Stargardt disease (STGD) is one of the most prevalent inherited macular dystrophy, and most often occurs in child or adolescence. Irreversible vision loss is observed in almost all cases. Type 1 (STGD1) is one of the most common type. It is an autosomal recessive condition, caused by mutations in the Abca4 gene. In recent years, encouraging progress has been made in the treatment of STGD1. C20-D3-retinyl acetate (ALK- 001), fenretinide and ICR-14967 (A1120) as visual cycle modulators, StarGen as gene supplementation therapies, and the stem cell transplantation of human embryonic stem cell-derived retinal pigment epithelium cells are the most promising therapies. With the development of studies and clinical trials, the clinical application of various treatments of STGD1 are expected in the near feature, which are expected to save the vision of most patients.

Objective:To evaluate the effect of intestinal carbapenem-resistant Enterobacteriaceae (CRE) active screening combined with enhanced intervention in the prevention and control of nosocomial infection in patients admitted to the hematological ward.Methods:Patients who were admitted to the Department of Hematology in a tertiary-care general hospital from March 1, 2017 to December 31, 2019 and underwent chemotherapy or immunosuppressive therapy comprised the intervention group. They were screened for intestinal CRE at least thrice. From December 1, 2016 to February 28, 2017, patients who underwent chemotherapy or immunosuppressive therapy without active intestinal CRE screening in the Department of Hematology formed the control group. Both the patient groups were monitored for CRE infection in real time. The χ2 test was used to compare the changes in the CRE infection rate and mortality in high-risk patients before and after the active screening. Results:During the intervention period, the CRE colonization rate of patients was 16.46% (66/401) ; in terms of disease distribution, the colonization rate of acute leukemia was the highest 23.03% (26/113) . Of the 66 colonized patients, 27 (40.9%) patients were identified as positive for CRE at the first screening, 15 (22.7%) were identified at the time of the second screening, and the remaining 24 (36.4%) were identified at the third or subsequent screening; Carbapenem-resistant Klebsiella pneumoniae (CRPK) strains were dominant among the pathogens, accounting for 54.55% (36/66) . During the active screening period, the CRE infection rate (2.49%) and mortality rate (50.00%) of high-risk patients were significantly lower than those of the controls (11.30% and 69.23%, respectively) . The pathogens of 10 CRE infection patients during the intervention period were exactly the same as the previous active screening pathogens, and the coincidence rate was 100%.Conclusion:The CRE colonization rate was the highest in patients with acute leukemia who were admitted in the hematology wards. CRPK is the main pathogen of CRE colonization, infection, and death. Increasing the frequency of screening can significantly raise the positive rate of screening, Active screening can effectively reduce the incidence and subsequent mortality of CRE in high-risk patients admitted in the hematological wards. High coincidence rate between CRE screening positive pathogens and subsequent CRE infection pathogens. Intestinal CRE screening can serve as an indicator of CRE bloodstream infection in patients with hematological diseases as well as provide information for antibiotics therapy.

Objective To investigate the distribution of diarrhea pathogens in infants without rotavirus-detection in Hangzhou. Methods 605 stool samples of children with rotavirus-negative diarrhea were collected from Hangzhou First People's Hospital of Zhejiang University, Children's Hospital of Zhejiang University and Hangzhou Children's Hospital from March 2017 to June 2018. The routine test results were analyzed retrospectively and Bristol score was used for the characteristics of stool samples. DNA and/or RNA were extracted from fecal samples with DNA and RNA extraction kit. The extracted DNA and RNA-reversed cDNA were used as templates. 7 common pathogens DNA and/or RNA were amplified by polymerase chain reaction (PCR). The amplified products were detected by agarose gel electrophoresis. The positive rates of pathogens were analyzed by chi-square test. Results Among 605 children, 375 were male (28±11) months and 230 were female (29±10) months. Bristol score of stool samples was mainly in type 6 (496, 82％), followed by type 7 (85, 14％) and type 5 (24, 4％). Among 605 results 97 cases were occult blood positive (positive rate 16％) and 170 cases were white blood cell positive (positive rate 28％).452 of 605 stool samples were positive for pathogen target genes. The positive rate was 74.7％. 319 cases detected single pathogen gene fragments. 127 cases detected two pathogen genes and 6 cases detected three pathogen gene fragments. The positive rate of Clostridium difficile toxin B (48.9％, 296/605)was the highest than the others, followed by Salmonella (20.0％, 121/605) and Norovirus (10.9％, 66/605). The positive rate of Clostridium difficile toxin A was 1.0％ (6/605). The positive rates of pathogens in male and female children were 86.7％(325 / 375) and 86.5％ (199 / 230) respectively, with (χ2 =0.002, P=0.959). Conclusions Salmonella and Norovirus were the main pathogens in children with diarrhea who were negative for rotavirus detection in Hangzhou. The high positive rate of Clostridium difficile toxin B may be related to the colonization of Clostridium difficile in the gastrointestinal tract of infants rather than the pathogen of diarrhea because of the low positive rate of Clostridium difficile toxin A. There was no gender difference in the detection rate of diarrhea pathogens.

