Acetyl-CoA C-acetyltransferase (AACT) is the first enzyme in the terpene synthesis pathway, catalyzed two units of acetyl-CoA to acetoacetyl-CoA. In order to study the tanshinone biosynthesis in Salvia miltiorrhiza, a novel AACT gene, SmAACT, was cloned using cDNA microarray and RACE strategy. The full length cDNA of SmAACT is 1 623 bp (accession No. EF635969), which contained a 1 200 bp open reading frame (ORF) encoding a 399 amino acid protein. Nine introns were found in the genomic sequence. SmAACT was upregulated by YE and Ag+ elicitors both with cDNA microarray and quantitative RT-PCR analyses along with the accumulation of tanshinones. Sequence homology comparison and phylogenetic analysis all suggested that SmAACT belonged to the class of acetyl-CoA C-acetyltransferase. The transcription level of SmAACT was relatively higher in root than that in stem and leaf tissues. SNP analysis revealed that SmAACT was highly variable in the region of 6 to 9 introns with 33 SNPs in the 600 bp region, there are 5 SNPs in the cDNA region while they are all synonymous cSNPs. Some special genotypes were found in Salvia miltiorrhiza from different areas. SmAACT will be an useful gene for further analyze the mechanism of gene regulation among the tanshinones biosynthesis.

Objective To investigate the serotypes of human enterovirus B ( HEV-B) species cau-sing hand, foot and mouth disease ( HFMD) and to analyze the genetic characteristics of VP1 region in cox-sackievirus B2 ( CVB2 ) and coxsackievirus B5 ( CVB5 ) strains circulating in Anyang area during 2011 to 2015. Methods Real-time RT-PCR and semi-nested RT-PCR were performed to identify coxsackievirus A16 (CVA16), enterovirus 71 (EV71) and other serotypes of enterovirus in order to obtain the complete etiologic composition of HFMD. The numbers of HEV-B serotypes and the percentages of specimens positive for every serotype in all enterovirus-positive specimens were calculated. As CVB2 and CVB5 were the pre-dominant serotypes of HEV-B species, five pairs of primers targeting the VP1 regions of CVB2 and CVB5 were designed to obtain the complete nucleotide sequences of CVB2 and CVB5 VP1 regions. The phylogenet-ic trees were constructed based on the VP1 sequences obtained in this study and those submitted to GenBank by using MEGA7. 0 and BioEdit7. 2. The selection pressures on VP1 regions of CVB2 and CVB5 strains cir-culating in China in recent years were evaluated with the online program of DataMonkey. Results A total of 57 specimens that belonged to 14 serotypes of HEV-B species were detected in Anyang area from 2011 to 2015. The 14 serotypes of HEV-B species accounted for 56% of all serotypes of enterovirus and the speci-mens positive for HEV-B species accounted for 3. 06% of all enterovirus-positive specimens. The HFMD ca-ses caused by most of the HEV-B serotypes were sporadic cases. Small outbreaks of HFMD could also be caused by some serotypes of HEV-B such as CVB2 and CVB5. The complete sequences of VP1 region were obtained from 8 CVB2 strains and 9 CVB5 strains. The phylogenetic trees based on the VP1 sequences dem-onstrated that the CVB2 strains were classified into four genotypes ( A-D) . The mean evolutionary distances between different genotypes ranged from 0. 191 to 0. 208 and the similarities in nucleotide sequences ranged from 79. 7% to 85. 8%. The CVB5 strains were classified into 6 genotypes (A-F). The mean evolutionary distances and the similarities in nucleotide sequences between different genotypes of CVB5 strains ranged from 0. 170 to 0. 285 and 76. 0% to 86. 8%, respectively. Strains of different genotypes varied significantly in the residues on positons 157 and 263 in the VP1 region of CVB2 strains and on positions 75, 90 and 95 in the VP1 region of CVB5 strains. All of the CVB2 strains isolated in Anyang area belonged to D genotype and located intensively in one lineage. The CVB5 strains circulated in Anyang area belonged to F genotype and located in two lineages. The selection pressures on CVB2 strains of D genotype and CVB5 strains of F geno-type circulating in China in recent years were 0. 037 and 0. 036, respectively. Six positively selected amino acid sites were found in the VP1 region of CVB5 strains, but no positively selected amino acid site was found in the VP1 region of CVB2 strains. Conclusion HEV-B species was an essential component of the etiologic spectrum of HFMD in Anyang area during 2011 to 2015, of which CVB5 and CVB2 were the predominant se-rotypes. The VP1 region of CVB5 was more complex and active than that of CVB2 over the course of evolution.

