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Objective:To establish a rapid method to detect drug-resistance genotypes of extended spectrum-β-Lactamases (ESBLs) produced by gram negative bacillus using the real -time fluorescence quantitative PCR. Methods: According to clinical common genotypes of ESBLs, SHV, TEM.CTX-M.OXA and their homology, 9 pairs of specific primers were designed including SHV, TEM, CTX-M-1, CTX-M-2, CTX-M-8, CTX-M-9, OXA-1, OXA-2 and OXA-10. To extract DNA template by boiling assay, and then establish and grade up SYBR GREEN I real-time fluorescence quantitative PCR reaction system, finally definite real-time fluorescence quantitative PCR method. Its precision and range of linearity were tested. With established assay 51 multi- drug resistant ESBLs- E. coli K. pneumoniae were detected and compared with improved three dimensional extract tests. Results: Except OXA-2, 8 genotypes SHV, TEM, CTX-M-1, CTX-M-2, CTX-M-8, CTX-M-9, OXA-1 and OXA-10 were amplified by quantitative PCR from 39 ESBLs+ and 51 multi-drug resistant ESBLs-E. coli K. pneumoniae and confirmed by sequence testing. The range of linearity was 3×10~3-3×10~8 copies/mL, r =-0.994 7. Repetitive experiments showed that the average coefficient of variation between -runs was 9.6%. Comparing with three dimensional extract test, there was no significant difference (χ2 = 1.125,P> 0.05). Conclusion: Testing drug-resistance genotypes of ESBLs with SYBR GREEN I real-time fluorescence quantitative PCR is a rapid,specific and sensitive method, which is capable of inspecting genotypes of ESBLs from clinical strains.
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OBJECTIVE To investigate the genotype characteristics of extended-spectrum ?-lactamases(ESBLs) producing Escherichia coli and Klebsiella pneumoniae and drug resistance condition in Tianjin.METHODS A total of 218 strains of ESBLs-producing E.coli and K.pneumoniae were isolated from clinic.The drug-resistant CTX-M-1,CTX-M-2,CTX-M-8,CTX-M-9,TEM,SHV,OXA-1,OXA-2 and OXA-10 were tested by SYBR GREEN Ⅰ real-time fluorescent quantitative Polymerase chain reaction(PCR).RESULTS Among 109 strains of ESBLs-producing E.coli,the drug resistance of genotypes CTX-M-1,CTX-M-2,CTX-M-8,CTX-M-9,TEM,SHV and OXA-1 was 32.1%,0.9%,0.9%,33.9%,73.4%,27.5% and 15.6%,respectively.The strains with more than two genes were 66.1%.The positive rate of CTX-M-1,CTX-M-2,CTX-M-8,CTX-M-9,TEM,SHV,OXA-1 and OXA-10 was 49.5%,2.8%,1.8%,22.9%,78.9%,76.1%,33.0% and 0.9%,respectively.The strains with more than two genes were 82.2%.The strains were highly sensitive to imipenem.CONCLUSIONS Most of the genotypes of ESBLs-producing E.coli and K.pneumoniae in Tianjin are SHV,TEM,CTX-M-1 and CTX-M-9.The rate of OXA are increasing,and the drug resistance genes of K.pneumoniae from different hospitals are different.
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OBJECTIVE:To understand the status quo adverse drug reactions(ADR) occurred in our hospital and to promote clinical use of drugs.METHODS:By retrospective analysis,study 73 ADR cases collected from our hospital in 2009 were analyzed in respect of gender and age of patient,route of administration,category of drugs,organs and systems involved in ADR and its clinical manifestation.RESULTS:Of total 73 ADR cases,35.6% of cases were induced by anti-infective agents and 9.6% traditional Chinese medicines.Most of cases were caused by intravenous administration.Main clinical manifestations were lesion of skin and its appendents.CONCLUSION:Rational use of anti-infective agents and traditional Chinese medicine(TCM) should be strengthened.Quality control and standard application of TCM injection should be enhanced to reduced or avoid the incidence of ADR.
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Objective:To observe the effects of Astragalus polysaccharides on antigen presentation of dendritic cells(DC).Methods:DC and CIK cells were generated by culturing PBMC of healthy blood donor.The typical DC and DC pulsed by Astragalus polysaccharides were co-cultured with CIK cells,respectively.Then the changes in the phenotypes of the DCs pulsed with Astragalus polysaccharides were determined.Cell surface markers were analysed by FACS method.IFN-? and IL-12 secreted by co-cultured cells were detected by ELISA.The cytotoxicity of effector cells on A549 cells in vitro were measured by MTT assays.Results:Astragalus polysaccharides could increase the expression of CD40,CD80 and HLA-DR on DC surface.The Astragalus polysaccharides pulsed DC-CIK cells resulted in an enhanced killing activity to A549 cells than that of unpulsd DC-CIK cells(P