砄bjective:To observe streptococcus sanguis (S.s) and Actinomycetes viscosus (A.v) in oral plaque biofilm formation.Methods:20 ml of saliva obtained from a health adult was centrifuged at 4 ℃ and 10 000 r/min for 10 min.The supernatant was disinfected in 60 ℃ water bath for 30 min.Glass coverslips in the size of 24 mm?24 mm were immersed into the saliva supernatant for 2 h to obtain biofilm.100 ?l of S.s ATCC 34 and A.v ATCC 19246 mixture cultured in TSB at the density of 10 5～6 CFU/ml was added into 20 ml of TSB,and then,the coverslips with biofilm were put into the mixture.The biofilm and bacteria were observed by scanning confocal laser microscopy at various times.Resuts:The biofilm reached the thickness of 15.4 ?m in 8 h and the clumps of the bacteria were mostly in the midle layer of the biofilm.The biofilm increased to 34.3 ?m in 16 h and became tassle like in 48 h.Conclusion: S.s and A.v may play some roles in the oral biofilm formation.

Objective: To develop a new antimicrobial sensitivity test model for oral products in vitro. Methods: An artificial biofilm mouth model for antimicrobial sensitivity test was established by modified LKB chromatography chamber. Using sodium fluoride and tea polyphenol as antimicrobial agents and Streptococcus mutans as target, various methodologies on the sensitivity tests were compared. Results: Agar diffusion, agar dilution and broth macro-dilution were more sensitive than modeling biofilms method. The modeling biofilm assay resulted in MIC values of fluoride or tea polyphenol against S. mutans 32 and 12 times higher respectively than the MIC values generated by broth macro-dilution method. Conclusion :The biofilm artificial mouth model may simulate the environment for oral products test.

Objective To observe the expressions of efflux pump gene cluster adeAB in acinetobacter baumannii isolated from the burn patients and the expression changes of its upstream regulatory genes adeR and adeS and determine the influence of those genes on drug resistance of acinetobacter baumannii.Methods Nine drug-resistant strains and nine sensitive strains of acinetobacter baumannii isolated from the burn patients treated between June 2012 and March 2013 were used.After strain identification using 16SrDNA sequencing,acinetobacter baumannii standard strain ATCC19606 was employed as the control.mRNA expressions of efllux pump genes adeA and adeB and their upstream regulatory genes adeR and adeS were detected by real-time quantitative PCR.Results (1) adeA and adeB genes presented higher expressions in drug-resistant strains (3.71 ±0.95,76.16 ± 8.75) than in sensitive strains (0.92 ± 0.94,0.72 ± 0.78) (F =38.71,663.65 respectively,both P ＜ 0.05).(2) adeS and adeR genes showed higher expressions in drug resistant strains (18.02 ± 6.71,3.02 ± 2.69) than in sensitive strains (1.64±1.51,0.76±0.61) (F =51.04,5.57 respectively,bothP＜0.05).Conclusions The over-expression of efflux pump gene cluster adeAB induced by the expression alteration of its upstream regulatory genes adeR and adeS is closely associated with the drug resistance of acinetobacter baumannii in the burn patient.Besides,the regulatory genes may depend on adeS to sense the nosocomial environment condition,activate or inactivate adeR and hence regulate efflux pump expression.

Objective To investigate the influence of abaI expression on acinetobacter baumannii biofilm formation.Methods Acinetobacter baumannii strain S isolated from bums patients was collected for the study,while the standard strain ATCC19606 was served as the control.At 6,24 and 48 hours,the gene expressions of abaI,pgaA,pgaB and pgaC were detected by real-time fluorescent quantitative PT-PCR,secretion of N-acyl-homoserine lactones (AHLs) by biological sensor and biofilm formation by MTT method.Results (1) Gene expressions of abaI,pgaA,pgaB and pgaC at 6 hours were 8.63 ±5.93,1.98 ± 1.93,1.01 ± 1.32 and 2.67 ± 3.46 respectively,which showed a quick increase at 24 hours (22.81 ± 17.60,5.13 ± 4.32,5.66 ± 3.97,11.97 ± 7.75 respectively),followed by a rapid decline in 48 hours (3.43 ± 0.88,1.30 ± 0.24,3.01 ± 3.00,3.02 ± 3.29 respectively).Gene expressions of pgaB and pgaC at 6 hours and that of pgaA and pgaC at 48 hours revealed statistically significant differences from those at 24 hours (P ＜ 0.05).(2) AHLs showed a level of 18.49 ± 11.03 at 6 hours,reached a peak of 52.23 ± 15.95 at 24 hours,then descended to 5.53 ± 0.94 at 48 hours.AHLs level at 24 hours showed statistically significant difference from that at 6 hours and 48 hours (P ＜ 0.05).(3)Biofilm formation at 24 hours and 48 hours was 2.83 ±0.44 and 2.71 ±0.15 respectively,far higher than that at 6 hours (0.49 ± 0.11,P ＜ 0.05).(4) In the correlation analysis among AHLs,biofilm formation and gene abaI,pgaA,pgaB and pgaC expressions,significant positive correlation was found between abaI and pgaA and between AHLs and pgaC expression (P ＜ 0.05).Conclusion Acinetobacter baumannii may regulate gene expressions of pgaA and pgaC responsible for biofilm formation to adjust to the external environment by means of changing abaI gene expression and AHLs secretion.

