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Japanese Encephalitis Virus NS2B-3 Protein Complex Promotes Cell Apoptosis and Viral Particle Release by Down-Regulating the Expression of AXL
Shengda Xie, Zhenjie Liang, Xingmiao Yang, Junhui Pan, Du Yu, Tongtong Li, Ruibing Cao
doi: 10.1007/s12250-021-00442-3
Received: 14 May 2021 Accepted: 12 July 2021 Published: 06 September 2021
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Japanese encephalitis virus (JEV) is a flavivirus transmitted by mosquitoes that causes severe encephalitis in humans and animals. It has been suggested that AXL, a transmembrane protein, can promote the replication of various flaviviruses, such as dengue (DENV), Zika (ZIKV), and West Nile (WNV) viruses. However, the effect of AXL on JEV infection has not yet been determined. In the present study, we demonstrate that AXL is down-regulated after JEV infection in the late stage. JEV NS2B-3 protein specifically interacted with AXL, and promoted AXL degradation through the ubiquitin–proteasome pathway. AXL-degradation increased cell apoptosis by disrupting phosphatidylinositol 3-kinase (PI3K)/Akt signal transduction. In addition, the degradation of AXL promoted JEV release to supernatant, whereas the virus in the cell lysates decreased. The supplementation of AXL ligand Gas6 inhibited the JEV-mediated degradation of AXL. Altogether, we discover a new function of NS2B-3 during the process of JEV replication, and provide a new insight into the interactions between JEV and cell hosts.

Characterization of Episomal Replication of Bovine Papillomavirus Type 1 DNA in Long-Term Virion-Infected Saccharomyces Cerevisiae Culture
Quanmei Tu, Weixu Feng, Zhuo Chen, Qijia Li, Yu Zhao, Jun Chen, Pengfei Jiang, Xiangyang Xue, Lifang Zhang, Kong-Nan Zhao
doi: 10.1007/s12250-021-00439-y
Received: 22 February 2021 Accepted: 21 May 2021 Published: 30 August 2021
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We have previously reported that bovine papillomavirus type 1 (BPV-1) DNA can replicate its genome and produce infectious virus-like particles in short term virion-infected S. cerevisiae (budding yeast) cultures (Zhao and Frazer 2002, Journal of Virology, 76:3359–64 and 76:12265–73). Here, we report the episomal replications of BPV-1 DNA in long term virion-infected S. cerevisiae culture up to 108 days. Episomal replications of the BPV-1 DNA could be divided into three patterns at three stages, early active replication (day 3–16), middle weak replication (day 23–34/45) and late stable replication (day 45–82). Two-dimensional gel electrophoresis analysis and Southern blot hybridization have revealed further that multiple replication intermediates of BPV-1 DNA including linear form, stranded DNA, monomers and higher oligomers were detected in the virion-infected yeast cells over the time course. Higher oligomers shown as covalently closed circular DNAs (cccDNAs) are the most important replication intermediates that serve as the main nuclear transcription template for producing all viral RNAs in the viral life cycle. In this study, the cccDNAs were generated at the early active replication stage with the highest frequencies and then at late stable replication, but they appeared to be suppressed at the middle weak replication. Our data provided a novel insight that BPV-1 genomic DNA could replicate episomally for the long period and produce the key replication intermediates cccDNAs in S. cerevisiae system.

Three Novel Avastroviruses Identified in Dead Wild Crows
Chunge Zhang, Yongchun Yang, Tao Hu, Hong Zhou, Cheng Zhang, Jian Cao, Juan Li, Peihan Wang, Gary Wong, Xiaodu Wang, Houhui Song, George F. Gao, Weifeng Shi, Yuhai Bi
doi: 10.1007/s12250-021-00416-5
Received: 22 February 2021 Accepted: 27 April 2021 Published: 30 August 2021
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The Establishment and Spatiotemporal History of A Novel HIV-1 CRF01_AE Lineage in Shenyang City, Northeastern China in 2002–2019
Minghui An, Wei Song, Bin Zhao, Xue Dong, Lin Wang, Wen Tian, Xin Li, Lu Wang, Zhenxing Chu, Junjue Xu, Haibo Ding, Xiaoxu Han, Hong Shang
doi: 10.1007/s12250-021-00435-2
Received: 24 May 2021 Accepted: 28 June 2021 Published: 23 August 2021
[FullText HTML] [PDF 615KB] Springerlink
Generation and Characterization of a Nanobody Against SARS-CoV
Jiang-Fan Li, Lei He, Yong-Qiang Deng, Shu-Hui Qi, Yue-Hong Chen, Xiao-Lu Zhang, Shi-Xiong Hu, Rui-Wen Fan, Guang-Yu Zhao, Cheng-Feng Qin
doi: 10.1007/s12250-021-00436-1
Received: 28 March 2021 Accepted: 06 May 2021 Published: 17 August 2021
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The sudden emergence of severe acute respiratory syndrome coronavirus (SARS-CoV) has caused global panic in 2003, and the risk of SARS-CoV outbreak still exists. However, no specific antiviral drug or vaccine is available; thus, the development of therapeutic antibodies against SARS-CoV is needed. In this study, a nanobody phage-displayed library was constructed from peripheral blood mononuclear cells of alpacas immunized with the recombinant receptor-binding domain (RBD) of SARS-CoV. Four positive clones were selected after four rounds of bio-panning and subjected to recombinant expression in E. coli. Further biological identification demonstrated that one of the nanobodies, S14, showed high affinity to SARS-CoV RBD and potent neutralization activity at the picomole level against SARS-CoV pseudovirus. A competitive inhibition assay showed that S14 blocked the binding of SARS-CoV RBD to either soluble or cell-expressed angiotensinconverting enzyme 2 (ACE2). In summary, we developed a novel nanobody targeting SARS-CoV RBD, which might be useful for the development of therapeutics against SARS.

Structure and Function of N-Terminal Zinc Finger Domain of SARS-CoV-2 NSP2
Jun Ma, Yiyun Chen, Wei Wu, Zhongzhou Chen
doi: 10.1007/s12250-021-00431-6
Received: 14 June 2021 Accepted: 15 July 2021 Published: 16 August 2021
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SARS-CoV-2 has become a global pandemic threatening human health and safety. It is urgent to find effective therapeutic agents and targets with the continuous emergence of novel mutant strains. The knowledge of the molecular basis and pathogenesis of SARS-CoV-2 in host cells requires to be understood comprehensively. The unknown structure and function of nsp2 have hindered our understanding of its role in SARS-CoV-2 infection. Here, we report the crystal structure of the N-terminal of SARS-CoV-2 nsp2 to a high resolution of 1.96?. This novel structure contains three zinc fingers, belonging to the C2H2, C4, and C2HC types, respectively. Structure analysis suggests that nsp2 may be involved in binding nucleic acids and regulating intracellular signaling pathways. The binding to single or double-stranded nucleic acids was mainly through the large positively charged region on the surface of nsp2, and K111, K112, K113 were key residues. Our findings lay the foundation for a better understanding of the relationship between structure and function for nsp2. It is helpful to make full use of nsp2 as further research and development of antiviral targets and drug design.

A multi-center study on Molecular Epidemiology of Human Respiratory Syncytial Virus from Children with Acute Lower Respiratory Tract Infections in the Mainland of China between 2015 and 2019
Xiangpeng Chen, Yun Zhu, Wei Wang, Changchong Li, Shuhua An, Gen Lu, Rong Jin, Baoping Xu, Yunlian Zhou, Aihuan Chen, Lei Li, Meng Zhang, Zhengde Xie
doi: 10.1007/s12250-021-00430-7
Received: 29 January 2021 Accepted: 18 May 2021 Published: 16 August 2021
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Human respiratory syncytial virus (RSV) is a major pathogen of acute lower respiratory tract infection among young children. To investigate the prevalence and genetic characteristics of RSV in China, we performed a molecular epidemiological study during 2015-2019. A total of 964 RSV-positive specimens were identified from 5529 enrolled patients during a multi-center study. RSV subgroup A (RSV-A) was the predominant subgroup during this research period except in 2016. Totally, 535 sequences of the second hypervariable region (HVR-2) of the G gene were obtained. Combined with 182 Chinese sequences from GenBank, phylogenetic trees showed that 521 RSV-A sequences fell in genotypes ON1 (512), NA1 (6) and GA5 (3), respectively; while 196 RSV-B sequences fell in BA9 (193) and SAB4 (3). ON1 and BA9 were the only genotypes after December 2015. Genotypes ON1 and BA9 can be separated into 10 and 7 lineages, respectively. The HVR-2 of genotype ON1 had six amino acid changes with a frequency more than 10%, while two substitutions H258Q and H266L were co-occurrences. The HVR-2 of genotype BA9 had nine amino acid substitutions with a frequency more than 10%, while the sequences with T290I and T312I were all from 2018 to 2019. One N-glycosylation site at 237 was identified among ON1 sequences, while two N-glycosylation sites (296 and 310) were identified in the 60-nucleotide duplication region of BA9. To conclusion, ON1 and BA9 were the predominant genotypes in China during 2015-2019. For the genotypes ON1 and BA9, the G gene exhibited relatively high diversity and evolved continuously.

SARS-CoV-2 Genomic Sequencing Revealed N501Y and L452R Mutants of S/A Lineage in Tianjin Municipality, China
Xiaoyan Li, Xin Gao, Ming Zou, Zhichao Zhuang, Zhaolin Tan, Baolu Zheng, Aiping Yu, Xu Su
doi: 10.1007/s12250-021-00432-5
Received: 31 March 2021 Accepted: 26 May 2021 Published: 11 August 2021
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In Vitro Inhibition of Alphaviruses by Lycorine
Na Li, Zhen Wang, Rui Wang, Zhe-Rui Zhang, Ya-Nan Zhang, Cheng-Lin Deng, Bo Zhang, Lu-Qing Shang, Han-Qing Ye
doi: 10.1007/s12250-021-00438-z
Received: 08 May 2021 Accepted: 08 June 2021 Published: 10 August 2021
[FullText HTML] [PDF 787KB] Springerlink

Chikungunya virus (CHIKV) is a mosquito-borne alphavirus. As an emerging virus, CHIKV imposes a threat to public health. Currently, there are no vaccines or antivirals available for the prevention of CHIKV infection. Lycorine, an alkaloid from Amaryllidaceae plants, has antiviral activity against a number of viruses such as coronavirus, flavivirus and enterovirus. In this study, we found that lycorine could inhibit CHIKV in cell culture at a concentration of 10 μmol/L without apparent cytotoxicity. In addition, it exhibited broad-spectrum anti-alphavirus activity, including Sindbis virus (SINV), Semliki Forest virus (SFV), and Venezuelan equine encephalomyelitis virus (VEEV). The time of addition studies indicated that lycorine functions at an early post-entry stage of CHIKV life cycle. The results based on two different CHIKV replicons provided further evidence that lycorine exerts its antiviral activity mainly by inhibiting CHIKV translation. Overall, our study extends the antiviral spectrum of lycorine.

