Aim To investigate the effects of dalbinol on proliferation and apoptosis of human colon cancer HCT1 16 cells and its mechanisms.Methods Anti-proliferative effect of dalbinol was evaluated by MTT assay.The morphological changes of apoptosis were observed by Hoechst33342 staining.Apoptotic rate and ROS generation were analyzed by flow cytometry.The related proteins of Wnt/β-catenin pathway and the ap-optosis-associated proteins expression were measured by Western blot.Results The growth of HCT1 16 treated with dalbinol was inhibited in a dose and time dependent manner with IC50 (4.8 ±0.53 ),(2.5 ± 0.43)and (0.6 ±0.22)μmol·L-1 at 24,48 and 72 h,respectively.Typical morphological changes of ap-optosis such as cell shrinkage,karyopyknosis and nu-clear condensation were observed by Hoechst33342 staining.Meanwhile,the apoptotic rate and intracellu-lar ROS generation of dalbinol were both increased dose-dependently. Western blot results showed that dalbinol could activate the expression of cleaved Caspase-3 and cleaved PARP by decreasing anti-apop-totic protein levels such as Bcl-2 and Mcl-1 and in-creasing pro-apoptotic protein levels such as Bax and Bim,which induced further apoptosis.Moreover,dal-binol can reduce the protein expression of the total and nuclear β-catenin,but not cytoplasmic β-catenin by suppressing the protein expression of Dvl-2 and GSK-3β(pS9 ),as well as its target proteins c-Myc and Sur-vivin.Conclusion dalbinol can induce apoptosis in colon cancer HCT1 16 cells by upregulating the intra-cellular ROS generation and suppressing Dvl/GSK-3β/β-catenin pathway.

We analyzed the genetic characteristics of coxsackievirus A4 (CV-A4) based on the entire VP1 coding region. Samples were isolated from patients with acute flaccid paralysis (AFP) in Shaanxi, China from 2006 to 2010. We wished to ascertain the predominant genotype and the relationship between CV-A4 infection and AFP. Sixty-eight non-polio enteroviruses were inoculated onto RD cells (to increase the virus titer) and molecular typing was undertaken. The entire VP1 coding region was amplified. Percentage of CV-A4 was 10.3% (7/68). Analyses of genetic identify and creation of phylogenetic trees revealed that CV-A4 could be classified into A, B and C genotypes. Seven CV-A4 strains from Shaanxi and other CV-A4 strains from China formed an independent evolution lineage located in group 4 and belonged to the C2 sub-genotype. These data suggested that CV-A4 strains of sub-genotype C2 were the predominant genotypes in China. These strains co-evolved and co-circulated with those from other provinces in China, so continued monitoring of CV-A4 (by clinical and genetic surveillance) should be enhanced.
China ; Enterovirus A, Human ; classification ; genetics ; isolation & purification ; Enterovirus Infections ; virology ; Genotype ; Humans ; Paralysis ; virology ; Phylogeny ; Viral Proteins ; genetics

The World Health Organization redefined the type 2 vaccine-derived poliovirus (VDPV) in 2010. To study the genetic characteristics and evolution of type 2 VDPV under this new definition, we conducted genome sequencing and analyses of type 2 VDPVs isolated from one patient with acute flaccid paralysis in Shanxi province (China) in 2014. Nucleotide sequencing revealed that the full-length of type 2 VDPV is 7439 bases encoding 2207 amino acids with no insertion or deletion of nucleotides compared with Sabin2. One nucleotide substitution identified as a key determinant of the attenuated phenotype of the Sabin 2 strain (A-G reversion at nucleotide nt 481 in the 5-end of the untranslated region) had reverted in the Shanxi type 2 VDPV. The other known key determinant of the attenuated phenotype of the Sabin 2 strain (U-->C reversion at nt2909 in the VP1 coding region that caused a Ile143Thr substitution in VP1) had not reverted in the Shanxi VDPV. The Shanxi type 2 VDPV was S2/S1 recombinant, the crossover site of which mapped to the 3-end of the 3D region (between nt 6247 and nt 6281). A phylogentic tree based on the VP1 coding region showed that evolution of the Shanxi type 2 VDPV was independent of other type 2 VDPVs detected worldwide. We estimated that the strain circulated for approximately = 11 months in the population according to the known evolution rate. The present study confirmed that the Chinese Polio Laboratory Network could discover the VDPV promptly and that it played an important part in maintenance of a polio-free China.
Amino Acid Sequence ; Base Sequence ; Capsid Proteins ; chemistry ; genetics ; China ; Humans ; Infant ; Male ; Molecular Sequence Data ; Phylogeny ; Poliomyelitis ; virology ; Poliovirus ; chemistry ; genetics ; metabolism ; Poliovirus Vaccines ; adverse effects ; chemistry ; genetics ; metabolism ; Sequence Alignment

