Objective To determine the structures of resistance transposons and muhilocus sequencing typing(MLST)in the vancomycin resistant enterococcus(VRE).Methods Twenty-one VRE strains were isolated from five hospitals in Hangzhou.The resistance to antimicrobial agents was determined by Etest.Polymerase chain reaction(PCR),conjugation,plasmid extract,transposon structures,pulse field gel electrophoresis(PFGE),muhilocus sequencing typing(MLST),and multiple-locus variable-number tandem repeat analysis(MLVA)were carried out.Results All of the 21 VRE strains harbored the vanA gene.These strains were divided into 10 PFGE types,7 sequence types(STs)and 5 MLVA types.All of these VRE strains were susceptible to linezolid and tigecycline.The vanA genes in two VRE strains were located in transposon Tnl546,and those in the other 19 VRE strains were located in transpeson Tnl546- like,with ISl485 inserted in vanXY.Vancomycin resistance of 1 8 VRE isolates was transferred by filter mating. All of these conjugants had a plasmid containing a molecular size of about 54 000 bo.Conclusions These 21 VRE strains were all caused by the vanA gene and divided into 7 MIST types.A novel trasnposon was detected.Most of these VRE isolates belonged to the clonal complex(CC17)by MIST,which was the hospital-adapted and pandemic VRE clonal complex.

Objective To evaluate the clinical application of HLA-peptides tetramer staining flow cytometry for determining HBV specific CD8+ cells.Methods HBV specific CD8+ cells in whole blood samples of chronic hepatitis B patients were stained with tetramer complex of HLA-A2 and HBV core 18-27 peptide and counted by flow cytometry. Results The medians of percentages of HBV specific CD8+ cells of total CD8+ cells were 0.20%(0.02%～2.04%) in 11 acute hepatitis B patients and 0.05%(

Since the outbreak of COVID-19 caused by the 2019-nCoV (SARS-CoV-2), with its high pathogenicity and contagiousness, it has posed a serious threat to global public health security. Up to now, the pathogenesis of 2019-nCoV is unclear, and there is no effective treatment. Vaccine as one of the most effective strategies to prevent virus infection has become a hot area. Based on the current understanding of 2019-nCoV, the development of 2019-nCoV vaccines covers all types: inactivated virus vaccine, recombinant protein vaccine, viral vector-based vaccine, mRNA vaccine, and DNA vaccine, etc. In this review, we focus on the candidate targets of the novel coronavirus, and the types, development status and progress of 2019-nCoV vaccines in order to provide information for further research and prevention.

Objective Some blood donors were negative for the surface antigen but positive for the core antibody,leading to acute hepatitis.This study was to determine the seroprevalence of the hepatitis B core antibody in voluntary blood donors.Methods A total of 9100 donor samples were screened for hepatitis B surface antigen and Hepatitis B core antibody with enzyme-linked immunosorbent assay(ELISA).The samples which were positive for the core antibody were subjected to real-time polymerase chain reaction (PCR) for the hepatitis B DNA detection.Results Among the 9100 donors,911 (10.01％) donors were positive for the core antibody.The hepatitis B surface antigen was positive in 199 (2.19％) donors.Among the 911 donors who were positive for the core antibody,820 (90.01％) donors were negative for the HB-sAg,and 35 donors were positive for hepatitis B DNA.Conclusions If a routine screening of the sera for the core antibody is not done,the HBV DNA viraemia may not be identified.The absence of the surface antigen in the blood of apparently healthy individuals may not be sufficient to ensure the lack of the circulating virus.It is necessary to attach importance to the blood donors screening HBsAg-negative population further serological testing,suspicious specimens should detect HBV DNA.