Inhibition of the Neddylation Pathway Suppresses Enterovirus Replication
Zhe Zhang, Haoran Guo, Jing Wang, Yan Li, Yanhang Gao, Quan Liu, Junqi Niu, Wei Wei
doi: 10.1007/s12250-021-00427-2
Received: 04 February 2021 Accepted: 26 May 2021 Published: 05 August 2021
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Genomic Characterization of a New Coronavirus from Migratory Birds in Jiangxi Province of China
Wentao Zhu, Wentao Song, Guoyin Fan, Jing Yang, Shan Lu, Dong Jin, Xue-lian Luo, Ji Pu, Haiying Chen, Jianguo Xu
doi: 10.1007/s12250-021-00402-x
Received: 25 January 2021 Accepted: 29 March 2021 Published: 08 July 2021
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Consecutive Monitoring of Interleukin-6 Is Needed for COVID-19 Patients
Xiaohua Chen, Juan Zhou, Chen Chen, Baidong Hou Hou, Ashaq Ali, Feng Li, Zhaolin Hua, Yingtao Wu, Qin Yang, Min Chen, Rong Zhang, Qianchuan Huang, Jinya Ding, Xian-En Zhang, Dong Men
doi: 10.1007/s12250-021-00425-4
Received: 26 January 2021 Accepted: 21 May 2021 Published: 07 July 2021
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Two Inhibitors Against the 3C-Like Proteases of Swine Coronavirus and Feline Coronavirus
Mengxin Zhou, Yutong Han, Mengxia Li, Gang Ye, Guiqing Peng
doi: 10.1007/s12250-021-00415-6
Received: 19 February 2021 Accepted: 06 May 2021 Published: 06 July 2021
[FullText HTML] [PDF 1998KB] Springerlink

Coronaviruses (CoVs) are important human and animal pathogens that cause respiratory and gastrointestinal diseases. Porcine epidemic diarrhoea (PED), characterized by severe diarrhoea and vomiting in pigs, is a highly lethal disease caused by porcine epidemic diarrhoea virus (PEDV) and causes substantial losses in the swine industry worldwide. However, currently available commercial drugs have not shown great therapeutic effects. In this study, a fluorescence resonance energy transfer (FRET)-based assay was applied to screen a library containing 1, 590 compounds and identified two compounds, 3-(aminocarbonyl)-1-phenylpyridinium and 2, 3-dichloronaphthoquinone, that target the 3C-like protease (3CLpro) of PEDV. These compounds are of low molecular weight (MW) and greatly inhibited the activity of this enzyme (IC50 values were obtained in this study). Furthermore, these compounds exhibited antiviral capacity against another member of the CoV family, feline infectious peritonitis virus (FIPV). Here, the inhibitory effects of these compounds against CoVs on Vero cells and feline kidney cells were identified (with EC50 values) and cell viability assays were performed. The results of putative molecular docking models indicate that these compounds, labeled compound 1 and compound 2, contact the conserved active sites (Cys144, Glu165, Gln191) of 3CLpro via hydrogen bonds. These findings provide insight into the antiviral activities of compounds 1 and 2 that may facilitate future research on anti-CoV drugs.

Non-Structural Protein 5 of Zika Virus Interacts with p53 in Human Neural Progenitor Cells and Induces p53-Mediated Apoptosis
Ping Li, Hualian Jiang, Hong Peng, Weijie Zeng, Yongheng Zhong, Miao He, Luyang Xie, Junhai Chen, Deyin Guo, Junyu Wu, Chun-Mei Li
doi: 10.1007/s12250-021-00422-7
Received: 05 April 2021 Accepted: 08 June 2021 Published: 05 July 2021
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Zika virus (ZIKV) infection could disrupt neurogenesis and cause microcephaly in neonates by targeting neural progenitor cells (NPCs). The tumor suppressor p53-mediated cell cycle arrest and apoptotic cell death have been suggested to be activated upon ZIKV infection,yet the detailed mechanism is not well understood. In the present study,we investigated the effects of ZIKV-encoded proteins in the activation of p53 signaling pathway and found that,among the ten viral proteins,the nonstructural protein 5 (NS5) of ZIKV most significantly activated the transcription of p53 target genes. Using the immunoprecipitation-coupled mass spectrometry approach,we identified that ZIKV-NS5 interacted with p53 protein. The NS5-p53 interaction was further confirmed by co-immunoprecipitation and GST pull-down assays. In addition,the MTase domain of NS5 and the C-terminal domain of p53 were mapped to be responsible for the interaction between these two proteins. We further showed that ZIKV-NS5 was colocalized with p53 and increased its protein level in the nuclei and able to prolong the half-life of p53. Furthermore,lentivirus-mediated expression of ZIKV-NS5 in hNPCs led to an apparent cell death phenotype. ZIKV-NS5 promoted the cleavage of PARP1 and significantly increased the cell apoptosis of hNPCs. Taken together,these findings revealed that ZIKV-NS5 is a previously undiscovered regulator of p53-mediated apoptosis in hNPCs,which may contribute to the ZIKV-caused abnormal neurodevelopment.

Desmoglein 2 (DSG2) Is A Receptor of Human Adenovirus Type 55 Causing Adult Severe Community-Acquired Pneumonia
Jing Zhang, Kui Ma, Xiangyu Wang, Yinbo Jiang, Shan Zhao, Junxian Ou, Wendong Lan, Wenyi Guan, Xiaowei Wu, Heping Zheng, Bin Yang, Chengsong Wan, Wei Zhao, Jianguo Wu, Qiwei Zhang
doi: 10.1007/s12250-021-00414-7
Received: 12 February 2021 Accepted: 06 May 2021 Published: 05 July 2021
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Human adenovirus type 55 (HAdV-B55) is a re-emergent acute respiratory disease pathogen that causes adult community-acquired pneumonia (CAP). Previous studies have shown that the receptor of HAdV-B14,which genome is highly similar with HAdV-B55,is human Desmoglein 2 (DSG2). However,whether the receptor of HAdV-B55 is DSG2 is undetermined because there are three amino acid mutations in the fiber gene between HAdV-B14 and HAdV-B55. Here,firstly we found the 3T3 cells,a mouse embryo fibroblast rodent cell line which does not express human DSG2,were able to be infected by HAdV-B55 after transfected with pcDNA3.1-DSG2,while normal 3T3 cells were still unsusceptible to HAdV-B55 infection. Next,A549 cells with hDSG2 knock-down by siRNA were hard to be infected by HAdV-B3/-B14/-B55,while the control siRNA group was still able to be infected by all these types of HAdVs. Finally,immunofluorescence confocal microscopy indicated visually that Cy3-conjugated HAdV-B55 viruses entered A549 cells by binding to DSG2 protein. Therefore,DSG2 is a major receptor of HAdV-B55 causing adult CAP. Our finding is important for better understanding of interactions between adenoviruses and host cells and may shed light on the development of new drugs that can interfere with these processes as well as for the development of potent prophylactic vaccines.

Antibody Cocktail Exhibits Broad Neutralization Activity Against SARS-CoV-2 and SARS-CoV-2 Variants
Yuanyuan Qu, Xueyan Zhang, Meiyu Wang, Lina Sun, Yongzhong Jiang, Cheng Li, Wei Wu, Zhen Chen, Qiangling Yin, Xiaolin Jiang, Yang Liu, Chuan Li, Jiandong Li, Tianlei Ying, Dexin Li, Faxian Zhan, Youchun Wang, Wuxiang Guan, Shiwen Wang, Mifang Liang
doi: 10.1007/s12250-021-00409-4
Received: 16 March 2021 Accepted: 06 May 2021 Published: 05 July 2021
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Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has precipitated multiple variants resistant to therapeutic antibodies. In this study,12 high-affinity antibodies were generated from convalescent donors in early outbreaks using immune antibody phage display libraries. Of them,two RBD-binding antibodies (F61 and H121) showed high-affinity neutralization against SARS-CoV-2,whereas three S2-target antibodies failed to neutralize SARS-CoV-2. Following structure analysis,F61 identified a linear epitope located in residues G446–S494,which overlapped with angiotensin-converting enzyme 2 (ACE2) binding sites,while H121 recognized a conformational epitope located on the side face of RBD,outside from ACE2 binding domain. Hence the cocktail of the two antibodies achieved better performance of neutralization to SARS-CoV-2. Importantly,these two antibodies also showed efficient neutralizing activities to the variants including B.1.1.7 and B.1.351,and reacted with mutations of N501Y,E484K,and L452R,indicated that it may also neutralize the recent India endemic strain B.1.617. The unchanged binding activity of F61 and H121 to RBD with multiple mutations revealed a broad neutralizing activity against variants,which mitigated the risk of viral escape. Our findings revealed the therapeutic basis of cocktail antibodies against constantly emerging SARS-CoV-2 variants and provided promising candidate antibodies to clinical treatment of COVID-19 patients infected with broad SARS-CoV-2 variants.

CRISPR/Cas12a Technology Combined with RT-ERA for Rapid and Portable SARS-CoV-2 Detection
Sihua Liu, Mengqian Huang, Yanan Xu, Jun Kang, Sheng Ye, Si Liu, Zhiyun Wang, Hongyun Liu, Jibin Yu, Kongxin Hu, Tao Wang
doi: 10.1007/s12250-021-00406-7
Received: 24 November 2020 Accepted: 28 March 2021 Published: 02 July 2021
[FullText HTML] [PDF 751KB] Springerlink ESM
ANXA2 Facilitates Enterovirus 71 Infection by Interacting with 3D Polymerase and PI4KB to Assist the Assembly of Replication Organelles
Qiuhan Zhang, Siliang Li, Ping Lei, Zixian Li, Feifei Chen, Qi Chen, Yulu Wang, Jiami Gong, Qi Tang, Xinjin Liu, Ke Lan, Shuwen Wu
doi: 10.1007/s12250-021-00417-4
Received: 14 March 2021 Accepted: 27 April 2021 Published: 01 July 2021
[FullText HTML] [PDF 3140KB] Springerlink

Similar to that of other enteroviruses, the replication of enterovirus 71 (EV71) occurs on rearranged membranous structures called replication organelles (ROs). Phosphatidylinositol 4-kinase Ⅲ (PI4KB), which is required by enteroviruses for RO formation, yields phosphatidylinositol-4-phosphate (PI4P) on ROs. PI4P then binds and induces conformational changes in the RNA-dependent RNA polymerase (RdRp) to modulate RdRp activity. Here, we targeted 3D polymerase, the core enzyme of EV71 ROs, and found that the host factor Annexin A2 (ANXA2) can interact with 3D polymerase and promote the replication of EV71. Then, an experiment showed that the annexin domain of ANXA2, which possesses membrane-binding capacity, mediates the interaction of ANXA2 with EV71 3D polymerase. Further research showed that ANXA2 is localized on ROs and interacts with PI4KB. Overexpression of ANXA2 stimulated the formation of PI4P, and the level of PI4P was decreased in ANXA2-knockout cells. Furthermore, ANXA2, PI4KB, and 3D were shown to be localized to the viral RNA replication site, where they form a higher-order protein complex, and the presence of ANXA2 promoted the PI4KB-3D interaction. Altogether, our data provide new insight into the role of ANXA2 in facilitating formation of the EV71 RNA replication complex.

