The hidden diversity of ancient bornaviral sequences from X and P genes in vertebrate genomes

Endogenous viral elements are not only derived from retroviruses but also from bornaviruses. Existence of endogenous bornavirus-like elements (EBLs) is attributed to reverse transcription of bornavirus mRNA followed by integration into host genome by long interspersed nuclear element-1 retro-transposon. However, detection of EBLX/Ps is limited as compared to EBLs derived from bornaviral N, P, M, G and L genes. Low detection of EBLX/Ps was hypothesized to result from limitations of sequence similarity search using tBLASTn. A 1.9-kb miEBLN-1 insertion present in genome of miniopterid bat was used for analysis. The 1.9-kb insertion consisted of 1.2-kb miEBLN-1 region and 0.7-kb unannotated region, and was similar in size to N/X/P mRNA of present orthobornaviruses. The insertion is shown to have originated from ancient N/X/P transcript which integrated into the genome of miniopterid bat. The findings illustrate existence of hidden EBLN/X/Ps or EBLX/Ps in vertebrate genomes. Despite of high evolutionary rates of X and P genes, the genes are likely to have been maintained for million years as demonstrated by intact X and P-like open reading frames of miEBLX/P which were expressed as a fusion transcript with cellular gene, ZNF451. Expression of ZNF451/miEBLP-4 chimeric transcript suggested potential co-option of EBLX/P in miniopterid bats.
(LML)

The Ebola virus VP40 matrix layer undergoes endosomal disassembly essential for membrane fusion

The notable filamentous shape and stability of Ebola virus (EBOV) depend on matrix viral protein 40 (VP40). The virus enters host cells by macropinocytosis where it fuses with endosomal membrane. This study evaluated conformational changes of VP40 during virion entry and factors initiating EBOV disassembly. Characterization of EBOV using endosome mimicking conditions and in situ cryo-electron tomography showed EBOV entering host cells via the endosomal route, and the VP40 matrices were disassembled inside the endosomes. Considering the EBOV nucleocapsids were intact in all endosomes, the study postulated that uncoating of EBOV starts with disassembly of the VP40 matrix in late endosomal compartments and detachment of the matrix from viral envelope. Disassembly is followed by membrane fusion via glycoproteins and release of nucleocapsid into the cytoplasm. The Disassembly of VP40 matrix is triggered by low endosomal pH which results in weakening of VP40 interactions with negatively charged lipids. The pH-mediated VP40 disassembly was noted to be independent of other viral proteins. Using membrane modeling approach, the study reveals that disassembly of the VP40 matrix is critical for membrane fusion. The study hypothesized that the findings unveil promising target for development of virus matrix-specific weak base inhibitors for late penetrating viruses.

(LML)

Canine distemper virus N protein induces autophagy to facilitate viral replication

Canine distemper virus (CDV) contains 6 transcription-units (N-P-M-F-H-L) and causes fatal disease in Canidae. Although autophagy plays roles in degradation and activation of immune response against pathogens, viruses have developed strategies to evade autophagy. Induction of autophagy in members of Paramyxoviridae family has been confirmed. Therefore, this study evaluated effect of autophagy mechanisms on CDV infection. The study confirmed onset of autophagy in CDV infected Vero cells by elevated autophagy associated proteins (LC3-II) with progression of CDV infection. Vero cells were treated with rapamycin to induce autophagy and Wortmannin and Chloroquine to inhibit autophagy. Autophagy induction resulted in increased viral RNA titer while inhibition of autophagy resulted in decreased viral RNA titer. Onset of autophagy in Vero cells infected with infectious and UV-inactivated CDV implicated structural proteins of CDV in virus induced autophagy. Further analysis using flag-tagged proteins, revealed that flag-nucleoprotein had the most significant increase in LC3-II level. Flag-nucleoprotein further promoted degradation of p62 protein. These results suggested that CDV nucleoprotein induced complete autophagic flux. The mammalian target of rapamycin (mTOR), a regulator of autophagy, was detected in low levels in Vero cells expressing flag-nucleoprotein, suggesting that CDV nucleoprotein induced autophagy by inhibiting mTOR pathway to facilitate viral replication. 

(LML)

The rVSV-EBOV vaccine provides limited cross-protection against Sudan virus in guinea pig

Sudan virus (SUDV), belonging to the Filoviridae family, caused high mortality among humans during the 2022 pandemic in Uganda. The research and manufacturing of vaccine is of critical urgency to mitigate SUDV infection. The clinical efficacy of the filovirus vaccines based on recombinant vesicular stomatitis virus (rVSV) has been demonstrated and evaluated on Ebola virus (EBOV), and rVSV-EBOV exhibit cross-protective effect against other filoviruses. Meanwhile, efficacy of other rVSV-based filovirus vaccine remains uncertain at the laboratory level due to interference from various factors in animal experiment, rendering it challenging to evaluate. Thus, this study aimed to investigate rVSV-SUDV efficacy and rVSV-EBOV cross-protection to SUDV on guinea pig models that are used for countermeasure evaluation in filovirus vaccine.  In conclusion, the administration of rVSV-SUDV demonstrated protective effect against SUDV infection in guinea pig. rVSV-EBOV vaccination implied cross-protective effect in guinea pig infected with SUDV. However, this study indicated that rVSV-SUDV vaccination administrated to guinea pigs that infected SUDV did not confer cross-protective effect against EBOV.  This research provides evidence to support that rVSV-based filovirus vaccine has the capability to induce cross-protective immune response in guinea pig models. However, certain rVSV-based filovirus vaccines elicit different cross-reactive immune response to various filovirus infection.

