Innovative toolbox for the quantification of Drosophila C virus, Drosophila A virus, and Nora virus

Several viruses, including picornaviruses, are known to establish persistent infections, but the mechanisms involved are poorly understood. Here, a novel picorna-like virus, Nora virus, which causes a persistent infection in Drosophila melanogaster, is described. It has a single-stranded, positive-sense genomic RNA of 11879 nt, followed by a poly(A) tail. Unlike other picorna-like viruses, the genome has four open reading frames (ORFs). One ORF encodes a picornavirus-like cassette of proteins for virus replication, including an iflavirus-like RNA-dependent RNA polymerase and a helicase that is related to those of mammalian picornaviruses. The three other ORFs are not closely related to any previously described viral sequences. The unusual sequence and genome organization in Nora virus suggest that it belongs to a new family of picorna-like viruses. Surprisingly, Nora virus could be detected in all tested D. melanogaster laboratory stocks, as well as in wild-caught material. The viral titres varied enormously, between 104 and 1010 viral genomes per fly in different stocks, without causing obvious pathological effects. The virus was also found in Drosophila simulans, a close relative of D. melanogaster, but not in more distantly related Drosophila species. It will now be possible to use Drosophila genetics to study the factors that control this persistent infection.

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The Novel Agrotis ipsilon Nora Virus Confers Deleterious Effects to the Fitness of Spodoptera frugiperda (Lepidoptera: Noctuidae)

Frontiers in Microbiology ◽

10.3389/fmicb.2021.727202 ◽

2021 ◽

Vol 12 ◽

Author(s):

Tong Li ◽

Ruobing Guan ◽

Yuqing Wu ◽

Su Chen ◽

Guohui Yuan ◽

...

Keyword(s):

Spodoptera Frugiperda ◽

Rna Virus ◽

Open Reading Frames ◽

Agrotis Ipsilon ◽

The Novel ◽

Black Cutworm ◽

A Genome ◽

Nora Virus ◽

Novel Host ◽

Lepidoptera Noctuidae

In the present study, we identified a novel, positive-sense single-stranded RNA virus in the Chinese black cutworm, Agrotis ipsilon. It has a genome length of 11,312 nucleotides, excluding the poly(A) tails, and contains five open reading frames. The ORF2 encodes the conserved domains of RNA helicase and RNA-dependent RNA polymerase, while ORF4 and 5 encode three viral proteins. Herein, the A. ipsilon virus was clustered with a Helicoverpa armigera Nora virus and was thus provisionally named “Agrotis ipsilon Nora virus” (AINV). AINV was successfully transmitted into a novel host, Spodoptera frugiperda, through injection, causing a stable infection. This found the possibility of horizontal AINV transmission among moths belonging to the same taxonomic family. Nonetheless, AINV infection was deleterious to S. frugiperda and mainly mediated by antiviral and amino acid metabolism-related pathways. Furthermore, the infection significantly increased the S. frugiperda larval period but significantly reduced its moth eclosion rate. It suggests that AINV is probably to be a parasitic virus of S. frugiperda.

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Nora Virus VP4b and ORF1 Circulate in Hemolymph of Infected D. melanogaster with Coordinate Expression of Vago and Vir-1

Vaccines ◽

10.3390/vaccines8030491 ◽

2020 ◽

Vol 8 (3) ◽

pp. 491

Author(s):

Amanda Macke ◽

Wilfredo Lopez ◽

Darby J. Carlson ◽

Kimberly A. Carlson

Keyword(s):

Rna Virus ◽

Primary Site ◽

The Novel ◽

Qrt Pcr ◽

Basal Expression ◽

Virus Rna ◽

Nora Virus ◽

Candidate Target ◽

The Brain ◽

Insight Into

Study of the novel RNA virus, Nora virus, which is a persistent, picorna-like virus that replicates in the gut of Drosophila melanogaster offers insight into human innate immunity and other picorna-like viruses. Nora virus infection leads to a locomotor abnormality and upregulation of two candidate target proteins, Vago and Virus-induced RNA 1 (Vir-1). These proteins are uncharacterized in response to Nora virus. We hypothesize that Nora virus is circulating in the hemolymph of Nora virus-infected D. melanogaster, allowing for migration beyond the primary site of replication in the gut. Analysis by qRT-PCR demonstrated biphasic viral load and corresponding vago and vir-1 transcription levels, suggesting transcription of vago and vir-1 occurs in response to viral infection. However, Vir-1 is also present in virus-free D. melanogaster suggesting basal expression or alternative functions. Presence of Nora virus RNA and the Viral Protein 4b (VP4b), in hemolymph of infected D. melanogaster supports the hypothesized circulation of Nora virus in the hemolymph. The study suggests that impaired locomotor function may be due to transport of Nora virus from the gut to the brain via the hemolymph.

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Drosophila Nora virus capsid proteins differ from those of other picorna-like viruses

Virus Research ◽

10.1016/j.virusres.2011.05.006 ◽

2011 ◽

Vol 160 (1-2) ◽

pp. 51-58 ◽

Cited By ~ 13

Author(s):

Jens-Ola Ekström ◽

Mazen S. Habayeb ◽

Vaibhav Srivastava ◽

Thomas Kieselbach ◽

Gunnar Wingsle ◽

...