Retinitis pigmentosa (RP) is a group of hereditary blinding fundus diseases caused by abnormalities in photoreceptors of the retina.RP is highly heterogeneous in hereditary and cdinical phenotypes.It can be divided into simple type RP and syndrome type RP.The main inheritance patterns are autosomal dominant,autosomal recessive inheritance and X-linked inheritance.With the popularization and clinical application of gene sequencing technology,more and more disease-causing genes have been discovered,and these genes are mainly expressed in photoreceptor cells and retinal pigment epithelial cell.ln-depth understanding of RP pathogenic genes not only provides a theoretical basis for RP diagnosis and genetic counseling,but also provides guidance for RP gene therapy.

Objective To investigate whether the population pharmacokinetics (PPK)models can optimize the initial dosage of individualized Valproic acid (VPA)in children with epilepsy. Methods The epileptic children without taking VPA previously were recruited from October 2015 to May 2017 at the Department of Pediatrics,Peking University First Hospital,and they were divided into the PPK model group and the traditional empirical method group by randomized method. The initial VPA dosages for the PPK model group were calculated by PPK model,whereas those of the traditional empirical method group were dosed at 20-25 mg/(kg·d)regularly. The steady-state serum trough concentrations of VPA were extracted,and then the number and percentage of the patients whose serum trough concen-trations of VPA were 50-100 mg/L in the 2 groups were analyzed and compared with prospectively randomized me-thod. Results Totally 65 epileptic children were recruited and they were randomly divided into the traditional empirical method group (32 cases)and the PPK model group (33 cases). Twenty-seven children in the traditional empirical method group were observed,and 12 children had local epilepsy attack and 15 had generalized seizures;whereas among 29 cases in the PPK model group,there were 12 local attack of epilepsy and 17 had generalized seizures. VPA add-on therapy was administrated in 9 cases and 15 cases in the traditional empirical method group and the PPK model group, respectively. There were 5 cases,21 cases and 1 case with VPA serum concentrations of ＜50 mg/L,50-100 mg/L and＞100 mg/L in the traditional empirical group;while there were 9 cases,20 cases and 0 case in the PPK model group. The VPA serum concentrations of 21 cases (77. 8%,21/27 cases)in the traditional empirical method group and 20 ca-ses (69. 0%,20/29 cases)in the PPK model group were 50-100 mg/L,respectively,and the difference was not sta-tistically significant(P＞0. 05). Conclusion Although the study doesn't suggest that the established PPK model of VPA in Chinese epileptic children is superior to the traditional empirical method,the PPK model might be potentially valuable for optimized individualized dosage adjustment for those with serum trough concentrations not in the reasonable range by the traditional empirical method and with clinical seizure or brain firing activities.

Objective To study the antiviral effects of Pudilan xiaoyan oral liquid on Hep-2 cell models infected with respiratory syncytial virus (RSV), adenoviruses serotype 3 strains (ADV3) in vitro. Methods The cell cytotoxic and inhibition effect of Pudilan xiaoyan oral liquid on RSV or ADV3 were investigated by MTT assay and the inhibitory assay for cytopathic effect (CPE) in Hep-2 cell cultures to detect its antiviral effects. Results The median toxic concentration (TC50) of Pudilan xiaoyan oral liquid on Hep-2 cells was 776.97 mg/L. The median effective concentration (EC50) of inhibiting RSV and ADV3 were 28.08, 28.10 mg/L,whose therapeutic index (TI) were 27.67 and 27.65 respectively. The safety coefficient of Pudilan xiaoyan oral liquid is higher than that of control, ribavirin. Compared with the virus control group, Pudilan xiaoyan oral liquid can obviously produce actions of a dose-dependent relationship to inhibit CPE in Hep-2 cells infected with RSV or ADV3 virus. Conclusions Pudilan xiaoyan oral liquid significantly improves the protection against RSV and ADV3 virus infection in Hep-2 cells. And the inhibition of CPE induced by viruses infection increased with the elevation of higher drug concentration for its antiviral effect augmented in vitro.

ObjectiveTo optimize the condition of multiple fluorescence quantitative PCR and establish a new assay of four human herpes virus (HHV) detected by AllGlo quadruple fluorescence quantitative PCR.MethodsFour HHV including HSV-1,HSV-2,EBV and CMV were identified by sequence analysing the qualitative PCR production.Furthermore,they were quantitatively detected by AllGlo and TaqMan multiple fluorescence quantitative PCR respectively.ResultsBoth the positive rate and specificity of AllGlo and TaqMan in detecting single HHV achieved 100％.And AllGlo single fluorescence quantitative PCR prevailed over TaqMan's by Ct of 1-3.Four HHV can be simultaneously detected by AllGlo quadruple fluorescence quantitative PCR,comparing to the only two by TaqMan.ConclusionAllGlo fluorescence quantitative PCR assay allows a higher throughput,sensitivity and specificity than TaqMan in detection and thus provides a board prospect.

