Objective To establish a method of high resolution melting analysis (HRM) with unlabeled probe for detection of rt204 mutation in HBV P gene.Methods Plasmids with wild strain rt204M,mutant strains rt204I and rt204V were constructed,and the probes were designed and optimized.HRM plots were established by the constructed plasmids.A total of 185 samples were collected from patients with chronic hepatitis B in the Affiliated Hospital of School of Medicine of Ningbo University during May 2010 and May 2012.All samples were detected by HRM,and matched with characteristic melting pattern.rt204 mutations were screened,and then verified by DNA sequencing.Paired x2 test was used for the comparison of the detection of mutations.Results The melting temperatures for rt204I,rt204V and rt204M were 58.0℃,60.6℃ and 62.5℃,respectively.Among 185 samples,168 samples (90.8％) could be analyzed by HRM method,and 155 samples (83.8％) coule be successfully sequenced (P ＜0.01).In the 155 samples which were completely analyzed by HRM assay and sequencing,75 samples were rt204M,55samples were rt204I,and 25 samples were rt204V by using HRM method,with an overall mutation detection rate of 51.6％ (80/155) ; and by sequencing,110 samples were rt204M,30 samples were rt204I,and 15samples were rt204V,with an overall mutation detection rate of 29.0％ (45/155).The difference onmutation rates detected by the above two methods was of statistical significance (P ＜ 0.01).Conclusion HRM with unlabeled probe is simple,sensitive,rapid and specific for detection of rt204 mutation in HBV P gene.

The experiment contents were integrated into three parts such as basic skills and verification experiment, systematic experiment and designing experiment. Basic skills and confirmato-ry experiments were performed alone with theory teaching by combination of modern teaching methods and traditional teaching methods, which consisted of “speaking”(experimental principles, methods and main technical points using multimedia), “looking”(demonstrating related operation on teaching website), teachers' demonstration, students' doing experiment independently and summarizing. In this part, the experimental operation skills such as the sterile operation technology, staining technology, microscopy technology and pure culture were emphasized. Systemic experiments would be carried out after completion of the most theory, the experiment time could be adjusted according to the experiment content, and the PBL teaching method was taken in this stage. After the theory teaching of Medical Microbiology was finished, students voluntarily participated in design experiments in the last stage, which were the fusion of scientific research subject and the experimental teaching. From the preview experiment, experiment operation, experiment report, to the final test, the multi-dimensional evalua-tion was implemented throughout the course of experiment teaching.

0bjective To analyze the factors related tO the changes of liver pathology in hepatitis B e antigen (HBeAg) negative chronic hepatitis B (CHB) patients.Methods Two hundred and eighty-eight HBeAg negative CHB patients were enrolled.Bivariate Pearson correlation was employed to analyze the correlation of different age,gender,alanine aminotransferase (ALT) level,the immunohistochemistry results of hepatitis B surface antigen (HBsAg) and hepatitis B core antigen (HBcAg) in liver tissue with hepatitis B virus (HBV) DNA level,changes of liver pathology.Results The inflammation grades and fibrosis stages of livertissuesinmale were 1.72±1.23 and 1.71±1.24,respectively;1.25±1.39 and 1.21±1.40 in female,respectively,which were significantly different (t=2.398,t-2.551;both P40 years old were all significantly higher than those<40 years old(t一2.060,t一2.536,t=2.808;all P<0.05).In patients with normal ALT levels,75 cases (52.03%) showed HBV DNA level≤1×103 copy/mL,56(38.89%) showed≥G2 of liver tissues; the ALT level and inflammation grade of liver tissues was significantly correlated(r:O.244,P-O.004).In patients with elevated ALT 1evels,42 cases (31.57%) showed HBV DNA level≤1×10s copy/mL,89 (66.92%) showed≥G2 of liver tissues; the ALT level and inflammation grade of liver tissues was not significantly correlated (r=0.007,P=0.939) .The inflammation/fibrosis of liver tissues,HBV DNA levels in patients with both HBsAg and HBcAg positive in liver tissues were significantly higher than those in other patients (P

Objective To explore the prevalence of 11 efflux pumps in isolates of multidrugresistant Escherichia coli(MDR-ECO). Methods Efflux pumps emrB, emrD , emrE, mdfA, sugE, mdtl,tehA, oqxA, qacE△1, qacE and smr-2 were detected by polymerase chain reaction(PCR)in 20 MDR-ECOs isolated from clinical samples. Results Efflux pumps emrB, emrD, emrE, mdfA, sugE, mdtI, qacE △1, tehA and oqxA were detected, and 8 efflux pumps were found in the same strain. Conclusion Multidrug- resistance in Escherichia coli may be related with efflux pumps.