Objective To analyze the patterns of amino acid changes in liver failure patients treated with non-bioartificial liver support system (ALSS), and to explore the efficacy of ALSS in liver failure treatment. Methods A total of 146 liver failure patients treated with ALSS from June 2009 to August 2010 were recruited in this study. Paired blood samples were collected from every patient and serum amino acids and ammonia were tested by automatic amino acid analyzer. The changes of amino acids in patients with different prognoses, different types/phases of liver failure were evaluated.Measurement data were compared by paired t test. Results After ALSS treatment, liver failure patients experienced a significant decrease in serum glutamic acid and lysine [(395.62±200.24)μmol/Lvs (260. 05±169.56) μmol/L and (436. 73±326. 18)μmol/L vs (407. 12±292.01) μmol/L,respectively; t= 8. 611 and 2. 659, respectively; both P＜0.01)], while experienced greatly increases in threonine and branched-chain amino acids/aromatic amino acid ratio [( 1302. 90 ±1288.70) μmol/L vs (1406.70 ±1272. 34) μmol/L and 1. 23 ± 0. 53 vs 1. 36 ± 0.57, respectively; t = 2. 895 and 1. 061,respectively; both P＜0. 01)]. The changes of glutamic acid, tyrosine, arginine and methionine before and after ALSS treatment in patients with different prognoses, different types/phases of liver failure were all significantly different. Conclusions ALSS treatment could improve the serum amino acid disorder in liver failure patients. The amino acids in patients with different types/phases or different prognoses of liver failure change significantly after ALSS treatment.

Objective To establish and improve the acute hepatic failure model in pigs induced with D-galactosamine (D-gal),and explore the feasibility of evaluating preclinical artificial liver devices.Methods Nineteen Duroc breeding pigs were divided into 4 groups.Fifteen unanesthetic Duroc breeding pigs out of 19 (5 of each group) received intravenously administration of D-gal at a dose of 1.0,1.25 and 1.5 g/kg body weight,respectively.The remaining 4 pigs which received the same volume of 5% dextrose in water served as controls. Clinical data and survival time of pigs were recorded.Blood samples were collected for dynamic testing of plasma ammonia,prothrombin time,liver and renal functions,blood glucose and L-lactate;liver tissues were sampled for pathological examination.The differences between groups were compared using t test and F test.The survival time of pigs was compared by Kaplan-Meier survival analysis and Log Rank test.Results Twelve hours after administration of D-gal,all pigs presented as acute hepatic failure characterized by progressive increases of levels of plasma ammonia,aspartate aminotransferase (AST),total bilirubin (TBil) and L-lactate,the level of blood glucose marked decreased and prothrombin time prolonged (F= 32.33,F=27.817,F=50.097,F=88.382,F=8.211,F=21.227;all P＜0.01);especially in the pigs which received D-gal at a dose of 1.5 g/kg.Except 2 pigs survived for 168 h,the other 3 pigs which received D-gal at a dose of 1.0 g/kg died within 68-84 h,while all pigs which received D-gal at a dose of 1.25 and 1.5 g/kg died within 33-89 h and 23-47 h,respectively.All pigs presented coma before death and liver histopathological examination indicated massive hepatic necrosis with severe hemorrhage.Conclusions D-gal induced acute hepatic failure model in unanesthetic Duroc breeding pig appears potential reversibility and high reproducibility,which has proper therapeutic window.Thus,this model could be applied to evaluate the therapeutic effects and safety of artificial liver devices.

Objective To investigate the ecological structure of intestinal fungal flora of patients with chronic hepatitis B infection. Methods HBV-liver cirrhosis patients, chronic hepatitis B patients, HBV carriers, and healthy volunteers were selected as research subjects to extract the total DNA from stool of each subject, 18S rRNA genes of intestinal fungi was cloned and sequence was analyzed for the construction of 18S rDNA clone libraries and phylogenetic tree, diversity and structural characteristics of intestinal fungal flora in each group was then analyzed. Results A total of 29 operational taxonomic units (OTUs) of intestinal fungal flora in all research subjects was acquired. All positive clones belonged to three fungal taxa: Zygomycetes (3.4%) , Ascomycetes (82. 8%) and Basidiomycetes (13. 8%) . The dominant fungal community was Candida spp. , uncultured fungus and Saccharomyces spp. , which accounted for 29. 2% , 15. 9%, 15.0% respectively. The OTUs which belonged to HBV-liver cirrhosis patients, chronic hepatitis B patients, HBV carriers, and healthy volunteers was 20,16,12,14 respectively. Conclusion There is an abundant fungal flora in the human intestine. Ecological structure of intestinal fungal flora appears to be various in patients with chronic hepatitis B infection, which indicate the close relationship between the alteration of ecological structure of fungal flora and the stage of chronic hepatitis B infection.