Profiles of SARS-CoV-2 RNA and Antibodies in Inpatients with COVID-19 not Related with Clinical Manifestation: A Single Centre Study
Li Zhao, Ruqin Gao, Roujian Lu, Huijuan Wang, Yao Deng, Peihua Niu, Fachun Jiang, Baoying Huang, Jiwei Liang, Jing Jia, Feng Zhang, Wenling Wang, Guizhen Wu, Wenjie Tan
doi: 10.1007/s12250-021-00411-w
Received: 13 August 2020 Accepted: 30 April 2021 Published: 01 July 2021
[FullText HTML] [PDF 331KB] Springerlink
Recovery of a Far-Eastern Strain of Tick-Borne Encephalitis Virus with a Full-Length Infectious cDNA Clone
Penghui Li, Chen Yao, Ting Wang, Tong Wu, Wenfu Yi, Yue Zheng, Yuanjiu Miao, Jianhong Sun, Zhongyuan Tan, Yan Liu, Xiaowei Zhang, Hanzhong Wang, Zhenhua Zheng
doi: 10.1007/s12250-021-00396-6
Received: 17 July 2020 Accepted: 15 March 2021 Published: 30 June 2021
[FullText HTML] [PDF 3290KB] Springerlink ESM

Tick-borne encephalitis virus (TBEV) is a pathogenic virus known to cause central nervous system (CNS) diseases in humans, and has become an increasing public health threat nowadays. The rates of TBEV infection in the endemic countries are increasing. However, there is no effective antiviral against the disease. This underscores the urgent need for tools to study the emergence and pathogenesis of TBEV and to accelerate the development of vaccines and antivirals. In this study, we reported an infectious cDNA clone of TBEV that was isolated in China (the WH2012 strain). A beta-globin intron was inserted in the coding region of nonstructural protein 1 (NS1) gene to improve the stability of viral genome in bacteria. In mammalian cells, the inserted intron was excised and spliced precisely, which did not lead to the generation of inserted mutants. High titers of infectious progeny viruses were generated after the transfection of the infectious clone. The cDNA-derived TBEV replicated efficiently, and caused typical cytopathic effect (CPE) and plaques in BHK-21 cells. In addition, the CPE and growth curve of cDNA-derived virus were similar to that of its parental isolate in cells. Together, we have constructed the first infectious TBEV cDNA clone in China, and the clone can be used to investigate the genetic determinants of TBEV virulence and disease pathogenesis, and to develop countermeasures against the virus.

New Simian Enterovirus 19 (EV-A122) Strains in China Reveal Large-Scale Inter-Serotype Recombination between Simian EV-As
Zhenzhi Han, Jinbo Xiao, Yang Song, Shuangli Zhu, Dongyan Wang, Huanhuan Lu, Tianjiao Ji, Dongmei Yan, Wenbo Xu, Yong Zhang
doi: 10.1007/s12250-021-00412-9
Received: 06 December 2020 Accepted: 13 April 2021 Published: 29 June 2021
[FullText HTML] [PDF 307KB] Springerlink ESM
Ectopic Expression of TRIM25 Restores RIG-I Expression and IFN Production Reduced by Multiple Enteroviruses 3Cpro
Huimin Xiao, Jingliang Li, Xu Yang, Zhaolong Li, Ying Wang, Yajuan Rui, Bin Liu, Wenyan Zhang
doi: 10.1007/s12250-021-00410-x
Received: 31 January 2021 Accepted: 12 April 2021 Published: 25 June 2021
[FullText HTML] [PDF 1309KB] Springerlink

Enteroviruses (EVs) 3C proteins suppress type Ⅰ interferon (IFN) responses mediated by retinoid acid-inducible gene I (RIG-I), while an E3 ubiquitin ligase, tripartite motif protein 25 (TRIM25)-mediated RIG-I ubiquitination is essential for RIG-I antiviral activity. Therefore, whether the effect of EVs 3C on RIG-I is associated with TRIM25 expression is worth to be further investigated. Here, we demonstrate that 3C proteins of EV71 and coxsackievirus B3 (CVB3) reduced not only RIG-I expression but also TRIM25 expression through protease cleavage activity, while overexpression of TRIM25 restored RIG-I expression and IFN-β production reduced by 3C proteins. Further investigation confirmed that the two amino acids and functional domains in TRIM25 required for RIG-I ubiquitination and TRIM25 structural conformation were essential for the recovery of RIG-I expression. Moreover, we also observed that TRIM25 could rescue RIG-I expression reduced by 3C proteins of CVA6 and EV-D68 but not CVA16. Our findings provide an insightful interpretation of 3C-mediated host innate immune suppression and support TRIM25 as an attractive target against multiple EVs infection.

Clinical Characteristics of Human Adenovirus Plastic Bronchitis in 10 Pediatric Cases: A Retrospective Study of Seven Years
Lingjian Zeng, Jianhua Wei, Yuyi Tang, Enmei Liu, Qubei Li, Na Zang
doi: 10.1007/s12250-021-00394-8
Received: 02 November 2020 Accepted: 25 April 2021 Published: 22 June 2021
[FullText HTML] [PDF 358KB] Springerlink
Histone Deacetylase Inhibitor SAHA Induces Expression of Fatty Acid-Binding Protein 4 and Inhibits Replication of Human Cytomegalovirus
Zhongshun Liu, Baoqin Xuan, Shubing Tang, Zhikang Qian
doi: 10.1007/s12250-021-00382-y
Received: 10 July 2020 Accepted: 11 March 2021 Published: 22 June 2021
[FullText HTML] [PDF 754KB] Springerlink ESM

Suberoylanilide hydroxamic acid (SAHA) is a histone deacetylase inhibitor that shows marked efficacy against many types of cancers and is approved to treat severe metastatic cutaneous T-cell lymphomas. In addition to its anticancer activity, SAHA has significant effects on the growth of many viruses. The effect of SAHA on replication of human cytomegalovirus (HCMV) has not, however, been investigated. Here, we showed that the replication of HCMV was significantly suppressed by treatment with SAHA at concentrations that did not show appreciable cytotoxicity. SAHA reduced transcription and protein levels of HCMV immediate early genes, showing that SAHA acts at an early stage in the viral life-cycle. RNA-sequencing data mining showed that numerous pathways and molecules were affected by SAHA. Interferon-mediated immunity was one of the most relevant pathways in the RNA-sequencing data, and we confirmed that SAHA inhibits HCMV-induced IFN-mediated immune responses using quantitative Real-time PCR (qRT-PCR). Fatty acid-binding protein 4 (FABP4), which plays a role in lipid metabolism, was identified by RNA-sequencing. We found that FABP4 expression was reduced by HCMV infection but increased by treatment with SAHA. We then showed that knockdown of FABP4 partially rescued the effect of SAHA on HCMV replication. Our data suggest that FABP4 contributes to the inhibitory effect of SAHA on HCMV replication.

Current Status of Human Papillomavirus-Related Head and Neck Cancer: From Viral Genome to Patient Care
Haoru Dong, Xinhua Shu, Qiang Xu, Chen Zhu, Andreas M. Kaufmann, Zhi-Ming Zheng, Andreas E. Albers, Xu Qian
doi: 10.1007/s12250-021-00413-8
Received: 21 January 2021 Accepted: 18 May 2021 Published: 21 June 2021
[FullText HTML] [PDF 619KB] Springerlink

Human papillomavirus (HPV) infection identified as a definitive human carcinogen is increasingly being recognized for its role in carcinogenesis of human cancers. Up to 38%–80% of head and neck squamous cell carcinoma (HNSCC) in oropharyngeal location (OPSCC) and nearly all cervical cancers contain the HPV genome which is implicated in causing cancer through its oncoproteins E6 and E7. Given by the biologically distinct HPV-related OPSCC and a more favorable prognosis compared to HPV-negative tumors, clinical trials on de-escalation treatment strategies for these patients have been studied. It is therefore raised the questions for the patient stratification if treatment de-escalation is feasible. Moreover, understanding the crosstalk of HPV-mediated malignancy and immunity with clinical insights from the proportional response rate to immune checkpoint blockade treatments in patients with HNSCC is of importance to substantially improve the treatment efficacy. This review discusses the biology of HPV-related HNSCC as well as successful clinically findings with promising candidates in the pipeline for future directions. With the advent of various sequencing technologies, further biomolecules associated with HPV-related HNSCC progression are currently being identified to be used as potential biomarkers or targets for clinical decisions throughout the continuum of cancer care.

The Application of a Safe Neutralization Assay for Ebola Virus Using Lentivirus-Based Pseudotyped Virus
Zengguo Cao, Hongli Jin, Gary Wong, Ying Zhang, Cuicui Jiao, Na Feng, Fangfang Wu, Shengnan Xu, Hang Chi, Yongkun Zhao, Tiecheng Wang, Weiyang Sun, Yuwei Gao, Songtao Yang, Xianzhu Xia, Hualei Wang
doi: 10.1007/s12250-021-00405-8
Received: 28 July 2020 Accepted: 19 April 2021 Published: 21 June 2021
[FullText HTML] [PDF 543KB] Springerlink
Dynamic Host Immune and Transcriptomic Responses to Respiratory Syncytial Virus Infection in a Vaccination-Challenge Mouse Model
Yu Zhao, Chen Ma, Jie Yang, Xiufen Zou, Zishu Pan
doi: 10.1007/s12250-021-00418-3
Received: 08 April 2021 Accepted: 20 April 2021 Published: 17 June 2021
[FullText HTML] [PDF 1891KB] Springerlink ESM

Respiratory syncytial virus (RSV) is the major cause of lower respiratory tract infections in children. Inactivated RSV vaccine was developed in the late 1960's,but the vaccine-enhanced disease (VED) occurred to vaccinated infants upon subsequent natural RSV infection. The excessive inflammatory immunopathology in the lungs might be involved in the VED,but the underlying mechanisms remain not fully understood. In this study,we utilized UV-inactivated RSV in the prime/boost approach followed by RSV challenge in BALB/c mice to mimic RSV VED. The dynamic virus load,cytokines,histology and transcriptome profiles in lung tissues of mice were investigated from day 1 to day 6 post-infection. Compared to PBS-treated mice,UV-RSV vaccination leads to a Th2 type inflammatory response characterized by enhanced histopathology,reduced Treg cells and increased IL4+CD4 T cells in the lung. Enhanced production of several Th2 type cytokines (IL-4,IL-5,IL-10) and TGF-β,reduction of IL-6 and IL-17 were observed in UV-RSV vaccinated mice. A total of 5582 differentially expressed (DE) genes between PBS-treated or vaccinated mice and naïve mice were identified by RNA-Seq. Eleven conserved high-influential modules (HMs) were recognized,majorly grouped into regulatory networks related to cell cycle and cell metabolism,signal transduction,immune and inflammatory responses. At an early time post-infection,the vaccinated mice showed obvious decreased expression patterns of DE genes in 11 HMs compared to PBS-treated mice. The extracellular matrix (HM5) and immune responses (HM8) revealed tremendous differences in expression and regulation characteristics of transcripts between PBS-treated and vaccinated mice at both early and late time points. The highly connected genes in HM5 and HM8 networks were further validated by RT-qPCR. These findings reveal the relationship between RSV VED and immune responses,which could benefit the development of novel RSV vaccines.

