2020 Vol.35(4)
Flaviviruses, including the dengue, Zika, West Nile, tick-borne encephalitis and yellow fever viruses (DENV, ZIKV, WNV, TBEV and YFV, respectively), are a genus of mostly arthropod-borne RNA viruses that cause a range of pathologies in humans. The success of productive infection of human cells by flaviviruses depends in part on the antiviral activity of a heterogeneous group of cellular antiviral proteins called restriction factors. Restriction factors are the effector proteins of the cell-autonomous innate response against viruses, an immune pathway that also includes virus sensors as well as intracellular and extracellular signal mediators. In this issue, Prof. Lionel Berthoux reviewed the recent progress toward the identification and characterization of flavivirus restriction factors and summarized several important unanswered questions in the restriction of flaviviruses. The cover image shows early flavivirus infection stages affected by IFN-I-inducible restriction factors. Please see page 363-377 for details.
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The Restrictome of Flaviviruses
2020, 35(4): 363 doi: 10.1007/s12250-020-00208-3
Received: 30 November 2019
Accepted: 03 February 2020
Published: 09 March 2020
Flaviviruses are a genus of mostly arthropod-borne RNA viruses that cause a range of pathologies in humans. Basic knowledge on flaviviruses is rapidly expanding, partly due to their status as frequent emerging or re-emerging pathogens. Flaviviruses include the dengue, Zika, West Nile, tick-borne encephalitis and yellow fever viruses (DENV, ZIKV, WNV, TBEV and YFV, respectively). As is the case with other families of viruses, the success of productive infection of human cells by flaviviruses depends in part on the antiviral activity of a heterogeneous group of cellular antiviral proteins called restriction factors. Restriction factors are the effector proteins of the cell-autonomous innate response against viruses, an immune pathway that also includes virus sensors as well as intracellular and extracellular signal mediators such as type Ⅰ interferons (IFN-I). In this review, I summarize recent progress toward the identification and characterization of flavivirus restriction factors. In particular, I focus on IFI6, Schlafen 11, FMRP, OAS-RNase L, RyDEN, members of the TRIM family of proteins (TRIM5α, TRIM19, TRIM56, TRIM69 and TRIM79α) and a new mechanism of action proposed for viperin. Recent and future studies on this topic will lead to a more complete picture of the flavivirus restrictome, defined as the ensemble of cellular factors with demonstrated anti-flaviviral activity.
New Gene Variants Associated with the Risk of Chronic HBV Infection
2020, 35(4): 378 doi: 10.1007/s12250-020-00200-x
Received: 05 March 2019
Accepted: 16 January 2020
Published: 15 April 2020
Some patients with chronic hepatitis B virus (HBV) infection failed to clear HBV, even persistently continue to produce antibodies to HBV. Here we performed a two stage Genome Wide Association Study (GWAS) in a cohort of Chinese patients designed to discover single nucleotide variants that associate with HBV infection and clearance of HBV. The first stage involved genome wide exome sequencing of 101 cases (HBsAg plus anti-HBs positive) compared with 102 control patients (anti-HBs positive, HBsAg negative). Over 80% of individual sequences displayed 20× sequence coverage. Adapters, uncertain bases > 10% or low-quality base calls (> 50%) were filtered and compared to the human reference genome hg19. In the second stage, 579 chronic HBV infected cases and 439 HBV clearance controls were sequenced with selected genes from the first stage. Although there were no significant associated gene variants in the first stage, two significant gene associations were discovered when the two stages were assessed in a combined analysis. One association showed rs506121-'T' allele [within the dedicator of cytokinesis 8 (DOCK8) gene] was higher in chronic HBV infection group than that in clearance group (P= 0.002, OR = 0.77, 95%CI [0.65, 0.91]). The second association involved rs2071676 –A allele within the Carbonic anhydrase (CA9) gene that was significantly elevated in chronic HBV infection group compared to the clearance group (P = 0.0003, OR = 1.35, 95%CI [1.15, 1.58]). Upon replication these gene associations would suggest the influence of DOCK8 and CA9 as potential risk genetic factors in the persistence of HBV infection.
