Organization of the Structural Protein Region of La Jolla Virus Isolated from the Invasive Pest Insect Drosophila suzukii.

Drosophila suzukii (Ds) is an invasive pest insect that infests ripening fruit, causing severe economic losses. Control measures based on chemical pesticides are inefficient and undesirable, so biological alternatives have been considered, including native Ds viruses. We previously isolated a strain of La Jolla virus (LJV-Ds-OS20) from Ds in Germany as a candidate biopesticide. Here we characterized the new strain in detail, focusing on the processing of its capsid proteins. We tested LJV growth during Ds development to optimize virus production, and established a laboratory production system using adult flies. This system was suitable for the preparation of virions for detailed analysis. The LJV-Ds-OS20 isolate was cloned by limiting dilution and the complete nucleotide sequence was determined as a basis for protein analysis. The terminal segments of the virus genome were completed by RACE-PCR. LJV virions were also purified by CsCl gradient centrifugation and analyzed by SDS-PAGE and electron microscopy. The capsid proteins of purified LJV virions were resolved by two-dimensional SDS-PAGE for N-terminal sequencing and peptide mass fingerprinting. The N-terminal sequences of VP1 and VP2, together with MS data representing several capsid proteins, allowed us to develop a model for the organization of the LJV structural protein region. This may facilitate the development of new viral strains as biopesticides.

ICTV Virus Taxonomy Profile: Polycipiviridae.

Polycipiviridae is a family of picorna-like viruses with non-segmented, linear, positive-sense RNA genomes of approximately 10-12 kb. Unusually for viruses within the order Picornavirales, their genomes are polycistronic, with four (or more) consecutive 5'-proximal open reading frames (ORFs) encoding structural (and possibly other) proteins and a long 3' ORF encoding the replication polyprotein. Members of species within the family have all been detected in ants or via arthropod transcriptomic datasets. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the Polycipiviridae, which is available at www.ictv.global/report/polycipiviridae.

Confocal Imaging of Double-Stranded RNA and Pattern Recognition Receptors in Negative-Sense RNA Virus Infection.

Double-stranded (ds) RNA is produced as a replicative intermediate during RNA virus infection. Recognition of dsRNA by host pattern recognition receptors (PRRs) such as the retinoic acid (RIG-I) like receptors (RLRs) RIG-I and melanoma differentiation-associated protein 5 (MDA-5) leads to the induction of the innate immune response. The formation and intracellular distribution of dsRNA in positive-sense RNA virus infection has been well characterized by microscopy. Many negative-sense RNA viruses, including some arenaviruses, trigger the innate immune response during infection. However, negative-sense RNA viruses were thought to produce low levels of dsRNA, which hinders the imaging study of PRR recognition of viral dsRNA. Additionally, infection experiments with highly pathogenic arenaviruses must be performed in high containment biosafety level facilities (BSL-4). The interaction between viral RNA and PRRs for highly pathogenic RNA virus is largely unknown due to the additional technical challenges that researchers need to face in the BSL-4 facilities. Recently, a monoclonal antibody (Mab) (clone 9D5) originally used for pan-enterovirus detection has been found to specifically detect dsRNA with a higher sensitivity than the traditional J2 or K1 anti-dsRNA antibodies. Herein, by utilizing the 9D5 antibody, we describe a confocal microscopy protocol that has been used successfully to visualize dsRNA, viral protein and PRR simultaneously in individual cells infected by arenavirus. The protocol is also suitable for imaging studies of dsRNA and PRR distribution in pathogenic arenavirus infected cells in BSL4 facilities.

Molecular and biological characterization of a novel botybirnavirus identified from a Pakistani isolate of Alternaria alternata.