The C/EBPb-Dependent Induction of TFDP2 Facilitates Porcine Reproductive and Respiratory Syndrome Virus Proliferation
Min Zhu, Xiaoyang Li, Ruiqi Sun, Peidian Shi, Aiping Cao, Lilin Zhang, Yanyu Guo, Jinhai Huang
doi: 10.1007/s12250-021-00403-w
Received: 07 February 2021 Accepted: 28 April 2021 Published: 17 June 2021
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Antigenic Drift of the Hemagglutinin from an Influenza A (H1N1) pdm09 Clinical Isolate Increases its Pathogenicity In Vitro
Lei Xing, Yunbo Chen, Boqian Chen, Ling Bu, Ying Liu, Zhiqi Zeng, Wenda Guan, Qigao Chen, Yongping Lin, Kun Qin, Honglin Chen, Xilong Deng, Xinhua Wang, Wenjun Song
doi: 10.1007/s12250-021-00401-y
Received: 06 January 2021 Accepted: 12 April 2021 Published: 09 June 2021
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Development and Characterization of SYBR Green Ⅰ Based RT-PCR Assay for Detection of Omsk Hemorrhagic Fever Virus
Ya-Nan Zhang, Si-Qing Liu, Cheng-Lin Deng, Zhi-Ming Yuan, Bo Zhang, Xiao-Dan Li, Han-Qing Ye
doi: 10.1007/s12250-021-00389-5
Received: 10 January 2021 Accepted: 17 March 2021 Published: 02 June 2021
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Safety and Considerations of the COVID-19 Vaccine Massive Deployment
Junwei Li, Mingyue Song, Deyin Guo, Yongxiang Yi
doi: 10.1007/10.1007/s12250-021-00408-5
Received: 20 February 2021 Accepted: 26 April 2021 Published: 01 June 2021
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Cholesterol-25-Hydroxylase Suppresses Seneca Valley Virus Infection via Producing 25-Hydroxycholesterol to Block Adsorption Procedure
Hui Li, Zekai Zhao, Xiangmin Li, Liuxing Qin, Wei Wen, Huanchun Chen, Ping Qian
doi: 10.1007/s12250-021-00377-9
Received: 03 June 2020 Accepted: 22 September 2020 Published: 01 June 2021
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Cholesterol-25-hydroxylase (CH25H) is a membrane protein associated with endoplasmic reticulum, and it is an interferon-stimulated factor regulated by interferon. CH25H catalyzes cholesterol to produce 25-hydroxycholesterol (25HC) by adding a second hydroxyl to the 25th carbon atom of cholesterol. Recent studies have shown that both CH25H and 25HC could inhibit the replication of many viruses. In this study, we found that ectopic expression of CH25H in HEK-293T and BHK-21 cell lines could inhibit the replication of Seneca Valley virus (SVV) and that there was no species difference. On the other hand, the knockdown of CH25H could enhance the replication of SVV in HEK-293T and BHK-21 cells, indicating the importance of CH25H. To some extent, the CH25H mutant without hydroxylase activity also lost its ability to inhibit SVV amplification. Further studies demonstrated that 25HC was involved in the entire life cycle of SVV, especially in repressing its adsorption process. This study reveals that CH25H exerts the advantage of innate immunity mainly by producing 25HC to block virion adsorption.

Modulation of Antiviral Immunity and Therapeutic Efficacy by 25-Hydroxycholesterol in Chronically SIV-Infected, ART-Treated Rhesus Macaques
Chunxiu Wu, Jin Zhao, Ruiting Li, Fengling Feng, Yizi He, Yanjun Li, Runhan Huang, Guangye Li, Heng Yang, Genhong Cheng, Ling Chen, Feng Ma, Pingchao Li, Caijun Sun
doi: 10.1007/s12250-021-00407-6
Received: 29 January 2021 Accepted: 19 April 2021 Published: 31 May 2021
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Cholesterol-25-hydroxylase (CH25H) and its enzymatic product 25-hydroxycholesterol (25HC) exert broadly antiviral activity including inhibiting HIV-1 infection. However, their antiviral immunity and therapeutic efficacy in a nonhuman primate model are unknown. Here, we report that the regimen of 25HC combined with antiretroviral therapy (ART), provides profound immunological modulation towards inhibiting viral replication in chronically SIVmac239-infected rhesus macaques (RMs). Compared to the ART alone, this regimen more effectively controlled SIV replication, enhanced SIV-specific cellular immune responses, restored the ratio of CD4/CD8 cells, reversed the hyperactivation state of CD4+ T cells, and inhibited the secretion of proinflammatory cytokines by CD4+ and CD8+ T lymphocytes in chronically SIV-infected RMs. Furthermore, the in vivo safety and the preliminary pharmacokinetics of the 25HC compound were assessed in this RM model. Taken together, these assessments help explain the profound relationship between cholesterol metabolism, immune modulation, and antiviral activities by 25HC. These results provide insight for developing novel therapeutic drug candidates against HIV-1 infection and other related diseases.

The Functional Characterization of Bat and Human P[3] Rotavirus VP8*s
Dandi Li, Mengxuan Wang, Tongyao Mao, Mingwen Wang, Qing Zhang, Hong Wang, Lili Pang, Xiaoman Sun, Zhaojun Duan
doi: 10.1007/s12250-021-00400-z
Received: 06 January 2021 Accepted: 12 April 2021 Published: 31 May 2021
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P[3] rotavirus (RV) has been identified in many species, including human, simian, dog, and bat. Several glycans, including sialic acid, histo-blood group antigens (HBGAs) are reported as RV attachment factors. The glycan binding specificity of different P[3] RV VP8*s were investigated in this study. Human HCR3A and dog P[3] RV VP8*s recognized glycans with terminal sialic acid and hemagglutinated the red blood cells, while bat P[3] VP8* showed neither binding to glycans nor hemagglutination. However, the bat P[3] VP8* mutant of C189Y obtained the ability to hemagglutinate the red blood cells, while human P[3] HCR3A/M2-102 mutants of Y189C lost the ability. Sequence alignment and structural analysis indicated that residue 189 played an important role in the ligand recognition and may contribute to the cross-species transmission. Structural superimposition exhibited that bat P[3] VP8* model was quite different from the simian P[3] Rhesus rotavirus (RRV) P[3] VP8*, indicating that bat P[3] RV was relatively distinct and partially contributed to the no binding to tested glycans. These results promote our understanding of P[3] VP8*/glycans interactions and the potential transmission of bat/human P[3] RVs, offering more insight into the RV infection and prevalence.

Re-isolation of Wuxiang Virus from Wild Sandflies Collected from Yangquan County, China
Qinyan Wang, Shihong Fu, Jingxia Cheng, Xiuyan Xu, Jing Wang, Bin Wu, Xiaodong Tian, Yan Li, Ying He, Fan Li, Kai Nie, Songtao Xu, Bin Wang, Huanyu Wang, Xiaoqing Lu, Guodong Liang
doi: 10.1007/s12250-021-00398-4
Received: 17 November 2020 Accepted: 25 March 2021 Published: 31 May 2021
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We previously isolated a new species of the genus Phlebovirus from wild sandflies collected from Wuxiang County in central China, which named the Wuxiang virus (WUXV). In this study, we re-isolated the WUXV from wild sandflies collected from two villages in Yangquan County, China in 2019. Four virus isolates that caused cytopathic effects in BHK-21 cells were successfully isolated from sandfly specimens collected from chicken pens and sheep pens. Phylogenetic analyses of the L, M and S gene segments of the viruses revealed that the four virus strains represented the previously isolated WUXV. The minimum infection rate (MIR) of the virus isolated from the sheep pen was 3.21, and the MIR of the virus isolated from the chicken pen was 3.45. The positive rates of Wuxiang virus neutralizing antibodies in serum samples of local healthy people and domestic chickens were 8.7% (4/46) and 100% (4/4), respectively, suggesting that Wuxiang virus can infect human and animal. In view of the fact that Wuxiang virus is infectious to humans and animals and has a relatively wide geographical distribution in China, it is of great public health significance to strengthen the investigation and study on the infection status of Wuxiang virus in humans and animals.

Computational Viromics: Applications of the Computational Biology in Viromics Studies
Congyu Lu, Yousong Peng
doi: 10.1007/s12250-021-00395-7
Received: 02 June 2020 Accepted: 14 April 2021 Published: 31 May 2021
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PML Suppresses Influenza Virus Replication by Promoting FBXW7 Expression
Hai-Yan Yan, Hui-Qiang Wang, Ming Zhong, Shuo Wu, Lu Yang, Ke Li, Yu-Huan Li
doi: 10.1007/s12250-021-00399-3
Received: 16 December 2020 Accepted: 29 March 2021 Published: 27 May 2021
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Influenza A viruses (IAV) are responsible for seasonal flu epidemics, which can lead to high morbidity and mortality each year. Like other viruses, influenza virus can hijack host cellular machinery for its replication. Host cells have evolved diverse cellular defense to resist the invasion of viruses. As the main components of promyelocytic leukemia protein nuclear bodies (PML-NBs), PML can inhibit the replication of many medically important viruses including IAV. However, the mechanism of PML against IAV is unclear. In the present study, we found PML was induced in response to IAV infection and ectopic expression of PML could inhibit IAV replication, whereas knockdown of endogenous PML expression could enhance IAV replication. Further studies showed that PML increased the expression of FBXW7 by inhibiting its K48-linked ubiquitination and enhanced the interaction between FBXW7 and SHP2, which negatively regulated IAV replication during infection. Moreover, PML stabilized RIG-I to promote the production of type Ⅰ IFN. Collectively, these data indicated that PML inhibited IAV replication by enhancing FBXW7 expression in the antiviral immunity against influenza virus and extended the mechanism of PML in antiviral immunity.

Cucurbit[7]uril as a Broad-Spectrum Antiviral Agent against Diverse RNA Viruses
Jia Quan, Xiangjun Zhang, Yuanfu Ding, Shengke Li, Yang Qiu, Ruibing Wang, Xi Zhou
doi: 10.1007/s12250-021-00404-9
Received: 25 March 2021 Accepted: 06 April 2021 Published: 26 May 2021
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The emergence and re-emergence of RNA virus outbreaks highlight the urgent need for the development of broad-spectrum antivirals. Polyamines are positively-charged small molecules required for the infectivity of a wide range of RNA viruses, therefore may become good antiviral targets. Cucurbit[7]uril (CB[7]), a synthetic macrocyclic molecule, which can bind with amine-based organic compounds with high affinity, has been shown to regulate bioactive molecules through competitive binding. In this study, we tested the antiviral activity of CB[7] against diverse RNA viruses, including a panel of enteroviruses (i.e. human enterovirus A71, coxsackievirus A16, coxsackievirus B3, and echovirus 11), some flaviviruses (i.e. dengue virus and Zika virus), and an alphavirus representative Semliki forest virus. CB[7] can inhibit virus replications in a variety of cell lines, and its mechanism of action is through the competitive binding with polyamines. Our findings not only for the first time provide evidence that CB[7] can be a promising broad-spectrum antiviral agent, but more importantly, offer a novel therapeutic strategy to fight against RNA viruses by supramolecular sequestration of polyamines.

