PcoI-PcoR is a quorum-sensing (QS) system influencing the biofilm formation and rhizosphere colonization in Pseudomonas fluorescens 2P24. The expression of the pcoI, a N-acyl-homoserne lactone (AHL) biosynthase gene, is under the regulation of a number of chromosomal factors, such as the GacS-GacA two-component system. In this paper, we investigated the upstream regulators that influence the transcription of pcoI gene using a chromosomal pcoI-lacZ fusion reporter strain PM203. Cosmids containing genomic DNA of the wild-type strain 2P24 were introduced into the reporter strain PM203 (gacA—, pcoI-lacZ) to screen positive transcriptional regulators of pcoI gene. One of them named pP32-24, which contained a 5-kb Pst I functional fragment was selected. Further analysis identified that the pcoI was the gene responsible for the increase of the pcoI-lacZ expression. The expression of pcoI-lacZ reporter was alsoimproved in both PM101 (pcoI-lacZ) and its gacAmutant PM203 after addition of exogenous AHL, indicating that the expression of pcoI is positively regulated by AHL (autoinduction) in strain 2P24. In addition, deletion mutagenesis and complementation experiments demonstrated that the transcriptional regulator PcoR positively controlled the expression of pcoI and the formation of biofilm. These results suggest that, in strain 2P24, the expression of PcoI-PcoR QS system is auto-inducted, and the transcriptional factor PcoR is involved in the regulation of pcoI transcription and the biofilm formation.

Objective To investigate drug resistance genes and epidemic characteristics of β-lactamase carried by carbapenem-resistant Acinetobacter baumannii (CRAB) in the respiratory intensive care unit(RICU) in a hospital.Methods Clinically isolated CRAB from RICU patients in October-December 2015 were collected.Five drug resistance genes (KPC-2,IMP,VIM,NDM-1,OXA-23) were specifically amplified by polymerase chain reaction (PCR),amplified products were performed agarose gel electrophoresis and sequencing analysis,the homology was analyzed with pulsed-field gel electrophoresis (PFGE).Results A total of 22 CRAB strains were isolated in October-December 2015,19 (86.36％) of which were isolated from sputum.The resistance rate of 22 CRAB strains to compound sulfamethoxazole was 59.09 ％,resistance rate to minocycline was 9.09 ％,all were sensitive to polymyxin B,resistance rates to other antimicrobial agents were more than 80％.Three kinds of resistance genes KPC-2,IMP and NDM-1 were not found by PCR amplification,positive rates of VIM and OXA-23 were both 100％.PFGE homology analysis revealed that 22 strains were divided into 13 different types,each type contained 1-5 strains,9 types(69.23％) contained only 1 strain respectively,the other 4 types (30.77％) contained 2-5 strains.A5,A7,and A8;A9,A11,A14,A19 and A22;A4,A10 and A12;A16 and A18 were of the same type respectively.Conclusion The main types of β-lactamase-resistant genes of CRAB in RICU are VIM and OXA-23.Homology analysis shows a small parts are of the same clone strains,which reveals epidemic of a small scale.

Objective To compare the therapeutic efficacy between emergency and non-emergency operation for ruptured intracranial aneurysms. Method A retrospective analysis of 184 patients with ruptured intracranial aneurysms the Second Affiliated Hospital Zhejiang University College of Medicine, admitted from Dec 2008 to Sep 2009, was carried out to evaluate the efficacy of operation to be done earlier. The patients were divided into 2groups according to the time of surgery. In the early operation group ( n = 102), the patients were operated on within 3 days of rupture of aneurysms, and in the delayed operation group ( n = 82), the patients were operated on after 3 days. The comparison in the rate of rebleeding before surgery, rate of complete occlusion of the ruptured aneurysm and rate of major complications such as cerebral infarction and hydrocephalus between two groups was made. The Glasgow outcome scale (GOS) scores of these patients were also evaluated by 6- 12 months follow-up after operation. Results Preoperative re-bleeding happened in 2 patients of the early operation group and in 7 patients of the delayed operation group. The rates were significantly different ( P ＜ 0.05). The complete occlusion rate in the early operation group was 91.2 % ( 93/102 ), while was 80.5 % ( 66/82 ) in the delayed operation group (P＜0.05). There were no statistically significant differences in post-operative cerebral infarction rate, post-operative hydrocephalus rate or GOS scores on follow-up between two groups. Conchusions Early operation can significantly reduce the re-bleeding before surgery, reducing the risk of death and disability. In early operation, the continuous lumbar drainage by cannulation and other methods can be used to reduce intracranial pressure, significantly increasing the rate of complete occlusion, and promoting rehabilitation.

