2023 Vol.38(3)
Commensal microbiota is closely related to hepatitis B virus (HBV) infection. However, existing studies have not elucidated the effect of gut bacteria on HBV immune tolerant phase yet. In this issue, Bu et al. altered gut bacteria richness and diversity using broad spectrum antibiotic mixtures and generated HBV immune tolerance mouse model via recombinant adeno-associated virus-HBV transduction to investigate the effects of gut bacteria on HBV replication. Their study provides new thoughts for elucidating the correlation between gut bacteria dysbiosis caused by antibiotic abuse and clinical chronic HBV infection. The cover image describes that antibiotic-induced gut bacteria depletion fails to affect HBV replication in HBV immune tolerance mouse model but contributes to increase HBV replication after immune activation (kindly designed and provided by Yuchen Xia). See page 335–343 for details.
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Antibiotic-induced gut bacteria depletion has no effect on HBV replication in HBV immune tolerance mouse model
2023, 38(3): 335 doi: 10.1016/j.virs.2023.04.010
Received: 29 January 2023
Accepted: 26 April 2023
Commensal microbiota is closely related to Hepatitis B virus (HBV) infection. Gut bacteria maturation accelerates HBV immune clearance in hydrodynamic injection (HDI) HBV mouse model. However, the effect of gut bacteria on HBV replication in recombinant adeno-associated virus (AAV)-HBV mouse model with immune tolerance remains obscure. We aim to investigate its role on HBV replication in AAV-HBV mouse model. C57BL/6 mice were administrated with broad-spectrum antibiotic mixtures (ABX) to deplete gut bacteria and intravenously injected with AAV-HBV to establish persistent HBV replication. Gut microbiota community was analyzed by fecal qPCR assay and 16S ribosomal RNA (rRNA) gene sequencing. HBV replication markers in blood and liver were determined by ELISA, qPCR assay and Western blot at indicated time points. Immune response in AAV-HBV mouse model was activated through HDI of HBV plasmid or poly(I:C) and then detected by quantifying the percentage of IFN-γ+/CD8+ T cells in the spleen via flow cytometry as well as the splenic IFN-γ mRNA level via qPCR assay. We found that antibiotic exposure remarkably decreased gut bacteria abundance and diversity. Antibiotic treatment failed to alter the levels of serological HBV antigens, intrahepatic HBV RNA transcripts and HBc protein in AAV-HBV mouse model, but contributed to HBsAg increase after breaking of immune tolerance. Overall, our data uncovered that antibiotic-induced gut bacteria depletion has no effect on HBV replication in immune tolerant AAV-HBV mouse model, providing new thoughts for elucidating the correlation between gut bacteria dysbiosis by antibiotic abuse and clinical chronic HBV infection.
A novel bat coronavirus with a polybasic furin-like cleavage site
2023, 38(3): 344 doi: 10.1016/j.virs.2023.04.009
Received: 07 January 2023
Accepted: 24 April 2023
The current pandemic of COVID-19 caused by a novel coronavirus, severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), threatens human health around the world. Of particular concern is that bats are recognized as one of the most potential natural hosts of SARS-CoV-2; however, coronavirus ecology in bats is still nascent. Here, we performed a degenerate primer screening and next-generation sequencing analysis of 112 bats, collected from Hainan Province, China. Three coronaviruses, namely bat betacoronavirus (Bat CoV) CD35, Bat CoV CD36 and bat alphacoronavirus CD30 were identified. Bat CoV CD35 genome had 99.5% identity with Bat CoV CD36, both sharing the highest nucleotide identity with Bat Hp-betacoronavirus Zhejiang2013 (71.4%), followed by SARS-CoV-2 (54.0%). Phylogenetic analysis indicated that Bat CoV CD35 formed a distinct clade, and together with Bat Hp-betacoronavirus Zhejiang2013, was basal to the lineage of SARS-CoV-1 and SARS-CoV-2. Notably, Bat CoV CD35 harbored a canonical furin-like S1/S2 cleavage site that resembles the corresponding sites of SARS-CoV-2. The furin cleavage sites between CD35 and CD36 are identical. In addition, the receptor-binding domain of Bat CoV CD35 showed a highly similar structure to that of SARS-CoV-1 and SARS-CoV-2, especially in one binding loop. In conclusion, this study deepens our understanding of the diversity of coronaviruses and provides clues about the natural origin of the furin cleavage site of SARS-CoV-2.
