To clone,express and purify the E Protein and EDⅢ of Zika virus in E.coli and prepare two kinds of polyclonal antibodies,the virus was amplified by Vero E6 cell culture.Total RNA was extracted by RT-PCR and reverse transcribed into cDNA.The prokaryotic expression vectors pET32a/E and pET28a/EDⅢ were constructed by cDNA sequence of E and EDⅢ gene.Then,recombinant plasmids were transformed into E.coli BL21 and induced by IPTG,and purified by Ni+ column affinity chromatography.BALB/C mice were immunized with purified recombinant proteins.Antiserum was collected and titer was determined by indirect ELISA.Western blot was used to detect the specificity.Results showed that the recombinant proteins were successfully expressed and purified.The titer of the polyclonal antibodies both reached 1:409 600.Western Blot analysis showed that the polyclonal antibodies could specifically recognize the recombinant proteins.Thus,the specific polyclonal antibody were successfully prepared,laying a foundation for further study on the pathogenesis,detection methods and immune strategies of Zika virus.

OBJECTIVETo study the clinical efficacy of porcine pulmonary surfactant (PS) combined with budesonide suspension intratracheal instillation in the treatment of neonatal meconium aspiration syndrome (MAS).

METHODSSeventy neonates with MAS were enrolled for a prospective study. The neonates were randomly assigned to PS alone treatment group and PS+budesonide treatment group (n=35 each). The PS alone treatment group was given PS (100 mg/kg) by intratracheal instillation. The treatment group was given budesonide suspension (0.25 mg/kg) combined with PS (100 mg/kg).

RESULTSThe rate of repeated use of PS in the PS+ budesonide group was significantly lower than that in the PS alone group 12 hours after treatment (p<0.05). The improvement of PaO/FiO, TcSaO, PaO, and PaCOin the PS+ budesonide group was significantly greater than that in the PS alone group 6, 12, and 24 hours after treatment (p<0.05). The chest X-ray examination showed that the pulmonary inflammation absorption in the PS+ budesonide group was significantly better than that in the PS alone group 48 hours after treatment (p<0.05). The incidence of complications in the PS+budesonide group was significantly lower than that in the PS alone group (p<0.05), and the average hospitalization duration was significantly shorter than that in the PS alone group (p<0.01).

CONCLUSIONSPS combined with budesonide suspension intratracheal instillation for the treatment of neonatal MAS is effective and superior to PS alone treatment.

Animals ; Budesonide ; administration & dosage ; Female ; Humans ; Infant, Newborn ; Length of Stay ; Male ; Meconium Aspiration Syndrome ; complications ; drug therapy ; Prospective Studies ; Pulmonary Surfactants ; administration & dosage ; Suspensions ; Swine ; Trachea

This article reported the clinical manifestations, steroid profiles and adrenal ultrasound findings in two unrelated Chinese girls with lipoid congenital adrenal hyperplasia (LCAH). Direct DNA sequencing and restriction fragment length polymorphism (RFLP) analysis were used to identify the mutations of steroidogenic acute regulatory protein (StAR) gene. The two patients with 46,XX karyotype, presented hyperpigmentation, growth retardation, and hyponatremia. Steroid profiles analysis revealed elevated plasma adrenocorticotrophic hormone levels, decreased or normal serum cortisol levels and low levels of androgens. Ultrasound examinations revealed that enlarged adrenals in patient 1 and normal adrenals in patient 2. Direct DNA sequencing of StAR gene showed a reported homozygous for c.772C>T(p.Q258X) in patient 1. Compound heterozygous for c.367G>A(p.E123K) and IVS4+2T>A (both novel mutations) were found in patient 2, inherited from her mother and father respectively. The amino acid of mutant position of the novel p.E123K was highly conserved in ten different species and was predicted to have impacts on the structure and function of StAR protein by the PolyPhen-2 prediction software. RFLP analysis revealed three bands (670, 423 and 247 bp) in patient 2 and her father and two bands (423 and 247 bp) in her mother and 50 controls. It is concluded that LCAH should be considered in girls with early onset of adrenal insufficiency and that steroid profiles, karyotype analysis, adrenal ultrasound and StAR gene analysis may be helpful for the definite diagnosis of LCAH.
46, XY Disorders of Sex Development ; diagnosis ; genetics ; Adrenal Hyperplasia, Congenital ; diagnosis ; genetics ; Amino Acid Sequence ; Child ; Female ; Humans ; Infant ; Infant, Newborn ; Molecular Sequence Data ; Mutation ; Phosphoproteins ; genetics ; Polymorphism, Restriction Fragment Length

In order to reveal the cause of disease occurred in the process of Coptis chinensis growth, this paper studied the bacterial species diversity index of different aged rhizospheric and non-rhizospheric soil planting normal or sick C. chinensis by using PCR-DGGE technique. The representative DGGE bands were chosen to be cloned, and sequenced, the phylogeny were constructed. The results showed that the bacterial communities were very different between the normal and diseased soil samples of C. chinensis, and the diversity index (H) of diseased soil samples were higher than that of normal soil samples. Sequencing analysis of representative cloned DGGE bands showed that the unculturable bacteria were the dominant groups, and bacteria belonged to genus Bacillus, Acidovorax, Acinetobacter, uncultured Kluyvera, and uncultured Comamonas were also existing, but the reported plant pathogenic bacteria were not found in the C. chinensis planting soil. The density and brightness of clone band d in diseased soil samples was higher than that in normal soil sample, and sequencing analysis showed that it belonged to genus Acidovorax. Obviously, during the process of C. chinensis growth, the rhizospheric bacteria population changed, and the quantity of bacteria belong Acidovorax increased, which probably resulted in the disease occurred during C. chinensis growth.
Bacteria ; classification ; genetics ; isolation & purification ; Biodiversity ; Coptis ; growth & development ; microbiology ; Denaturing Gradient Gel Electrophoresis ; Molecular Sequence Data ; Phylogeny ; Polymerase Chain Reaction ; Rhizosphere ; Soil Microbiology