Mycoviruses ubiquitously infect a wide range of fungal hosts in the world. The current study reports a novel double stranded RNA (dsRNA) virus, termed Alternaria alternata botybirnavirus 1 (AaBbV1), infecting a Pakistani strain, 4a, of a phytopathogenic ascomycetous fungus Alternaria alternata. A combined approach of next generation and conventional terminal end sequencing of the viral genome revealed that the virus is a distinct member of the genus Botybirnavirus. This virus comprised of two segments (dsRNA1 and dsRNA2) of sizes 6127 bp and 5860 bp respectively. The dsRNA1-encoded protein carrying the RNA-dependent RNA polymerase domain showed 61% identity to the counterpart of Botrytis porri botybirnavirus 1 and lower levels of amino acid similarity with those of other putative botybirnaviruses and the fungal dsRNA viruses such as members of the families Totiviridae, Chrysoviridae and Megabirnaviridae. The dsRNA2-encoded protein showed resemblance with corresponding proteins of botybirnaviruses. Electron microscopy showed AaBbV1 to form spherical particles of 40 nm in diameter. Biochemical analyses showed that two structural proteins encoded by dsRNA1 and dsRNA2 underwent processing to some extent during particle purification, resulting in the appearance of multiple smaller products. Phylogenetic analyses of structural proteins suggested that their coding region might have been duplicated once and maintained without recombination. Protoplast fusion technique allowed for the introduction of AaBbV1 into a virus free Japanese strain of A. alternata and demonstrated its symptomless infection by the virus. Interesting similarities and dissimilarities between AaBbV1 and other previously reported botybirnaviruses are also discussed.

ICTV Virus Taxonomy Profile: Nyamiviridae.

The Nyamiviridae is a family of viruses with unsegmented, negative-sense RNA genomes of 11.3-12.2 kb that produce enveloped, spherical virions. Viruses of the genus Nyavirus are tick-borne and some also infect birds. Other nyamiviruses infecting parasitoid wasps and plant parasitic nematodes have been classified into the genera Peropuvirus and Socyvirus, respectively. This is a summary of the current International Committee on Taxonomy of Viruses (ICTV) Report on the taxonomy of Nyamiviridae, which is available at www.ictv.global/report/nyamiviridae.

Structure of a pentameric virion-associated fiber with a potential role in Orsay virus entry to host cells.

Despite the wide use of Caenorhabditis elegans as a model organism, the first virus naturally infecting this organism was not discovered until six years ago. The Orsay virus and its related nematode viruses have a positive-sense RNA genome, encoding three proteins: CP, RdRP, and a novel δ protein that shares no homology with any other proteins. δ can be expressed either as a free δ or a CP-δ fusion protein by ribosomal frameshift, but the structure and function of both δ and CP-δ remain unknown. Using a combination of electron microscopy, X-ray crystallography, computational and biophysical analyses, here we show that the Orsay δ protein forms a ~420-Å long, pentameric fiber with an N-terminal α-helical bundle, a ß-stranded filament in the middle, and a C-terminal head domain. The pentameric nature of the δ fiber has been independently confirmed by both mass spectrometry and analytical ultracentrifugation. Recombinant Orsay capsid containing CP-δ shows protruding long fibers with globular heads at the distal end. Mutant viruses with disrupted CP-δ fibers were generated by organism-based reverse genetics. These viruses were found to be either non-viable or with poor infectivity according to phenotypic and qRT-PCR analyses. Furthermore, addition of purified δ proteins to worm culture greatly reduced Orsay infectivity in a sequence-specific manner. Based on the structure resemblance between the Orsay CP-δ fiber and the fibers from reovirus and adenovirus, we propose that CP-δ functions as a cell attachment protein to mediate Orsay entry into worm intestine cells.

Construction and Rescue of a Molecular Clone of Deformed Wing Virus (DWV).

European honey bees are highly important in crop pollination, increasing the value of global agricultural production by billions of dollars. Current knowledge about virulence and pathogenicity of Deformed wing virus (DWV), a major factor in honey bee colony mortality, is limited. With this study, we close the gap between field research and laboratory investigations by establishing a complete in vitro model for DWV pathogenesis. Infectious DWV was rescued from a molecular clone of a DWV-A genome that induces DWV symptoms such as crippled wings and discoloration. The expression of DWV proteins, production of infectious virus progeny, and DWV host cell tropism could be confirmed using newly generated anti-DWV monoclonal antibodies. The recombinant RNA fulfills Koch's postulates circumventing the need of virus isolation and propagation of pure virus cultures. In conclusion, we describe the development and application of a reverse genetics system for the study of DWV pathogenesis.

