Objective To investigate the effect of livzon weisanlian(bismuth agent 0.22g,bid+tinidazole,0.5g,bid+clarithromycin 0.25g,bid) for 7 days for eradication of H.pylori and infection of patients suffering from chronic gastritis and peptic ulcer in Huannai area.Methods Four pieces of gastric mucosa were taken out by gastroscopy,two pieces of them from gastric antrum 3～5cm near pylori and the others from gastric body.All of them were detected for H.pylori and biopsy and smear.If both the former and the one of the later are all positive the patient was diagnosed with H.pylori and treated with livzon weisanlian.Results The positive of CSG is 81.6%,CAG is 63%,GU is 90.8% and DU is 93.3%.Conclusion The positive rate of patients suffering from chronic gastritis and peptic ulcer in Huainan area is high.The eradicative rate with livzon weisanlian is also high and it shows that livzon weisanlian is feasible to treat chronic gastritis and peptic ulcer.

Objective To establish mixed-sandwich ELISA detection system by screening aptam-ers of MPT64 antibodies with SELEX to detect clinical serum samples, and explore the potential laboratory diagnosis value of this method. Methods To detect the affinity of the final round ssDNA library to MPT64 antibodies inhibited by MPT64 antigen with the competitive ELISA method, optimize the mixed-sandwich ELISA detection method that was aptamer-serum-horseradish peroxidase labeled goat anti-human IgG anti-body detection system to detect 230 cases of clinical serum samples as well as the lowest concentration of MPT64 antibodies and the linear range. Results In competitive ELISA test results, the percentage of inhi-bition effect of MPT64 antigen to final round ssDNA library is from 0.25% to 80% when the MPT64 antigen concentration rised from 2 μg/ml to 256 μg/ml. The Optimized detection system of mixed-sandwich ELISA was constitute of the concentration of ssDNA coated with 0.1μg/hole, serum dilution of 1/200, horseradish peroxidase labeled goat anti-human IgG antibody concentration of 1/40 000. The lowest concentration of MPT64 antibody is 3 mg/L and the linear range is between 10 mg/L and 1000 mg/L. The serum samples of 100 cases of tuberculosis patients, 100 healthy individuals and 30 cases of non-tuberculesis were tested in this system and the test result was analyzed with Graphpad Prism, the difference of tuberculosis group and healthy group was statistically significant (P<0.001 ), the difference of TB group and non-TB control group was also statistically significant (P<0.001). The specificity and the sensitivity was 96.1% and 31.0% re-spectively. Conclusion The aptamer mixed-sandwich ELISA method will play an important role in the sero-logical diagnosis of tuberculosis.

OBJECTIVE To study the prevalence of aac(6′)-Ⅰb with its subtypes,one kind of gene for aminoglycoside-modifying enzymes in Escherichia coli.METHODS DNA sequences were analyzed in those strains with aac(6′)-Ⅰb,compared to its family marked in NCBI.Thus,its subtypes were determined.RESULTS Four strains were classical on genotype,the other one with aac(6′)-Ⅰb-Cr,and the last one with a new subtype among the 6 strains of E.coli with aac(6′)-Ⅰb.CONCLUSIONS There exist at least 3 subtypes of aac(6′)-Ⅰb,one kind of gene for aminoglycoside-modifying enzymes in local strains of E.coli.Among those subtypes,the classical type is the main,accompanied by aac(6′)-Ⅰb-Cr and its new subtype.