Objective To investigate the construction of Streptococcus mutans luxS gene knockout mutant which can act as the technical platform for following researches on luxS quorum sensing function in oral ecosystem.Methods Erythromycin resistance gene was inserted between two 1 kb fragments containing regions of DNA immediately upstream and downstream of the luxS translational start and stop codons.The resuhing construct was linearized and electro-transformed into Streptococcus mutans cells.After allelic exchange,the luxS gene knockout mutant strains were selected on 10μg/ml erythromycin plates,and compared the growth and biofilms formation of luxS knockout mutant with wild type strains.Results The luxSknockout mutant was confirmed by PCR,and it was also confirmed that this gene mutant could be stably passed through in vitro.The growth mode of luxS knockout mutant showed obvious difierences against that of wild type at stationary phase,the knockout mutant gained more bacteria cells growth.Conclusion Streptococcus mutans luxS gene has been successfully disrupted with allelic exchange.This mutant strains showed higher growth abilitv which could be the consequence of quorum sensing mutant.

Objective To evaluate the efficacy and safety of recombinant human brain natriuretic peptide (rh-BNP) combined with levosimendan on acute myocardial infarction complicated by heart failure.Methods Patients who suffered from anterior wall acute myocardial infarction (AMI) with heart failure (KillipⅡ ～ Ⅲ) within 12 to 24 hours after the onset of chest pain were randomized into two groups: the control group (n=30, receiving dobutamine and/or cedilanid) and the experimental group (n=30, receiving rh-BNP combined with levosimendan).The hemodynamics, parameters of laboratory tests and adverse events were observed before and after treatment.Results The experimental group showed that the respiratory rate (RR), heart rate (HR), systolic blood pressure (SBP), arterial blood gas oxygen saturation (SaO2), cardiac index (CI), extravascular lung water index (EVLWI) were significantly different between 2 h and sequential time points after treatment and pre-treatment (allP＜0.05).The control group showed that RR, HR, SaO2, CI, EVLWI were significantly different between 6 h and sequential time points after treatment and pre-treatment (P＜0.05 for all).There were significant differences in RR, HR, SBP, SaO2, CI, EVLWI at 2 h and 6 h after treatment between the two groups (P＜0.05 for all).Parameters of RR, HR, CI, EVLWI at 72 h after treatment had differences between the experimental group and controls.Patients in the experimental group presented larger urine volume, lower level of plasma NT-pro BNP, higher left ventricular ejection fraction (LVEF) and shorter length of stay in CCU as compared with patients in the control group (P＜0.05 for all).In adverse events monitoring in hepatic parameters, electrolyte level and coagulation function before and after treatment, there was no significant difference between the two groups.Conclusions Compared with the conventional treatment, the combination therapy with rh-BNP and levosimendon can improve the hemodynamics, increase the urine volume, decrease the level of plasma NT-proBNP and elevate LVEF significantly, so as to improve the clinical symptoms and shorten the hospital stay in patients with acute myocardial infarction complicated by heart failure.