Heterodimers as the Structural Unit of the T=1 Capsid of the Fungal Double-Stranded RNA Rosellinia necatrix Quadrivirus 1.

Most double-stranded RNA (dsRNA) viruses are transcribed and replicated in a specialized icosahedral capsid with a T=1 lattice consisting of 60 asymmetric capsid protein (CP) dimers. These capsids help to organize the viral genome and replicative complex(es). They also act as molecular sieves that isolate the virus genome from host defense mechanisms and allow the passage of nucleotides and viral transcripts. Rosellinia necatrix quadrivirus 1 (RnQV1), the type species of the family Quadriviridae, is a dsRNA fungal virus with a multipartite genome consisting of four monocistronic segments (segments 1 to 4). dsRNA-2 and dsRNA-4 encode two CPs (P2 and P4, respectively), which coassemble into ∼450-Å-diameter capsids. We used three-dimensional cryo-electron microscopy combined with complementary biophysical techniques to determine the structures of RnQV1 virion strains W1075 and W1118. RnQV1 has a quadripartite genome, and the capsid is based on a single-shelled T=1 lattice built of P2-P4 dimers. Whereas the RnQV1-W1118 capsid is built of full-length CP, P2 and P4 of RnQV1-W1075 are cleaved into several polypeptides, maintaining the capsid structural organization. RnQV1 heterodimers have a quaternary organization similar to that of homodimers of reoviruses and other dsRNA mycoviruses. The RnQV1 capsid is the first T=1 capsid with a heterodimer as an asymmetric unit reported to date and follows the architectural principle for dsRNA viruses that a 120-subunit capsid is a conserved assembly that supports dsRNA replication and organization. IMPORTANCE: Given their importance to health, members of the family Reoviridae are the basis of most structural and functional studies and provide much of our knowledge of dsRNA viruses. Analysis of bacterial, protozoal, and fungal dsRNA viruses has improved our understanding of their structure, function, and evolution, as well. Here, we studied a dsRNA virus that infects the fungus Rosellinia necatrix, an ascomycete that is pathogenic to a wide range of plants. Using three-dimensional cryo-electron microscopy and analytical ultracentrifugation analysis, we determined the structure and stoichiometry of Rosellinia necatrix quadrivirus 1 (RnQV1). The RnQV1 capsid is a T=1 capsid with 60 heterodimers as the asymmetric units. The large amount of genetic information used by RnQV1 to construct a simple T=1 capsid is probably related to the numerous virus-host and virus-virus interactions that it must face in its life cycle, which lacks an extracellular phase.

Isolation and characterization of Nylanderia fulva virus 1, a positive-sense, single-stranded RNA virus infecting the tawny crazy ant, Nylanderia fulva.

We report the discovery of Nylanderia fulva virus 1 (NfV-1), the first virus identified and characterized from the ant, Nylanderia fulva. The NfV-1 genome (GenBank accession KX024775) is 10,881 nucleotides in length, encoding one large open reading frame (ORF). Helicase, protease, RNA-dependent RNA polymerase, and jelly-roll capsid protein domains were recognized within the polyprotein. Phylogenetic analysis placed NfV-1 in an unclassified clade of viruses. Electron microscopic examination of negatively stained samples revealed particles with icosahedral symmetry with a diameter of 28.7±1.1nm. The virus was detected by RT-PCR in larval, pupal, worker and queen developmental stages. However, the replicative strand of NfV-1 was only detected in larvae. Vertical transmission did not appear to occur, but horizontal transmission was facile. The inter-colonial field prevalence of NfV-1 was 52±35% with some local infections reaching 100%. NfV-1 was not detected in limited samples of other Nylanderia species or closely related ant species.

Exploring the architecture of viral RNA genomes.