Keyword(s):

Capsid Proteins ◽

Virus Capsid ◽

Nora Virus

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Analysis of immune-related genes during Nora virus infection of Drosophila melanogaster using next generation sequencing

AIMS Microbiology ◽

10.3934/microbiol.2018.1.123 ◽

2018 ◽

Vol 4 (1) ◽

pp. 123-139 ◽

Cited By ~ 6

Author(s):

Wilfredo Lopez ◽

◽

Alexis M. Page ◽

Darby J. Carlson ◽

Brad L. Ericson ◽

...

Keyword(s):

Drosophila Melanogaster ◽

Next Generation Sequencing ◽

Virus Infection ◽

Next Generation ◽

Nora Virus ◽

Immune Related Genes ◽

Generation Sequencing

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Differential gene expression related to Nora virus infection of Drosophila melanogaster

Virus Research ◽

10.1016/j.virusres.2013.03.021 ◽

2013 ◽

Vol 175 (2) ◽

pp. 95-100 ◽

Cited By ~ 20

Author(s):

Ethan J. Cordes ◽

Kellie D Licking-Murray ◽

Kimberly A. Carlson

Keyword(s):

Gene Expression ◽

Drosophila Melanogaster ◽

Virus Infection ◽

Differential Gene Expression ◽

Nora Virus ◽

Differential Gene

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Nora Virus Persistent Infections Are Not Affected by the RNAi Machinery

PLoS ONE ◽

10.1371/journal.pone.0005731 ◽

2009 ◽

Vol 4 (5) ◽

pp. e5731 ◽

Cited By ~ 21

Author(s):

Mazen S. Habayeb ◽

Jens-Ola Ekström ◽

Dan Hultmark

Keyword(s):

Rnai Machinery ◽

Persistent Infections ◽

Nora Virus

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Structure of Nora virus at 2.7 Å resolution and implications for receptor binding, capsid stability and taxonomy

Scientific Reports ◽

10.1038/s41598-020-76613-1 ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Pasi Laurinmäki ◽

Shabih Shakeel ◽

Jens-Ola Ekström ◽

Pezhman Mohammadi ◽

Dan Hultmark ◽

...

Keyword(s):

Capsid Protein ◽

Rna Virus ◽

Virus Entry ◽

Amino Acid Sequences ◽

Structural Elements ◽

Helical Bundles ◽

Signalling Molecule ◽

Nora Virus ◽

Virus Genomes ◽

Capsid Stability

AbstractNora virus, a virus of Drosophila, encapsidates one of the largest single-stranded RNA virus genomes known. Its taxonomic affinity is uncertain as it has a picornavirus-like cassette of enzymes for virus replication, but the capsid structure was at the time for genome publication unknown. By solving the structure of the virus, and through sequence comparison, we clear up this taxonomic ambiguity in the invertebrate RNA virosphere. Despite the lack of detectable similarity in the amino acid sequences, the 2.7 Å resolution cryoEM map showed Nora virus to have T = 1 symmetry with the characteristic capsid protein β-barrels found in all the viruses in the Picornavirales order. Strikingly, α-helical bundles formed from the extended C-termini of capsid protein VP4B and VP4C protrude from the capsid surface. They are similar to signalling molecule folds and implicated in virus entry. Unlike other viruses of Picornavirales, no intra-pentamer stabilizing annulus was seen, instead the intra-pentamer stability comes from the interaction of VP4C and VP4B N-termini. Finally, intertwining of the N-termini of two-fold symmetry-related VP4A capsid proteins and RNA, provides inter-pentamer stability. Based on its distinct structural elements and the genetic distance to other picorna-like viruses we propose that Nora virus, and a small group of related viruses, should have its own family within the order Picornavirales.

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Characterization of Nora Virus Structural Proteins via Western Blot Analysis

Scientifica ◽

10.1155/2016/9067848 ◽

2016 ◽

Vol 2016 ◽

pp. 1-8 ◽

Cited By ~ 1

Author(s):

Brad L. Ericson ◽

Darby J. Carlson ◽

Kimberly A. Carlson

Keyword(s):

Western Blot ◽

Open Reading Frames ◽

Structural Proteins ◽

Polypeptide Composition ◽

Virus Particles ◽

E Coli ◽

Nora Virus ◽

Monospecific Antisera ◽

Reading Frames

Nora virus is a single stranded RNA picorna-like virus with four open reading frames (ORFs). The coding potentials of the ORFs are not fully characterized, but ORF3 and ORF4 are believed to encode the capsid proteins (VP3, VP4a, VP4b, and VP4c) comprising the virion. To determine the polypeptide composition of Nora virus virions, polypeptides from purified virus were compared to polypeptides detected in Nora virus infectedDrosophila melanogaster. Nora virus was purified from infected flies and used to challenge mice for the production of antisera.ORF3,ORF4a,ORF4b, andORF4cwere individually cloned and expressed inE. coli; resultant recombinant proteins purified and were used to make monospecific antisera. Antisera were evaluated via Western blot against whole virus particles and Nora virus infected fly lysates. Viral purification yielded two particle types with densities of ~1.31 g/mL (empty particles) and ~1.33 g/mL (complete virions). Comparison of purified virus polypeptide composition to Nora virus infectedD. melanogasterlysate showed the number of proteins in infected cell lysates is less than purified virus. Our results suggest the virion is composed of 6 polypeptides, VP3, VP4a, two forms of VP4b, and two forms of VP4c. This polypeptide composition is similar to other small RNA insect viruses.

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