Epidemiology and genetic diversity of norovirus GII genogroups among children in Hubei, China, 2017–2019
2023, 38(3): 351 doi: 10.1016/j.virs.2023.04.002
Received: 08 October 2022
Accepted: 03 April 2023
Norovirus (NoV) is an important cause of viral acute gastroenteritis (AGE). To gain insights into the epidemiological characteristics and genetic diversity of NoV among children in Hubei, 1216 stool samples from children (≤ 5 years) obtained under AGE surveillance from January 2017 to December 2019 were analyzed. The results showed that NoV was responsible for 14.64% of AGE cases, with the highest detection rate in children aged 7–12 months (19.76%). Statistically significant differences were found between male and female infection rates (χ2 = 8.108, P = 0.004). Genetic analysis of RdRp and VP1 sequences showed that NoV GII genotypes were GII.4 Sydney [P31] (34.35%), GII.3 [P12] (25.95%), GII.2 [P16] (22.90%), GII.4 Sydney [P16] (12.98%), GII.17 [P17] (2.29%), GII.6 [P7] and GII.3 [P16] (each at 0.76%). GII.17 [P17] variants were divided into the Kawasaki323-like lineage and the Kawasaki308-like lineage. A unique recombination event was detected between strains of GII.4 Sydney 2012 and GII.4 Sydney 2016. Significantly, all GII.P16 sequences associated with GII.4/GII.2 obtained in Hubei were correlated with novel GII.2 [P16] variants that re-emerged in Germany in 2016. Antigenic site analysis of complete VP1 sequences from all GII.4 variants from Hubei identified notable variable residues of antibody epitopes. Genotyping under continuous AGE surveillance and observation of the antigenic sites of VP1 are important monitoring strategies for emerging NoV strains.
Prevalence, variation, and transmission patterns of human respiratory syncytial virus from pediatric patients in Hubei, China during 2020–2021
2023, 38(3): 363 doi: 10.1016/j.virs.2023.05.001
Received: 17 February 2022
Accepted: 26 April 2023
Human respiratory syncytial virus (RSV) is a severe threat to children and a main cause of acute lower respiratory tract infections. Nevertheless, the intra-host evolution and inter-regional diffusion of RSV are little known. In this study, we performed a systematic surveillance in hospitalized children in Hubei during 2020–2021, in which 106 RSV-positive samples were detected both clinically and by metagenomic next generation sequencing (mNGS). RSV-A and RSV-B groups co-circulated during surveillance with RSV-B being predominant. About 46 high-quality genomes were used for further analyses. A total of 163 intra-host nucleotide variation (iSNV) sites distributed in 34 samples were detected, and glycoprotein (G) gene was the most enriched gene for iSNVs, with non-synonymous substitutions more than synonymous substitutions. Evolutionary dynamic analysis showed that the evolutionary rates of G and NS2 genes were higher, and the population size of RSV groups changed over time. We also found evidences of inter-regional diffusion from Europe and Oceania to Hubei for RSV-A and RSV-B, respectively. This study highlighted the intra-host and inter-host evolution of RSV, and provided some evidences for understanding the evolution of RSV.
