In the present study, 89 porcine reproductive and respiratory syndrome virus (PRRSV) isolates in China during 2007 to 2012 were randomly selected from the GenBank genetic sequence database. Evolutionary characteristics of these isolates were analyzed based on the sequences of non-struc-tural protein 2 (Nsp2) and glycoprotein 5 (GP5). The genetic variations of the isolates were also compared with six representative strains. The results showed that a high degree of genetic diversity exists among the PRRSV population in China. Highly pathogenic PRRSV isolates, with a discon-tinuous deletion of a 30 amino acid residue in the Nsp2 region, remained the most dominant virus throughout 2007-2012 in China. Owing to the extensive use of representative vaccine strains, natu-ral recombination events occurred between strains. Three isolates-HH08, DY, and YN-2011-were more closely related to vaccine strains than the other isolates. Both YN-2011 and DY were the evolu-tionary products of recombination events between strains SP and CH-1R. The results of the present study provide useful information for the epidemiology of PRRSV as well as for vaccine development.

Sweepovirus is an important monopartite begomovirus that infects plants of the genus Ipomoea worldwide. Development of artiifcial infection methods for sweepovirus using agroinoculation is a highly efficient means of studying infectivity in sweet potato. Unlike other begomoviruses, it has proven difficult to infect sweet potato plants with sweepoviruses using infectious clones. A novel sweepovirus, called Sweet potato leaf curl virus-Jiangsu (SPLCV-JS), was recently identiifed in China. In addition, the infectivity of the SPLCV-JS clone has been demonstrated in Nicotiana benthamiana. Here we describe the agroinfection of the sweet potato cultivar Xushu 22 with the SPLCV-JS infectious clone using vacuum inifltration. Yellowing symptoms were observed in newly emerged leaves. Molecular analysis confirmed successful inoculation by the detection of viral DNA. A synergistic effect of SPLCV-JS and the heterologous betasatellite DNA-βof Tomato yellow leaf curl China virus isolate Y10 (TYLCCNV-Y10) on enhanced symptom severity and viral DNA accumulation was conifrmed. The development of a routine agroinoculation system in sweet potato with SPLCV-JS using vacuum inifltration should facilitate the molecular study of sweepovirus in this host and permit the evaluation of virus resistance of sweet potato plants in breeding programs.

VC2002, isolated from postweaning multisystemic wasting syndrome (PMWS)-affected pig, is a mixture of two porcine circovirus genotype 2b (PCV2b) viruses, K2 and K39. Preliminary experiments disclosed short-term adverse effects of K39, but not K2, on porcine foetuses. These findings led to the hypothesis that infection of immuno-incompetent foetuses with K2 confers a status of immunotolerance, and postnatal super-infection with K39 triggers PMWS. To explore this hypothesis, nine 55-day-old foetuses were inoculated in utero (three with K2-104.3TCID50, three with K39-104.3TCID50 and three with medium), and foeto-pathogenicity examined. At 21 days post-inoculation (dpi), K2 did not induce pathology, whereas pathological effects of K39 were evident. Twenty-four 45-day-old foetuses were subsequently inoculated to examine the long-term effect of K2, including six with K2-high dose-104.3TCID50, six with K2-low dose-102.3TCID50 and 12 mock-inoculated controls. Both doses resulted in ifve mummiifed foetuses and one live-born piglet each (69dpi). K2 was recovered from all mummies. K2 and K2-speciifc antibodies were not detected in serum of the two live-born piglets at birth, indicating full control of K2 infection. The K2-low dose-infected piglet was immunostimulated at day 2, but not the K2-high dose-infected piglet. Both non-stimulated and stimulated K2-infected piglets were super-inoculated with K39 at day 6 or 8 (taken as 0 days post super-inoculation). Low viral replication was observed in the non-stimulated K2-K39 piglet (up to 103.3 TCID50/g;identiifed as K39). In contrast, viral replication was extremely high in the stimulated K2-K39 piglet (up to 105.6TCID50/g) and identiifed as K2, indicating that K2 infection is controlled during foetal life, but emerges after birth upon immunostimulation. However, none of the piglets showed any signs of PMWS.

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.

Tobacco mosaic virus (TMV) causes significant yield loss in susceptible crops irrigated with contaminated water. However, detection of TMV in water is difficult owing to extremely low concentrations of the virus. Here, we developed a simple method for the detection and quantiifcation of TMV in irrigation water. TMV was reliably detected at concentrations as low as 10 viral copies/μL with real-time PCR. The sensitivity of detection was further improved using polyethylene glycol 6000 (PEG6000, MW 6000) to concentrate TMV from water samples. Among the 28 samples from Shaanxi Province examined with our method, 17 were tested positive after virus concentration. Infectivity of TMV in the original water sample as well as after concentration was conifrmed using PCR. The limiting concentration of TMV in water to re-infect plants was determined as 102 viral copies/mL. The method developed in this study offers a novel approach to detect TMV in irrigation water, and may provide an effective tool to control crop infection.