Objective To investigate the correlation between acquired drug resistance-related genes and mobile genetic elements from pandrug-resistant Acinetobacter baumannii. Methods Fifty-three horizontal transfer drug resistance-related genes ( β-lactamases,aminoglycoside and quinolones resistance related) and 12 mobile genetic elements (including zygosity plasmid,transposon,insertion sequence and integron) were detected by polymerase chain reaction (PCR) in 20 clinical isolates of pandrug-resistant Acinetobacter baumannii.Index cluster analysis was performed to explore the correlation.Results In 20 strains of pandrug-resistant Acinetobacter baumannii,there were 3 types of β-lactamases related genes (TEM-1,ADC-30,OXA-23 ),4 types of aminoglycoside modifying enzyme genes [ aac (3)-Ⅰ,aac(6′)-Ⅰ b,ant( 3″)-Ⅰ and aph( 3′)-Ⅰ ],and 5 kinds of mobile genetic elements ( int Ⅰ 1,tnpU,tnp513,IS26 and ISAba1 ). Index cluster analysis showed high correlations between resistance genes [TEM-1,ADC-30,aac( 6′)-Ⅰ b,ant( 3″)-Ⅰ,abeB,qacE Δ1] and mobile genetic elements ( int Ⅰ 1,tnpU,tnp513,IS26,ISAba1 ).Conclusion Clinical isolated pandrug-resistant Acinetobacter baumannii carries several acquired drug resistance-related genes and mobile genetic elements,and there may be a close association between them.

Objective To study effects of low molecular weight heparin on liver fibrosis and the serum levels of TGF-?_1 in patients with chronic hepatitis B virus.Methods 45 patients with chronic hepatitis B virus were randomized into control group(the routine strategy)and trial group(the routine strategy + low molecular weight heparin).The period of treatment is 3 weeks.Serum hepatic fibrosis indices before and after heparin treatment were examined by RIA,the levels of serum TGF-?_1 before and after heparin treatment were examined by ELISA.Results Hepatic functions became significantly better in trial groups,serum PⅢP and type IV collage levels and the levels of serum TGF-?_1 in trial group decreased significantly after treatment.Conclusions The mechanism of anti-fibrosis action of LMWH may inhibited production of TGF-?_1 in patients with HBV.

Objective To explore the depression status of the liver transplant recipients after the surgery and the impact factors.Method Forty two liver transplant recipients were assessed at 5 different time points(1 month,6 months,12 months, 24 months,36 months after the transplantation)using general information questionnaire(GIQ),social support rating scale(SSRS) and self-rating depression scale(SDS).Results After the transplantation,the longer recipients lived,the less depression they underwent.Age and post-operative objective support are the two major factors on depression status of the liver transplant recipients Conclusion The clinical and community nurses may pay attention to the mind state of the recipients after liver transplantation,and provide mental nursing in time and social support to prevent and improve the state of their depression.

Objective To explore the effect of virtual laboratory (VL) + flipped classroom in teaching of virus infection diagnosis.Methods 40 students of Class One from clinical medical undergraduates of Grade 2014 were randomly taken as the experimental group,with 40 students of Class Two as the control group.The experiment group adopted flipped classroom teaching by virtual lab platform and classroom activities,while the control group adopted traditional classroom teaching such as watching video and lecturing.Finally post-test scores were compared by the independent samples t-test of SPSS 18.0 statistical software between the two groups.The teaching effects were evaluated through questionnaires survey in experimental group.Results The scores (82.73 ± 2.62) of comprehensive assessments were superior to the control scores (57.94 ± 4.65).Difference between the two groups was statistically significant (t=29.380,P=0.000).Students' satisfaction concerning the teaching methods and effects of the flipped classroom in experimental group was up to 85％.Conclusion Flipped classroom based on internet virtual lab platform in teaching of virus infection diagnosis can enhance the teaching quality and improve students' learning enthusiasm and thinking ability.