Objective To analyze the evolution and genetic characteristics of Echovirus 11 (Echo11) strains isolated from patients with hand food and mouth disease ( HFMD) in Henan province. Methods Enterovirus strains were isolated from stool samples of patients with HFMD from year 2010 to 2012.The sequences of VP1 region of all positive strains were amplified , sequenced and then analyzed by BLSAT to determine their genotypes .The entire VP1 coding regions of 10 Echo11 strains were amplified by RT-PCR, and the homology analysis was conducted among them and other Echo 11 strains published in Gen-Bank.A phylogenetic tree was constructed to evaluate the evolution of Henan isolates and the prevalence of different genotypes .Results From year 2010 to 2012 , 184 non-Enterovirus 71 ( EV71 ) strains and non-Coxsakievirus A16 (CA16) strains causing HFMD were isolated, 10 (5.43%) of with were Echo11 strains. There were 876 nucleotides ( nt) in the complete VP1 gene sequence , encoding 292 amino acids ( aa) .The 10 Echo11 strains shared 93.1%-100% homologies in nt sequences and 97.3%-100% in aa sequences. The homology analysis indicated that Henan strains and prototype strains were highly similar but different with the homology of 77.8%-78.8% in nt and 90.8%-91.8% in aa, respectively.Conclusion Echo11 was one of the pathogens causing HFMD in He-nan province and genotype A was the dominant genotype .

Objective: To investigate the correlation between the expression of vascular endothelial growth factor(VEGF) and angiogenesis in tumor. Methods: VEGF 165 sense and antisense gene recombinants were introduced into human gastric cancer cells, respectively. Then the growth of transfected cells in nude mice and the microvascular density and histological change were examined. Results: The growth rate of tumor in nude mice inoculated with sense VEGF cells was markedly higher than that in nude mice inoculated with antisense-VEGF cells. In histological examination, the microvascular density in tumor caused by sense-VEGF cells was greatly higher than that in tumor caused by antisense VEGF cells. Conclusion: As starting angiogenesis, VEGF might promote the growth of tumor, and the inhibition of VEGF production might prevent solid tumor from growing.

China ; Electrophoresis, Gel, Pulsed-Field ; Escherichia coli O157 ; classification ; genetics ; Genes, Bacterial ; Humans ; Molecular Typing ; Virulence

Objective To evaluate different detection methods in the diagnosis of severe fever with thrombocytopenia syndrome (SFTS), and find the most quick and accurate one for the identification of new bunyavirus infection. Methods Real-time PCR and ELISA-IgM were used to detect serum samples of 158 patients with acute phase of SFTS, which were collected from the special monitoring system of SFTS in Henan Province in 2014. IgM and IgG antibodies were detected by ELISA in 109 acute and convalescent paired serum specimens. The differences of the positive rates were compared between the three methods, and the influence of the collected interval time on the detection results was analyzed. Results For 158 acute phase serum samples of SFTS patients, the positive rate detected by real-time PCR (76.58%) was higher than that of ELISA-IgM (47.47%), and the difference was statistically significant (χ2=34.13, P < 0.05). For 109 cases with acute and convalescent paired serum samples, there was no significant difference in the positive rates between ELISA-IgG ( 75.23%) and real-time PCR (72.48%) detections (χ2=0.18, P>0.05). In both the acute phase and convalescent phase, the positive rate of IgM was higher than that of IgG, and the difference was statistically significant (χ2=41.68 and 6.25, P<0.05). With the extension of collected interral time, the positive rates of IgM and IgG antibodies were both increased ( Z=6.42 and 10.08, P < 0.05). Conclusion Real-time PCR is the most sensitive method for the early diagnosis of the SFTS. ELISA-IgG is suitable for the detection of SFTS at recovery period. ELISA-IgM can be used as an assistant method to guide clinical diagnosis.

To investigate the animals infection situation of novel bunyavirus in Xinyang City ,Henan Province ,China , animal serum samples such as cattle ,dog ,swine ,mice were collected in Shangcheng County and Guangshan County in Xinyang City .All the serum samples were detected by novel bunyavirus ELISA and real time RT-PCR method .A total of 292 animal serum samples were collected including 5 kinds of animals .The result of all the animal serum samples were negative by using real time RT-PCR ,and the positive rate was 45 .19% (141/312) by ELISA method .Of the 5 animal serum samples including mice ,cattle ,goats ,swine and dogs ,the positive rate were detected to be 1 .06% ,100 .00% ,76 .27% ,3 .57% ,and 75 .00%respectively .There was significant difference in results among 5 kind of animal serum antibodies .Animals such as cattle and dog may be the host of novel bunyavirus which were detected novel bunyavirus antibodies in cattle and dog in Xinyang City , Henan Province ,China .

