Objective To investigate the metabolism and function of the intestinal microbiota from liver cirrhosis patients.Methods Sixteen cases of liver cirrhosis and twenty normal individuals were selected , whose intestinal microbiota metagenomic DNA was extracted , followed by high-throughput Solexa sequencing and the bioinformatics analysis of metabo-lism and function annotation to compare the differences between the patients and normal subjects and find out about the cir -rhosis-related functions .Results The functional diversity was significantly reduced in the intestinal microbiota of cirrhotic patients.At the module or pathway level , the intestinal microbiota of patients showed an enrichment in metabolisms of drugs, essential amino acid , propanoate metabolism and inflammatory reaction , whereas an opposite tendency was observed in the metabolic ability of butyrate , bile acid and cell cycle .Conclusion Under the influence of liver cirrhosis , the growth environment in the intestine is destroyed , causing, the intestinal microbiota the exhibit some compensation to adapt to the changed intestinal micro-environment .

Objective To construct eukaryotic expression vector of wild type E 4F1 and the mutant deleting amino acid region 32-81, and to detect the interaction between wild type or mutant E 4F1 and p53 and to study the effect of E4F1 on the expression level of p21.Methods Wild type and mutant sequences of E 4F1 were amplified from the mammary library using standard PCR and recombinant PCR .The sequences were cloned into pXJ 40-MYC vector to generate the MYC-E4F1 and MYC-E4F1(Δ32-81) recombinant plasmids that were transfected into 293T cells and identified by Western blotting . FLAG-p53 and MYC-E4F1 or MYC-E4F1(Δ32-81) were co-transfected into 293T cells and immunoprecipitation assay was performed to detect the interaction of wild type or mutant E 4F1 with p53.Wild type and mutant E4F1 expressing vec-tors were co-transfected into osteosarcoma U2OS cells and the expression of p21was detected.Results Recombinant plas-mids of MYC-E4F1 and MYC-E4F1(Δ32-81) were successfully constructed.Both wild type and mutant E4F1 interacted with p53.Deletion of amino acid region 32-81 of E4F1 increased the interaction .The expression level of p21 was in-creased by wild-type E4F1, but not by mutant E4F1.Conclusion The eukaryotic expression vector of wild type E4F1 and its deletion mutant is successfully constructed .Both of them interact with p53.Deletion of amino acid region 32-81 of E4F1 increases the interaction .This study contributes to further studies on the regulation and mechanism of E 4F1 on p53.

Objective To establish a multiplex loop-mediated isothermal amplification(mLAMP)method for simultaneous detection of Salmonella,Vibrio parahaemolyticus (VPH)and Listeria monocytogenes (LM).Methods Three sets of mLAMP primers were designed to specifically target bcfD of Salmonella and tlh of VPH and iap of LM.The respective single LAMP assay of the three kinds of bacteria was developed,and the ratio of primer concentration was optimized to develop a multiplex LAMP system.The specificity and sensitivity of multiplex LAMP were observed.Results Turbidity monitoring results in real time suggests that the mLAMP was highly specific and amplification could be obtained within 45 min under isothermal conditions.The sensitivity of this mLAMP was found to be 300 fg/μl genomic DNAs for Salmonella and 4.2 pg/μl for VPH and 4.5 pg/μl for LM,which was consistent with conventional PCR.Conclusion The mLAMP described can potentially facilitate simultaneous detection of three kinds of bacteria in a large number of food samples, which could be used as a primary screening method and as a supplement to classical detection methods.