Objective To study thegenetic and molecula repid emiological characteristics of ECHO29 virusVP1 region coding in Tibet under 5-year-old children in 1999-2000.Method 7 strains of ECHO29 viruses isolated from 659 stool specimens of AFP cases and under 5-year-old patients and some healthy children in Tibet in 1999-2002 were used to extract ribonucleic acid (RNA) while VP1 coding area were amplified by reverse transcription-polymerase chain reaction (RT-PCR) and the nucleotide sequence of PCR products were determined and analyzed.Result A total of 7 strains ECHO29 viruses were isolated from the stool specimens and identified.5 strains were isolated from specimens in 1999 while the other 2 strains isolated from samples in 2000.The complete nucleotides sequence of VP1 region of 7 strains ECHO29 viruses were 876nt coding 292 amino acids.The sequence homology of 7 strains detected ECHO29 viruses was between 75.41％-99.3％ while amino acid homology was between 71.26％-79.69％.The phylogenetic tree of ECHO29 viruses showed 4 genotypes which the isolates from Tibet were belong to Genotype A and C.Conclusion It was first reported that the popular ECHO29 viruses in children in Tibet were proposed to 2 different genotypes in 1999 and 2000.The popular ECHO29 virus epidemic in 1999 was belong to Genotype C,while popular ECHO29 virus in 2000 belong to Genotype A.ECHO29 virus increasingly popular was not found in 2001 and 2002.

Objective: To evaluate 2017 poliovirus surveillance in Qinghai Province. Methods: According to the World Health Organization (WHO) 4 th edition of the polio laboratory manual procedure for virus isolation, the isolated L20B positive strain was identified as intratypic differentiation (ITD) by the China Center for Disease Control and Prevention, CDC). The National Polio Laboratory performed the nucleotide sequence determination of the capsid protein VP1 coding region of poliovirus (PV) and analyzed the poliovirus surveillance and the result of analysis of the cases with acute flaccid paralysis (AFP) reported in Qinghai Province in 2017 and stool samples of healthy children. Results: In 2017, Qinghai CDC Polio Laboratory received specimens of 211 AFP cases and healthy stool samples. PV2 strains were isolated with a separation rate of 0.95%. Non-polio enterovirus (NPEV) strains were isolated from 25 strains with the isolation rate of 11.85%. Two PVs were used for ITD. All of them were vaccine-associated strains. Conclusions In 2017, the Qinghai CDC Polio Laboratory did not find any poliovirus and vaccine-derived poliovirus in the AFP cases and stool samples from healthy persons, maintaining the polio-free status.

Objective To reveal the genetic characteristics of VP1 encoding region of Coxsackievirus A16 (CA16) in Ningxia Hui Autonomous Region,2010.Methods With specific primers,reverse transcriptase polymerase chain reaction (RT-PCR) were performed to amplify the complete VP1 encoding sequence from CA16 isolates,which were isolates from hand,foot,and mouth disease (HFMD) patients in Ningxia in 2010.The VP1 sequences were determined,and analyzed with bioinformatic software.Results Totally,VP1 of 62 CA16 strains were determined,showing 90.1％-100％ (average,94.5％) identities of nucleotide sequences and 98.7％-100％ (average,99.6％) identities of amino acid sequences.On phylogenetic tree,all of the viruses in this study clustered together with subgenotype B1,which could be divided into clade B1a and B1b further.And viruses from each prefecture in Ningxia included both clade B1a and clade B1b.Conclusions Consistent with CA16 from other provinces,CA16 circulating in Ningxia in 2010 included both clade B1a and B1b.These viruses showed high diversity of VP1 nucleotide sequences,suggesting heterologous CA16 strains were co-circulating in Ningxia in year 2010.

Objective To evaluate the inactivation effect of heat inactivation and three common chemical disinfectants on enterovirus group A 71 (EV-A71),provide basic data for disinfection work after laboratory activities in enterovirus secondary biosafety laboratory.Methods EV-A71 virus was treated at different temperatures and different disinfectant concentrations according to different time.The inactivated virus was inoculated into 96-well cell culture plates,and the cytopathic effect was observed and recorded.Finally,the virus titer of the virus treatment solution was calculated.Results After inactivation at 56 ℃ for 30 min,the inactivation logarithm of EV-A71 was more than 6,and inactivation at 65 ℃ for 30 rin,70 ℃ for 10 min,no infection was observed.Three chemical disinfectants could effectively inactivate a certain titer of EV-A71 at a certain concentration and treatment time.Conclusions The inactivation effect of EV-A71 virus is related to factors such as thermal inactivation temperature,chemical disinfectant concentration,and treatment time.Studying the inactivation curve of EV-A71 is beneficial to the selection of inactivation conditions in different inactivation environments.