The genomes of RNA viruses contain local structural elements and long-range interactions that control various steps in virus replication. While many individual RNA elements have been characterized, it remains less clear how the structure and activity of such elements are integrated and regulated within the complex context of complete viral genomes. Recent technical advances, particularly the development of high-throughput solution structure mapping methods, have made secondary structural analysis of entire viral RNA genomes feasible. As a consequence, whole-genome structural models have been deduced for a number of plus-strand RNA viruses and retroviruses and these structures have provided intriguing functional and evolutionary insights into global genome architecture.

Viral gastroenteritis in children in Colorado 2006-2009.

Acute gastroenteritis accounts for a significant burden of medically attended illness in children under the age of five. For this study, four multiplex reverse transcription PCR assays were used to determine the incidence of adenovirus, astrovirus, coronavirus, norovirus GI and GII, rotavirus, and sapovirus in stool samples submitted for viral electron microscopy (EM) to the Children's Hospital Colorado. Of 1105 stool samples available, viral RNA/DNA was detected in 247 (26.2%) of 941 pediatric samples (median age = 2.97 years, 54% male) with 28 (3.0%) positive for more than one virus. Adenovirus, astrovirus, norovirus GI, norovirus GII, rotavirus, and sapovirus were detected in 95 (10.0%), 33 (3.5%), 8 (0.9%), 90 (9.6%), 49 (5.2%), and 2 (0.2%) of the pediatric samples, respectively. No coronaviruses were identified. Sequencing of norovirus positive samples indicated an outbreak of norovirus strain GII.4 in 2006 with evidence of numerous circulating strains. Multiple samples from the same immunocompromised patients demonstrated symptomatic shedding of norovirus for up to 32 weeks and astrovirus for 12 weeks. RT-PCR detected 99 of 111 (89%) adenovirus-positive samples versus 12 (11%) by EM, and 186 of 192 (97%) sapovirus/astrovirus/norovirus-positive samples versus 21 (11%) by EM. Noroviruses and adenoviruses are common causes of gastroenteritis in children. Immunocompromised patients can be infected with multiple viruses and shed viruses in their stools for prolonged periods. This data support the superiority of RT-PCR compared to EM for diagnosis of viral gastroenteritis.

The evidence of Tobacco rattle virus impact on host plant organelles ultrastructure.

Tobraviruses, like other (+) stranded RNA viruses of plants, replicate their genome in cytoplasm and use such usual membranous structures like endoplasmic reticulum. Based on the ultrastructural examination of Tobacco rattle virus (TRV)-infected potato and tobacco leaf tissues, in this work we provide evidence of the participation of not only the membranous and vesicular ER structures but also other cell organelles during the viral infection cycle. Non-capsidated TRV PSG particles (potato isolate from the Netherlands) (long and short forms) were observed inside the nucleus while the presence of TRV capsid protein (CP) was detected in the nucleus caryolymph and within the nucleolus area. Both capsidated and non-capsidated viral particles were localized inside the strongly disorganized chloroplasts and mitochondria. The electron-dense TRV particles were connected with vesicular structures of mitochondria as well as with chloroplasts in both potato and tobacco tissues. At 15-30 days after infection, vesicles filled with TRV short particles were visible in mitochondria revealing the expanded cristae structures. Immunodetection analysis revealed the TRV PSG CP epitope inside chloroplast with disorganized thylakoids structure as well as in mitochondria of different tobacco and potato tissues. The ultrastructural analysis demonstrated high dynamics of the main cell organelles during the TRV PSG-Solanaceous plants interactions. Moreover, our results suggest a relationship between organelle changes and different stages of virus infection cycle and/or particle formation.

Lymantria dispar iflavirus 1 (LdIV1), a new model to study iflaviral persistence in lepidopterans.

