Human noroviruses (HuNoVs) are the leading cause of viral gastroenteritis worldwide; yet currently, no vaccines or FDA-approved antiviral drugs are available to counter these pathogens. To understand HuNoV biology and the epithelial response to infection, we performed transcriptomic analyses, RT-qPCR, CRISPR-Cas9 modification of human intestinal enteroid (HIE) cultures, and functional studies with two virus strains (a pandemic GII.4 and a bile acid-dependent GII.3 strain). We identified a predominant type III interferon (IFN)-mediated innate response to HuNoV infection. Replication of both strains is sensitive to exogenous addition of IFNs, suggesting the potential of IFNs as therapeutics. To obtain insight into IFN pathway genes that play a role in the antiviral response to HuNoVs, we developed knockout (KO) HIE lines for IFN alpha and lambda receptors and the signaling molecules, MAVS, STAT1, and STAT2 An unexpected differential response of enhanced replication and virus spread was observed for GII.3, but not the globally dominant GII.4 HuNoV in STAT1-knockout HIEs compared to parental HIEs. These results indicate cellular IFN responses restrict GII.3 but not GII.4 replication. The strain-specific sensitivities of innate responses against HuNoV replication provide one explanation for why GII.4 infections are more widespread and highlight strain specificity as an important factor in HuNoV biology. Genetically modified HIEs for innate immune genes are useful tools for studying immune responses to viral or microbial pathogens.

https://www.pnas.org/content/pnas/early/2020/09/08/2010834117.full.pdf

Human norovirus exhibits strain-specific sensitivity to host interferon pathways in human intestinal enteroids.

Epidemiología y complicaciones neurológicas de la infección por el virus del Zika: un nuevo virus neurotropo emergente

The central nervous system (CNS) is vulnerable to viral infection, yet few host factors in the CNS are known to defend against invasion by neurotropic viruses. Long noncoding RNAs (lncRNAs) have been revealed to play critical roles in a wide variety of biological processes and are highly abundant in the mammalian brain, but their roles in defending against invasion of pathogens into the CNS remain unclear. We report here that multiple neurotropic viruses, including rabies virus, vesicular stomatitis virus, Semliki Forest virus, and herpes simplex virus 1, elicit the neuronal expression of a host-encoded lncRNA EDAL. EDAL inhibits the replication of these neurotropic viruses in neuronal cells and rabies virus infection in mouse brains. EDAL binds to the conserved histone methyltransferase enhancer of zest homolog 2 (EZH2) and specifically causes EZH2 degradation via lysosomes, reducing the cellular H3K27me3 level. The antiviral function of EDAL resides in a 56-nt antiviral substructure through which its 18-nt helix-loop intimately contacts multiple EZH2 sites surrounding T309, a known O-GlcNAcylation site. EDAL positively regulates the transcription of Pcp4l1 encoding a 10-kDa peptide, which inhibits the replication of multiple neurotropic viruses. Our findings show that a neuronal lncRNA can exert an effective antiviral function via blocking a specific O-GlcNAcylation that determines EZH2 lysosomal degradation, rather than the traditional interferon-dependent pathway.

/pdf/a-novel-antiviral-lncrna-edal-shields-a-t309-o-glcnacylation-qk5glvt7z1.pdf

A novel antiviral lncRNA, EDAL, shields a T309 O -GlcNAcylation site to promote EZH2 lysosomal degradation

Differential roles of interferons in innate responses to mucosal viral infections.

Construction and evaluation of recombinant Lactobacillus plantarum NC8 delivering one single or two copies of G protein fused with a DC-targeting peptide (DCpep) as novel oral rabies vaccine.

Rabies, caused by rabies virus (RABV), is a fatal neurological disease that still causes more than 59,000 human deaths each year. Type III interferon IFN-λs are cytokines with type I IFN-like antiviral activities. Although IFN-λ can restrict the infection for some viruses, especially intestinal viruses, the inhibitory effect against RABV infection remains undefined. In this study, the function of type III IFN against RABV infection was investigated. Initially, we found that IFN-λ2 and IFN-λ3 could inhibit RABV replication in cells. To characterize the role of IFN-λ in RABV infection in a mouse model, recombinant RABVs expressing murine IFN-λ2 or IFN-λ3, termed as rB2c-IFNλ2 or rB2c-IFNλ3, respectively, were constructed and rescued. It was found that expression of IFN-λ could reduce the pathogenicity of RABV and limit viral spread in the brains by different infection routes. Furthermore, expression of IFN-λ could induce the activation of the JAK-STAT pathway, resulting in the production of interferon-stimulated genes (ISGs). It was also found that rRABVs expressing IFN-λ could reduce the production of inflammatory cytokines in primary astrocytes and microgila cells, restrict the opening of the blood-brain barrier (BBB), and prevent excessive infiltration of inflammatory cells into the brain, which could be responsible for the neuronal damage caused by RABV. Consistently, IFN-λ was found to maintain the integrity of tight junction (TJ) protein ZO-1 of BBB to alleviate neuroinflammation in a transwell model. Our study underscores the role of IFN-λ in inhibiting RABV infection, which potentiates IFN-λ as a possible therapeutic agent for the treatment of RABV infection.

https://www.mdpi.com/1999-4915/12/4/405/pdf

Interferon-λ Attenuates Rabies Virus Infection by Inducing Interferon-Stimulated Genes and Alleviating Neurological Inflammation.

