Evolution of influenza A(H3N2) virus hemagglutinin (HA1) gene in Shangluo, China, 2014-2015 / 中国人兽共患病学报

Comparative analysis of the variations in HA 1 gene of the influenza A (H3N2) virus and the vaccine recommended were conducted in Shangluo city of China,during the surveillance year of 2014-2015.In this study,we collected the samples of H3N2 subtype strain from the Shanglou City of China during the surveillance period of 2014-2015.The strain was cultured in MDCK cells,HA gene fragment was amplified by RT-PCR and the nucleotide sequence was determined.Sequence alignment was performed using the clustax2.1 software.The phylogenetic tree was constructed by Mega6.0 software and was analyzed by Neighboring-joining method.Results showed that the homology of isolated strain during 2014-2015 was 97.2 ％-99.9％ and homology with the recommended vaccine strain A/Texas/50/2012 was 97.3％-98.5％.The amino acid sequence of the HA 1 gene of the isolated strain was compared with that of the vaccine strain.The major antigenic determinants of the isolates in 2014,having mutations were section B,Y159F,S198P,while the major antigenic determinants of isolates in 2015,having amino acid mutations were A zone G142R,B region S159F,S198P.These results indicated that the key antigenic determinant of influenza H3N2 subtype strain in Shangluo City has changed in 2014-2015 and A/Texas/50/2012 vaccine component is no more effective.Hence,there is an urgent need to update the influenza H3N2 subtype vaccine components and in future we should be deeply concerned about the evolution ofinfluenza H3N2 gene trends.

Analysis the viral etiology of fever and respiratory tract infection syndrome in Shaanxi province / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>Analysis the viral pathogenic spectrum for patients with fever and respiratory tract infection syndrome in Shaanxi province during 2010 and investigate the molecular epidemiology characteristics of respiratory syncytial virus.</p><p><b>METHODS</b>A total of 208 patients' pharyngeal swabs were collected based on surveillance definition from January 2010 to January 2011 and screened for sixteen human respiratory virus types/subtypes by Qiaxcel-based multiplex reverse transcription-PCR assay, including HRV,HCoV, Flu, HPIV, ADV, HRSV, HMPV and HBoV and investigate molecular epidemiology of HRSV by sequencing and phylogenetic analysis of the C-terminal second hypervariable region of the G gene.</p><p><b>RESULTS</b>109 out of 208 specimens (53%) were positive for one or more viruses. HRSV(42. 2%) was the dominant pathogen detected, followed by Flu(24. 5%), PIV(20%), HRV(13.6%) and ADV( 10.9%),there were also 8 strains of HCoV, 5 strains of HMPV and 3 strains of HBoV detected. The results showed that 22 specimens were positive for two or more viruses, PIV (14/22) was the most frequently detected viral agent among co-infection specimens, and the highest incidence of mixed infection is aged 15-39 years group (P < 0.05). The overall viral detection rate was no related to age. In addition to Flu, HMPV and PIV, other viruses (HRV, HBoV, HCoV, ADV, RSV) mainly infected 0 to 4 years old children. Among 46 HRSV positive specimens, 42 HRSV-A strains clustered into NA1 genotype and two HRSV-B strains clustered into two genotypes, BA9 and GB2.</p><p><b>CONCLUSION</b>HRSV is the dominate pathogen collected from patients with fever and respiratory tract infection syndrome in Shaanxi and HRSV A is the predominant subtype. For most viruses, infection was most prevalent among children aged <4 years. PIV was the most common pathogen in co-infection.</p>

Comparison of direct immune-fluorescent assay and real-time quantitative PCR in detecting the Hantavirus / 中华预防医学杂志

<p><b>OBJECTIVE</b>To compare the differences between the direct immuno-fluorescent assay (DFA) and real-time quantitative PCR in detecting the Hantavirus (HV) in rat lungs.</p><p><b>METHODS</b>From April to October in 2012, a total of 479 rats were caught by mouse-trap in residential or wild areas in Huxian, Jingyang, and Meixian of Shaanxi province, where haemorrhagic fever with renal syndrome (HFRS) was highly prevalent. The rats were dissected to take the two lungs, one was frozen and applied immuno-fluorescent assay to detect HV antigen while the other one was extracted its RNA and detected HV nucleic acid by real-time quantitative PCR. Then we compared the positive rate of the two methods.</p><p><b>RESULTS</b>Out of the 479 rats, 105 were caught from residential areas and the other 374 were caught from wild areas. Among the 105 rats caught from residential areas, no HV were detected out neither by DFA nor by real-time quantitative PCR. Among the 374 wild rats, 13.1% (49/374) were detected HV positive by DFA and 14.7% (55/374) were detected HV positive by real-time quantitative PCR. The difference showed no statistical significance (χ(2) = 0.402, P = 0.526). When detecting each lung sample, the HV positive rate was 10.2% (49/479) under the detection by DFA while the HV positive rate was 11.5% (55/479) under the detection by real-time quantitative PCR. The difference had no statistical significance (χ(2) = 1.286, P = 0.257) and the consistency coefficient was 68.2% under the paired chi-square test analysis, which showed high consistency (u = 11.759, P < 0.05). The sensitivity of real-time quantitative PCR to detect HV was 77.6% (38/49) comparing with DFA as standard, and the specificity was 96.1% (413/430). Out of the 9 suspected HV positive sample detected by DFA, 6 were confirmed positive by real-time quantitative PCR and 3 were denied.</p><p><b>CONCLUSION</b>Compared with the DFA, real-time quantitative PCR could also be used to detect the infection of HV in rats, and the result might be more stable.</p>