Anti-SARS-CoV-2 IgY Isolated from Egg Yolks of Hens Immunized with Inactivated SARS-CoV-2 for Immunoprophylaxis of COVID-19
Haiyan Shen, Yanxing Cai, Huan Zhang, Jie Wu, Lin Ye, Penghui Yang, Xiaojun Lin, Shibo Jiang, Ming Liao
doi: 10.1007/s12250-021-00371-1
Received: 29 November 2020 Accepted: 08 February 2021 Published: 21 May 2021
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Epidemiological Characteristics of Influenza A and B in Macau, 2010–2018
HoiMan Ng, Teng Zhang, Guoliang Wang, SiMeng Kan, Guoyi Ma, Zhe Li, Chang Chen, Dandan Wang, MengIn Wong, ChioHang Wong, Jinliang Ni, Xiaohua Douglas Zhang
doi: 10.1007/s12250-021-00388-6
Received: 04 December 2020 Accepted: 04 March 2021 Published: 20 May 2021
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Influenza is one of the major respiratory diseases in humans. Macau is a tourist city with high density of population and special population mobility. The study on the epidemiological characteristics of influenza in Macau should bring great value for preventing influenza in tourist cities like Macau in the world. In this study, we collected a total of 104, 874 samples with influenza-like illness (ILI) in Macau from 2010 to 2018. Chi-square test and binary multivariable logistic regression were used to investigate the epidemiological characteristics of influenza A and B in Macau. Among these ILI samples, the overall positive rate is 17.17% for influenza A and 6.97% for influenza B. The epidemics of influenza in three years (i.e., 2012, 2017 and 2018) differ from the remaining years (i.e., normal years). In a normal year, influenza A occurs year-round whereas influenza B is seasonal. Our research shows significant differences in influenza infections between different age groups in normal years. Interestingly, our analysis shows no significant difference between locals and tourists in influenza A and B infection in a normal year, whereas the odds of influenza A in tourists were significantly higher than those in locals in July 2017 and the odds of influenza B in tourists were significantly higher than those in locals in January–February 2012 and January–February 2018. This is possibly attributed by the policy of free vaccination to everyone in Macau. These findings should be valuable for preventing influenza in not only Macau but also the world.

Transcriptome Analyses Implicate Endogenous Retroviruses Involved in the Host Antiviral Immune System through the Interferon Pathway
Miao Wang, Liying Wang, Haizhou Liu, Jianjun Chen, Di Liu
doi: 10.1007/s12250-021-00370-2
Received: 10 October 2020 Accepted: 08 February 2021 Published: 19 May 2021
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Human endogenous retroviruses (HERVs) are the remains of ancient retroviruses that invaded our ancestors' germline cell and were integrated into the genome. The expression of HERVs has always been a cause for concern because of its association with various cancers and diseases. However, few previous studies have focused on specific activation of HERVs by viral infections. Our previous study has shown that dengue virus type 2 (DENV-2) infection induces the transcription of a large number of abnormal HERVs loci; therefore, the purpose of this study was to explore the relationship between exogenous viral infection and HERV activation further. In this study, we retrieved and reanalyzed published data on 21 transcriptomes of human cells infected with various viruses. We found that infection with different viruses could induce transcriptional activation of HERV loci. Through the comparative analysis of all viral datasets, we identified 43 key HERV loci that were up-regulated by DENV-2, influenza A virus, influenza B virus, Zika virus, measles virus, and West Nile virus infections. Furthermore, the neighboring genes of these HERVs were simultaneously up-regulated, and almost all such neighboring genes were interferon-stimulated genes (ISGs), which are enriched in the host's antiviral immune response pathways. Our data supported the hypothesis that activation of HERVs, probably via an interferon-mediated mechanism, plays an important role in innate immunity against viral infections.

A Convenient and Biosafe Replicon with Accessory Genes of SARS-CoV-2 and Its Potential Application in Antiviral Drug Discovery
Yun-Yun Jin, Hanwen Lin, Liu Cao, Wei-Chen Wu, Yanxi Ji, Liubing Du, Yiling Jiang, Yanchun Xie, Kuijie Tong, Fan Xing, Fuxiang Zheng, Mang Shi, Ji-An Pan, Xiaoxue Peng, Deyin Guo
doi: 10.1007/s12250-021-00385-9
Received: 02 March 2021 Accepted: 11 March 2021 Published: 17 May 2021
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SARS-CoV-2 causes the pandemic of COVID-19 and no effective drugs for this disease are available thus far. Due to the high infectivity and pathogenicity of this virus, all studies on the live virus are strictly confined in the biosafety level 3 (BSL3) laboratory but this would hinder the basic research and antiviral drug development of SARS-CoV-2 because the BSL3 facility is not commonly available and the work in the containment is costly and laborious. In this study, we constructed a reverse genetics system of SARS-CoV-2 by assembling the viral cDNA in a bacterial artificial chromosome (BAC) vector with deletion of the spike (S) gene. Transfection of the cDNA into cells results in the production of an RNA replicon that keeps the capability of genome or subgenome replication but is deficient in virion assembly and infection due to the absence of S protein. Therefore, such a replicon system is not infectious and can be used in ordinary biological laboratories. We confirmed the efficient replication of the replicon by demonstrating the expression of the subgenomic RNAs which have similar profiles to the wild-type virus. By mutational analysis of nsp12 and nsp14, we showed that the RNA polymerase, exonuclease, and cap N7 methyltransferase play essential roles in genome replication and sgRNA production. We also created a SARS-CoV-2 replicon carrying a luciferase reporter gene and this system was validated by the inhibition assays with known anti-SARS-CoV-2 inhibitors. Thus, such a one-plasmid system is biosafe and convenient to use, which will benefit both fundamental research and development of antiviral drugs.

Enriched Opportunistic Pathogens Revealed by Metagenomic Sequencing Hint Potential Linkages between Pharyngeal Microbiota and COVID-19
Dongyan Xiong, Caroline Muema, Xiaoxu Zhang, Xinming Pan, Jin Xiong, Hang Yang, Junping Yu, Hongping Wei
doi: 10.1007/s12250-021-00391-x
Received: 15 December 2020 Accepted: 15 March 2021 Published: 12 May 2021
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As a respiratory tract virus, SARS-CoV-2 infected people through contacting with the upper respiratory tract first. Previous studies indicated that microbiota could modulate immune response against pathogen infection. In the present study, we performed metagenomic sequencing of pharyngeal swabs from eleven patients with COVID-19 and eleven Non-COVID-19 patients who had similar symptoms such as fever and cough. Through metagenomic analysis of the above two groups and a healthy group from the public data, there are 6502 species identified in the samples. Specifically, the Pielou index indicated a lower evenness of the microbiota in the COVID-19 group than that in the Non-COVID-19 group. Combined with the linear discriminant analysis (LDA) and the generalized linear model, eighty-one bacterial species were found with increased abundance in the COVID-19 group, where 51 species were enriched more than 8 folds. The top three enriched genera were Streptococcus, Prevotella and Campylobacter containing some opportunistic pathogens. More interestingly, through experiments, we found that two Streptococcus strains, S. suis and S. agalactiae, could stimulate the expression of ACE2 of Vero cells in vitro, which may promote SARS-CoV-2 infection. Therefore, these enriched pathogens in the pharynxes of COVID-19 patients may involve in the virus-host interactions to affect SARS-CoV-2 infection and cause potential secondary bacterial infections through changing the expression of the viral receptor ACE2 and/or modulate the host's immune system.

Accelerated Evolution of H7N9 Subtype Influenza Virus under Vaccination Pressure
Yifan Wu, Jingkai Hu, Xuanjiang Jin, Xiao Li, Jinfeng Wang, Mengmeng Zhang, Jianglin Chen, Shumin Xie, Wenbao Qi, Ming Liao, Weixin Jia
doi: 10.1007/s12250-021-00383-x
Received: 21 October 2020 Accepted: 17 March 2021 Published: 11 May 2021
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No avian H7N9 outbreaks have occurred since the introduction of H7N9 inactivated vaccine in the fall of 2017. However, H7N9 is still prevalent in poultry. To surveil the prevalence, genetic characteristics, and antigenic changes of H7N9, over 7000 oropharyngeal and cloaca swab specimens were collected from live poultry markets and farms in 15 provinces of China from 2017 to 2019. A total of 85 influenza virus subtype H7N9 strains were isolated and 20 representative strains were selected for genetic analysis and antigenicity evaluation. Results indicated the decreased prevalence of low-pathogenic H7N9 strains while highly-pathogenic H7N9 strains became dominated since the introduction of vaccine. Phylogenetic analysis showed that strains from 2019 formed an independent small branch and were genetically distant to strains isolated in 2013–2018. Analysis of key amino acid sites showed that the virus strains may adapt to the host environment evolutionally through mutation. Our analysis predicted additional potential glycosylation sites for HA and NA genes in the 2019 strains. Sequence analysis of HA gene in strains isolated from 2018 to 2019 showed that there were an increased nucleotide substitution rate and an increased mutation rate in the first and second nucleotides of coding codons within the open reading frame. The hemagglutination inhibition (HI) assay showed that H7-Re1 and H7-Re2 exhibited a lower HI titer for isolates from 2019, while H7-Re3 and rLN79 showed a high HI titer. The protective effect of the vaccine decreased after 15 months of use. Overall, under vaccination pressure, the evolution of influenza virus subtype H7N9 has accelerated.

Expansion of GARP-Expressing CD4+CD25-FoxP3+ T Cells and SATB1 Association with Activation and Coagulation in Immune Compromised HIV-1-Infected Individuals in South Africa
Eman Teer, Danzil E. Joseph, Leanne Dominick, Richard H. Glashoff, M. Faadiel Essop
doi: 10.1007/s12250-021-00386-8
Received: 17 September 2020 Accepted: 23 February 2021 Published: 11 May 2021
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Although antiretroviral treatment lowers the burden of human immunodeficiency virus (HIV)-related disease, it does not always result in immunological recovery. This manifests as persistent chronic inflammation, immune activation or exhaustion that can promote the onset of co-morbidities. As the exact function of regulatory T (Treg) cells in HIV remains unclear, this cross-sectional study investigated three expression markers (Forkhead box protein P3 [FOXP3], glycoprotein A repetitions predominant [GARP], special AT-rich sequence binding protein 1 [SATB1]) and compared their expansion between CD4+CD25- and CD4+CD25++ T cells. Age-matched study subjects were recruited (Western Cape, South Africa) and sub-divided: HIV-negative subjects (n = 12), HIV-positive naïve treated (n = 22), HIV-positive treated based on CD4 count cells/μL (CD4 > 500 and CD4 < 500) (n = 34) and HIV-treated based on viral load (VL) copies/mL (VL < 1000 and VL > 1000) (n = 34). Markers of immune activation (CD38) and coagulation (CD142) on T cells (CD8) were assessed by flow cytometry together with FOXP3, GARP and SATB1 expression on CD4+CD25- and CD4+CD25++ T cells. Plasma levels of interleukin-10 (IL-10; anti-inflammatory marker), IL-6 (inflammatory marker) and D-dimer (coagulation marker) were assessed. This study revealed three major findings in immuno-compromised patients with virological failure (CD4 < 500; VL > 1000): (1) the expansion of the unconventional Treg cell subset (CD4+CD25-FOXP3+) is linked with disease progression markers; (2) increased GARP expression in the CD4+CD25- and CD4+CD25++ subsets; and (3) the identification of a strong link between CD4+CD25-SATB1+ cells and markers of immune activation (CD8+CD38+) and coagulation (CD8+CD142+ and D-dimer).