Conservative Evolution of Hepatitis B Virus Precore and Core Gene During Immune Tolerant Phase in Intrafamilial Transmission
2020, 35(4): 388 doi: 10.1007/s12250-020-00194-6
Received: 02 August 2019
Accepted: 06 December 2019
Published: 02 March 2020
Hepatitis B virus (HBV) is characterized with high mutations, which is attributed to the lack of proof-reading of the viral reverse transcriptase and host immune pressure. In this study, 31 HBV chronic carriers from 14 families were enrolled to investigate the evolution of the same original HBV sources in different hosts. Sequences of pre-C and C (pre-C/C) genes were analyzed in eight pairs of HBV-infected mothers with longitudinal sera (at an interval of 6.0–7.2 years) and their children (5.5–6.7 years old), and in 15 adults (21–78 years old) from six families with known intrafamilial HBV infection. The pre-C/C sequences had almost no change in eight mothers during 6.0–7.2 years and their children who were in immune tolerant phase. The pre-C/C sequences from the 15 adults of six families, mostly in the immune-clearance phase or the low replicative phase, showed various diversified mutations between individuals from each family. Compared to a reference stain (GQ205441) isolated nearby, the pre-C/C in individuals in immune tolerant phase showed 98.56%–99.52% homology at nucleotide level and 99.5%–100% homology at amino acid level. In contrast, multiple mutations were developed in the immune-clearance phase or the low replicative phase, affecting immune epitopes in core gene and G1896 in pre-C gene. The results indicate that the evolution of new HBV variants is not mainly resulted from the spontaneous error rate of viral reverse transcription, but from the host immune pressure.
Comparison of Viromes in Ticks from Different Domestic Animals in China
2020, 35(4): 398 doi: 10.1007/s12250-020-00197-3
Received: 08 May 2019
Accepted: 11 December 2019
Published: 10 March 2020
Ticks are involved in the transmission of various arboviruses and some tick-borne viruses pose significant threats to the health of humans or livestock. This study aimed to investigate the geographical distribution of tick species and tick-associated viruses in central and eastern China. Total 573 ticks from domestic animals including dogs, sheep and cattle were collected in 2017. Two genera of ticks were identified including Rhipicephalus and Haemaphysalis. Sequencing was performed on Miseq Illumina platform to characterize the tick viromes from the four different sampling locations. Following trimming, 13, 640 reads were obtained and annotated to 19 virus families. From these sequences, above 37.74% of the viral reads were related to the RNA viruses. Virome comparison study revealed that the tick viral diversity was considerably different in the two identified tick genera. The viral diversity of R. microplus was significantly different from that of other Rhipicephalus species. On the other hand, substantial overlap in viral species was observed between the same genera. In addition, we found no evidence that the natural host played a major role in shaping virus diversity based on the comparison of their viromes. Rather, the geographic location seems to significantly influence the viral families. Phylogenetic study indicated that the novel negative-sense RNA viruses identified in this study was closely related to Bole tick virus 1 and 3 viruses. In conclusion, the present study provides a baseline for comparing viruses detected in ticks, according to species, natural hosts, and geographic locations.
Visualizing the Transport of Porcine Reproductive and Respiratory Syndrome Virus in Live Cells by Quantum Dots-Based Single Virus Tracking
2020, 35(4): 407 doi: 10.1007/s12250-019-00187-0
Received: 20 August 2019
Accepted: 09 October 2019
Published: 23 December 2019
Quantum dots (QDs)-based single particle analysis technique enables real-time tracking of the viral infection in live cells with great sensitivity over a long period of time. The porcine reproductive and respiratory syndrome virus (PRRSV) is a small virus with the virion size of 40–60 nm which causes great economic losses to the swine industry worldwide. A clear understanding of the viral infection mechanism is essential for the development of effective antiviral strategies. In this study, we labeled the PRRSV with QDs using the streptavidin–biotin labeling system and monitored the viral infection process in live cells. Our results indicated that the labeling method had negligible effect on viral infectivity. We also observed that prior to the entry, PRRSV vibrated on the plasma membrane, and entered the cells via endosome mediated cell entry pathway. Viruses moved in a slow–fast–slow oscillatory movement pattern and finally accumulated in a perinuclear region of the cell. Our results also showed that once inside the cell, PRRSV moved along the microtubule, microfilament and vimentin cytoskeletal elements. During the transport process, virus particles also made contacts with non-muscle myosin heavy chain Ⅱ-A (NMHC Ⅱ-A), visualized as small spheres in cytoplasm. This study can facilitate the application of QDs in virus infection imaging, especially the smaller-sized viruses and provide some novel and important insights into PRRSV infection mechanism.
The Establishment of an In Vivo HIV-1 Infection Model in Humanized B-NSG Mice
2020, 35(4): 417 doi: 10.1007/s12250-019-00181-6
Received: 12 July 2019
Accepted: 02 December 2019
Published: 21 December 2019
Suitable animal models for human immunodeficiency virus type 1 (HIV-1) infection are important for elucidating viral pathogenesis and evaluating antiviral strategies in vivo. The B-NSG (NOD-PrkdcscidIl2rgtm1/Bcge) mice that have severe immune defect phenotype are examined for the suitability of such a model in this study. Human peripheral blood mononuclear cells (PBMCs) were engrafted into B-NSG mice via mouse tail vein injection, and the repopulated human T-lymphocytes were observed at as early as 3-weeks post-transplantation in mouse peripheral blood and several tissues. The humanized mice could be infected by HIV-1, and the infection recapitulated features of T-lymphocyte dynamic observed in HIV-1 infected humans, meanwhile the administration of combination antiretroviral therapy (cART) suppressed viral replication and restored T lymphocyte abnormalities. The establishment of HIV-1 infected humanized B-NSG mice not only provides a model to study virus and T cell interplays, but also can be a useful tool to evaluate antiviral strategies.