Objective To investigate drug resistance,especially the resistance to aminoglycoside and the prevalence of the genes for aminoglycoside-modifying enzymes in Escherichia coli producing ESBLs.Methods VITEK-32 GNI+ cards and K-B methods were used to identify E.coli and detect the resistance to 18 kinds of antibiotics such as aminoglycoside and ?-lactams respectively in E.coli isolated from our hospital from January to December,2006.PCR method was used to detect the genes for aminoglycoside-modifying enzymes such as aac(3)-Ⅰ,aac(3)-Ⅱ,aac(6')-Ⅰb,aac(6')-Ⅱ,ant(3″)-Ⅰ and ant(2″)-Ⅰ in those 17 strains producing ESBLs.Results Resistance rates were 100.0%,100.0%,41.6%,100.0%,69.5%,30.5%,98.2%,81.0%,81.0%,79.6%,79.2%,34.8%,15.4%,7.1%,7.9%,5.4% and 81.0% for ampicillin,ampicillin/sulbactam,aztreonam,cefazolin,cefepime,ceftazidime,ceftriaxone,ciprofloxacin,levofloxacin,gentamicin,tobramycin,amikacin,cifoxitin,nitrofurantoin,piperacillin/tazobactam,cefoperazone/sulbactam and co-trimoxazole respectively in E.coli producing ESBLs.No drug-resistant strain imepenem was found.Positive rate was 46.4% for ESBLs.Resistance rates were 52.9%(9/17),100.0%(17/17) and 100%(17/17) for amikacin,gentamycin and tobmycin respectively in those 17 strains producing ESBLs.The most genotype for aminoglycoside-modifying enzymes was aac(3)-Ⅱ(94.1%),the second was aac(6')-Ⅰb(35.3%).The positive rates of aac(3″)-Ⅰ and ant(2″)-Ⅰ were 11.8% and 5.9% respectively.of them,one strain was resistant to aminoglycoside on phenotype,but no genes mentioned above was detected.4strains were classical on genotype,one with aac(6')-Ⅰb-Cr,the other one with a new subtype among the 6 strains of E.coli with aac(6')-Ⅰb.Conclusions Drug resistance in E.coli is rather serious.Typically,E.coli producing ESBLs shows multi-drug resistance.Positive rate for aac(3)-Ⅱ takes the first place,followed by aac(6')-Ⅰb among the genes for aminoglycoside-modifying enzymes.Aac(3″)-Ⅰand ant(2″)-Ⅰ show both lower positive rates.Aac(3)-Ⅰ and aac(6')-Ⅱ were not found.There exist at least 3 sub-types of aac(6')-Ⅰb,one kind of gene for aminoglycoside-modifying enzymes in strains of E.coli.Among those sub-types,the classical type is the main,accompanied by aac(6')-Ⅰb-Cr and its new sub-type.

OBJECTIVE To investigate drug resistance of Escherichia coli,especially the prevalence of the genes for TEM and SHV type ?-lactams in ESBLs-producing E.coli. METHODS A total of 2277 strains of E.coli collected in our hospital from Jan 2006 to Nov 2008 were analyzed for drug resistance to 21 kinds of antibiotics such as aminoglycoside,?-lactams and quinolones using VITEK-32 GNS-145 or GNS-448 card following GNI + for identification.PCR methods were used to analyze TEM and SHV type ?-lactams genes in 36 strains of E.coli with ESBLs so as to study their prevalence. RESULTS From them 1025 strains of E.coli were found positive for ESBLs,with a positive rate of 45.0%.Drug resistance rates were 83.0%,74.0%,52.0%,52.0%,29.1%,27.8%,16.5%,47.0%,31.0%,54.5%,53.0%,56.8%,10.0%,53.9%,38.5%,5.0%,4.0%,5.0% and 5.0%,respectively,for ampicillin,ampicillin/sulbactam,cefazolin,cefuroxime,aztreonam,cefepime,ceftazidime,ceftriaxone,cefotaxime,ciprofloxacin,levofloxacin,gentamicin,amikacin,tobramycin,Co-trimoxazole,nitrofurantoin,piperacillin /tazobactam,cefoperazone/sulbactam and cefoxitin.None was found resistant to imipenem or meropenem.Twenty-five and 14 strains of E.coli were found positive for TEM and SHV genes,respectively(69.4% and 38.9%). CONCLUSIONS Drug resistance of E.coli is severe.The local E.coli with ESBLs shows high positive rates of TEM and SHV genes,which play an important role in its severe drug resistance.