Objective: To investigate patients' perception of service quality at hospitals in nine Chinese cities and propose some measures for improvement. Methods: The ServQual scale method was used in a survey involving patients at outpatient and in-patient facilities in Shanghai, Chongqing, Chengdu, Nanning, Guilin and Laibin of Guangxi, Honghezhou of Yunnan, Wulumuqi of Xinjiang and Zhongshan of Guangdong. The data collected were entered and analyzed using SPSS 20.0. Statistical analyses included descriptive statistics, factor analyses, reliability analyses, productmoment correlations, independent-sample t-tests, One-way ANOVA and regression analyses. Results: The Kaiser-Meyer-Olkin value for the factor analysis of the scale was 0.979. The Cronbach's α for the reliability analysis was 0.978. All the Pearson correlation coefficients were positive and statistically significant. Visitors to out-patient facilities reported more positive perception than visitors to in-patient facilities on tangibles (t = 4.168, P < 0.001) and reliability (t = 1.979, P < 0.05). Patients of 60 years of age and above reported more positive perception than those between 40 and 49 on reliability (F = 3.311, P = 0.010), assurances (F = 2.751, P < 0.05) and empathy (F = 4.009, P = 0.003). For the five dimensions of the scale, patients in Laibin, Guangxi reported the most positive perceived service quality, followed by patients in Shanghai. On the other hand, patients in Chongqing and Nanning and Guilin of Guangxi reported relatively poor perceptions of service quality. Standardized regression coefficients showed statistically significant (P < 0.001) positive values for all ServQual dimensions. Empathy (β = 0.267) and reliability (β = 0.239) most strongly predicted perception of service quality. Conclusions: Chinese patients perceived service quality as satisfactory. Hospitals in various regions of China should enhance their awareness and ability to serve their patients.

Background Retinal retinoic acid (RA) plays an important role in the formation of the lensinduced myopia.However,it is not clear how RA transfer the myopic signal to choroid throughout the retinal pigment epithelium (RPE) barrier.Objective The aim of this study was to investigate the effect of all-trans retinoic acid (atRA) on the barrier of RPE in lens-induced myopic eye of guinea pig.Methods Thirty left eyes of 30 21-dayold clean guinea pigs were randomized into normal control group and the model group.The models of out of focus were induced by covering of-6.00 D concave lens on the left eyes for 15 days.Radius of corneal curvature was measured using corneal curvimeter,and diopeter of the guinea pig was examined by mydriatic optometry.The length of ocular axis was detected by A-sonography.The animals were sacrificed and the retinas of the left eyes were isolated for the culture and passage of RPE cells.The third generation of cells were incubated Millcell-PET microporous film,and atRA at the concentration of 1 × 10-6,1 × 10-7,1 × 10-8 and 1 × 10-9 mol/L was added to the micropore respectively for 12 hours,and the micropores with equal-solvent served as negative control group.Methyl thiazolyl tetrazolium (MTT)colorimetric method was used to detect the survival rate of the cells.Subsequently,the transepithelial electrical resistance (TER) of the monolayer cells was determined with CN10-EVOM2 resistance measuring meter.The vesicular transport change of RPE membrane in different groups was evaluated by FM1-43 fluorescence staining.Results The mean diopter was (-2.20±0.95) D in the models,and that in the controls was (+ 1.15 ±0.30) D,with a significant difference between them (t =14.57,P＜ 0.01).The axial length was (8.24 ± 0.09) mm in the models and it was significantly longer than (7.81±0.05) mm in the controls (t=17.20,P＜0.01).RPE cells grew well to form a monolayer in Millcell culture pool after one week.After 24 hours of the atRA treatment,the survival rate of RPE cells reduced gradually with the increase of atRA concentration with the highest rate in the 1 × 10-9 mol/L atRA group (93.3 ％) and followed by the 1 × 10-8 mol/L atRA group (88.2％).More than half of the cells dead in the 1 × 10-6mol/L and 1 × 10-7mol/L atRA groups (53.8％ and 47.1％).Significant differences in the TER value and fluorescence staining intensity of the cells were seen among the various groups (F =43.89,P =0.00 ; F =26.13,P =0.00),with the maximal values in the 1 × 10-8mol/L atRA group.The FM1-43 fluorescence located on the cellular membrane and cytoplasm.Conclusions AtRA can increase the functional state of tight junction and vesicular transport,which regulated the RPE cell barrier in the guinea pig.

Objective To study the rule of aerosol deposition in human upper respiratory tract and analyze the impact of respiratory pattern on aerosol deposition. Methods A computer model of human upper respiratory tract was established first. CFD (computational fluid dynamics) method was then used to numerically simulate the aerosol deposition within the human upper respiratory tract and the rule of aerosol deposition was analyzed. Results The efficiency of aerosol deposition in human upper respiratory tract was improved with the increase of inertial parameter. The breathing intensity and aerosol property had little impact on the pattern of aerosol deposition, which was at most in larynx due to the inertial impact and turbulent dispersion. Under the mode of cyclic inhalation, the aerosol deposition efficiency was higher at unsteady respiratory than that at steady respiratory, at cyclic inhalation than at cyclic exhalation. Conclusions Inertial impact is the main key deposition mechanism for micro aerosol, while turbulent dispersion, secondary flow and recirculation flow have an equally important impact on aerosol deposition in human respiratory tract.