The cell line IPLB-LD-652Y, derived from the gypsy moth (Lymantria dispar L.), is routinely used to study interactions between viruses and insect hosts. Here we report the full genome sequence and biological characteristics of a small RNA virus, designated Lymantria dispar iflavirus 1 (LdIV1), that was discovered to persistently infect IPLB-LD-652Y. LdIV1 belongs to the genus Iflavirus. LdIV1 formed icosahedral particles of approx. 30 nm in diameter and contained a 10, 044 nt polyadenylated, positive-sense RNA genome encoding a predicted polyprotein of 2980 aa. LdIV1 was induced by a viral suppressor of RNA silencing, suggesting that acute infection is restricted by RNA interference (RNAi). We detected LdIV1 in all tested tissues of gypsy-moth larvae and adults, but the virus was absent from other L. dispar-derived cell lines. We confirmed LdIV1 infectivity in two of these cell lines (IPLB-LD-652 and IPLB-LdFB). Our results provide a novel system to explore persistent infections in lepidopterans and a new model for the study of iflaviruses, a rapidly expanding group of viruses, many of which covertly infect their hosts.

Barley stripe mosaic virus: structure and relationship to the tobamoviruses.

Barley stripe mosaic virus (BSMV) is the type member of the genus Hordeivirus, rigid, rod-shaped viruses in the family Virgaviridae. We have used fiber diffraction and cryo-electron microscopy to determine the helical symmetry of BSMV to be 23.2 subunits per turn of the viral helix, and to obtain a low-resolution model of the virus by helical reconstruction methods. Features in the model support a structural relationship between the coat proteins of the hordeiviruses and the tobamoviruses.

Orchid fleck virus structural proteins N and P form intranuclear viroplasm-like structures in the absence of viral infection.

Orchid fleck virus (OFV) has a unique two-segmented negative-sense RNA genome that resembles that of plant nucleorhabdoviruses. In infected plant cells, OFV and nucleorhabdoviruses induce an intranuclear electron-lucent viroplasm that is believed to be the site for virus replication. In this study, we investigated the molecular mechanism by which OFV viroplasms are produced in vivo. Among OFV-encoded proteins, the nucleocapsid protein (N) and the putative phosphoprotein (P) were present in nuclear fractions of OFV-infected Nicotiana benthamiana plants. Transient coexpression of N and P, in the absence of virus infection, was shown to be sufficient for formation of an intranuclear viroplasm-like structure in plant cells. When expressed independently as a fluorescent protein fusion product in uninfected plant cells, N protein accumulated throughout the cell, while P protein accumulated in the nucleus. However, the N protein, when coexpressed with P, was recruited to a subnuclear region to induce a large viroplasm-like focus. Deletion and substitution mutagenesis demonstrated that the P protein contains a nuclear localization signal (NLS). Artificial nuclear targeting of the N-protein mutant was insufficient for formation of viroplasm-like structures in the absence of P. A bimolecular fluorescence complementation assay confirmed interactions between the N and P proteins within subnuclear viroplasm-like foci and interactions of two of the N. benthamiana importin-α homologues with the P protein but not with the N protein. Taken together, our results suggest that viroplasm formation by OFV requires nuclear accumulation of both the N and P proteins, which is mediated by P-NLS, unlike nucleorhabdovirus viroplasm utilizing the NLS on protein N.

Class II enveloped viruses.

A number of viruses transport their genomic material from cell to cell enclosed within a lipid bilayer that is in turn encased within a symmetric protein shell. This review focuses in a group of RNA viruses that have this type of virions. This group includes several of important human pathogenic viruses, such as the hepatitis C virus, dengue virus, chikungunya virus, rubella virus and the bunyaviruses. The best studied are the flaviviruses and the alphaviruses, which have a ß-sheet rich class II viral fusion protein used for entry into susceptible cells. We extend here the class II concept to encompass symmetric viruses in which the envelope proteins are derived from a precursor polyprotein containing two transmembrane glycoproteins arranged in tandem. The first glycoprotein acts as chaperone for the folding of the second one, which carries the membrane fusion function. Since the bunyaviruses, included here, are very similar to the class I arenaviruses in other respects, this analysis highlights the patchwork nature of the various viral functional modules acting at different stages of the virus cycle, which appear assembled from genes of different origins.