Objective To construct the recombinant plasmid of protein CFP10-MPT48-TB8.4 of Mycobacterium tuberculosis and to investigate the diagnosis potential of this fusion protein in tuberculosis serodiagnosis.Methods The recombinant fusion protein CFP10-MPT48-TB8.4 was expressed, and identified by Western blot.The ELSIA based on the purified fusion protein was done,and used for screening in 230 cases of clinical serum samples including pulmonary tuberculosis patients ( n =150 ),pulmonary disease patients other than tuberculosis (n =70) and health controls (n =103 ).The test result was analyzed by Medcale11.5 software.Results The fusion protein CFP10-MPT48-TB8.4 was successfully expressed with a purity over 95％.Specific immunogenicity of the recombinant protein was confirmed by Western blot.The overall sensitivity and specificity obtained of ELISA were 56.7％ (85/150) and 90.8％ ( 157/173 ),respectively.The specificity was 85.7 ％ (60/70) in non-tuberculosis group and 94.2％ (97/103 ) in healthy group,respectively.Conclusion The recombinant protein of CFP10-MPT48-TB8.4 has a high sensitivity and specificity and may be a potential candidate antigen in tuberculosis serodiagnosis.

Objective To obtain DNA oligonucleotide aptamers which can specifically bind to MPT64 protein from Mycobacterium tuberculosis by SELEX(systematic evolution of ligands by exponential enrichment)technology. Methods A random ssDNA library with in vitro synthesized 78 nucleotides in length was subiected to 12 rounds of selection by SELEX method against MPT64 protein. The binding ability of the aptamers to the protein was examined by biotin-streptavidin-horseradish peroxidase system. Results The selective system used was as follows:in PCR amplification,annealing temperature was 65℃ and the concentration of Mg2+ was 1.5 mmol/L in optimizing library, and when preparation of ssDNA with asymmetrical PCR amplification, 0.75 mmol/L of Mg2+ was used. When using the plate for ELISA as the substrate for the selection, the pattern of electrophoretic band of PCR product after the tenth round selection became unitary and denser than that of the first round. The binding assay demonstrated that A value at 450 nm of the tenth round increased by 9.18 times as compared with that of the first round. The results showed that the affinities of the aptamers were different. The highest A value at 450 nm was 1.606, and the lowest 0.572. Conclusion A set of aptamers with considerable binding affinity to MPT64 protein are successfully picked out from the initial random DNA library.

Objective To develop an assay to determine Mycobacterium tuberculosis resistance to rifampin, isoniazid, ofloxacin and amikacin by pyrosequencing and evaluate the value of this method in clinical application. Methods Eighty-nine clinical isolates of Mycobacterium tuberculosis from tuberculosis patients were collected from Shanghai Pulmonary Hospital during 2008 to 2009. All strains were isolated from decontaminated sputum, cultured on Lowenstein-Jensen media and identified by traditional biochemical tests with standard methods. Ten Mycobacterium tuberculosis were selected from the strain bank of Shanghai Pulmonary Hospital. The optimal conditions of detection of rpoB, katG, gyrA and rrs gene by pyroseuencing were determined, using the 10 Mycobacterium tuberculosis strains whose drug susceptibility of Bactec 960 and sequence of rpoB, katG, gyrA, rrs gene were known. Then the drug susceptibility of 89 Mycobacterium tuberculosis clinical isolate strains were detected by pyrosequencing using this conditions and the results were compared with that of the Bactec 960 methods. Results The pyrosequencing program of sequence analysis was suitable for the detection of the mutations of rpoB and gyrA genes, and the program of single nucleotide polymorphism was suitable for katG and rrs genes. Among the 89 Mycobacterium tuberculosis clinical isolates,when using the drug susceptibility of Bactec 960 as the standard, the sensitivity of rifampin, isoniazid,ofloxacin and amikacin is 98.0%, 64. 1%, 79.5%, 78. 3% respectively, the specificity is 97.5%,100. 0%, 90. 0%, 100. 0% respectively, the accuracy is 97.8%, 77. 5%, 85.4%, 94. 4% respectively,tested by pyrosequencing. The results of the detection of resistance to rifampin, isoniazid, ofloxacin and amikacin in Mycobacterium tuberculosis using pyrosequencing technique were almost the same with that of Bactec 960, and Kappa ≥0. 7 in each detection. Conclusion Pyrosequencing is thus a rapid, accurate and high throughput method for detecting Mycobacterium tuberculosis resistance to these four drugs.