Objective To establish a sensitive real-time quantitative PCR assay with TaqMan probe for rapid detection of mcr-1 gene in clinical isolated strains. Methods According to the mcr-1 gene sequence, a pair of specific primers and a TaqMan probe were designed. Moreover, a recombinant plasmid with mcr-1 gene was constructed as the positive standard. TaqMan probe-based fluorescence quantitative PCR assay was used to detect the colistin resistance gene mcr-1. The sensitivity, repeatability and specificity of the assay were evaluated. Results There was a good linear relationship between the initial template amount and Ct value (R2>0. 999). The lower limit of detection was 10 copies/μL, which was 100 times more sensitive than the conventional PCR. Results of test for specificity showed that only the strains carrying the mcr-1 gene were positive, while the remaining strains were negative. Coefficients of variation of intra-and inter-group repeatability tests were less than 1％. Two out of 150 clinical isolated strains carried mcr-1 re-sistance gene and both of them were identified as Escherichia coli. Conclusion TaqMan probe-based fluo-rescence quantitative PCR for the detection of colistin resistance gene mcr-1 was established with strong spe-cificity, high sensitivity and good repeatability. It could be used for the specific detection of clinical drug-re-sistant strains positive for mcr-1 gene and provide reference for pharmacotherapy.

Objective To establish a polymerase spiral reaction (PSR) method for rapid detection of influenza A/H1N1 virus.Methods Six sets of primers were designed for influenza A/H1N1 virus HA gene, and the results were determined with real time kinetic turbidimetric assay and colorimetry method.Results and Conclusion The best primers were selected from six sets of primers, and the best temperature was determined as 65 degrees Celsius.Further experiments showed that the best primer had good specificity for detection of influenza A/H1N1 virus,without cross reactions with 14 other respiratory tract pathogenic nucleic acids.The sensitivity was up to 100 copies,and consistent with that of PCR.So a PSR method is established for rapid detection of the influenza A/H1N1 virus, which is simple, quick, highly specific and sensitive,and especially applicable to field and grass-roots units.

Objective In this study, a strain of colistin and tigecycline-resistant bacteria isolated in 2009 was analyzed, and the structure of drug-resistant plasmid and genetic environment were discussed, so as to provide basis for the prevention and control of multidrug-resistant bacteria. Methods A strain (GZ12244) with positive mcr and tet(M) was obtained by screening colistin and tigecycline resistance genes. Vitek-2 was used for strain identification, and the drug sensitivity test was carried out by broth dilution method. The molecular typing, drug resistance genes, insertion sequences, plasmid structure and genetic background were analyzed by genome-wide sequencing and bioinformatics. Results Strain GZ12244 is Klebsiella pneumoniae, which is resistant to colistin B, tigecycline, cefuroxime and tetracycline, and carries a variety of drug-resistant related genes such as mcr-1 and tet(M), and some of the drug-resistant genes with antibiotic efflux and antibiotic target change have amino acid substitution mutations. Mcr-1 and tet(M) coexist in a plasmid, and mcr-1 flanked by two insertion sequences ISApl1. There are insertion sequences such as IS15, IS1D and ISEc63 in the upstream and downstream of tet(M) gene. Conclusion Klebsiella pneumoniae GZ12244 is a multidrug-resistant strain. The drug-resistant gene exists in plasmid, and the mobile elements in upstream and downstream may spread the drug-resistant gene.

Objective To explore the research progress, research hotspot and development trend of tigecycline resistance based on the quantitative analysis and visualization function of CiteSpace. Methods The data were collected from 4,263 Chinese and English articles on tigecycline resistance in CNKI, Wanfang, VIP and Web of Science (WOS) databases from 2012 to 2022. CiteSpace 5.8.R3 software was used to analyze the cooperative network of authors, the cooperative network of countries and institutions, the total citation times of journals, and keywords included in the literature, to reveal the hotspots and trends of tigecycline resistance research. Results The number of articles published in English literature was higher than that in Chinese literature. China had the largest number of published documents, showing a significant international academic influence in this research field. Countries all over the world were concerned about the resistance of tigecycline, but Chinese literatures focused more on the clinical infection and prevention of tigecycline resistance, while English literatures placed special emphasis on the research about the drug resistance mechanism of tigecycline. Conclusion The research direction at home and abroad is basically the same, but the research focus has gradually shifted from the clinical treatment and monitoring of tigecycline to the molecular level of drug resistance mechanism.