(HW)

Structure and antigenicity of divergent Henipavirus fusion glycoproteins

 The novel Henipavirus (HNV), Langya virus (LayV) was found in China, caused severe pneumonic disease in human. The discovery of LayV that causes infection in human beside Nipah virus (NiV) and Hendra virus (HeV), highlights the emergence of HNVs threat to global health. Despite the significant mortality and morbidity, there are currently no licensed vaccines or treatment for HNVs. This study was performed to identify the prefusion structures of Mòjiāng virus (MojV) and LayV fusion (F) proteins using cryogenic electron microscopy. The results showed that LayV and MojV F have a similar structure with NiV F, indicating the tendency of RNA viruses that prone to significant rates of mutation while maintaining overall structure for functionality. This study discovered that prefusion NiV F polyclonal sera had minimal to no cross-reactivity with MojV and LayV F, which has significance for the development of broad-spectrum vaccines. The HNV F glycoproteins possess higher sequence conservation in comparison to HNV G glycoproteins, indicating that F glycoprotein may be a better candidate for a broad-spectrum vaccine. Glycoproteomic analysis showed a significant less glycosylation of F protein in LayV, however, LayV F contains a glycan that protect a susceptible site in NiV. Despite being structurally similar with NiV, LayV and MojV F have a significant difference in the antigenic profile. The lack of glycans on the main surface of glycoproteins may translate into higher susceptibility to antibody neutralization, which may have consequences for vaccine and treatment design.

(MA)

Development of SARS‐CoV‐2 animal vaccines using a stable and efficient NDV expression system

SARS CoV-2, a coronavirus, infects humans and animals. Animal species that are vulnerable would probably end up serving as mixing vessels. An alternative to preventing animals from acting as mixing vessels for the SARS CoV-2 is to vaccinate them. In this study, recombinant ND viruses expressing SARS CoV-2's spike or subunit protein were employed. The immunologic reactivity of rNDV was confirmed using Western blotting, neutralization, and other immunological techniques. The surface of the rNDV-vectored virus has SARS-CoV2 structural protein-1 (S-1) and the spike (S) protein inserted. In contrast to rNDV-S1, which induced a broad spectrum of T-cell responses but few antibodies, rNDV-S induced robust antibodies and an S-specific T-cell response after intramuscular vaccination. An rNDV-vectored vaccine candidate that is stable and effective and has strong humoral and cellular protection has been established.

(MMR)

Recombinant vesicular stomatitis vaccine against Nipah virus has a favorable safety profile: Model for assessment of live vaccines with neurotropic potential

Nipah virus (NiV) is a neurotropic bat-borne virus that causes inflammation of the brain. The glycoprotein of NiV binds to ephrin B2 and B3 receptors which are expressed on neural cells. A recombinant vesicular stomatitis virus-Nipah (rVSV-Nipah) vaccine was developed using reverse genetics system. The candidate live vaccine was constructed by deleting vesicular stomatitis virus (VSV) glycoprotein gene and replacing with glycoprotein genes of negative-sense single-strand RNA viruses, Ebola and Nipah virus. The Ebola glycoprotein is required for fusion and viral entry. The addition of NiV glycoprotein with ability to bind neural cells has a potential to increase neurovirulence. Therefore, safety tests for neurovirulence were conducted to assess whether the NiV glycoprotein changes tropism of the rVSV vector and if virulence factor is retained. The candidate vaccine was directly inoculated into mice and hamsters through intracerebral route. Correspondingly, rVSV-EBOV and yellow fever (YF) commercial vaccines were also inoculated as active controls. Furthermore, monkey neurovirulence test (MNVT) was conducted using monkeys. The results of MNVT showed that the candidate vaccine was significantly less neurovirulent than the live YF vaccine. The study postulated that the candidate vaccine is likely to be safer than YF which has record of safe use in humans. 

(LML)

Plasticity, Paralogy and Pseudogenization: Rhabdoviruses of Freshwater Mussels Elucidate Mechanisms of Viral Genome Diversification and the Evolution of the Finfish-Infecting Rhabdoviral Genera

Killamcar Virus 1 (KILLV-1) from Lampsilis cardium and Chemarfal Virus 1 (CHMFV-1) from Margarutufera falcata were the first two Rhabdovirus reported in mollusks. These two novel viruses highlight an uncommon example of Rhabdoviral plasticity among negative-sense RNA viruses. Based on sequence analysis with phylogenetic interferences and analyses of transcriptional regulation, KILLV-1 is an Alpharhabdovirus. KILLV-1 developed and diverged due to glycoprotein gene duplication processing to a new gene paralog, increasing its genome size and complexity, which separates KILLV-1 from Scophravirus, a closely related virus. CHMFV-1 belongs to the family of Gammarhabdovirinae. It closely relates to Novirhabdovirus owing to its comparatively longer genome size and lower GC content. Intriguingly, CHMFV-1 also carries a pseudogenized version of Non virion (NV) genes that encoded for NV proteins, which is essential for the establishment of antiviral state during the Novirhabdovirus infection of fish. This discovery suggests that the origin of NV protein occurred beforehand to the divergence of Novirhabdovirus and CHMFV-1 because these viruses contain NV gene. Furthermore, because most Rhabdoviruses are mostly found in vertebrate animals like finfish, host switching from finfish to freshwater mussels theoretically occurs when freshwater mussels’ larvae parasitize on finfish and consume fish blood and tissues.

(HW)