Objective To analyze risk factors and complication characteristics of healthcare-associated infection (HAI)in patients with lung cancer,and provide evidence for the formulation of HAI management strategy. Methods HAI-related articles were retrieved from China Biology Medicine (CBM),China National Knowledge Infrastructure (CNKI),Wanfang database,Vip database,PubMed,and Embase,all data were conducted Meta-analysis.Results A total of 19 articles involving 8 069 hospitalized patients with lung cancer (1 280 had HAI)were included.Meta-analysis on combined values of medical factors for HAI were as follows:OR(95%CI )of anti-tumor therapy(radiotherapy and chemotherapy),number of chemotherapy (≥ 2 times ),antimicrobial prophylaxis, immunosuppressant therapy,and invasive operation were 3.13 (1 .82,5.39),9.20 (3.04,27.87),3.23 (1 .77, 5.91),2.00(1 .56,2.57),and 2.28(1 .81 ,2.88),respectively;Meta-analysis on combined values of complication factors for HAI were as follows:OR (95% CI )of pulmonary diseases,chronic obstructive pulmonary disease (COPD),diabetes,renal dysfunction,malnutrition,hypoalbuminemia,neutropenia,and leukopenia were 2.65 (1 .74,4.02),2.40 (1 .76,3.27),2.25 (1 .85,2.73 ),2.56 (1 .18,5.52),5.51 (1 .70,17.89),2.05 (1 .56, 2.70),3.38(1 .40,8.18),and 2.10 (1 .22,3.62),respectively.Conclusion HAI-related factors of medical treat-ment and complications in patients with lung cancer are diversity,risk factors for HAI in patients with lung cancer are anti-tumor therapy,immunosuppressant therapy,antimicrobial prophylaxis,invasive operation,pulmonary dis-eases,COPD,diabetes,renal dysfunction,malnutrition,hypoalbuminemia,neutropenia,and leucopenia.

Objective To clone and express of Rv0091 encoding protein in Mycobacterium tuberculosis,identify and characterize of the enzyme activities.Methods Construct the Rv0091 prokaryotic expression plasmid,the vector was transformed into E.coli strain BL21trxB.After induced by IPTG,recombinant protein was purified by Ni2+-NTA chromatography and analyzed for purity by SDS-PAGE gels stained with Coomassie Blue.Immunological activity was identified by Western blot.The recombinant protein molecular weight was identified by Mass spectrometry.The enzyme-coupled assay detectes enzyme activity.Results The expression plasmid pET32a-Rv0091 was constructed and expressed in E.coli.BL21trxB,and the optimum expression system was conformed.The purity of the recombinant protein was more than 95％.Western blot analysis confirmed that recombinant protein was one of Mycobacterium tuberculosis proteins.Mass spectrometry identified the relative molecular weight and theoretical molecular weight was basically the same.Enzyme assay showed the recombinant protein able to catalyze the substrate MTA.Enzymatic properties showed that the optimal buffer for the phosphate and Hepes buffer,the poor thermal stability of the enzyme,the optimal temperature of 37℃,optimal pH10-12,when the pH ≤7,the protein denaturation and loss of some vitality.Conclusion The recombinant protein methylthioadenosine nucleosidase(MTAN) was obtained and enzyme activity was detected and plays a key role in the metabolism of Mycobacterium tuberculosis.