OBJECTIVETo investigate the mechanism of clinical haemorrhage in an inherited coagulation factor VII (FVII) deficiency and tissue factor abnormality pedigree.

METHODSAll exons, exon-intron boundaries and the 3', 5' untranslated sequences of FVII and tissue factor (TF) genes were amplified by PCR and sequenced directly. Any mutation identified by direct sequencing was confirmed by reverse sequencing. FVII cDNA of the proband was synthesized with random primers and amplified by nest PCR.

RESULTS55C-->T heterozygous mutation located in promoter of FVII gene was identified in the proband. The heterozygous mutation was derived from his mother. Tracing the other pedigree members found that his sister had the same heterozygous mutation and the others had wild-type FVII genes. A 9363 C-->T (Arg131Trp) heterozygous polymorphism in TF gene, which was 2.63% frequency of T allele polymorphism, was found in all of the pedigree members.

CONCLUSIONIt was the first report that the -55C-->T heterozygous mutation in FVII gene and the Arg131Trp heterozygous polymorphism in TF gene explained the clinical symptom of the proband.

Adult ; DNA Mutational Analysis ; Factor VII ; genetics ; Factor VII Deficiency ; genetics ; Heterozygote ; Humans ; Male ; Pedigree ; Polymorphism, Genetic ; Thromboplastin ; genetics

OBJECTIVETo evaluate the expression of recombinant plasmid pcDNA3.1/GBD of glucan binding protein of Streptococcus mutans in mammalian cells COS-7.

METHODSEukaryotic plasmid carrying encoding gene of GBD of Streptococcus mutans gbpA was constructed and the plasmid was introduced into COS-7 cells by Lipofectamine reagent. The transient expressed protein in COS-7 cells was detected by immunochemistry technique.

RESULTSThe positive expression was detected in plasma of the cells which were transfected with recombinant plasmid pcDNA3.1/GBD. The cells which were transfected with pcDNA3.1 were negative.

CONCLUSIONGBD can translate and express in COS-7 cells after transfected with recombinant plasmid pcDNA3.1/GBD. The expressed protein locates in the plasma and the protein is able to combine with anti-GbpA antibody. The expressed protein has the antigenicity and is a candidate gene vaccine.

Animals ; Antigens, Surface ; biosynthesis ; genetics ; immunology ; Bacterial Proteins ; biosynthesis ; genetics ; immunology ; COS Cells ; Carrier Proteins ; biosynthesis ; genetics ; immunology ; Dental Caries ; prevention & control ; Eukaryotic Cells ; metabolism ; Gene Expression ; Genetic Vectors ; Humans ; Lectins ; Mammals ; Plasmids ; genetics ; immunology ; Recombinant Proteins ; Streptococcus mutans ; genetics ; metabolism ; Transfection ; Vaccines, DNA

OBJECTIVETo explore the regulatory effect of clearing-Heat secreting-bile regulating-Qi flow and activating blood circulation (CSRA) principle on cholecystokinin receptor (CCK-R) and its mechanism.

METHODSCholecystokinin (CCK) in serum of portal venous blood, maximum binding capacity (Bmax) and affinity (Kd) of CCK-R levels in gallbladder of guinea pigs allocated in four groups (control, high cholesterol, natural recovery and treated groups) were determined using radioimmunoassay and radioligand receptor assay (RRA). At the same time, changes of fasting volume (FV) and postprandial volume (PV) of gallbladder, fasting and postprandial bile (FB and PB) in gallbladder, gallbladder contraction rate (GCR) and cholesterol concentration (CC) in bile were observed.

RESULTSCompared with the control group, after two weeks of high cholesterol feeding, increase of FV, FB, PV, PB and CC (P < 0.05), and decrease of GCR (P < 0.01) and Bmax were found in cholesterol group, but with no significant change in Kd and CCK level. The above-mentioned criteria were restored to normal range in the treated group.

CONCLUSIONCSRA principle could promote the recovery of gallbladder contraction by regulating CCK-R expression in it, its mechanism is possibly correlated with reduction of cholesterol concentration in bile.