Objective To establish a rapid,accurate and specific method to detect the common mycobacteria based on multiplex real-time PCR.Methods The dual priming oligonucleotide ( DPO)primers and TaqMan probes labeled with FAM,ROX,HEX or JOE fluoresceins at 5' end and eclipse at 3' end respectively were designed to detect the 16S rRNA of mycobacteria.Both specificity and sensitivity were estimated on multiplex real-time PCR detecting genome DNA from 4 mycobacterial model species.Sixty eight early morning sputum specimens collected from hospitalized patients in the Red Cross Hospital of Hangzhou were detected by multiplex real-time PCR,bacterial culture and smear microscopy simultaneously.The positive rates were analyzed by chi-square.Results Mycobacteria including Mycobacterium tuberculosis and three common non-tuberculosis mycobacteria spp.were identified by multiplex real-time PCR accurately and specifically,with the limited load at 101 cfu/ml.In 68 sputum specimens,31 were positive (positive rate 45.6％ ) by this method,which was significant higher than that by smear microscopy ( positive rate 14.7％,x2 =15.4,P ＜0.05 ).The positive cases were identified as 28 Mycobacterium tuberculosis,1 Mycobacterium avium and 2 Mycobacterium intracellulare in agreement with the culture results.One case,which is detected by culture,but not by PCR,was identified as Mycobacterium chelonae by sequencing.Conclusion The multiplex real-time PCR characterizing as sensitive,specific and time-saving for Mycobacterium tuberculosis and common non-tuberculosis mycobacteria could be chosen as the rapid laboratory test of mycobacterial infection.

Objective To determine the effect of cheA gene of Helicobacter pylori in the bacterial chemotaxis in vitro and colonization in vivo. Methods The entire cheA and cheY genes were amplified and cloned from genomic DNA of H. pylori NCTC11637 strain. Subsequently, the prokaryotic expression systems of cheA and cheY genes were generated and the target recombinant proteins rCheA and rCheY were extracted by Ni-NTA affinity chromatography. Rabbits were immunized with either rCheA or rCheY for obtaining antisera, and rCheA-IgG and rCheY-IgG in the antisera were prepared using ammonium sulfate precipitation plus DEAE-52 column chromatography. A suicide plasmid of cheA gene was constructed and then a cheA gene knock-out mutant ( cheA - ) was generated based on homologous recombinant exchange using the suicide plasmid. The cheA- mutant was identified using PCR and sequencing. The phosphorylation levels of CheA and CheY molecules of cheA - and wild-type strain were determined by using rCheA-IgG and rCheY-IgG anchoring the target proteins and protein phosphorylation detection kit. The differences of chemotaxis in vitro and colonization in vivo between cheA- mutant and wild-type strain were compared using chemotactic model and BALB/c infection model of H. pylori. Results The cheA gene knock-out in genome of cheA- mutant was confirmed by the results of PCR and sequencing. After treated with 0. 001-0. 1 mol/L HCI for 10 min, the phosphorylation levels of CheA and CheY molecules of wild-type strain were rapidly descended from ( 59.6 ±11.5) μmol and (55.5 ± 10.2) μmol to ( 10.8 ± 2.6) and (5. 5 ± 1.2) μmol (P ＜ 0.05 ), while the phosphorylation of CheY molecule of cheA - mutant was no markedly changed with a persistent lower level ( P ＞0.05). The diameters [(10-20) ± (2-3) mm] of chemotactic aggregative rings of cheA- mutant were significantly less than those [(16-24) ± (2-3)mm] of wild-type strain (P ＜0.05). The positive isolation rate (90%) of H. pylori in gastric biopsy specimens of mice that infected with wild-type strain was remarkably higher than that (40%) of mice that infected with cheA- mutant (P ＜0.05). The result of fluorescence quantitative was also showed that the numbers (6.3 × 103 ±2.1 × 103 copies/mg) of H. pylori in gastric biopsy specimens of wild-type strain infected mice were significantly larger than those (8.3 × 101 ±3. 1 × 101 copies/mg) in gastric biopsy specimens ofcheA- mutant infected mice (P＜0.05). Conclusion The cheA gene of H. pylori has an important role in the bacterial chemotaxis in vitro and colonization in vivo.

Objective To evaluate the application of fluorescence PCR in detecting ureA gene of Helicobacter pylori(HP)in feces.Methods Fluorescence PCR was used to detect ureA gene of HP in feces from 50 patients,including 23 confirmed by gastric biopsy urease test and histological staining.Bacterium culture and serum antibody detection were also performed, and chi-square test was used to compare the sensitivity,specificity,positive predictive value and negative predictive value among three methods.Results The sensitivity,specificity,positive predictive value and negative predictive value of fluorescence PCR were 1.00,0.96,96.O%and 100.0%,while those for HP culture were 0.78,1.00,100.0%and 84.0%,and thee for serum antibody detection Was 0.96,0.74,76.O%and 95.0%.There were significant differences in sensitivity and negative predictive value between PCR and bacterium culture (X2=5.60 and 4.44,P<0.05),and significant differences in specificity and positive predictive value between PCR and serum antibody detection(X2=5.28 and 4.08,P<0.05).Conclusion ureA gene detection in feces by fluorescence PCR is of value for the diagnosis of HP infection.