Development of RNA Polymerase III-Driven Reverse Genetics System for the Rescue of a Plant Rhabdovirus
Xiaoyan Zhang, Kai Sun, Yan Liang, Chenglu Zhao, Zhenghe Li
doi: 10.1007/s12250-021-00390-y
Received: 23 November 2020 Accepted: 25 March 2021 Published: 03 May 2021
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Detection of SARS-CoV-2 RNA in Medical Wastewater in Wuhan During the COVID-19 Outbreak
Jun-Bo Zhou, Wen-Hua Kong, Sheng Wang, Yi-Bing Long, Lian-Hua Dong, Zhen-Yu He, Man-Qing Liu
doi: 10.1007/s12250-021-00373-z
Received: 28 July 2020 Accepted: 14 January 2021 Published: 03 May 2021
[FullText HTML] [PDF 223KB] Springerlink ESM
Porcine Bocavirus: A 10-Year History since Its Discovery
Manita Aryal, Guangliang Liu
doi: 10.1007/s12250-021-00365-z
Received: 04 July 2020 Accepted: 11 December 2020 Published: 28 April 2021
[FullText HTML] [PDF 507KB] Springerlink

Porcine bocavirus (PBoV) is a single-stranded DNA virus, belongs to the genus Bocaparvovirus of family Parvoviridae. It was discovered along with porcine circovirus 2 (PCV 2) and torque tenovirus (TTV) in the lymph nodes of pigs suffering from postweaning multisystemic wasting syndrome (PMWS) in Sweden in 2009. PBoV has been reported throughout the world, mostly in weaning piglets, and has a broad range of tissue tropism. Since PBoV is prevalent in healthy as well as clinically infected pigs and is mostly associated with coinfection with other viruses, the pathogenic nature of PBoV is still unclear. Currently, there are no cell lines available for the study of PBoV, and animal model experiments have not been described. This review summarizes the current state of knowledge about PBoV, including the epidemiology, evolution analysis, detection methods, pathogenesis and public health concerns.

A Virulent PEDV Strain FJzz1 with Genomic Mutations and Deletions at the High Passage Level Was Attenuated in Piglets via Serial Passage In Vitro
Pengfei Chen, Xiongwei Zhao, Shuting Zhou, Tianxing Zhou, Xiangmei Tan, Xia Wu, Wu Tong, Fei Gao, Lingxue Yu, Yifeng Jiang, Hai Yu, Zhibiao Yang, Guangzhi Tong, Yanjun Zhou
doi: 10.1007/s12250-021-00368-w
Received: 28 September 2020 Accepted: 28 December 2020 Published: 28 April 2021
[FullText HTML] [PDF 2561KB] Springerlink ESM

Highly virulent porcine epidemic diarrhea virus (PEDV) strains re-emerged and circulated in China at the end of 2010, causing significant economic losses in the pork industry worldwide. To understand the genetic dynamics of PEDV during its passage in vitro,the PEDV G2 strain FJzz1 was serially propagated in Vero cells for up to 200 passages. The susceptibility and adaptability of the FJzz1 strain increased gradually as it was serially passaged in vitro. Sequence analysis revealed that amino acid (aa) changes were mainly concentrated in the S glycoprotein,which accounted for 72.22%–85.71% of all aa changes. A continuous aa deletion (55I56G57E→55K56Δ57Δ) occurred in the N-terminal domain of S1 (S1-NTD). To examine how the aa changes affected its virulence,FJzz1-F20 and FJzz1-F200 were selected to simultaneously evaluate their pathogenicity in suckling piglets. All the piglets in the FJzz1-F20-infected group showed typical diarrhea at 24 h postinfection,and the piglets died successively by 48 h postinfection. However,the clinical signs of the piglets in the FJzz1-F200-infected group were significantly weaker,and no deaths occurred. The FJzz1-F200-infected group also showed a lower level of fecal viral shedding and lower viral loads in the intestinal tissues,and no obvious histopathological lesions. Type I and III interferon were induced in the FJzz1-F200 infection group,together with pro-inflammatory cytokines,such as TNF-α,IL-1β and IL-8. These results indicate that the identified genetic changes may contribute to the attenuation of FJzz1 strain,and the attenuated FJzz1-F200 may have the potential for developing PEDV live-attenuated vaccines.

Human Endogenous Retroviruses as Biomedicine Markers
Yuhe Song, Xiang Li, Xiaoman Wei, Jie Cui
doi: 10.1007/s12250-021-00387-7
Received: 22 August 2020 Accepted: 15 March 2021 Published: 27 April 2021
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Human endogenous retroviruses (HERVs) were formed via ancient integration of exogenous retroviruses into the human genome and are considered to be viral "fossils". The human genome is embedded with a considerable amount of HERVs, witnessing the long-term evolutionary history of the viruses and the host. Most HERVs have lost coding capability during selection but still function in terms of HERV-mediated regulation of host gene expression. In this review, we summarize the roles of HERV activation in response to viral infections and diseases, and emphasize the potential use of HERVs as biomedicine markers in the early diagnosis of diseases such as cancer, which provides a new perspective for the clinical application of HERVs.

Genetic Mutation of SARS-CoV-2 during Consecutive Passages in Permissive Cells
Ying Chen, Mei-Qin Liu, Yun Luo, Ren-Di Jiang, Hao-Rui Si, Yan Zhu, Bei Li, Xu-Rui Shen, Hao-Feng Lin, Kai Zhao, Ben Hu, Zheng-Li Shi, Xing-Lou Yang
doi: 10.1007/s12250-021-00384-w
Received: 08 December 2020 Accepted: 15 March 2021 Published: 26 April 2021
[FullText HTML] [PDF 1429KB] Springerlink ESM
Beagle Dogs Have Low Susceptibility to Florida Clade 2 H3N8 Equine Avian Influenza
Pei Zhou, Xiangyu Xiao, Xinkai Hu, Jie Dong, Haoyao Zhang, Yanchao Li, Shoujun Li
doi: 10.1007/s12250-021-00366-y
Received: 03 August 2020 Accepted: 25 January 2021 Published: 15 April 2021
[FullText HTML] [PDF 1298KB] Springerlink
Rapid Acquisition of High-Quality SARS-CoV-2 Genome via AmpliconOxford Nanopore Sequencing
Yi Yan, Ke Wu, Jun Chen, Haizhou Liu, Yi Huang, Yong Zhang, Jin Xiong, Weipeng Quan, Xin Wu, Yu Liang, Kunlun He, Zhilong Jia, Depeng Wang, Di Liu, Hongping Wei, Jianjun Chen
doi: 10.1007/s12250-021-00378-8
Received: 28 October 2020 Accepted: 18 February 2021 Published: 13 April 2021
[FullText HTML] [PDF 698KB] Springerlink ESM

Genome sequencing has shown strong capabilities in the initial stages of the COVID-19 pandemic such as pathogen identification and virus preliminary tracing. While the rapid acquisition of SARS-CoV-2 genome from clinical specimens is limited by their low nucleic acid load and the complexity of the nucleic acid background. To address this issue, we modified and evaluated an approach by utilizing SARS-CoV-2-specific amplicon amplification and Oxford Nanopore PromethION platform. This workflow started with the throat swab of the COVID-19 patient, combined reverse transcript PCR, and multi-amplification in one-step to shorten the experiment time, then can quickly and steadily obtain high-quality SARS-CoV-2 genome within 24 h. A comprehensive evaluation of the method was conducted in 42 samples: the sequencing quality of the method was correlated well with the viral load of the samples; high-quality SARS-CoV-2 genome could be obtained stably in the samples with Ct value up to 39.14; data yielding for different Ct values were assessed and the recommended sequencing time was 8 h for samples with Ct value of less than 20; variation analysis indicated that the method can detect the existing and emerging genomic mutations as well; Illumina sequencing verified that ultra-deep sequencing can greatly improve the single read error rate of Nanopore sequencing, making it as low as 0.4/10, 000 bp. In summary, high-quality SARS-CoV-2 genome can be acquired by utilizing the amplicon amplification and it is an effective method in accelerating the acquisition of genetic resources and tracking the genome diversity of SARS-CoV-2.

Recombinant GII.4[P31] Was Predominant Norovirus Circulating in Beijing Area, China, 2018–2020
Junhong Ai, Meng Zhang, Fang Jin, Zhengde Xie
doi: 10.1007/s12250-021-00381-z
Received: 28 November 2020 Accepted: 20 February 2021 Published: 09 April 2021
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Protective Efficacy of Inactivated Vaccine against SARS-CoV-2 Infection in Mice and Non-Human Primates
Yan-Feng Yao, Ze-Jun Wang, Ren-Di Jiang, Xue Hu, Hua-Jun Zhang, Yi-Wu Zhou, Ge Gao, Ying Chen, Yun Peng, Mei-Qin Liu, Ya-Nan Zhang, Juan Min, Jia Lu, Xiao-Xiao Gao, Jing Guo, Cheng Peng, Xu-Rui Shen, Qian Li, Kai Zhao, Lian Yang, Xin Wan, Bo Zhang, Wen-Hui Wang, Jia Wu, Peng Zhou, Xing-Lou Yang, Shuo Shen, Chao Shan, Zhi-Ming Yuan, Zheng-Li Shi
doi: 10.1007/s12250-021-00376-w
Received: 23 January 2021 Accepted: 08 February 2021 Published: 09 April 2021
[FullText HTML] [PDF 7920KB] Springerlink ESM

The ongoing coronavirus disease 2019 (COVID-19) pandemic caused more than 96 million infections and over 2 million deaths worldwide so far. However, there is no approved vaccine available for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the disease causative agent. Vaccine is the most effective approach to eradicate a pathogen. The tests of safety and efficacy in animals are pivotal for developing a vaccine and before the vaccine is applied to human populations. Here we evaluated the safety, immunogenicity, and efficacy of an inactivated vaccine based on the whole viral particles in human ACE2 transgenic mouse and in non-human primates. Our data showed that the inactivated vaccine successfully induced SARS-CoV-2-specific neutralizing antibodies in mice and non-human primates, and subsequently provided partial (in low dose) or full (in high dose) protection of challenge in the tested animals. In addition, passive serum transferred from vaccine-immunized mice could also provide full protection from SARS-CoV-2 infection in mice. These results warranted positive outcomes in future clinical trials in humans.