Background Gut microbiome plays a crucial role in modulating human and animal immune responses. Rabies is a fatal zoonosis causing encephalitis in mammals and vaccination is the most effective method to control and eliminate rabies. The relationship between the gut microbiome and humoral immunity post rabies vaccination has not been investigated yet. Methods Mice orally administrated with a cocktail of broad-spectrum antibiotics were inoculated with rabies vaccines, and humoral immune response was analyzed at indicated time points. The 16S ribosomal RNA (16S rRNA) gene sequencing was performed on fecal samples from groups in vancomycin-treated and untreated mice. Mice were immunized with rabies vaccines and virus-neutralizing antibody (VNA) levels were measured, resulting in VNA high (H) and low (L) groups. Then 16S rRNA gene sequencing was performed on fecal samples from H and L group mice. Results After antibiotic (Abx) treatment, mice had decreased levels of rabies virus (RABV)-specific IgM, IgG, and virus-neutralizing antibody compared with untreated mice. Abx-treated mice had fewer T follicular helper cells, germinal center B cells, and antibody secreting cells (ASCs) in lymph nodes than did untreated mice. Gut microbiome facilitated secondary immune responses by increasing the generation of ASCs. Treatment with vancomycin alone had a similarly impaired effect on the humoral immune responses compared with Abx-treated mice. From the natural population group of mice received rabies vaccines, VNA titers vary significantly and the abundance of Clostridiales and Lachnospiraceae was positively associated with the antibody titers in mice. Conclusions Our results provide the evidence that the gut microbiome impacts humoral immunity post rabies vaccination, and further investigation of the mechanism will help the development of novel adjuvants and vaccines.

https://onlinelibrary.wiley.com/doi/pdf/10.1002/ctm2.161

Composition of the murine gut microbiome impacts humoral immunity induced by rabies vaccines.

Rabies virus (RABV) infection can initiate the host immune defence response and induce an antiviral state characterized by the expression of interferon (IFN)-stimulated genes (ISGs), among which the family of genes of IFN-induced protein with tetratricopeptide repeats (Ifits) are prominent representatives. Herein, we demonstrated that the mRNA and protein levels of Ifit1, Ifit2 and Ifit3 were highly increased in cultured cells and mouse brains after RABV infection. Recombinant RABV expressing Ifit3, designated rRABV-Ifit3, displayed a lower pathogenicity than the parent RABV in C57BL/6 mice after intramuscular administration, and Ifit3-deficient mice exhibited higher susceptibility to RABV infection and higher mortality during RABV infection. Moreover, compared with their individual expressions, co-expression of Ifit2 and Ifit3 could more effectively inhibit RABV replication in vitro. These results indicate that murine Ifit3 plays an essential role in restricting the replication and reducing the pathogenicity of RABV. Ifit3 acts synergistically with Ifit2 to inhibit RABV replication, providing further insight into the function and complexity of the Ifit family.

Murine Ifit3 restricts the replication of Rabies virus both in vitro and in vivo.

Rabies, caused by rabies virus (RABV), is a fatal zoonosis, which still poses a threat to public health in most parts of the world. Glycoprotein of RABV is the only viral surface protein, which is critical for the induction of virus-neutralizing antibodies (VNA). In order to improve the production of VNA, recombinant RABVs containing two copies of G gene and codon-optimized G gene were constructed by using reverse genetics, named LBNSE-dG and LBNSE-dOG, respectively. After being inoculated into the mouse brains, LBNSE-dOG induced more apoptosis and recruited more inflammatory cells than LBNSE-dG and LBNSE, resulting in reduced virulence in vivo. After intramuscular (im) immunization in mice, LBNSE-dOG promoted the formation of germinal centres (GCs), the recruitment of GC B cells and the generation of antibody-secreting cells (ASCs) in the draining lymph nodes (LNs). Consistently, LBNSE-dOG boosted the production of VNA and provided better protection against lethal RABV challenge than LBNSE-dG and LBNSE when it was used as both live and inactivated vaccines. Our results demonstrate that the codon-optimized RABV LBNSE-dOG displays attenuated pathogenicity and enhanced immunogenicity, therefore it could be a potential candidate for the next generation of rabies vaccines.

Fei Huang

Papers

A novel antiviral lncRNA, EDAL, shields a T309 O -GlcNAcylation site to promote EZH2 lysosomal degradation

Interferon-λ Attenuates Rabies Virus Infection by Inducing Interferon-Stimulated Genes and Alleviating Neurological Inflammation.

Composition of the murine gut microbiome impacts humoral immunity induced by rabies vaccines.

Murine Ifit3 restricts the replication of Rabies virus both in vitro and in vivo.

Codon optimization of G protein enhances rabies virus-induced humoral immunity.