Viral research in Brazilian owls (Tyto alba and Rhinoptynx clamator) by transmission electron microscopy / Investigación viral en buhos brasileños (Tyto alba y Rhinoptynx clamator) a través de microscopía electrónica de transmisión

The barn-owl (Tyto Alba) and striped-owl (Rhinoptynx clamator) belong respectively to the families Tytonidae and Strigidae. Avian paramyxoviruses have been isolated from a variety of species of wild and domestic birds wordlwide causing diverse clinical symptoms and signs. Paramyxoviruses belong to the family Paramyxoviridae and Avulovirus genus, including nine serotypes (APMV 1 to 9). The lymphoid leukosis is a retrovirus-induced neoplasia. The avian retroviruses belong to the Retroviridae family and to the Alpharetrovirus genus. Coronaviruses can cause respiratory and enteric disease in several species of birds. They belong to the Coronaviridae family and to the groups 3a e 3c. In this study, we describe the presence of viruses in four owls, two barn owls (Tyto alba) and two striped owls (Rhinoptynx clamator), rescued from tree-lined streets of Sao Paulo, Brazil and sent to the Recovery Center of Wild Animals of the Tietê Ecological Park, where the animals died. Fragments of lung, liver and small intestine of these birds were processed for transmission electron microscopy utilizing negative staining (rapid preparation), immunoelectron microscopy and immunocitochemistry techniques. Under the transmission electron microscopy paramyxovirus particles, pleomorphic, roughly spherical or filamentous, measuring 100 to 500 nm of diameter containing an envelope covered by spikes, an herring-bone helical nucleocapsid-like structure, measuring 15 to 20 nm in diameter, were visualized in the samples of lung, liver and small intestine of all owls. In small intestine samples of the two striped-owl (owls 3 and 4) it was detected pleomorphic coronavirus particles with a diameter of 75-160 nm containing a solar corona-shaped envelope, with projections of approximately 20 nm of diameter. In liver fragments of one striped-owl (owl 4) pleomorphic particles of retrovirus with a diameter of 80-145 nm containing an envelope with short projections and diameter of 9 nm were....

La lechuza (Tyto Alba) y el búho de orejas (Rhinoptynx clamator) pertenecen respectivamente a las familias Strigidae y Tytonidae. El paramixovirus aviario se ha aislado de especies de vida silveste como las aves domésticas por todo el mundo, causando diversos síntomas clínicos. El paramixovirus pertenece a la familia Paramyxoviridae y al Avulovirus genus que incluye nueve serotipos (APMV 1 a 9). La leucosis linfoide es una neoplasia inducida por retrovirus. Los retrovirus aviarios pertenecen a la familia Retroviridae y el género Alpharetrovirus. Los coronavirus pueden causar enfermedades respiratorias y entéricas en varias especies de aves. Ellos pertenecen a la familia Coronaviridae y a los grupos 3a y 3c. En este estudio, se describe la presencia del virus en cuatro búhos, dos lechuzas (Tyto alba) y dos búhos de orejas (Rhinoptynx clamator), rescatados de las calles arboladas de São Paulo, Brasil y enviados al Centro de Recuperación de Animales Silvestres del Parque Ecológico de Tietê, donde hubo murieron los animales. Fragmentos de pulmón, delhígado y del intestino delgado de estas aves fueron procesados para microscopía electrónica de transmisión utilizando tinción negativa (preparación rápida), inmunomicroscopía y técnicas de inmunocitoquímica. Bajo microscopía electrónica de transmisión, partículas de paramixovirus, pleomórficas, aproximadamente esféricas o filamentosas, de 100 a 500 nm de diámetro con un sobre cubierto por espigas, y nucleocápside helicoidal con características de espiga, midiendo 15 a 20 nm de diámetro, fueron visualizadas en las muestras de pulmón, hígado e intestino delgado de todos los búhos. En muestras de intestino delgado de dos búho de orejas (búhos 3 y 4) se detectaron partículas pleomórficas con coronavirus de un diámetro de 75-160 nm con un sobre con forma de corona solar, con proyecciones de aproximadamente 20 nm de diámetro. En el hígado de un búho de orejas (búho 4) se observaron partículas pleomórficas de retrovirus con ...

Isolation and characterization of a single-stranded RNA virus infecting the bloom-forming diatom Chaetoceros socialis.

