- Volume 103, Issue 3, 2022

The infectious pancreatic necrosis virus (IPNV) is responsible for significant economic losses in the aquaculture industry. It is an unenveloped virus with an icosahedral capsid. Its viral genome comprises two dsRNA segments, A and B. Segment A contains a small ORF, which encodes VP5, and a large ORF, which encodes a polyprotein that generates the structural proteins and the viral protease. Segment B encodes the RNA-dependent RNA polymerase (RdRp), called VP1 in this free form, or Vpg when it covalently attaches to the viral RNA. The viral genome does not have cap or poly(A). Instead, each 5′ end is linked to the Vpg. Recently, we demonstrated that mRNA-A contains an internal ribosome entry site (IRES) to command polyprotein synthesis. However, the presence of Vpg on IPNV mRNAs and its impact on cellular translation has not been investigated. This research demonstrates that IPNV mRNAs are linked to Vpg and that this protein inhibits cap-dependent translation on infected cells. Also, it is demonstrated that Vpg interacts with eIF4E and that rapamycin treatment partially diminishes the viral protein synthesis. In addition, we determined that an IRES does not command translation of IPNV mRNA-B. We show that VPg serves as a cap substitute during the initiation of IPNV translation, contributing to understanding the replicative cycle of Birnaviruses. Our results indicate that the viral protein VP1/Vpg is multifunctional, having a significant role during IPNV RNA synthesis as the RdRp and the primer for IPNV RNA synthesis and translation as the viral protein genome, acting as a cap substitute.

Intraspecific variation in pathogen shedding impacts disease transmission dynamics; therefore, understanding the host factors associated with individual variation in pathogen shedding is key to controlling and preventing outbreaks. In this study, ileum and bursa of Fabricius tissues of wild-bred mallards (Anas platyrhynchos) infected with low-pathogenic avian influenza (LPAIV) were evaluated at various post-infection time points to determine genetic host factors associated with intraspecific variation in viral shedding. By analysing transcriptome sequencing data (RNA-seq), we found that LPAIV-infected wild-bred mallards do not exhibit differential gene expression compared to uninfected birds, but that gene expression was associated with cloacal viral shedding quantity early in the infection. In both tissues, immune gene expression was higher in high/moderate shedding birds compared to low shedding birds, and significant positive relationships with viral shedding were observed. In the ileum, expression for host genes involved in viral cell entry was lower in low shedders compared to moderate shedders at 1 day post-infection (DPI), and expression for host genes promoting viral replication was higher in high shedders compared to low shedders at 2 DPI. Our findings indicate that viral shedding is a key factor for gene expression differences in LPAIV-infected wild-bred mallards, and the genes identified in this study could be important for understanding the molecular mechanisms driving intraspecific variation in pathogen shedding.

Rabies, caused by rabies lyssavirus (RABV), is a fatal disease among humans and almost all warm-blooded animals. Our previous study showed that the long non-coding RNA (lncRNA) EZH2 degradation-associated lncRNA (EDAL) effectively inhibits RABV infection both in vitro and in vivo by degrading EZH2 and promoting the transcription of an antiviral gene, Pcp4l1. Herein, we found that recombinant RABV expressing EDAL (rRABV-EDAL) restricts RABV replication in primary granule neurons but not in primary cortical neurons or astrocytes. Further study revealed that EDAL induced EZH2 protein degradation and thereby decreased trimethylation of lysine 27 on the histone 3 (H3K27me3) level in granule neuron cells but not in cortical neurons or astrocytes. Furthermore, rRABV-EDAL infection induces more Pcp4l1 mRNA transcription in granule neurons, while there are almost no obvious changes in cortical neurons or astrocytes. Consistently, compared with the parent virus RABV, reduced pathogenicity of rRABV-EDAL was observed in mice post-intranasal infection but not intramuscular infection. These results suggest that the lncRNA EDAL restricts RABV replication in a cell-specific and infection route-dependent manner.

Hepatitis C virus (HCV) infection affects more than 71 million people worldwide. The disease slowly progresses to chronic, long-term liver injury which leads to hepatocellular carcinoma (HCC) in 5 % of infections. The alternative reading frame protein (ARFP/core+1) is encoded by a sequence overlapping the HCV core gene in the +1 reading frame. Its role in hepatitis C pathogenesis and the viral life cycle is unclear, although some observers have related its production to disease progression and the development of HCC. The aim of this study was to determine whether ARFP is immunogenic in patients with chronic HCV genotype 3 infection and to assess whether sero-reactivity is associated with disease progression, particularly to HCC. Immunogenic epitopes within the protein were predicted by a bioinformatics tool, and three −20 aa length-peptides (ARFP-P1, ARFP-P2 and ARFP-P3) were synthesized and used in an avidin-biotin ARFP/core+1 peptide ELISA. Serum samples from 50 patients with chronic HCV genotype 3 infection, 50 genotype-1 patients, 50 HBV patients and 110 healthy controls were tested. Sero-reactivity to the ARFP peptides was also tested and compared in 114 chronic HCV genotype-3 patients subdivided on the basis of disease severity into non-cirrhotic, cirrhotic and HCC groups. Chronic HCV genotype-3 patients showed noticeable rates of reactivity to ARFP and core peptides. Seropositivity rates were 58% for ARFP-P1, 47 % for ARFP-P2, 5.9 % for ARFP-P3 and 100 % for C22 peptides. There was no significant difference between these seroreactivities between HCV genotype-3 patients with HCC, and HCV genotype-3 patients with and without liver cirrhosis. Patients with chronic HCV genotype-3 infection frequently produce antibodies against ARFP/core+1 protein. ARFP peptide reactivity was not associated with disease severity in patients with HCV genotype-3. These results support the conclusion that ARFP/core+1 is produced during HCV infection, but they do not confirm that antibodies to ARFP can indicate HCV disease progression.