Molecular characteristics of the untranslated segment-specific region of the hemagglutinin segments in H5N6 and H7N9 influenza viruses in Shenzhen / 中华实验和临床病毒学杂志

Objective@#Exploring the molecular characteristic of global and Shenzhen district H5N6 and H7N9 influenza viruses HA untranslated regions(UTRs).@*Methods@#Mega7.0 and DNAStar 7.1.0 were used to construct phylogenetic tree and nucleotide analysis.@*Results@#From 2014 to 2015, 3 strains of H5N6 influenza virus from Shenzhen were compared with the other H5NX influenza viruses, the nucleotide homology of HA-3’UTR was 77.4%-100%, which did not have obvious mutated sites. The nucleotide homology of H5N6-HA-5’UTR was 91.7%-100%, and the sites of 24 and 31 sites were mutated. From 2013 to 2014, 11 strains of H7N9 influenza virus from Shenzhen were compared with the other H7NX influenza viruses, the nucleotide homology of H7N9-HA-5’UTR was 76.8%-100%, which had multi-mutated sites on 2-6, 9, 10, 12 and 15-17 positions.@*Conclusions@#HA-UTR from human-infected H5N6 and H7N9 influenza viruses isolated in Shenzhen district has unique molecular characteristics, its conserved region has relatively high homology and the segment-specific region has genetic polymorphism.

Molecular identification of the species of Biomphalaria snails in Shenzhen reservoir based on 16S rDNA sequences / 中华地方病学杂志

Objective To identify the species of Biomphalaria snails collected in Shenzhen reservoir,based on the mitochondrial 16S rDNA sequences.Methods The 16S rDNA fragments were amplified by PCR from the genome DNA of Biomphalaria snails,and inserted in plasmid pMD-18T for sequencing.The sequence of 16S rDNA fragment and its phylogenetic relationships with those of other species of Biomphalaria snails were analyzed with BLAST and MEGA4 software.Results The amplified 16S rDNA fragment of the Biomphalaria snails was about 466 bp in length.As aligned with the corresponding sequences of the related Biomphalaria species,the identity of nucleotides was 99％ with 1 isolate of Biomphaltria straminea (B.straminea),98％ with 3 isolates of B.kuhniana,95％ with 1 isolate of B.intermedia,and 94％ with 1 isolate of B.edisoni.Based on the 16S rDNA sequence,the results of phylogenetic analysis with neighbor-joining (NJ) and unweighted pair-group method with arithmetic means (UPGMA) indicated that the snails had close genetic relationships with the B.straminea isolate (Genbank accession NO.AY030213.1) Conclusion The Biomphalaria snails collected in Shenzhen reservoir could be classified as B.straminea based on the characteristics of 16S rDNA sequence.

Study on distribution of cohere antigen of 12D5 and 21B7 monoclonal antibodies of Angiostrongylus cantonensis and its application of immnodiagnosis / 中华微生物学和免疫学杂志

Objective To detect infection of Angiostrongylus cantonensis and examine effection of treatment to prepare monoclonal antibodies(McAbs). Methods Six-week-old BALB/c mice were imrnunized by the intraperitoneal injection of e/s antigens of Angiostrongylus cantonensis. Fusion of splecn cells from immunized mice with prepared SP2/0-Ag14 myeloma cells was performed in RPMI 1640. Fused cells were suspended in RPMI 1640 containing 1% HAT and 20% fetal calf serum and dispensed into 96-well cell culture plates. The supernatants of clones were screened by ELISA with sera of patients of angiostrongyliasis.Distribution of cohere antigen of 12D5 and 21B7 monoclonal antibodies was analyzed with immunohistochemistry. Two McAbs ( 12D5 and 21B7) were applied to detect the circulating antigen (CAg) in the sera of rats infected with A. cantonensis and angiostrongyliasis patients respectively by double antibody sandwich ELISA.Results 12D5 McAb was identified as IgG1 and 21 B7 McAb was IgM. Western blot result showed two McAbs could used to identified 55 × 103 protein of adult worms of A. cantonensis. Cohere antigen of 12D5 and 21B7 monoclonal antibodies were distributed on intestine surface of A. cantonensis. The detection rates of CAg in the sera of infected rats 100% (48/48), the detection rates of CAg in the sera of angiostrongyliasis patients was 100% (32/32). No cross-reaction to sera of patients with other infection of parasites, such as clonochiasis, fasiolopsiasis, ancylostomiasis, trichinosis, anisakiasis as well as schsitosomiasis, and health srea did not reacted with 12D5 and 21B7 McAbs,and detaction rate of antibody of angiostrongyliasis patients only reached 75% (24/32) with antigen of A. cantonensis. Conclusion Cohere antigen of 12D5 and 21B7monoclonal antibodies were antigens of enteric epithelium. Sandwich ELISA with 12D5 and 21B7 McAbs showed high specificity act as detecting CAg of A. cantonensis in sera of infection animal and patients. It is apparent that Sandwich ELISA with 12D5 and 21 B7 is not only rapid and simple without requirement of special instrument, but also rather sensitive and specific for the detection of current infection with A. cantonensis.

Identification of the gene differentially expressed in female Anopheles anthropophagus and cloning of its full-length cDNA / 中国人兽共患病学报

To identify the gene differentially expressed in female Anopheles anthropophagus and to analyze its gene sequence, this gene amplified by PCR was identified by real-time PCR and its cDNA was then amplified with rapid amplification of cDNA ends (RACE) technology. It was found that the expression ratio of the female differentially expressed gene in female and male mosquitoes was 267.49 according to the formula F=2~(-⊿⊿CT).The size of mRNA of the gene was 364 bp, and the amino acid sequence deduced from the open reading frame (ORF) was found to be similar to the sequence of tectin protein of Culex quinquefasciatus and proteins of other species. The mRNA sequence of this gene was submitted to NCBI with a accession number of FJ907236. This gene may provide a foundation for further studies on the biological functions of mosquitoes.