDa. The 180 kDa protein, which contains consensus motifs of a putative methyltransferase, helicase and RNA-dependent RNA polymerase, shows significant similarity to those of other tetraviruses. The 17 kDa protein is a PEST (Pro/Glu/Ser/Thr) protein of unknown function. The 70 kDa protein is the coat protein precursor and is predicted to be cleaved at an Asn-Phe site located after residue 570. The 70 kDa protein shows 86 and 66 % identity to its homologues in NomegaV and Helicoverpa armigera stunt virus, respectively. Secondary-structure analysis revealed that the RNAs of DpTV have tRNA-like structures at their 3' termini.