Construction of Non-infectious SARS-CoV-2 Replicons and Their Application in Drug Evaluation
Bei Wang, Chongyang Zhang, Xiaobo Lei, Lili Ren, He Huang, Jianwei Wang, Zhendong Zhao
doi: 10.1007/s12250-021-00369-9
Received: 07 December 2020 Accepted: 09 February 2021 Published: 09 April 2021
[FullText HTML] [PDF 3177KB] Springerlink ESM

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a devastating pandemic worldwide. Vaccines and antiviral drugs are the most promising candidates for combating this global epidemic, and scientists all over the world have made great efforts to this end. However, manipulation of the SARS-CoV-2 should be performed in the biosafety level 3 laboratory. This makes experiments complicated and time-consuming. Therefore, a safer system for working with this virus is urgently needed. Here, we report the construction of plasmid-based, non-infectious SARS-CoV-2 replicons with turbo-green fluorescent protein and/or firefly luciferase reporters by reverse genetics using transformation-associated recombination cloning in Saccharomyces cerevisiae. Replication of these replicons was achieved simply by direct transfection of cells with the replicon plasmids as evident by the expression of reporter genes. Using SARS-CoV-2 replicons, the inhibitory effects of E64-D and remdesivir on SARS-CoV-2 replication were confirmed, and the half-maximal effective concentration (EC50) value of remdesivir and E64-D was estimated by different quantification methods respectively, indicating that these SARS-CoV-2 replicons are useful tools for antiviral drug evaluation.

3H-31, A Non-structural Protein of Heliothis virescens ascovirus 3h, Inhibits the Host Larval Cathepsin and Chitinase Activities
Huan Yu, Yi-Yi Ou-Yang, Chang-Jin Yang, Ni Li, Madoka Nakai, Guo-Hua Huang
doi: 10.1007/s12250-021-00374-y
Received: 29 August 2020 Accepted: 16 November 2020 Published: 08 April 2021
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3h-31 of Heliothis virescens ascovirus 3h (HvAV-3h) is a highly conserved gene of ascoviruses. As an early gene of HvAV-3h, 3h-31 codes for a non-structural protein (3H-31) of HvAV-3h. In the study, 3h-31 was initially transcribed and expressed at 3 h post-infection (hpi) in the infected Spodoptera exigua fat body cells (SeFB). 3h-31 was further inserted into the bacmid of Autographa californica nucleopolyhedrovirus (AcMNPV) to generate an infectious baculovirus (AcMNPV-31). In vivo experiments showed that budded virus production and viral DNA replication decreased with the expression of 3H-31, and lucent tubular structures were found around the virogenic stroma in the AcMNPV-31-infected SeFB cells. In vivo, both LD50 and LD90 values of AcMNPV-31 were significantly higher than those of the wild-type AcMNPV (AcMNPV-wt) in third instar S. exigua larvae. An interesting finding was that the liquefaction of the larvae killed by the infection of AcMNPV-31 was delayed. Chitinase and cathepsin activities of AcMNPV-31-infected larvae were significantly lower than those of AcMNPV-wt-infected larvae. The possible regulatory function of the chitinase and cathepsin for 3H-31 was further confirmed by RNAi, which showed that larval cathepsin activity was significantly upregulated, but chitinase activity was not significantly changed due to the RNAi of 3h-31. Based on the obtained results, we assumed that the function of 3H-31 was associated with the inhibition of host larval chitinase and cathepsin activities, so as to restrain the hosts in their larval stages.

Host Interferon-Stimulated Gene 20 Inhibits Pseudorabies Virus Proliferation
Xiaoyong Chen, Dage Sun, Sujie Dong, Huanjie Zhai, Ning Kong, Hao Zheng, Wu Tong, Guoxin Li, Tongling Shan, Guangzhi Tong
doi: 10.1007/s12250-021-00380-0
Received: 29 October 2020 Accepted: 23 February 2021 Published: 08 April 2021
[FullText HTML] [PDF 1006KB] Springerlink ESM

Host interferon-stimulated gene 20 (ISG20) exerts antiviral effects on viruses by degrading viral RNA or by enhancing IFN signaling. Here, we examined the role of ISG20 during pseudorabies virus (PRV) proliferation. We found that ISG20 modulates PRV replication by enhancing IFN signaling. Further, ISG20 expression was upregulated following PRV infection and poly(I: C) treatment. Ectopic expression of ISG20 inhibited PRV proliferation in PK15 cells, whereas knockdown of ISG20 promoted PRV proliferation. In addition, ISG20 expression upregulated IFN-β expression and enhanced IFN downstream signaling during PRV infection. Notably, PRV UL24 suppressed the transcription of ISG20, thus antagonizing its antiviral effect. Further domain mapping analysis showed that the N terminus (amino acids 1-90) of UL24 was responsible for the inhibition of ISG20 transcription. Collectively, these findings characterize the role of ISG20 in suppressing PRV replication and increase the understanding of host-PRV interplay.

Stability of SARS-CoV-2 on the Surfaces of Three Meats in the Setting That Simulates the Cold Chain Transportation
Xiao-Li Feng, Bei Li, Hao-Feng Lin, Hong-Yi Zheng, Ren-Rong Tian, Rong-Hua Luo, Mei-Qin Liu, Ren-Di Jiang, Yong-Tang Zheng, Zheng-Li Shi, Yu-Hai Bi, Xing-Lou Yang
doi: 10.1007/s12250-021-00367-x
Received: 09 October 2020 Accepted: 01 February 2021 Published: 08 April 2021
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Ozone Water Is an Effective Disinfectant for SARS-CoV-2
Xiao Hu, Zhen Chen, Zhengyuan Su, Fei Deng, Xinwen Chen, Qi Yang, Pan Li, Quanjiao Chen, Jun Ma, Wuxiang Guan, Rongjuan Pei, Yun Wang
doi: 10.1007/s12250-021-00379-7
Received: 13 January 2021 Accepted: 23 February 2021 Published: 31 March 2021
[FullText HTML] [PDF 428KB] Springerlink
Human Endogenous Retrovirus Type W Envelope from Multiple Sclerosis Demyelinating Lesions Shows Unique Solubility and Antigenic Characteristics
Benjamin Charvet, Justine Pierquin, Joanna Brunel, Rianne Gorter, Christophe Quétard, Branka Horvat, Sandra Amor, Jacques Portoukalian, Hervé Perron
doi: 10.1007/s12250-021-00372-0
Received: 02 February 2021 Accepted: 08 February 2021 Published: 26 March 2021
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In multiple sclerosis (MS), human endogenous retrovirus W family (HERV-W) envelope protein, pHERV-W ENV, limits remyelination and induces microglia-mediated neurodegeneration. To better understand its role, we examined the soluble pHERV-W antigen from MS brain lesions detected by specific antibodies. Physico-chemical and antigenic characteristics confirmed differences between pHERV-W ENV and syncytin-1. pHERV-W ENV monomers and trimers remained associated with membranes, while hexamers self-assembled from monomers into a soluble macrostructure involving sulfatides in MS brain. Extracellular hexamers are stabilized by internal hydrophobic bonds and external hydrophilic moieties. HERV-W studies in MS also suggest that this diffusible antigen may correspond to a previously described high-molecular-weight neurotoxic factor secreted by MS B-cells and thus represents a major agonist in MS pathogenesis. Adapted methods are now needed to identify encoding HERV provirus(es) in affected cells DNA. The properties and origin of MS brain pHERV-W ENV soluble antigen will allow a better understanding of the role of HERVs in MS pathogenesis. The present results anyhow pave the way to an accurate detection of the different forms of pHERV-W ENV antigen with appropriate conditions that remained unseen until now.

Role of Intracellular Distribution of Feline and Bovine SAMHD1 Proteins in Lentiviral Restriction
Chu Wang, Lina Meng, Jialin Wang, Kaikai Zhang, Sizhu Duan, Pengyu Ren, Yingzhe Wei, Xinyu Fu, Bin Yu, Jiaxin Wu, Xianghui Yu
doi: 10.1007/s12250-021-00351-5
Received: 03 August 2020 Accepted: 28 December 2020 Published: 22 March 2021
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Human SAMHD1 (hSAM) restricts lentiviruses at the reverse transcription step through its dNTP triphosphohydrolase (dNTPase) activity. Besides humans, several mammalian species such as cats and cows that carry their own lentiviruses also express SAMHD1. However, the intracellular distribution of feline and bovine SAMHD1 (fSAM and bSAM) and its significance in their lentiviral restriction function is not known. Here, we demonstrated that fSAM and bSAM were both predominantly localized to the nucleus and nuclear localization signal (11KRPR14)-deleted fSAM and bSAM relocalized to the cytoplasm. Both cytoplasmic fSAM and bSAM retained the antiviral function against different lentiviruses and cytoplasmic fSAM could restrict Vpx-encoding SIV and HIV-2 more efficiently than its wild-type (WT) protein as cytoplasmic hSAM. Further investigation revealed that cytoplasmic fSAM was resistant to Vpx-induced degradation like cytoplasmic hSAM, while cytoplasmic bSAM was not, but they all demonstrated the same in vitro dNTPase activity and all could interact with Vpx as their WT proteins, indicating that cytoplasmic hSAM and fSAM can suppress more SIV and HIV-2 by being less sensitive to Vpx-mediated degradation. Our results suggested that fSAM- and bSAM-mediated lentiviral restriction does not require their nuclear localization and that fSAM shares more common features with hSAM. These findings may provide insights for the establishment of alternative animal models to study SAMHD1 in vivo.

Significant Inhibition of Porcine Epidemic Diarrhea Virus In Vitro by Remdesivir, Its Parent Nucleoside and β-D-N4-hydroxycytidine
Yuanchao Xie, Xiaozhen Guo, Tianwen Hu, Daibao Wei, Xiuli Ma, Jiaqiang Wu, Bing Huang, Jingshan Shen
doi: 10.1007/s12250-021-00362-2
Received: 15 October 2020 Accepted: 22 January 2021 Published: 22 March 2021
[FullText HTML] [PDF 3152KB] Springerlink

Porcine epidemic diarrhea (PED) caused by porcine epidemic diarrhea virus (PEDV) is widespread in the world. In recent years, the increased virulence of the virus due to viral variations, has caused great economic losses to the pig industry in many countries. It is always worthy to find effective therapeutic methods for PED. As an important class of antivirals, nucleoside drugs which target viral polymerases have been applied in treating human viral infections for half a century. Herein, we evaluated the anti-PEDV potential of three broad-spectrum antiviral nucleoside analogs, remdesivir (RDV), its parent nucleoside (RDV-N) and β-D-N4-hydroxycytidine (NHC). Among them, RDV-N was the most active agent in Vero E6 cells with EC50 of 0.31 μmol/L, and more potent than RDV (EC50 = 0.74 μmol/L) and NHC (EC50 = 1.17 μmol/L). The activity of RDV-N was further confirmed using an indirect immuno-fluorescence assay. Moreover, RDV-N exhibited a good safety profile in cells and in mice. The high sequence similarity of the polymerase functional domains of PEDV with other five porcine coronaviruses indicated a broader antiviral spectrum for the three compounds. Generally, RDV-N is a promising broad-spectrum antiviral nucleoside, and it would be worthy to make some structural modifications to increase its oral bioavailability.

