We constructed a recombinant plasmid encoding VP1 gene of O type foot-and-mouth disease virus fused to a molecular adjuvant, goat complement C3d gene. The goat C3d gene was cloned and three copies were tandem-linked with the linker (G4S)2 sequence. VP1 gene of O type foot-and-mouth disease virus was linked to three tandem repeats of C3d through the linker sequence and cloned into pUC19 to obtain the recombinant plasmid pUC19-VP1-C3d3. The VP1-C3d3 fusion gene was then subcloned into the eukaryotic vector pcDNA3.1(+) that had been modified to contain the tissue plasminogen activator (tPA) leader sequence to obtain pcDNA3.1-tPA-VP1-C3d3. HeLa cells were transfected with pcDNA3.1-tPA-VP1-C3d3 by Lipofectamine 2000. Indirect immunofluorescent assay and Western blot assay showed that VP1-C3d3 fusion gene was successfully expressed in HeLa cells. The fusion protein with the expected size 133 kD could be secreted outside the cells. This study laid a good foundation to further research on the novel vaccine against foot-and-mouth disease virus by using goat C3d as a molecular adjuvant to enhance the immunogenicity of VP1.
Animals ; Capsid Proteins ; biosynthesis ; genetics ; Cloning, Molecular ; Complement C3d ; biosynthesis ; genetics ; immunology ; Female ; Foot-and-Mouth Disease Virus ; genetics ; Goats ; HeLa Cells ; Humans ; Immunologic Factors ; biosynthesis ; genetics ; immunology ; Plasmids ; genetics ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; immunology ; Transfection

Objective To investigate the effect of multimedia technology on disease management capability in patients with chronic obstructive pulmonary disease (COPD).Methods 77 patients with COPD were randomly divided into control group ( 38 cases ) and observation group ( 39 cases ) .The control group was educated and managed on the basis of authorization education .While the observation group was cooperated with multimedia technology for COPD education and management , on the basis of authorization education .The intervention time lasted for 3 months.And the self -management capacity and quality of life of patients in the two groups were evaluated by utilizing self-management scale and St .George Respiratory Questionnaire ( SGRQ) respectively before intervention and 3 months after intervention .Results There were no statistically significant differences in self -management capacity and SGRQ score in all subjects before intervention (all P>0.05).After 3 months of intervention,the scores of self-management capacity on symptom management , daily life management , emotional management , information management and self -efficacy of the observation group were (30.71 ±6.79)points,(42.10 ±5.92)points,(39.51 ± 5.64) points,(30.92 ±6.08) points and (31.58 ±5.43) points,respectively,which were significantly higher than those in the control group [(27.76 ±4.12) points,(39.31 ±5.11) points,(36.28 ±6.38) points,(28.39 ± 4.70)points and (29.21 ±3.3)points],and the differences were statistically significant (t=-2.299,-2.207,-2.348,-2.037,-2.299,all P<0.05).The SGRQ scores of respiratory symptoms,disease effects and motion limitation of the observation group were (40.87 ±7.27) points,(41.17 ±7.15) points and (40.48 ±5.91) points, respectively,which were significantly lower than those in the control group [( 44.7 ±8.13 ) points, ( 45.21 ± 7.60)points,(43.60 ±5.87)points],and the differences were statistically significant (t=2.199,2.395,2.321,all P<0.05).Conclusion Application of advanced digital equipment and demonstration equipment to the disease education and management of patients with COPD based on patients 'needs provides effective ways and models to patients'health education ,which promotes the establishment of patients'healthy behaviors and the management of their own diseases ,and significantly improves the patients'self-management capacity and quality of life .