Objective:To evaluate the effects of vaccine switch on the levels of neutralizing antibody (NA) of poliovirus in children aged<12 years.Methods:Subjects aged<12 years from 9 districts in Fujian province were chosen by stratified cluster and complete random sampling method. Blood samples were collected for testing NA of poliovirus by microcell neutralization.Results:A total of 2 134 subjects aged<12 years were selected. The positive rate of NA against PVⅠand Ⅲ were 98.64% and 95.83%; and geometric mean titers (GMTs) were 1∶259.35 and 1∶105.14, respectively. The GMTs presented a trend of decreasing as age increased. Compared to trivalent oral poliovirus vaccine (tOPV), bivalent oral poliovirus vaccine (bOPV) and inactivated poliovirus vaccine (IPV) induced higher GMTs of NA against poliovirusⅠand Ⅲ respectively. Among 182 children aged<5 years, the positive rate of NA against PVⅠ, Ⅱ, Ⅲ were 97.25%, 76.37% and 92.86%. There were statistical differences among the three types ( χ2=44.44, P=0.000). The rate of NA against PVⅡwas significantly lower than those ofⅠand Ⅲ (Ⅱ vs. I: χ2=34.65, P=0.000; Ⅱ vs. Ⅲ: χ2=18.99, P=0.000). And the GMTs of NA against PV Ⅰ, Ⅱ, Ⅲ were 1∶368.96、1∶23.06 and 1∶183.10, which were significantly different ( F=156.54, P=0.000). The GMT of PVⅠwas the highest, PV Ⅲ was the second, PVⅡ was lowest (pairwise comparison showed values of P all were 0.000). The analysis of general linear model showed that the interval between the date of last immunization and the date of sample collection would affect the GMTs of PVⅠand Ⅲ, different vaccine models would affect only the GMT of PV Ⅰ. The age maybe was a confounding factor. But no factors would affect the GMT of PV Ⅱ. Conclusions:After the vaccine switch, the level of NA against PVⅠand PV Ⅲ were still maintaining high level, but the level of PVⅡamong children<5 years was at relatively low level, which indicated that the surveillance should be strengthened.

Objective:Objective The inhibitory effect of Niuhuang Qinggan Capsule on 2019-nCoV virus was studied to provide experimental evidence for drug screening and treatment of 2019-nCoV acute respiratory disease.Methods:The 2019-nCoV virus GS048 strain was isolated and cultured by Vero-E6 cells. The solution of Niuhuang Qinggan capsule was incubated with virus in vitro to determine the change of virus titer. The effect of Niuhuang Qinggan capsule on the proliferation of 2019-nCoV virus was determined by cell culture medium method.Results:It was found that the concentration of 274.3 μg/ml was not toxic to the cells when the concentration of drugs was lower. When the drug was incubated with virus, the titer of virus decreased from 10 4.4TCID 50/ml to 10 2.8TCID 50/ml. In vitro 2019-nCoV virus culture experiment showed that the addition of the drugs reduced the cytopathy, and the logarithmic value of virus titer decreased by 40% -50%. Conclusion:In this study, in vitro inhibition experiments suggest that Niuhuang Qinggan capsule has the potential to inhibit the proliferation of 2019-nCoV. It can be used as a candidate drug to further verify the therapeutic effect of the 2019-nCoV acute respiratory disease.

Objective:To study the genomic sequence of Coxsackievirus A8 (CV-A8) associated with hand, foot and mouth disease (HFMD) from 2013 to 2018 in China and to analyze the genetic evolution of each coding region of the full-length genome.Methods:The genome sequences of 11 CV-A8 strains isolated from patients with HFMD in different regions of China from 2013 to 2018 were determined. Sequence alignment and genetic evolution analysis were performed by Sequencher 5.0 and MEGA 7.0 software, etc.Results:Sequence alignment showed that the genome length of 11 CV-A8 strains ranged from 7 393 bp to 7 400 bp. There was no base insertion or deletion in the coding region compared with the prototype strain, but there were individual base insertion or deletion in the non-coding region. The nucleotide and amino acid similarities in the VP1 region of 11 CV-A8 strains were 78.3%-98.6% and 92.6%-99.7%, respectively, and the nucleotide and amino acid sequences identities with the CV-A8 prototype strain were 78.3%-98.2% and 92.6%-99.7%, respectively. Based on the phylogenetic analysis of VP1 region sequences, the CV-A8 can be divided into five genotypes: A, B, C, D and E. The 11 CV-A8 strains in this study belonged to genotypes C (1 strain), D (2 strains) and E (8 strains). The nucleotide and amino acid similarities of 11 CV-A8 full-length genomes were 81.3%-98.8% and 95.9%-99.5%, respectively. The phylogenetic tree of the P2 region showed that the eight E genotypes CV-A8 had the closest evolutionary distance with CV-A4, CV-A14, and CV-A16. The phylogenetic tree of the P3 region showed that the eight E genotypes CV-A8 had a close evolutionary distance with CV-A5, CV-A16, CV-A14 and CV-A4.Conclusions:The 11 CV-A8 stains in this study showed significant intra-genotype diversity in capsid region and recombinant diversity in non-capsid region which indicated that CV-A8 quasispecies were still undergoing dynamics variation. CV-A8 may become an important pathogen of HFMD and the monitoring of CV-A8 needs to be further strengthened.