Diatoms are very significant primary producers in the world's oceans. Various environmental factors affect the depletion of diatom populations. The importance of viruses as a potential mortality source has recently been recognized. We isolated and characterized a new diatom virus (Chaetoceros socialis f. radians RNA virus [CsfrRNAV]) causing the lysis of the bloom-forming species Chaetoceros socialis Lauder f. radians (Schütt) Proschkina-Lavrenko. The virus infectious to C. socialis f. radians was isolated from water samples collected in Hiroshima Bay. Here we show the physiology, morphology, and genome characteristics of the virus clone. Virions were 22 nm in diameter and accumulated in the cytoplasm of the host cells. The latent period and the burst size were estimated to be <48 h and 66 infectious units per host cell, respectively. CsfrRNAV harbors a single-stranded RNA (ssRNA) genome and encodes at least three polypeptides of 32.0, 28.5, and 25.0 kDa. Sequencing analysis shows the length of the genome is 9,467 bases, excluding a poly(A) tail. The monophyly of CsfrRNAV and other diatom-infecting RNA viruses, Rhizosolenia setigera RNA virus and Chaetoceros tenuissimus RNA virus, was strongly supported by phylogenetic analysis based on the amino acid sequence of the RNA-dependent RNA polymerase domains. This suggested a new ssRNA virus family, Bacillariornaviridae. This discovery of CsfrRNAV may aid in further understanding the ecological dynamics of the C. socialis f. radians population in nature and the relationships between ssRNA diatom viruses and their hosts.

Picornavirales, a proposed order of positive-sense single-stranded RNA viruses with a pseudo-T = 3 virion architecture.

Despite the apparent natural grouping of "picorna-like" viruses, the taxonomical significance of this putative "supergroup" was never addressed adequately. We recently proposed to the ICTV that an order should be created and named Picornavirales, to include viruses infecting eukaryotes that share similar properties: (i) a positive-sense RNA genome, usually with a 5'-bound VPg and 3'-polyadenylated, (ii) genome translation into autoproteolytically processed polyprotein(s), (iii) capsid proteins organized in a module containing three related jelly-roll domains which form small icosahedral, non-enveloped particles with a pseudo-T = 3 symmetry, and (iv) a three-domain module containing a superfamily III helicase, a (cysteine) proteinase with a chymotrypsin-like fold and an RNA-dependent RNA polymerase. According to the above criteria, the order Picornavirales includes the families Picornaviridae, Comoviridae, Dicistroviridae, Marnaviridae, Sequiviridae and the unassigned genera Cheravirus, Iflavirus and Sadwavirus. Other taxa of "picorna-like" viruses, e.g. Potyviridae, Caliciviridae, Hypoviridae, do not conform to several of the above criteria and are more remotely related: therefore they are not being proposed as members of the new order. Newly described viruses, not yet assigned to an existing taxon by ICTV, may belong to the proposed order.

Molecular and biological characterization of deformed wing virus of honeybees (Apis mellifera L.).

Deformed wing virus (DWV) of honeybees (Apis mellifera) is closely associated with characteristic wing deformities, abdominal bloating, paralysis, and rapid mortality of emerging adult bees. The virus was purified from diseased insects, and its genome was cloned and sequenced. The genomic RNA of DWV is 10,140 nucleotides in length and contains a single large open reading frame encoding a 328-kDa polyprotein. The coding sequence is flanked by a 1,144-nucleotide 5' nontranslated leader sequence and a 317-nucleotide 3' nontranslated region, followed by a poly(A) tail. The three major structural proteins, VP1 (44 kDa), VP2 (32 kDa), and VP3 (28 kDa), were identified, and their genes were mapped to the N-terminal section of the polyprotein. The C-terminal part of the polyprotein contains sequence motifs typical of well-characterized picornavirus nonstructural proteins: an RNA helicase, a chymotrypsin-like 3C protease, and an RNA-dependent RNA polymerase. The genome organization, capsid morphology, and sequence comparison data indicate that DWV is a member of the recently established genus Iflavirus.