Construction and characterization of a synthesized herpes simplex virus H129-Syn-G2
2023, 38(3): 373 doi: 10.1016/j.virs.2023.03.005
Received: 27 December 2022
Accepted: 15 March 2023
Herpes simplex virus type 1 (HSV-1) causes lifelong infections worldwide, and currently there is no efficient cure or vaccine. HSV-1-derived tools, such as neuronal circuit tracers and oncolytic viruses, have been used extensively; however, further genetic engineering of HSV-1 is hindered by its complex genome structure. In the present study, we designed and constructed a synthetic platform for HSV-1 based on H129-G4. The complete genome was constructed from 10 fragments through 3 rounds of synthesis using transformation-associated recombination (TAR) in yeast, and was named H129-Syn-G2. The H129-Syn-G2 genome contained two copies of the gfp gene and was transfected into cells to rescue the virus. According to growth curve assay and electron microscopy results, the synthetic viruses exhibited more optimized growth properties and similar morphogenesis compared to the parental virus. This synthetic platform will facilitate further manipulation of the HSV-1 genome for the development of neuronal circuit tracers, oncolytic viruses, and vaccines.
Deletion of the first glycosylation site promotes Lassa virus glycoprotein-mediated membrane fusion
2023, 38(3): 380 doi: 10.1016/j.virs.2023.04.003
Received: 11 January 2023
Accepted: 10 April 2023
The Lassa virus (LASV) is endemic in West Africa and causes severe hemorrhagic Lassa fever in humans. The glycoprotein complex (GPC) of LASV is highly glycosylation-modified, with 11 N-glycosylation sites. All 11 N-linked glycan chains play critical roles in GPC cleavage, folding, receptor binding, membrane fusion, and immune evasion. In this study, we focused on the first glycosylation site because its deletion mutant (N79Q) results in an unexpected enhanced membrane fusion, whereas it exerts little effect on GPC expression, cleavage, and receptor binding. Meanwhile, the pseudotype virus bearing GPCN79Q was more sensitive to the neutralizing antibody 37.7H and was attenuated in virulence. Exploring the biological functions of the key glycosylation site on LASV GPC will help elucidate the mechanism of LASV infection and provide strategies for the development of attenuated vaccines against LASV infection.
3Cpro of FMDV inhibits type II interferon-stimulated JAK-STAT signaling pathway by blocking STAT1 nuclear translocation
2023, 38(3): 387 doi: 10.1016/j.virs.2023.03.003
Received: 26 October 2022
Accepted: 08 March 2023
Foot-and-mouth disease virus (FMDV) has developed various strategies to antagonize the host innate immunity. FMDV Lpro and 3Cpro interfere with type I IFNs through different mechanisms. The structural protein VP3 of FMDV degrades Janus kinase 1 to suppress IFN-γ signaling transduction. Whether non-structural proteins of FMDV are involved in restraining type II IFN signaling pathways is unknown. In this study, it was shown that FMDV replication was resistant to IFN-γ treatment after the infection was established and FMDV inhibited type II IFN induced expression of IFN-γ-stimulated genes (ISGs). We also showed for the first time that FMDV non-structural protein 3C antagonized IFN-γ-stimulated JAK-STAT signaling pathway by blocking STAT1 nuclear translocation. 3Cpro expression significantly reduced the ISGs transcript levels and palindromic gamma-activated sequences (GAS) promoter activity, without affecting the protein level, tyrosine phosphorylation, and homodimerization of STAT1. Finally, we provided evidence that 3C protease activity played an essential role in degrading KPNA1 and thus inhibited ISGs mRNA and GAS promoter activities. Our results reveal a novel mechanism by which an FMDV non-structural protein antagonizes host type II IFN signaling.