Objective To identify the pathogen that caused an outbreak of viral encephalitis in Henan area in 2011.Phylogenic analysis was carried out on Coxsackie-virus B5 (CVB5) which was isolated during this outbreak.Methods Five throat swab,21 stool and 14 cerebrospinal fluid (CSF) specimens were collected from 29 inpatients during this outbreak.Viral isolation and real time RT-PCR were then performed for all specimens.Viral nucleic acid of enterovirus 71 (EV71),coxsackievirus A 16 (CA16) and pan-enterovirus (PE)were detected by real time RT-PCR.Phylogenetic tree based on entire VP1 sequences was constructed among CVB5 isolates from 2 stool and 3 CSF specimens of 5 inpatients and others published data retrieved from GenBank.Results The real time RT-PCR results showed that the PE nucleic acid positive rates of throat swab,stool and CSF specimens were 60.0％ (3/5),61.9％ (13/21) and 85.7％ (12/14) respectively.All of these specimens were negative for EV71 and CA16.The isolation rates of throat swab,stool and CSF specimens were 20.0％ (1/5),25.0％ (5/21) and 29.0％ (4/14),respectively.BLAST with both VP1 and 5′-UTR sequences and molecular typing indicated that CVB5 was the main pathogen.Analysis among the 5 positve isolates based on the complete VP1 sequences showed 97.9％-99.5％ homology.Data from homologous comparisons indicated that these isolates had the highest nucleotide acid identity with the Changchun CVB5 CC10/10/Changchun strain (97.1％-98.1％) which caused hand,foot,and mouth disease (HFMD) outbreak in Changchun in 2010,and lower identity (89.0％-89.6％ and 91.8％-92.5％) with the COXB5/Henan/2010 and 03001N strain isolated from Pingdingshan,Henan in 2010 and 2012,respectively.Phylogenetic tree in VP1 region showed that isolates of this outbreak belonged to genotype D,the same clade with Changchun strain.Conclusion CVB5 was the major etiological agent correlated with this outbreak.The shift of predominant genotype might serve as one of the causes that associated with this outbreaks.

Objective:To study the effect of moxibustion at head points on vascular dementia in improving clinic symptoms and integration of scale and regulating blood fat. Methods: 63 cases of VaD patients were randomly divided into moxibustion treatment group and the Western medicine treatment group. After 3 periods of treatment, the integration changes of the examination scale (HDS), intelligent scale (MMSE), and scale of daily living (ADL) before and after treatment were compared. And the blood lipid cholesterol (TC), triglycerides (TG), low-density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDC-C) level were detected. Results: In the two groups, before and after treatment, the HDS, MMSE score compared with the ADL scale were significantly different (P

Aim To study the protective effects of Rhynchophyll a of total alkaloids ( RTA ) on cerebral ischemia/reperfusion injury and the possi ble mechanism of action. Methods The effects of RTA on decapit ated gasping model and model of middle cerebral artery ischemia 2 h/reperfusion 22 h were observed. The neurological scores, cerebral infarct volume and cerebr al water content after ischemia/reperfusion were observed in rats respectively. The activities of NOS and SOD and the content of MDA in rat's brain tissue were measured. Neuron apoptosis in ischemia penumbral area were detected by terminal depoxynucleotidyl transferase mediated dUTP-biotin nick end labeling ( TUNEL ) . Results The average gasping times in mice treated with RTA 50 , 75 mg?kg -1 was significantly prolonged. The cerebral infarct volume and cerebral water content in rats treated with RTA 40, 60 mg?kg -1 were sign ificantly decreased in ischemic rats. RTA 40, 60 mg?kg -1 increased the ac tivity of SOD ,and decreased the activity of NOS and the content of MDA in the i schemic brains of rats. The number of apoptotic neurons in ischemia penumbral ar ea of cerebral tissue of rats treated with RTA 40, 60 mg?kg -1 was signif icantly lower than that in control rats. Conclusions RTA has pr otective effect on cerebral ischemia/reperfusion injury; this may be related to inhibit the activity of NOS and lipoperoxidation, and increasing the activity of SOD and decreasing neuron apoptosis.