Objective:To explore the effects of protoporphyrin Ⅸ (PpⅨ)-mediated photodynamic therapy (PDT) on induction of apoptosis and death in colon cancer cell and the underlying mechanisms.Methods:The cell killing effect of PDT on HCT116 cell was determined by cell counting kit (CCK).The cells were divided into a control group,a single light group,a single PpⅨ group,and a PDT group.Hoechst 33342 and flow cytometry was used to assess the cell apoptosis.Western blot was employed to analyze the expressions ofbd-2,bax,and caspase-3.Reactive oxygen species (ROS) was detected by flow cytometry.Results:The viability of HCT116 cell was decreased gradually with the increase of irradiation dose (P<0.05).Compared to the other 3 groups,ROS production,the number of apoptotic cells and the protein expressions ofbax and caspase-3 in PDT group increased,while bcl-2 expression was decreased (P<0.05).Conclusion:PpⅨ-mediated PDT can enhance the apoptosis in HCT116 cell,which may be related to mitochondrial apoptosis pathway.

Objective To evaluate the clinical efficacy and safety of different doses of prednisone combined with topiramate(TPM) in the treatment of infantile spasms(IS),in order to provide a new choice of the therapy of IS.Methods Fifty-six cases were collected in the Department of Neurology of Jiangxi Children's Hospital from May.2011 to Dec.2012.They were randomly divided into 2 groups:control group and trial group.The patients in control group took prednisone tablet of 1 mg/kg,2 times a day for 2 weeks;and the patients in trial group took prednisone tablet of 10 mg/ d,4 times a day for 2 weeks.In addition,TPM was used in both groups by initial dose 1 mg/(kg · d) or 2 times a day,and then was gradually increased to 3-5 mg/(kg · d) within 2 weeks.For those children in whom the spasms seizure completely ceased after 2 weeks,prednisone was then reduced by degrees to be discontinued for a 7-weeks course(extending to 4 weeks with the initial doses if spasms continued after 2 weeks).All patients underwent the assessment of spasms seizure and a 3-12 h video-electroencephalogram monitoring including wake and sleep states,which were performed before treatment,after 2 weeks and the end of the courses (7 or 9 weeks after treatment),respectively.Meanwhile,the side effects of the drugs during the treatment were recorded.The developmental quotient (DQ) tests of children with complete cessation of spasms more than 6 months were performed before treatment and after 6 months.All patients had been followed up for 2-18 months.Results 1.After 2 weeks of the therapy,the rate of cessation of spasms were 75.00％ (21/28 cases) and 28.57％ (8/28 cases) in the trial group and the control group,respectively,there was significant difference (x2 =12.087,P =0.001).And in the same term,the rate of complete resolution of hypsrrhythmia were 60.71％ (17/28 cases) and 21.43 ％ (6/28 cases),respectively,there was significant difference (x2 =8.928,P =0.003).At the end of treatment,the rate of cessation of spasms were 67.86％ (19/28 cases) and 35.71％ (10/28 cases) in the trial group and control group,respectively,there was significant difference (x2 =5.793,P =0.016).And in the mean time,the rate of complete resolution of hypsrrhythmia were 57.14％ (16/28 cases) and 14.29％ (4/28 cases),respectively,there was significant difference (x2 =11.200,P =0.001).2.Weight gain and increased appetite were the most frequent side effects.The incidence of side effects were 82.14％ (23/28 cases) and 67.86％ (19/28 cases) in the trial group and control group,respectively,there was no significant difference between the 2 groups (x2 =1.524,P =0.217).No death occurred in this clinical trail and no one discontinued the treatment protocol as result of the adverse events.3.The recurrence rate in the trial group and the control group were 31.82％ and 72.73％,respectively.And there was significant difference between the 2 groups (x2 =4.950,P =0.026).In the trial group,there were 9 cases with cessation of spasms more than 6 months,whose average business development values had no significant difference before and after treatment (t =2.271,P =0.053).Conclusion The efficacy of large-dose prednisone combined with TPM for IS was significantly better than that of conventional dose prednisone combined with TPM.