Transcriptome analysis of CD4+ T cells from HIV-infected individuals receiving ART with LLV revealed novel transcription factors regulating HIV-1 promoter activity
2023, 38(3): 398 doi: 10.1016/j.virs.2023.03.001
Received: 29 September 2022
Accepted: 06 March 2023
Some HIV-infected individuals receiving ART develop low-level viremia (LLV), with a plasma viral load of 50–1000 copies/mL. Persistent low-level viremia is associated with subsequent virologic failure. The peripheral blood CD4+ T cell pool is a source of LLV. However, the intrinsic characteristics of CD4+ T cells in LLV which may contribute to low-level viremia are largely unknown. We analyzed the transcriptome profiling of peripheral blood CD4+ T cells from healthy controls (HC) and HIV-infected patients receiving ART with either virologic suppression (VS) or LLV. To identify pathways potentially responding to increasing viral loads from HC to VS and to LLV, KEGG pathways of differentially expressed genes (DEGs) were acquired by comparing VS with HC (VS-HC group) and LLV with VS (LLV-VS group), and overlapped pathways were analyzed. Characterization of DEGs in key overlapping pathways showed that CD4+ T cells in LLV expressed higher levels of Th1 signature transcription factors (TBX21), toll-like receptors (TLR-4, -6, -7 and -8), anti-HIV entry chemokines (CCL3 and CCL4), and anti-IL-1β factors (ILRN and IL1R2) compared to VS. Our results also indicated activation of the NF-κB and TNF signaling pathways that could promote HIV-1 transcription. Finally, we evaluated the effects of 4 and 17 transcription factors that were upregulated in the VS-HC and LLV-VS groups, respectively, on HIV-1 promoter activity. Functional studies revealed that CXXC5 significantly increased, while SOX5 markedly suppressed HIV-1 transcription. In summary, we found that CD4+ T cells in LLV displayed a distinct mRNA profiling compared to that in VS, which promoted HIV-1 replication and reactivation of viral latency and may eventually contribute to virologic failure in patients with persistent LLV. CXXC5 and SOX5 may serve as targets for the development of latency-reversing agents.
FOLR1-induced folate deficiency reduces viral replication via modulating APOBEC3 family expression
2023, 38(3): 409 doi: 10.1016/j.virs.2023.04.001
Received: 17 October 2022
Accepted: 30 March 2023
Folate receptor alpha (FOLR1) is vital for cells ingesting folate (FA). FA plays an indispensable role in cell proliferation and survival. However, it is not clear whether the axis of FOLR1/FA has a similar function in viral replication. In this study, we used vesicular stomatitis virus (VSV) to investigate the relationship between FOLR1-mediated FA deficiency and viral replication, as well as the underlying mechanisms. We discovered that FOLR1 upregulation led to the deficiency of FA in HeLa cells and mice. Meanwhile, VSV replication was notably suppressed by FOLR1 overexpression, and this antiviral activity was related to FA deficiency. Mechanistically, FA deficiency mainly upregulated apolipoprotein B mRNA editing enzyme catalytic subunit 3B (APOBEC3B) expression, which suppressed VSV replication in vitro and in vivo. In addition, methotrexate (MTX), an FA metabolism inhibitor, effectively inhibited VSV replication by enhancing the expression of APOBEC3B in vitro and in vivo. Overall, our present study provided a new perspective for the role of FA metabolism in viral infections and highlights the potential of MTX as a broad-spectrum antiviral agent against RNA viruses.
TRAF7 negatively regulates the RLR signaling pathway by facilitating the K48-linked ubiquitination of TBK1
2023, 38(3): 419 doi: 10.1016/j.virs.2023.04.005
Received: 10 December 2022
Accepted: 17 April 2023
TANK-binding kinase 1 (TBK1) is a nodal protein involved in multiple signal transduction pathways. In RNA virus-mediated innate immunity, TBK1 is recruited to the prion-like platform formed by MAVS and subsequently activates the transcription factors IRF3/7 and NF-κB to produce type I interferon (IFN) and proinflammatory cytokines for the signaling cascade. In this study, TRAF7 was identified as a negative regulator of innate immune signaling. TRAF7 interacts with TBK1 and promotes K48-linked polyubiquitination and degradation of TBK1 through its RING domain, impairing the activation of IRF3 and the production of IFN-β. In addition, we found that the conserved cysteine residues at position 131 of TRAF7 are necessary for its function toward TBK1. Knockout of TRAF7 could facilitate the activation of IRF3 and increase the transcript levels of downstream antiviral genes. These data suggest that TRAF7 negatively regulates innate antiviral immunity by promoting the K48-linked ubiquitination of TBK1.