As a fundamental component of the host cellular cytoskeleton, actin is routinely engaged by infecting viruses. Furthermore, viruses from diverse groups, and infecting diverse hosts, have convergently evolved an array of mechanisms for manipulating the actin cytoskeleton for efifcacious infection. An ongoing chorus of research now indicates that the actin cytoskeleton is critical for viral replication at many stages of the viral life cycle, including binding, entry, nuclear localization, genomic transcription and reverse transcription, assembly, and egress/dissemination. Speciifcally, viruses subvert the force-generating and macromolecular scaffolding properties of the actin cytoskeleton to propel viral surifng, internalization, and migration within the cell. Additionally, viruses utilize the actin cytoskeleton to support and organize assembly sites, and eject budding virions for cell-to-cell transmission. It is the purpose of this review to provide an overview of current research, focusing on the various mechanisms and themes of virus-mediated actin modulation described therein.

A novel avian influenza A (H7N9) virus recently emerged in the Yangtze River delta and caused diseases, often severe, in over 130 people. This H7N9 virus appeared to infect humans with greater ease than previous avian inlfuenza virus subtypes such as H5N1 and H9N2. While there are other potential explanations for this large number of human infections with an avian influenza virus, we investigated whether a lack of conserved T-cell epitopes between endemic H1N1 and H3N2 inlfuenza viruses and the novel H7N9 virus contributes to this observation. Here we demonstrate that a number of T cell epitopes are conserved between endemic H1N1 and H3N2 viruses and H7N9 virus. Most of these conserved epitopes are from viral internal proteins. The extent of conservation between endemic human seasonal inlfuenza and avian inlfuenza H7N9 was comparable to that with the highly pathogenic avian inlfuenza H5N1. Thus, the ease of inter-species transmission of H7N9 viruses (compared with avian H5N1 viruses) cannot be attributed to the lack of conservation of such T cell epitopes. On the contrary, our ifndings predict signiifcant T-cell based cross-reactions in the human population to the novel H7N9 virus. Our findings also have implications for H7N9 virus vaccine design.

Enterovirus 71 (EV71) is a member of the Entero-virus genus of the Picornaviridae family and is the major cause of Hand,foot,and mouth disease (HFMD) in children.Different strains from Gansu were cloned and the P1 protein was sequenced and analysed.Results indicate that there are three kinds of EV71 infections prevalent in Gansu.The VP1 protein from one of these strains,55F,was expressed.The recombinant protein was expressed with high level and reacted specifically with the EV71 patient antibody,the recombinant protein was also applied to raise antiserum in rabbits and after the fourth injection a high titer of antiserum was detected by ELISA assay.These data are useful for further clarification of prevalent EV71 strains in the north of China at the molecular level and provide a basis for EV71 diagnosis.

Protamines are a group of highly basic proteins first discovered in spermatozoon that allow for denser packaging of DNA than histones and will result in down-regulation of gene transcription[1].It is well recognized that the Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV) encodes P6.9,a protamine-like protein that forms the viral subnucleosome through binding to the viral genome[29].Previous research demonstrates that P6.9 is essential for viral nucleocapsid assembly,while it has no influence on viral genome replication[31].In the present study,the role of P6.9 in viral gene transcription regulation is characterized.In contrast to protamines or other protamine-like proteins that usually down-regulate gene transcription,P6.9 appears to up-regulate viral gene transcription at 12-24 hours post infection (hpi),whereas it is non-essential for the basal level of viral gene transcription.Fluorescence microscopy reveals the P6.9's co-localization with DNA is temporally and spatially synchronized with P6.9's impact on viral gene transcription,indicating the P6.9-DNA association contributes to transcription regulation.Chromatin fractionation assay further reveals an unexpected co-existence of P6.9 and host RNA polymerase Ⅱ in the same transcriptionally active chromatin fraction at 24 hpi,which may probably contribute to viral gene transcription up-regulation in the late infection phase.

The major immunogenic proteins (Ems,E2 and NS3) of classical swine fever virus (CSFV) (Shimen strain) were expressed in E.coli and purified by affinity chromatography.The recombinant antigens were applied to develop multiple enzyme-linked immunosorbent assays (ELISAs) for the detection of specific antibodies in pig sera.Optimum cut-off values were determined by receiver operating characteristic (ROC) analysis after testing 201 sera of vaccinated pigs and 64 negative sera of unvaccinated piglets.The multiple ELISAs were validated with 265 pig sera yielding high sensitivity and specificity in comparison with the virus neutralization results.The results demonstrated that multiple ELISAs can be a valuable tool for the detection of CSFV infection and serological surveys in CSFV-free countries or for the evaluation of the antibody responses in pigs induced by a live attenuated C-strain vaccination.