Porcine Picornavirus 3C Protease Degrades PRDX6 to Impair PRDX6-mediated Antiviral Function
Congcong Wang, Huanhuan Feng, Xiangle Zhang, Kangli Li, Fan Yang, Weijun Cao, Huisheng Liu, Lili Gao, Zhaoning Xue, Xiangtao Liu, Zixiang Zhu, Haixue Zheng
doi: 10.1007/s12250-021-00352-4
Received: 30 September 2020 Accepted: 17 December 2020 Published: 15 March 2021
[Abstract] [PDF 918KB] Springerlink ESM
Peroxiredoxin-6 (PRDX6) is an antioxidant enzyme with both the activities of peroxidase and phospholipase A2 (PLA2), which is involved in regulation of many cellular reactions. However, the function of PRDX6 during virus infection remains unknown. In this study, we found that the abundance of PRDX6 protein was dramatically decreased in foot-and-mouth disease virus (FMDV) infected cells. Overexpression of PRDX6 inhibited FMDV replication. In contrast, knockdown of PRDX6 expression promoted FMDV replication, suggesting an antiviral role of PRDX6. To explore whether the activity of peroxidase and PLA2 was associated with PRDX6-mediated antiviral function, a specific inhibitor of PLA2 (MJ33) and a specific inhibitor of peroxidase activity (mercaptosuccinate) were used to treat the cells before FMDV infection. The results showed that incubation of MJ33 but not mercaptosuccinate promoted FMDV replication. Meanwhile, overexpression of PRDX6 slightly enhanced type I interferon signaling. We further determined that the viral 3Cpro was responsible for degradation of PRDX6, and 3Cpro-induced reduction of PRDX6 was independent of the proteasome, lysosome, and caspase pathways. The protease activity of 3Cpro was required for induction of PRDX6 reduction. Besides, PRDX6 suppressed the replication of another porcine picornavirus Senecavirus A (SVA), and the 3Cpro of SVA induced the reduction of PRDX6 through its proteolytic activity as well. Together, our results suggested that PRDX6 plays an important antiviral role during porcine picornavirus infection, and the viral 3Cpro induces the degradation of PRDX6 to overcome PRDX6-mediated antiviral function.
Autographa Californica Multiple Nucleopolyhedrovirus orf13 Is Required for Efficient Nuclear Egress of Nucleocapsids
Xingang Chen, Xiaoqin Yang, Chengfeng Lei, Fujun Qin, Xiulian Sun, Jia Hu
doi: 10.1007/s12250-021-00353-3
Received: 20 October 2020 Accepted: 18 December 2020 Published: 15 March 2021
[Abstract] [PDF 2841KB] Springerlink ESM
Autographa californica multiple nucleopolyhedrovirus (AcMNPV) orf13 (ac13) is a conserved gene in all sequenced alphabaculoviruses. However, its function in the viral life cycle remains unknown. In this study, we found that ac13 was a late gene and that the encoded protein, bearing a putative nuclear localization signal motif, colocalized with the nuclear lamina. Deletion of ac13 did not affect viral genome replication, nucleocapsid assembly or occlusion body (OB) formation, but reduced virion budding from infected cells by approximately 400-fold compared with the wild-type virus. Deletion of ac13 substantially impaired the egress of nucleocapsids from the nucleus to the cytoplasm, while the OB morphogenesis was unaffected. Taken together, our results indicated that ac13 was required for efficient nuclear egress of nucleocapsids during virion budding, but was dispensable for OB formation.
Porcine Coronaviruses: Overview of the State of the Art
Hanna Turlewicz-Podbielska, Małgorzata Pomorska-Mól
doi: 10.1007/s12250-021-00364-0
Received: 26 May 2020 Accepted: 19 November 2020 Published: 15 March 2021
[Abstract] [PDF 1704KB] Springerlink
Like RNA viruses in general, coronaviruses (CoV) exhibit high mutation rates which, in combination with their strong tendency to recombine, enable them to overcome the host species barrier and adapt to new hosts. It is currently known that six CoV are able to infect pigs. Four of them belong to the genus Alphacoronavirus [transmissible gastroenteritis coronavirus (TEGV), porcine respiratory coronavirus (PRCV), porcine epidemic diarrhea virus (PEDV), swine acute diarrhea syndrome coronavirus (SADS-CoV)], one of them to the genus Betacoronavirus [porcine hemagglutinating encephalomyelitis virus (PHEV)] and the last one to the genus Deltacoronavirus (PDCoV). PHEV was one of the first identified swine CoV and is still widespread, causing subclinical infections in pigs in several countries. PRCV, a spike deletion mutant of TGEV associated with respiratory tract infection, appeared in the 1980s. PRCV is considered non-pathogenic since its infection course is mild or subclinical. Since its appearance, pig populations have become immune to both PRCV and TGEV, leading to a significant reduction in the clinical and economic importance of TGEV. TGEV, PEDV and PDCoV are enteropathogenic CoV and cause clinically indistinguishable acute gastroenteritis in all age groups of pigs. PDCoV and SADS-CoV have emerged in 2014 (US) and in 2017 (China), respectively. Rapid diagnosis is crucial for controlling CoV infections and preventing them from spreading. Since vaccines are available only for some porcine CoV, prevention should focus mainly on a high level of biosecurity. In view of the diversity of CoV and the potential risk factors associated with zoonotic emergence, updating the knowledge concerning this area is essential.
Establishment of a Reverse Genetic System of Severe Fever with Thrombocytopenia Syndrome Virus Based on a C4 Strain
Mingyue Xu, Bo Wang, Fei Deng, Hualin Wang, Manli Wang, Zhihong Hu, Jia Liu
doi: 10.1007/s12250-021-00359-x
Received: 18 December 2020 Accepted: 21 January 2021 Published: 15 March 2021
[Abstract] [PDF 795KB] Springerlink ESM
Severe fever with thrombocytopenia syndrome virus (SFTSV) is an emerging tick-borne bunyavirus that causes hemorrhagic fever-like disease (SFTS) in humans with a case fatality rate up to 30%. To date, the molecular biology involved in SFTSV infection remains obscure. There are seven major genotypes of SFTSV (C1–C4 and J1–J3) and previously a reverse genetic system was established on a C3 strain of SFTSV. Here, we reported successfully establishment of a reverse genetics system based on a SFTSV C4 strain. First, we obtained the 5′- and 3′-terminal untranslated region (UTR) sequences of the Large (L), Medium (M) and Small (S) segments of a laboratory-adapted SFTSV C4 strain through rapid amplification of cDNA ends analysis, and developed functional T7 polymerase-based L-, M- and S-segment minigenome assays. Then, full-length cDNA clones were constructed and infectious SFTSV were recovered from co-transfected cells. Viral infectivity, growth kinetics, and viral protein expression profile of the rescued virus were compared with the laboratory-adapted virus. Focus formation assay showed that the size and morphology of the foci formed by the rescued SFTSV were indistinguishable with the laboratory-adapted virus. However, one-step growth curve and nucleoprotein expression analyses revealed the rescued virus replicated less efficiently than the laboratory-adapted virus. Sequence analysis indicated that the difference may be due to the mutations in the laboratory-adapted strain which are more prone to cell culture. The results help us to understand the molecular biology of SFTSV, and provide a useful tool for developing vaccines and antivirals against SFTS.
Sustainability of SARS-CoV-2 Induced Humoral Immune Responses in COVID-19 Patients from Hospitalization to Convalescence Over Six Months
Yang Zheng, Qing Zhang, Ashaq Ali, Ke Li, Nan Shao, Xiaoli Zhou, Zhiqin Ye, Xiaomin Chen, Shanshan Cao, Jing Cui, Juan Zhou, Dianbing Wang, Baidong Hou, Min Li, Mengmeng Cui, Lihua Deng, Xinyi Sun, Qian Zhang, Qinfang Yang, Yong li, Hui Wang, Yake Lei, Bo Yu, Yegang Cheng, Xiaolin Tong, Dong Men, Xian-En Zhang
doi: 10.1007/s12250-021-00360-4
Received: 24 November 2020 Accepted: 13 January 2021 Published: 04 March 2021
[Abstract] [PDF 2768KB] Springerlink ESM
Understanding the persistence of antibody in convalescent COVID-19 patients may help to answer the current major concerns such as the risk of reinfection, the protection period of vaccination and the possibility of building an active herd immunity. This retrospective cohort study included 172 COVID-19 patients who were hospitalized in Wuhan. A total of 404 serum samples were obtained over six months from hospitalization to convalescence. Antibodies in the specimens were quantitatively analyzed by the capture chemiluminescence immunoassays (CLIA). All patients were positive for the anti-SARS-CoV-2 IgM/IgG at the onset of COVID-19 symptoms, and the IgG antibody persisted in all the patients during the convalescence. However, only approximately 25% of patients can detect the IgM antibodies, IgM against N protein (N-IgM) and receptor binding domain of S protein (RBD-IgM) at the 27th week. The titers of IgM, N-IgM and RBD-IgM reduced to 16.7%, 17.6% and 15.2% of their peak values respectively. In contrast, the titers of IgG, N-IgG and RBD-IgG peaked at 4–5th week and reduced to 85.9%, 62.6% and 87.2% of their peak values respectively at the end of observation. Dynamic behavior of antibodies and their correlation in age, gender and severity groups were investigated. In general, the COVID-19 antibody was sustained at high levels for over six months in most of the convalescent patients. Only a few patients with antibody reducing to an undetectable level which needs further attention. The humoral immune response against SARS-CoV-2 infection in COVID-19 patients exhibits a typical dynamic of acquired immunity.
Alterations in Phenotypes and Responses of T Cells Within 6 Months of Recovery from COVID-19: A Cohort Study
Bali Zhao, Maohua Zhong, Qingyu Yang, Ke Hong, Jianbo Xia, Xia Li, Ying Liu, Yao-Qing Chen, Jingyi Yang, Chaolin Huang, Huimin Yan
doi: 10.1007/s12250-021-00348-0
Received: 11 October 2020 Accepted: 30 November 2020 Published: 09 February 2021
[Abstract] [PDF 2088KB] Springerlink ESM
The COVID-19 pandemic, caused by the SARS-CoV-2 infection, is a global health crisis. While many patients have clinically recovered, little is known about long-term alterations in T cell responses of COVID-19 convalescents. In this study, T cell responses in peripheral blood mononuclear cells of a long-time COVID-19 clinically recovered (20–26 weeks) cohort (LCR) were measured via flow cytometry and ELISpot. The T cell responses of LCR were comparatively analyzed against an age and sex matched short-time clinically recovered (4–9 weeks) cohort (SCR) and a healthy donor cohort (HD). All volunteers were recruited from Wuhan Jinyintan Hospital, China. Phenotypic analysis showed that activation marker PD-1 expressing on CD4+ T cells of LCR was still significantly lower than that of HD. Functional analysis indicated that frequencies of Tc2, Th2 and Th17 in LCR were comparable to those of HD, but Tc17 was higher than that of HD. In LCR, compared to the HD, there were fewer IFN-γ producing T cells but more IL-2 secreting T cells. In addition, the circulating Tfh cells in LCR were still slightly lower compared to HD, though the subsets composition had recovered. Remarkably, SARS-CoV-2 specific T cell responses in LCR were comparable to that of SCR. Collectively, T cell responses experienced long-term alterations in phenotype and functional potential of LCR cohort. However, after clinical recovery, SARS-CoV-2 specific T cell responses could be sustained at least for six months, which may be helpful in resisting re-infection.