Vagal-mAChR4 signaling promotes Friend virus complex (FV)-induced acute erythroleukemia
2023, 38(3): 429 doi: 10.1016/j.virs.2023.05.005
Received: 04 July 2022
Accepted: 08 May 2023
Erythroleukemia belongs to acute myeloid leukemia (AML) type 6 (M6), and treatment remains difficult due to the poor prognosis of the disease. Friend virus (FV) is a complex of two viruses: Friend murine leukemia virus (F-MuLV) strain along with a defective spleen focus-forming virus (SFFV), which can induce acute erythroleukemia in mice. We have previously reported that activation of vagal α7 nicotinic acetylcholine receptor (nAChR) signaling promotes HIV-1 transcription. Whether vagal muscarinic signaling mediates FV-induced erythroleukemia and the underlying mechanisms remain unclear. In this study, sham and vagotomized mice were intraperitoneally injected with FV. FV infection caused anemia in sham mice, and vagotomy reversed this change. FV infection increased erythroblasts ProE, EryA, and EryB cells in the spleen, and these changes were blocked by vagotomy. In bone marrow, FV infection reduced EryC cells in sham mice, an effect that was counteracted by vagotomy. FV infection increased choline acetyltransferase (ChAT) expression in splenic CD4+ and CD8+ T cells, and this change was reversed by vagotomy. Furthermore, the increase of EryA and EryB cells in spleen of FV-infected wild-type mice was reversed after deletion of ChAT in CD4+ T cells. In bone marrow, FV infection reduced EryB and EryC cells in sham mice, whereas lack of ChAT in CD4+ T cells did not affect this change. Activation of muscarinic acetylcholine receptor 4 (mAChR4) by clozapine N-oxide (CNO) significantly increased EryB in the spleen but decreased the EryC cell population in the bone marrow of FV-infected mice. Thus, vagal-mAChR4 signaling in the spleen and bone marrow synergistically promotes the pathogenesis of acute erythroleukemia. We uncover an unrecognized mechanism of neuromodulation in erythroleukemia.
Temperature-regulated type II grass carp reovirus establishes latent infection in Ctenopharyngodon idella brain
2023, 38(3): 440 doi: 10.1016/j.virs.2023.04.006
Received: 28 October 2022
Accepted: 26 April 2023
Grass carp reovirus (GCRV) causes extensive infection and death in grass carp and black carp fingerlings, with a highly seasonal prevalence. Previous studies suggested that GCRV can become latent after primary infection. In this study, we investigated type II GCRV (GCRV-II) latency in asymptomatic grass carp with GCRV infection or exposure history. We found that during latent infection, GCRV-II was detectable only in the brain of grass carp, unlike the multi-tissue distribution observed in natural infection. GCRV-II only caused damage to the brain during latent infection, while in natural infection, brain, heart, and eye tissues had relatively higher viral loads. We also discovered viral inclusion bodies in infected fish brains. Additionally, GCRV-II distribution in grass carp was notably affected by ambient temperature, with the virus targeting the brain only during low temperatures and multi-tissue distribution during high temperatures. This study provides insights into the mechanisms of GCRV-II latent infection and reactivation and contributes to the prevention and control of GCRV pandemics.
Spastin is required for human immunodeficiency virus-1 efficient replication through cooperation with the endosomal sorting complex required for transport (ESCRT) protein
2023, 38(3): 448 doi: 10.1016/j.virs.2023.05.006
Received: 02 August 2022
Accepted: 08 May 2023
Human immunodeficiency virus-1 (HIV-1) encodes simply 15 proteins and thus depends on multiple host cellular factors for virus reproduction. Spastin, a microtubule severing protein, is an identified HIV-1 dependency factor, but the mechanism regulating HIV-1 is unclear. Here, the study showed that knockdown of spastin inhibited the production of the intracellular HIV-1 Gag protein and new virions through enhancing Gag lysosomal degradation. Further investigation showed that increased sodium tolerance 1 (IST1), the subunit of endosomal sorting complex required for transport (ESCRT), could interact with the MIT domain of spastin to regulate the intracellular Gag production. In summary, spastin is required for HIV-1 replication, while spastin-IST1 interaction facilitates virus production by regulating HIV-1 Gag intracellular trafficking and degradation. Spastin may serve as new target for HIV-1 prophylactic and therapy.