The Establishment of Infectious Clone and Single Round Infectious Particles for Coxsackievirus A10
2020, 35(4): 426 doi: 10.1007/s12250-020-00198-2
Received: 27 September 2019
Accepted: 24 December 2019
Published: 06 March 2020
Coxsackievirus A10 (CVA10) is one of the major etiological agents of hand, foot, and mouth disease. There are no vaccine and antiviral drugs for controlling CVA10 infection. Reverse genetic tools for CVA10 will benefit its mechanistic study and development of vaccines and antivirals. Here, two infectious clones for the prototype and a Myc-tagged CVA10 were constructed. Viable CVA10 viruses were harvested by transfecting the viral mRNA into human rhabdomyosarcoma (RD) cells. Rescued CVA10 was further confirmed by next generation sequencing and characterized experimentally. We also constructed the vectors for CVA10 subgenomic replicon with luciferase reporter and viral capsid with EGFP reporter, respectively. Co-transfection of the viral replicon RNA and capsid expresser in human embryonic kidney 293T (HEK293T) cells led to the production of single round infectious particles (SRIPs). Based on CVA10 replicon RNA, SRIPs with either the enterovirus A71 (EVA71) capsid or the CVA10 capsid were generated. Infection by EVA71 SRIPs required SCARB2, while CVA10 SRIPs did not. Finally, we showed great improvement of the replicon activity and SRIPs production by insertion of a cis-active hammerhead ribozyme (HHRib) before the 5′-untranslated region (UTR). In summary, reverse genetic tools for prototype strain of CVA10, including both the infectious clone and the SRIPs system, were successfully established. These tools will facilitate the basic and translational study of CVA10.
Hepatitis C Virus NS2 Protein Suppresses RNA Interference in Cells
2020, 35(4): 436 doi: 10.1007/s12250-019-00182-5
Received: 12 October 2019
Accepted: 01 November 2019
Published: 27 November 2019
RNAi interference (RNAi) is an evolutionarily conserved post-transcriptional gene silencing mechanism and has been well recognized as an important antiviral immunity in eukaryotes. Numerous viruses have been shown to encode viral suppressors of RNAi (VSRs) to antagonize antiviral RNAi. Hepatitis C virus (HCV) is a medically important human pathogen that causes acute and chronic hepatitis. In this study, we screened all the nonstructural proteins of HCV and found that HCV NS2 could suppress RNAi induced either by small hairpin RNAs (shRNAs) or small interfering RNAs (siRNAs) in mammalian cells. Moreover, we demonstrated that NS2 could suppress RNAi via its direct interaction with double-stranded RNAs (dsRNAs) and siRNAs, and further identified that the cysteine 184 of NS2 is required for the RNAi suppression activity through a serial of point mutation analyses. Together, our findings uncovered that HCV NS2 can act as a VSR in vitro, thereby providing novel insights into the life cycle and virus-host interactions of HCV.
Structure of the HRV-C 3C-Rupintrivir Complex Provides New Insights for Inhibitor Design
2020, 35(4): 445 doi: 10.1007/s12250-020-00196-4
Received: 21 August 2019
Accepted: 25 December 2019
Published: 26 February 2020
Human rhinoviruses (HRVs) are the predominant infectious agents for the common cold worldwide. The HRV-C species cause severe illnesses in children and are closely related to acute exacerbations of asthma. 3C protease, a highly conserved enzyme, cleaves the viral polyprotein during replication and assists the virus in escaping the host immune system. These key roles make 3C protease an important drug target. A few structures of 3Cs complexed with an irreversible inhibitor rupintrivir have been determined. These structures shed light on the determinants of drug specificity. Here we describe the structures of HRV-C15 3C in free and inhibitor-bound forms. The volume-decreased S1' subsite and half-closed S2 subsite, which were thought to be unique features of enterovirus A 3C proteases, appear in the HRV-C 3C protease. Rupintrivir assumes an "intermediate" conformation in the complex, which might open up additional avenues for the design of potent antiviral inhibitors. Analysis of the features of the three-dimensional structures and the amino acid sequences of 3C proteases suggest new applications for existing drugs.
