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Human noroviruses (HuNoVs) are a major cause of epidemic and sporadic cases of acute gastroenteritis worldwide. Recently, human intestinal enteroids (HIEs) have been shown to support the replication of HuNoVs, and be an excellent model to study HuNoV-host interactions. We implemented the HIE system in our laboratory and investigated the global molecular events associated with the mechanism of HuNoV-host interactions. Successful replication was observed for several norovirus GII genotypes, and totally 5,376 genes with different expression in HIEs were identified during infection. Bioinformatics analysis revealed that several important pathways, especially the "Signal transduction" and "Immune system" pathways, were involved in the HuNoV-host interaction. Quantitative PCR results validated that IFN-λ instead of IFN-β was elevated in HIEs after infection. Our study showed the holistic understanding of the transcriptome events in the HIE model infected by HuNoVs, and highlighted the important role of IFN-λ signaling in the HuNoV-host interactions., Competing Interests: None.

Objective: Study on the antiviral effect in vivo and in vitro of recombinant antibodies with neutralization activity to influenza virus HA antigen, IgG-IV-2 and IgG-IV-6 obtained from expression in Baculo/insert system., Methods: Viral titer of influenza virus in MDCK was compared before and after antibody application, and measured viral titer in mouse lung before and after antibodies applied on mucous membrane., Results: IgG-IV-2 and IgG-IV-6 could reduce the titer from 4.5 log10 TCID50 to half is by 50% at doses 0.8 microg and 0.5 microg respectively. When recombinant antibodies were used on mucous membrane, the IgG-IV-2 and IgG-IV-6 reduced the titer by half at doses 0.25 mg/kg weight and 0.1 mg/kg weight respectively. The dose was 0.08 mg/kg weight when the two antibodies were jointly used., Conclusions: The recombinant antibodies have antiviral effect in vivo and in vitro, they can neutralize viral virulence.

Objective: To increase prokaryotic expression level of IFN-alpha 1C gene through the quantitative theory of translational control and the translational enhancer sequence., Methods: Stepwise polymerase chain reaction (PCR) was used to alter the 5 terminal cDNA sequence of IFN-alpha 1C in three different grades of base mutation. In this way, the free energy (Delta G) of the secondary structure in translational initiation region (TIR) was decreased gradually. An expression plasmid (pBVE) was constructed to contain the translational enhancer cDNA sequence by modifying pBV220 upstream of the SD region., Results: The expression levels of three kinds of IFN-alpha 1C modified gene were all increased. Furthermore, it presented an increasing trend with decreasing in delta G varying from -50,241.6 to -22,190.0 J/mol. The highest expression was 2.43 x 10(8) U/L, covering twelve times more than its original cDNA. IFN-alpha 1C gene and its modified cDNA was inserted into pBVE as reporting genes. E.Coli cells harbouring pBVE/IFN-alpha 1Cs cDNA produced two to five times more IFN than cells harbouring pBV220/IFN-alpha 1Cs., Conclusions: pBVE containing translational enhancer is a high level prokaryotic expression vector. The theory of quantitative translational control can effectively be used to enhance the IFN-alpha 1C gene expression level in E.coli.

Objective: Constructing a plasmid containing the shRNA of luciferase to suppress the expression of luciferase in BHK-21 cell., Methods: A 334 bp human U6 snRNA promoter was amplified from human genomic DNA by PCR and ligated to a 21 bp reverse repeated motif of luciferase target sequence with 9 bp spacer and AAV plasmid pSNAV. The recombinant pSNAV/U6/Luc plasmid cotransfected with pMAMneoLuc or transfected luciferase cell line to detect the effect of luciferase expression separately., Results: pSNAV/U6/luc suppresses the luciferase expression from pMAMneoLuc by 50% and luciferase cell line by 70%., Conclusions: The results showed that the short hairpin RNA of luciferase can efficiently suppress its expression in BHK-21.

Background: Based on the earlier mapping of the epitope recognized by neutralizing antibody, the authors directly replaced binding domain of IFN in AB-loop for enhancement of biological activity., Methods: Two unique restriction sites (EcoR? and BsE?) were created into region flanking LoopAB. Casette mutagesis, restriction enzyme digestion, DNA sequencing, antiviral activity assay and antiproliferative activity assay have been used in the project., Results: The mutated residues M31?D,D32?P of LoopAB in parent IFN were produced. The recombinant phagemid pCANTAB5E/3132IFN 1c/86D and expression plasmid PBV322-132IFN 1c/86D were constructed respectively by replacing the corresponding LoopAB with DNA fragment mutated in the residues M31?D,D32?P, which have been confirmed. The recombinant protein has been expressed in E.coli JM103. The crude 3132IFN 1c/86D has been assayed on human WISH cells challenged with VSV and on HeLa cells by colorimetric MTT. 3132IFN 1c/86D showed 8-old antiviral activity compared to that of parent IFN 1c/86D, while IFN?induced growth inhibition of both types had no difference., Conclusions: The authors concluded that a mutant IFN with enhanced antiviral activity can be obtained via a targeted replacement of receptor binding domain in AB-loop.

Background: To construct a series of recombinant herpes simplex viruses that can provide replicating and packaging functions for recombinant adeno-associated virus (rAAV), and to select the strains possessing stronger functions for large-scale production of rAAV., Methods: A set of cosmids that represents the whole genome of HSV-1 was used to generate recombinant HSV-1 expressing rep and cap proteins of AAV-2. An ATG-to-ACG mutation in the start codon of AAV-2 rep protein was generated by site-directed mutagenesis. rep and cap genes, under control of their native promoters, with or without the ATG-to-ACG mutation in the start codon of rep, were inserted into the Xba site of UL2 or UL44 genes, respectively, resulting in the recombinant cosmids cos6-rmc/ UL2, cos56-rc/ UL44 and cos56-rmc/ UL44. Homologous recombination among the HSV-1 fragment on the recombinant cosmids and the rest fragments of HSV-1 genome resulted in three strains of recombinant HSV-1. Together with the one was constructed previously, there were four strains of recombinant HSV-1named HSV1-rc/ UL2, HSV1-rmc/ UL2, HSV1-rc/ UL44 and HSV1-rmc/ UL44 respectively., Results: PCR detection confirmed the existence of rep- gene in the genomes of all four strains of the recombinant HSV-1. Recombinant AAV was produced after infecting the AAV vector cell line that carrying the GFP expression cassette with the four strains of recombinant HSV-1 respectively. However, HSV1-rc/ UL2 and HSV1-rmc/ UL2 produced much more rAAV than HSV1-rc/ UL44 and HSV1c/ UL44 did., Conclusions: All the four strains of recombinant HSV-1 support rAAV replication and packaging. HSV1 UL2 and HSV1-rmc/ UL2 that provide much stronger functions may be useful for large-scale production of rAAV.