ASFV transcription reporter screening system identifies ailanthone as a broad antiviral compound
2023, 38(3): 459 doi: 10.1016/j.virs.2023.03.004
Received: 25 October 2022
Accepted: 15 March 2023
African swine fever (ASF) is an acute, highly contagious and deadly viral disease in swine that jeopardizes the worldwide pig industry. Unfortunately, there are no authoritative vaccine and antiviral drug available for ASF control. African swine fever virus (ASFV) is the etiological agent of ASF. Among the ASFV proteins, p72 is the most abundant component in the virions and thus a potential target for anti-ASFV drug design. Here, we constructed a luciferase reporter system driven by the promoter of p72, which is transcribed by the co-transfected ASFV RNA polymerase complex. Using this system, we screened over 3200 natural product compounds and obtained three potent candidates against ASFV. We further evaluated the anti-ASFV effects and proved that among the three candidates, ailanthone (AIL) inhibits the replication of ASFV at the nanomolar concentration (IC50 = 15 nmol/L). Our in vitro experiments indicated that the antiviral effect of AIL is associated with its inhibition of the HSP90-p23 cochaperone. Finally, we showed the antiviral activity of AIL on Zika virus and hepatitis B virus (HBV), which supports that AIL is a potential broad-spectrum antiviral agent.
Oridonin inhibits SARS-CoV-2 replication by targeting viral proteinase and polymerase
2023, 38(3): 470 doi: 10.1016/j.virs.2023.04.008
Received: 18 December 2022
Accepted: 27 April 2023
COVID-19 has become a global public health crisis since its outbreak in China in December 2019. Currently there are few clinically effective drugs to combat SARS-CoV-2 infection. The main protein (Mpro), papain-like protease (PLpro) and RNA-dependent RNA polymerase (RdRp) of SARS-CoV-2 are involved in the viral replication, and might be prospective targets for anti-coronavirus drug development. Here, we investigated the antiviral activity of oridonin, a natural small-molecule compound, against SARS-CoV-2 infection in vitro. The time-of-addition analysis showed that oridonin efficiently inhibited SARS-CoV-2 infection by interfering with the genome replication at the post-entry stage. Mechanistically, the inhibition of viral replication by oridonin depends on the oxidation activity of α, β-unsaturated carbonyl. Further experiments showed that oridonin not only effectively inhibited SARS-CoV-2 Mpro activity, but also had some inhibitory effects on PLpro-mediated deubiquitinating and viral polymerase-catalyzed RNA elongation activities at high concentrations. In particular, oridonin could inhibit the bat SARS-like CoV and the newly emerged SARS-CoV-2 omicron variants (BA.1 and BA.2), which highlights its potential as a pan-coronavirus antiviral agent. Overall, our data provide strong evidence that oridonin is an efficient antiviral agent against SARS-CoV-2 infection.
Due to our negligence, the original version of this article, published online on 12 January 2022, contained a mistake in Fig. 4A. The lane of β-actin in Western blotting was misused. The correct Fig. 4 is given below. We apologize for our oversight when preparing the figure and state that this does not change the scientific conclusions of the article in any way.
Due to our negligence, the original version of this article, published online on April 08, 2021, contained a mistake in Fig. 1E. The lane of β-actin in Western blotting was misused. The correct Fig. 1 is given below. We apologize for our oversight when preparing the figure and state that this does not change the scientific conclusions of the article in any way.