Objective To investigate the efficacy and safety of adefovir dipivoxil (ADV) in treatment of chronic hepatitis B (CHB) patients with lamivudine (LAM) resistance. Methods There were treatment group (32 CHB patients with LAM resistance) and historical control group (24 CHB patients with LAM resistance) in this study. The treatment group received ADV 10 mg/d and LAM 100 mg/d for 48 weeks; the historical control group continued to use LAM monotherapy. During the treatment causes, serum HBV DNA levels, liver function and HBV serology were monitored regularly, and safety assessments were also conducted. Results In treatment group, mean HBV DNA levels decreased by 2.56 log10 eopies/ml and 2.93 log10 copies/ml, virus response rates were 50. 0% and 75.0%, ALT normalization rates were 53.1% and 68.8% after 24 and 48 weeks of treatment, respectively. The histological improvement rate was 65.6% after 48 weeks. Comparing with those in control group, the differences were statistically significant ( P <0. 05), while there was no significant statistical differences in HBeAg loss rate and HBeAg seroconversion rate between two groups. There was no severe adverse event during the treatment. Conclusion ADV is effective and safe in treatment of lamivudine-resistant CHB.

Objective To study the dynamics changes of cerebrospinal fluid (CSF) bacterial load within 48 h after infection in a rabbit meningitis model, and provide information for diagnosis,treatment and prognosis of this disease. Methods Taking New Zealand white rabbit as the study object, meningitis model was established via cerebellar cistern puncture with different concentrations of Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) to explore the relationship between the mortality of animals and the subarachnoid inoculation dosage. The dynamics study of CSF bacterial load was conducted with proper inoculation bacterial dosage. Forty-eight rabbits were separated into four groups (12 each group): E. coli meningitis model group, E. coli meningitis + ceftriaxone treated group, S. aureus meningitis model group and S. aureus meningitis + vancomycin treated group. At 0,12, 24, 36 and 48 h of inoculation, CSF and blood samples were obtained for CSF bacterial quantitative culture, CSF leukocyte count and peripheral blood leukocyte count. Finally, the relationships between the early mortality of animals, the efficacy of antibiotics, CSF leukocyte counts and the dynamics changes of CSF bacterial load were analyzed in the bacterial meningitis rabbit model.The CSF bacterial load and the white blood cell count curve were compared by analysis of covariance (ANOVA). Correlation test was done using correlate partial analysis. Results The relationship between subarachnoid inoculation dosage and the mortality of rabbits presented S-curve correlation.The bacterial load in subarachnoid space peaked in 12-24 h after infection and then gradually decreased. Effective antibiotic therapy could significantly speed up the decline of this process. There were significantly different between E. coli meningitis model group and E. coli meningitis+ceftriaxone treated group (F= 27. 10, P<0. 01), between S. aureus meningitis model group and S. aureus meningitis + vancomycin treated group (F=5. 97, P = 0. 016). There was a positive correlation between CSF bacterial load and CSF leukocyte count in E. coli and S. aureus meningitis model groups (r=0. 89, 0.84, respectively; P = 0.046, 0.049, respectively). Conclusions In the treatment of bacterial meningitis, effective and sufficient antibiotics should be used as soon as possible to control the CSF bacterial load and reduce the mortality. The CSF leukocyte count can be used as indicator of CSF bacterial load and guide the antibiotic treatment in clinical bacterial meningitis.

Objective To establish normally immortalized porcine hepatocyte lines by ectopic expression of simian virus 40 large T (SV40LT) antigen and the human telomerase reverse transcriptase(hTERT). Methods Primary porcine Hepatoeyte cells were transfeeted with recombinant retrovirus containing SV40LT or hTERT respectively. Subsequently drug resistant cell clones were screened and expanded for further studies. Immortalized porcine hepatocyte was confirmed by examination. Results The morphological phenotype of the transfected cells was similar to the primary porcine hepatocyte. One clone, HepLP, has been maintained in cultue for half year, and expanded by more than 60 passages. SV40 LT and hTERT could be detected in transfected porcine hepatocyte. Pig albumin mRNA was also detected by RT-PCR. No tumor formation occurred when HepLP cells were injected into Balb/c nude mice. Conclusions The immortalized, nontumorigenic, porcine hepatoeytes maintained the properties of porcine primary hepatocytes such as the albumin secretion. This generation of immortalized porcine hepatocyte may be helpful for bioartifical liver support system, hepatocytes transplantation, drug/toxicological studies, and liver biologic studies.