Salmonella enterica serovar Choleraesuis strain C500 is a live, attenuated vaccine that has been used in China for over 40 years to prevent piglet paratyphoid. The objective of this study was to evaluate the potential of attenuated Salmonella enterica serovar Choleraesuis C500 strain with a delta asd mutant as an effective live vaccine vector by the Asd+ balanced-lethal host-vector system. Here, we compared the characteristics of S. enterica serovar Choleraesuis delta asdC500 strain with the parent C500 strain, including phenotype, growth rate, virulence, safety, and expression for heterologous antigen. The mean generation times of delta asdC500 mutant, the vector control delta asdC500 (pYA3493), and the parent avirulent C500 vaccine strain in Luria broth were 30.7, 28.1, and 27.9 min, respectively. The fermentation patterns of theses three strains on different carbohydrates, and the levels of production of H2S, were similar. The O and H antigens of delta asdC500 mutant, delta asdC500 (pYA3493) and delta asdC500 (pYA-F1P2) were 6,7:C:1,5, identical to the parent strain C500. By the method of Reed and Muench, groups of mice were challenged by the intraperitoneal route with different amounts of delta asdC500 (pYA3493) or the parent C500 strain, and the virulence of delta asdC500 (pYA3493) with LD50 of 1.1 x 10(7) CFU was a little lower than C500 with LD50 of 4.4 x 10(6) CFU. All piglets inoculated with delta asdC500 (pYA3493) or C500 survived, and no signs of disease were observed during the entire experimental period. No major differences were found in these two groups. In addition, the recombinant pYA-F1P2 plasmid was very stable in the recombinant delta asdC500 (pYA-F1P2) strain, which expressed secretorily a large amount of the recombinant filamentous hemagglutinin type I domain and pertactin region 2 domain antigen (rF1P2) of Bordetella bronchiseptica. In this study, we have shown that the delta asdC500 mutant had a series of biological characteristics similar to the parent vaccine strain C500. Furthermore, the strain could express secretorily a large amount of heterologous antigen. It is likely that this Salmonella expression and delivery system could be easily adapted to develop multivalent recombinant Salmonella vaccines against infectious agents using the Asd+ balanced-lethal host-vector system.
Amino Acid Oxidoreductases ; genetics ; Animals ; Bacterial Proteins ; genetics ; Gene Deletion ; Genetic Vectors ; Mice ; Mutation ; Salmonella Vaccines ; genetics ; immunology ; Salmonella enterica ; genetics ; immunology ; pathogenicity ; Swine ; Transduction, Genetic ; Vaccines, Attenuated ; genetics ; immunology ; Vaccines, Synthetic ; genetics ; immunology ; Virulence

Using mutation PCR, we cloned the target gene containing 421-480nt (141-160aa) and 598-639nt (200-213aa) of VP1 gene of foot and mouth disease virus (FMDV) into the deleted region (508-532aa) of Nsp2 gene of a highly pathogenic porcine reproductive and respiratory syndrome virus derived vaccine strain (HuN4-F112) that was used as vector. The recombinant cDNA was in vitro transcribed followed by transfection of BHK-21 cells for 36 h. Then, the supernatant of the cell culture was continuously seeded to monolayer of MARC-145 cells for recovery of the recombinant virus. CPE was obviously visible after a couple of passages in the seeded MARC-145, and the rescued virus (designated as rPRRSV-F112-O/VP1ep) was identified by Mlu I digestion, sequencing and immunofluorescence assay. Meanwhile, expression of inserted FMDV epitopes was also detected by indirect immunofluorescence assay with polyclonal antibodies against VP1 protein of FMDV. The analysis of biological characteristics shows that the titer of the rescued recombinant PRRSV (TCID50 = -log10(-6.75)/0.1 mL) was similar to its direct parental virus rHuN4-F112-delta508-532, but higher than rHuN4-F112.
Animals ; Antigens, Viral ; immunology ; Base Sequence ; Capsid Proteins ; immunology ; Cell Line ; Cysteine Endopeptidases ; genetics ; Epitopes ; genetics ; Foot-and-Mouth Disease ; immunology ; prevention & control ; Foot-and-Mouth Disease Virus ; genetics ; immunology ; Molecular Sequence Data ; Mutation ; Porcine respiratory and reproductive syndrome virus ; genetics ; immunology ; Recombination, Genetic ; Swine ; Transfection ; Vaccines, Attenuated ; genetics ; immunology ; Viral Envelope Proteins ; genetics ; immunology ; Viral Vaccines ; genetics ; immunology

Of all known small molecules targeting human immunodeficiency virus (HIV) capsid protein (CA), PF74 represents by far the best characterized chemotype, due to its ability to confer antiviral phenotypes in both early and late phases of viral replication. However, the prohibitively low metabolic stability renders PF74 a poor antiviral lead. We report herein our medicinal chemistry efforts toward identifying novel and metabolically stable small molecules targeting the PF74 binding site. Specifically, we replaced the inter-domain-interacting, electron-rich indole ring of PF74 with less electron-rich isosteres, including imidazolidine-2,4-dione, pyrimidine-2,4-dione, and benzamide, and identified four potent antiviral compounds (