Many flaviviruses are emerging and reemerging pathogens, such as West Nile virus (WNV), dengue virus (DENV), yellow fever virus (YFV), and Japanese encephalitis virus. Serological assay is the dominant method for diagnosis of flavivirus infections in human. Because antibodies generated during flavivirus infections cross-react with other flavivirus members, plaque reduction neutralization test (PRNT) is the only available assay to determine the infecting flavivirus type.Since PRNT requires culturing raw viruses, it must be performed in biosafety level-3 or level-4 containment for many flaviviruses, and takes more than ten days to complete. To overcome these problems, we have developed flavivirus viral-like particles (VLPs) that could be used to replace raw viruses in the neutralization assay. The VLPs were prepared by trans packaging a luciferase-reporting replicon with viral structural proteins. This novel assay involves three simple steps: (ⅰ) VLPs from a panel of flaviviruses are incubated with flavivirus-infected sera at 37℃ for 1 h; (ⅱ)the neutralized VLPs are used to infect Vero cells; and (ⅲ) the infected cells are measured for luciferase activities at 22 h post-infection. The virus type whose VLP is most efficiently neutralized by the serum specimen (as quantified by the luciferase activities) is the etiologic agent. As a proof-of-concept, we show that a WNV-infected mouse serum neutralized the WNV VLP more efficiently and selectively than the DENV and YFV VLPs. Our results demonstrate that the VLP neutralization assay maintains the "gold standard" of the classic PRNT; importantly, it shortens the assay time from ＞10 days to ＜1 day, and can be performed in biosafety level-2 facility.

Dengue virus (DENV) nonstructural protein 1 (NS1) is a highly conserved 46-kDa protein that contains 2 glycosylation sites (Asn-130 and Asn-207) and 12 conserved cysteine (Cys) residues. Here, we performed site-directed mutagenesis to generate systematic mutants of viral strain TSV01. The results of the subsequent analysis showed that an alanine substitution at the second N-linked glycan Asn-207 in NS1 delayed viral RNA synthesis, reduced virus plaque size, and weakened the cytopathic effect. Three mutants at Cys sites (Cys-4, Cys-55, Cys-291) and a C-terminal deletion (ΔC) mutant signiifcantly impaired RNA synthesis, and consequently abolished viral growth, whereas alanine mutations at Asn-130 and Glu-173 resulted in phenotypes that were similar to the wild-type (WT) virus. Further analysis showed that the Asn-207 mutation slightly delayed viral replication. These results suggest that the three conserved disulifde bonds and the second N-linked glycan in NS1 are required for DENV-2 replication.

ObjectiveTo study the effect of comprehensive rehabilitation therapy on activity of daily living(ADL) of old patients with stroke.Methods56 old patients with stroke were rehabilitated by the therapy of Bobath,Brunnstrom,Rood,PNF and psychotherapy for 3 months.The Modified Barthel Index (MBI) was applied to measure ADL before and after treatment.ResultsAfter treatment,the MBI of the patients was markedly higher than before(P<0.01),which was relative with the recovery of movement function and the psychology.ConclusionsThe rehabilitation therapy can improve ADL of old patients on stroke.

As a member of T-box family,Tbx3 (T-box3) is widely expressed in multiple organs and involves in the development of breast,limb,heart,eye,lung and pancreas during embryonic development stage.Moreover,Tbx3 plays an important role in maintaining the embryonic stem cells.Recent studies show that Tbx3 can work as a transcription factor to participate in the initiation and progression of tumor by epithelialmesenchymal transition,induction tumor stemness and methylation.A better understanding of Tbx3 will provide new insights into genetic diagnosis and targeted treatment of tumor.

The naturally occuring antibiotic resistance in twentyseven strains from 7 different serotypes of Bacillus sphaericus was investigated. The antibiotics used in this experiment are Streptomycin, Ampicilline, Tetracycline, Chloramphenicol, Kanamycin, Erythromycin, Qingdamycin, Midecamycin, Ribostamycin, Acriflavin, Rifampicin, Sulphanilamide and Azide. All strains are resist to streptomycin and azide at higher concentration and chloramphenicol at lower concentration. Most of strains are resist to tetracycline at lower concentration.

A bacterial cell surface display technique based on an ice nucleation protein has been employed for the development of live vaccine against viral infection.Due to its ubiquitous ability to invade host cells,Salmonella typhimurium might be a good candidate for displaying viral antigens.We demonstrated the surface display of domain III of Japanese encephalitis virus E protein and the enhanced green fluorescent protein on S.typhimurium BRD509 using the ice nucleation protein.The effects of the motif in the ice nucleation protein on the effective display of integral protein were also investigated.The results showed that display motifs in the protein can target integral foreign protein on the surface of S.typhimurium BRD509.Moreover,recombinant strains with surface displayed viral proteins retained their invasiveness,suggesting that the recombinant S.typhimurium can be used as live vaccine vector for eliciting complete immunogenicity.The data may yield better understanding of the mechanism by which ice nucleation protein displays foreign proteins in the Salmonella strain.