Chimeric Newcastle Disease Virus-like Particles Containing DC-Binding Peptide-Fused Haemagglutinin Protect Chickens from Virulent Newcastle Disease Virus and H9N2 Avian Influenza Virus Challenge
2020, 35(4): 455 doi: 10.1007/s12250-020-00199-1
Received: 03 February 2019
Accepted: 18 November 2019
Published: 09 April 2020
Newcastle disease virus (NDV) and H9N2 subtype Avian influenza virus (AIV) are two notorious avian respiratory pathogens that cause great losses in the poultry industry. Current inactivated commercial vaccines against NDV and AIV have the disadvantages of inadequate mucosal responses, while an attenuated live vaccine bears the risk of mutation. Dendritic cell (DC) targeting strategies are attractive for their potent mucosal and adaptive immune-stimulating ability against respiratory pathogens. In this study, DC-binding peptide (DCpep)-decorated chimeric virus-like particles (cVLPs), containing NDV haemagglutinin–neuraminidase (HN) and AIV haemagglutinin (HA), were developed as a DC-targeting mucosal vaccine candidate. DCpep-decorated cVLPs activated DCs in vitro, and induced potent immune stimulation in chickens, with enhanced secretory immunoglobulin A (sIgA) secretion and splenic T cell differentiation. 40 μg cVLPs can provide full protection against the challenge with homologous, heterologous NDV strains, and AIV H9N2. In addition, DCpep-decorated cVLPs could induce a better immune response when administered intranasally than intramuscularly, as indicated by robust sIgA secretion and a reduced virus shedding period. Taken together, this chimeric VLPs are a promising vaccine candidate to control NDV and AIV H9N2 and a useful platform bearing multivalent antigens.
A Mother-to-Child Transmission Study in Nigeria: The Impact of Maternal HIV Infection and HAART on Plasma Immunoglobulins, Cytokine Profiles and Infant Outcome
2020, 35(4): 468 doi: 10.1007/s12250-020-00202-9
Received: 09 October 2019
Accepted: 24 December 2019
Published: 10 March 2020
Prevention of mother-to-child transmission (PMTCT) of HIV with highly active antiretroviral therapy (HARRT) allows the HIV+ pregnant mothers to have vaginal delivery and breastfeed. Here we investigated the maternal plasma immunoglobulin, cytokine secretion and the outcome of the exposed infants among the HIV+ HAART treated pregnant women in Nigeria. In this study, different plasma immunoglobulins and cytokines were measured in the HIV+ HAART treated pregnant mothers. Pooled culture supernatants of B and T lymphocytes showed lower levels of IFN-c, IL-10 and IL-4. There were lower IFN-c and IL-10 secretions at 1st trimester; however, IL-10 continued to be lower throughout 2nd and 3rd trimesters. TNF-a secretion significantly decreased as pregnancy progressed to term. There were high plasma IgG and low IgM in the HIV+ HAART treated pregnant women. Plasma IgG was high during 1st and 3rd trimesters. After one year of follow up, all the exposed children were seronegative for HIV-1 and HIV-2. Vaginal delivery and breastfeeding among HIV+ HAART treated mothers have shown to be safe. The use of HAART by the infected mothers and the use of septrin and niverapin by the exposed infants prevented mother to-child transmission of HIV.
The Limitation of Rapid Tests for DENV2 Infection in Host with Unique Immune Status: Low NS Antigenemia and Deficient Antibody Responses
2020, 35(4): 478 doi: 10.1007/s12250-019-00183-4
Received: 25 April 2019
Accepted: 18 November 2019
Published: 14 January 2020
Hepatitis C Virus Infection Caused by Infrequent Exposure in China Should Be of Concern
2020, 35(4): 481 doi: 10.1007/s12250-019-00191-4
Received: 26 April 2019
Accepted: 02 December 2019
Published: 21 February 2020
Sero-Epidemiological Survey of Crimean-Congo Hemorrhagic Fever among the Human Population of the Punjab Province in Pakistan
2020, 35(4): 486 doi: 10.1007/s12250-020-00195-5
Received: 18 April 2019
Accepted: 07 November 2019
Published: 26 February 2020
Prion Protein Expression is Correlated with Glioma Grades
2020, 35(4): 490 doi: 10.1007/s12250-020-00209-2
Received: 08 October 2019
Accepted: 17 February 2020
Published: 31 March 2020
Re-emergence of Highly Pathogenic Avian Influenza H5N1 in Nigeria, 2014–2016: Role of Social Network and Value Chain Forces in Interstate Transmission
2020, 35(4): 494 doi: 10.1007/s12250-020-00201-w
Received: 07 August 2019
Accepted: 17 January 2020
Published: 31 March 2020