Genetic characterization of human parainfluenza virus 3 circulating in Gansu and Shaanxi Provinces from 2009 to 2011 / 病毒学报

To investigate the genetic characterization of Human parainfluenza virus-3 (HPIV-3) circulating in Gansu and Shaanxi Provinces of China, 719 throat swabs were collected from pediatric patients with acute respiratory infections from 2009-2011. Multiplex RT-PCR was used to screen common respiratory viral pathogens. For HPIV-3-positive specimens, nested RT-PCR was used to amplify the HN gene of HPIV-3. The nucleotides of Hemagglutinin-neuraminidase(HN)gene of 13 HPIV-3 positive strains identified in Gansu and Shaanxi Provinces were successfully sequenced and compared with those downloaded from GenBank. The phylogenetic analysis based on the nucleotides sequence of HN gene showed that 13 HPIV-3 strains belonged to sub-cluster C3 with little sequence variation (overall nucleotide divergence of 0.2%-2.3% and amino acid divergence at 0-1.1%). Compared with the complete gene of HPIV-3 strains from U.S.A., Canada, and Australia, the biggest divergence of the nucleotide and amino acid lovels was 6.0% and 3.4%, respectively. The nucleotide divergence between shaanxi09-2 and shaanxi10-H0091 was 0.9%, while the nucleotide divergence between shaanxi10-H005 and gansull-62110372 was 0.5%, between shaanxi09-2 and BJ/291/09 was 0.6%. However, there was no amino acid divergence among them. It is likely that HPIV-3 virus had been transmitting in Gansu and Shaanxi Provinces for several years. Human parainfluenza virus-3 (HPIV-3) circulated in Gansu and Shaanxi Provinces from 2009 to 2011 belonged to sub-cluster C3.

Long term epidemiological effects of vaccination on hemorrhagical fever with renal syndrome (HFRS) in Shaanxi provincial HFRS epidemic areas / 中华流行病学杂志

Objective To evaluate the protective rate and the variation of HFRS-IgG on hemorrhagical fever with renal syndrome (HFRS) vaccine.Methods Cluster,random sampling and cross-sectional study were used to assess the protective rate of HFRS vaccination.Level of HFRS-IgG was detected with ELISA in epidemic and non-epidemic areas of HFRS.Results Curve equation was obtained as Yprocective rate=(0.863+0.283/Xvaccination term) × 100％ by protective rate with vaccination term.Protective rates showed a reducing trend,90％ after 7-8 years of vaccination,88％ after 10 years,and 94％ on average.Absorbance (A) value of HFRS-IgG was 4 times higher in persons with vaccination than those without,in the epidemic area.Higher antibody level could be obtained after primary vaccination,but the level of antibody had a 50％ reduction after 5-10 years of vaccination,and a 60％ reduction after 10 years of vaccination.Conclusion HFRS antibody had a 50％ reduction after 5-10 years of vaccination.The protective rate of HFRS vaccination had a 90％ loss,after 7-8 years of vaccination.Booster dose was necessary after 7 years of vaccination.

Prokaryotic expression, purification and activity assay of recombinant vascular endothelial growth factor / 南方医科大学学报

<p><b>OBJECTIVE</b>To express human vascular endothelial growth factor (hVEGF(165)) in E. coli JM109 in the form of fusion protein by genetic engineering and test the biological activity and immunological competence of the expressed protein.</p><p><b>METHODS</b>hVEGF(165) gene was subcloned by PCR and inserted into pQE30 plasmid. hVEGF(165) fusion protein was expressed in E. coli JM109 and purified by Ni(2+)-NTA. The immunological competence of the expressed protein was tested by means of Western blotting and enzyme-linked immunosorbent assay (ELISA), and its biological activity was assayed by chicken chorioallantoic membrane (CAM) and Matrigel angiogenesis assay.</p><p><b>RESULTS</b>The recombinant hVEGF(165) fusion protein was successfully expressed in E. coli JM109 and its expression accounted for 30% of the total cellular protein. The purified protein presented a single band of 23 kD in SDS-PAGE. Western blotting, ELISA, CAM and matrigel angiogenesis assay showed excellent immunologic competence and biological activity of the recombinant protein.</p><p><b>CONCLUSION</b>Recombinant hVEGF(165) protein with excellent biological activity has been successfully expressed in E.coli JM109, which may facilitate future study in construction of prefabricated tissue-engineered bone graft.</p>