Objective To observe and analyze the effect of HbA1c level on macular microcirculation in patients with type 2 diabetes mellitus (T2DM).Methods A cross-sectional study.One hundred and twenty-four T2DM patients (124 eyes) without diabetic retinopathy who diagnosed by the examination of fundus color photography in Lixiang Eye Hospital Of Soochow University during September to December 2017 were enrolled in this study.There were 59 males (59 eyes) and 65 females (65 eyes),with the mean age of 65.06±7.99 years old.All patients underwent BCVA,fundus color photography,and OCT angiography (OCTA).The history of diabetes,hypertension and dyslipidemia were recorded in detail.According to the HbA 1 c level,patients were divided into three groups,HbA1c ideal control group (group A,HbA1c ＜7％,67 eyes),HbA1c control group (group B,7％≤HbA1c≤9％,44 eyes),and HbA1c poor control group (group C,HbA1c＞9％,13 eyes),respectively.The 3 mm × 3 mm range of the macular area was scanned by OCTA instrument.The vascular density (VD) and skeleton density (SD) of nonsegmented retinal layer (NRL),superficial retinal layer (SRL) and deep retinal layer (DRL) in the macular area and foveal avascular zone (FAZ) area,non-circularity index,axial rate (AR) of SRL were measured.The correlation between HbA1c,BCVA and VD,SD ofNRL,SRL,DRL was analyzed statistically with Spearman correlation test.The correlation between systemic factors and the above indicators was analyzed statistically with linear regression analysis.Results The results of linear regression analysis showed that HbA1 c was significantly correlated with VD (t=-3.237,-3.156,-2.050) and SD (t=-0.3.45,-3.034,-2.248) of NRL,SRL and DRL (P＜0.05);but no correlation with FAZ,non-circularity index and AR (t=1.739,0.429,1.155;P＞0.05).The differences of VD (F=6.349,5.981,3.709),SD (F=7.275,6.085,1.904) and AR (F=0.027) of NRL,SRL and DRL in group A,B and C were statistically significant (P＜0.05);but the differences of FAZ (F=1.904),non-circularity index (F=0.280) was not statistically significant (P＞0.05).Significant differences (P＜0.05) of VD and SD of NRL were found between group A and B (t=1.987,2.201),group A and C (t=3.365,3.572),group B and C (t=2.010,2.076).Significant differences (P＜0.05) of VD and SD of SRL were found between group A and B (t=2.087,2.168),group A and C (t=3.197,3.194).There were significant differences (P＜ 0.05) in SD of DRL between group A and B (t=2.239),group A and C (t=-2.519).There was significant difference in VD of DRL between group A and C (t=2.363).The results of Spearman correlation analysis showed that HbA1c was negatively correlated with VD (r=-0.273,-0.255,-0.222;P=0.002,0.004,0.013) and SD (r=-0.275,-0.236,-0.254;P＜0.05) ofNRL,SRL,DRL;positively correlated with FAZ and BCVA (r=0.221,0.183;P＜0.05).BCVA was negatively correlated with VD (r=-0.210,-0.190,-0.245) and SD (r=-0.239,-0.207,-0.296) of NRL,SRL,and DRL (P＜0.05),but not correlated with FAZ (r=0.099,P＞0.05).Conclusion The decrease of macular perfusion and the morphological change of FAZ accompanied by HbA1c increased.

OBJECTIVETo investigate the predominant contribution of methyl-metabolism pathway to the regulation of LuxS of Strecptococcus mutans.

METHODSThe differences in biofilm formation and aciduricity of Strecptococcus mutans among the methyl-metabolism-complementation strain (KO-S), the parental wide-type strain (WT) and the luxS null strain (KO) were observed by real-time PCR for monitoring the transcriptional level of genes related to biofilm formation (smu.238, gtfD) and aciduricity (smu.44, smu.46) of the studied strains, methyl thiazolyl tetrazolium (MTT) for quantifying the biofilm of the exhibited strains and confocal laser scanning microscopy for estimating the structure of the biofilm.

RESULTSThe transcriptional level of smu.44, smu.46, smu.238, gtfD in WT were 1.289 ± 0.051, 1.694 ± 0.140, 1.565 ± 0.107, 1.667 ± 0.196 respectively; in KO were 1.001 ± 0.045, 1.007 ± 0.151, 1.000 ± 0.021, 1.012 ± 0.196 respectively, downregulated compared with WT (P < 0.05); in KO-S were 4.662 ± 0.091, 5.019 ± 0.258, 3.462±0.029, 3.071 ± 0.136 respectively, upregulated compared both with KO and with WT (P < 0.05). The quantity of biofilms formed by the studied strains were WT (1.592 ± 0.213), KO (0.939 ± 0.029), KO- S (2.177 ± 0.226), KO- P (1.020 ± 0.093), respectively, representing a less quantity by KO and KO-P than WT (P < 0.05) and a more quantity by KO-S than other three stains (P < 0.05). According to the observation of biofilms texture by confocal laser scanning microscopy, the WT biofilm was condensed and even. In contrast, fissures and gaps were found scattered in biofilms of KO, KO-P while lessened in that of KO-S, in which high-density bacterial aggregates were observed. The acid assay indicated a smaller biofilm decrease by WT and KO-S than that by KO and KO- P(P < 0.05).

CONCLUSIONSThe methyl- metabolism pathway contributes to LuxS regulation on biofilm formation and auiduricity of Strecptococcus mutans.

Bacterial Proteins ; metabolism ; Biofilms ; Carbon-Sulfur Lyases ; metabolism ; Glucosyltransferases ; Microscopy, Confocal ; Real-Time Polymerase Chain Reaction ; Streptococcus mutans ; metabolism