Animals ; Bile ; metabolism ; Cholecystokinin ; metabolism ; Cholesterol ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Gallbladder ; physiopathology ; Guinea Pigs ; Hypercholesterolemia ; metabolism ; Male ; Medicine, Chinese Traditional ; Random Allocation ; Receptors, Cholecystokinin ; metabolism

Objective To investigate the effects of Nod-like receptor protein 3 inhibitor MCC950 on cognitive function in a mouse model of sepsis-associated encephalopathy (SAE).Methods Ninety adult male C57BL/6 mice were randomly (random number) divided into three groups:the sham + saline group (n=20,sham group),CLP + saline group (n=35,CLP group),and CLP + MCC950 group (n=35,MCC950 group).SAE mouse model was established by cecal ligation and puncture (CLP) surgery.Saline (10 mL/kg) or MCC950 (10 mg/kg) was intraperitoneally injected 30 min before surgery and on day 1,2,4 and 6 after surgery according the grouping.Seven days after surgery,six mice were taken from each group.Western blot was used to detect the hippocampal content of NLRP3,apoptosis-associated specklike protein (ASC),interleukin-1β (IL-1β) and IL-18.The number of NLRP3-positive cells in CA 1 region were detected by immunohistochemical analysis.The remaining mice in each group were used for open field and fear conditioning tests 14 days after surgery.One-way analysis of variance was used for inter-group comparison,and SNK-q test was used for pairwise comparison.A P value ＜0.05 was considered statistically significant.Results Compared the MCC950 group with the CLP group,the freezing time of context test was significantly increased [(137±21) s vs (84±15) s,P=0.013],the hippocampal content of NLRP3,IL-1β and IL-18 were significantly reduced (P＜0.01),and the number of NLRP3-immunoreactive cells per mm2 in CA region were significantly decreased (23±5 vs 74±13,P＜0.01).There was no significant changes in protein level of ASC and results of open field tests (P＞0.05).Conclusions MCC950 administration can improve cognitive function in a mouse model of SAE,which is probably due to the inhibition of NLRP3 inflammasome and downstream inflammatory cytokines IL-1β and IL-18.

Objective To determine the prevalence and genotype of hepatitis E virus (HEV)among commercial swine population in Eastern and Southern China. Methods Six hundred specimens of swine bile collected from 5 slaughterhouses in Eastern and Southern China from 2007 to 2009 were tested for HEV RNA using nested RT-PCR. PCR products were sequenced for phylogenetic analysis. Results Forty-seven out of the 600 samples (7.83%) were positive for HEV RNA. Based on the 150 nt fragment within HEV ORF2, data from phylogenetic analysis revealed that all the 47 HEV isolates were identified to be genotype Ⅳ, sharing 75.0%-83.4%, 75.0%-84.6%, 71.9%-80.7% and 88.1%-91.5% nucleotide identities with prototype Ⅰ,Ⅱ,Ⅲ and Ⅳ HEV strains respectively while majority of the isolates clustered within their respective isolation sites. Conclusion HEV was widespread in commercial swine population in Eastern and Southern China that raised a serious concern about the safety regarding the consumption of pork products.

Objective To optimize antimicrobial use process,ensure the rational use of preoperative antimicrobial prophylaxis during consecutive operations.Methods Antimicrobial use process in a hospital in December 2015 was optimized,6 072 cases of consecutive operations in May-November 2015 were selected as control group,5 832 cases of consecutive operations in December 2015-May 2016 were as trial group,the qualified rate of rational use of antimicrobial agents was compared between two groups,causes for delayed/prior use was analyzed.Results Before and after the optimization of antimicrobial use process,rates of antimicrobial use were 77.16％ and 78.80％ respectively,there was significant difference between two groups(x2 =8.305,P =0.004).After the optimization of antimicrobial use process,rate of antimicrobial use within 0.5-1 hour was significantly higher than that before the optimization (82.36％ vs 41.11％);rate of antimicrobial use ＜0.5 hour before skin incision decreased from 57.11％ before optimization to 4.32％ after optimization;but rate of antimicrobial use ＞1 hour before skin incision increased from 1.78％ to 13.32％.Causes for delay/prior use of antimicrobial agents was due to the lack of effective communication between doctors and nurses,which resulted in circuit nurses' inaccurate assessment on interval of consecutive operations(62.13％),the duration of intubation or puncture was too long for anesthesiologists (13.57％).Conclusion Optimizing antimicrobial use process in consecutive operations can improve prophylactic antimicrobial use rate within 0.5-1 hour,and is helpful for ensuring the efficacy of antimicrobial prophylaxis.