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Objective To understand the epidemiological characteristics, genomic variations and macrolide resistance of Bordetella pertussis ( B. pertussis) strains circulating in Shenzhen with clinical data analysis, genotype profiling, phylogenetic analysis and antimicrobial susceptibility test. Methods Clinical data of patients with pertussis in Shenzhen Children's Hospital were collected from the electronic medical re-cord system. Genome sequences of 31 B. pertussis isolates were analyzed with next-generation sequencing and de novo assembled. Multilocus sequence typing (MLST) was performed to identify their sequences types. Sequence alignment by BLASTn was used to identify virulence genotypes and mutations in 23S rRNA gene. A phylogenetic tree was constructed to analyze the relationships among them. E-test was used to identify ma-crolide resistance. Results All of the 31 B. pertussis strains were identified as sequence type-2 (ST-2) by MLST with diverse virulence genotypes. Two were prn-deficient strains. Based on the phylogenetic tree, all of the isolates were distant from vaccine strains. Nineteen isolates were resistant to erythromycin with A2047G mutation in 23S rRNA. Conclusions The virulence genotypes of B. pertussis strains in Shenzhen were diverse with increasing non-vaccine genotypes. Macrolide-resistant strains were prevalent. This study might provide reference for improving the prevention, management and vaccination strategy of pertussis.
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Objective To establish the pipeline and evaluate the feasibility of high-throughput sequencing method used in the detection of severe pneumonia pathogens.Methods Clinical control study was used.Bronchi alveolar lavage fluids (BALF) samples from 76 patients with severe pneumonia (treatment group) and 18 patients with tracheal malacia (control group) and without clinical detected pathogens were collected during March 2015 to December 2016 in Shenzhen Children′s Hospital.The pathogens in the samples were detected using clinical tests and high-throughput sequencing respectively.The results of high-throughput sequencing were confirmed by real-time quantitative PCR and the high-throughput sequencing method used in the detection of severe pneumonia pathogens was evaluated.The χ2 test was used to analyze the correlation of detection rate between the high-throughput sequencing group and the non high-throughput sequencing group.Results The pipeline and method of high-throughput sequencing used in the severe pneumonia pathogens detection was established.The pipeline included sample collection, DNA extraction, library construction, sequencing, and bioinformatic analysis.In 76 cases of patients with severe pneumonia, the results of high-throughput sequencing in 66 cases of bronchoalveolar lavage fluid specimens were positive.The sensitivity was 86.84%, which was significantly higher than the total sensitivity of traditional clinical detection methods including bacterial culture, immunofluorescence and quantitative PCR(68.42%,52/76),χ2=7.426,P<0.001.A total of 13 pathogens were detected in 66 positive samples of high-throughput sequencing, including Mycoplasma pneumoniae, Streptococcus pneumoniae, Haemophilus influenzae and adenovirus, etc.Nine kinds of pathogens were detected in these samples through non-high-throughput sequencing.In the experimental group, the results obtained by clinical test and high-throughput (80.26%) were entirely consistent in 61 samples and not completely consistent in 15 samples (19.74%) specimens.These inconsistent results were mainly concentrated in the detection of adenovirus, Streptococcus pneumoniae and Haemophilus influenzae through high-throughput sequencing, whereas clinical cultures and immunofluorescence methods were not able to detect these pathogens.PCR validation showed that there was no significant difference between the results of high-throughput sequencing and the PCR tests (χ2=0.517,P=0.472), and the difference between the results of high-throughput sequencing and traditional clinical detection methods was statistically significant (χ2=11.671,P<0.001).Conclusion The method for the detection of severe pneumonia pathogens based on high-throughput sequencing is highly sensitive and can detect unknown pathogens, which is superior to those used in traditional clinical detection.
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Objective To investigate the distribution of surfactant protein-B(SP-B) gene single nucleotide polymorphisms and to clarify the correlation between SP-B gene polymorphisms and idiopathic interstitial lung disease(ILD) in children.Methods Sixty-seven children with idiopathic ILD(case group) and 102 children without idiopathic ILD(control group)were selected from October 2013 to September 2016 in Shenzhen Children's Hospital and the First Affiliated Hospital of Guangxi Medical University.Total exons and flanking region of SP-B were detected by high-throughput sequencing,genotype and allele distribution of exon 4(T131I)were analyzed.Results SP-B exon 4(T131I) genotypes could check out three genotypes:namely CC,CT and TT.The frequencies of genotype CC,CT and TT of exon 4(T131I) in the case group were 67.16%,25.37%,7.46%,and in the control group were 56.86%,35.29%,7.84%,respectively.There was no significant difference in genotype distribution between the two groups(χ2=1.981,P=0.371).Frequency of allele C was 79.85% in the case group and 74.51% in the control group,no significant difference showed between the two groups(χ2=1.288,P=0.256).In the control group,the mutation frequency of SP-B exon 4(T131I) was 43.14%(44/102),compared to the frequency of mutations in the population data in the thousands of human genome programs was 52.00%,in European was 53.88%,in South Asia was 45.50%,and in American was 41.93%(P>0.05);but the frequency of gene mutations was 26.39% in East Asia and 80.18% in Africa,there were significant differences compared to the control group(P<0.05).Conclusion The genetic polymorphism of SP-B exon 4(T131I)is not correlated with the susceptibility of idiopathic ILD in children.The mutation frequency of SP-B exon 4(T131I)is related to the race and the region.
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Objective:To detect the proportion of lymphocyte subsets in peripheral blood of the ad-vanced breast cancer patients before and after chemotherapy with docetaxel and thiotepa,as well as the association between the proportion of peripheral blood lymphocyte subsets with the response rate and prog-nosis.Methods:The proportions of lymphocyte subsets (CD3 +T cell,CD3 +/CD4 +T cell,CD3 +/CD8 +T cell,CD3 -/CD16 +56 +NK cell,CD3 +/CD16 +56 +T cell,CD19 +B cell,CD4 +/CD25 +T cell,CD8 +/CD28 -T cell,CD8 +/CD28 +T cell)in the peripheral blood of 86 patients were analyzed with flowcytometry before and after chemotherapy.The result was analyzed in combination with clinico-pathological data.Results:The proportion of regulatory T cells (Treg)after chemotherapy in the disease control patients decreased significantly compared with that of the progressive patients (P=0.034).The difference of the proportions of Treg before and after chemotherapy affected significantly the overall survi-val (OS).The OS of the patients with decreased proportion of Treg was significantly longer than that of the patients with increased proportion of Treg,which was 23.5 and 9.4 months respectively (P<0.05). Conclusion:The patients with decreased proportion of Treg after chemotherapy had higher response rate and better survival benefit.
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Background and purpose:Nab-paclitaxel (Abraxane) is an albumin-bound form of paclitaxel that utilizes the natural properties of albumin to improve paclitaxel delivery to the tumor. It has recently been approved for treatment of breast cancer after failure of combination chemotherapy for metastatic disease or relapse within short time after adjuvant chemotherapy. The purpose of this study was to evaluate the efifcacy and safety of albumin-bound paclitaxel in patients with aggressive and refractory metastatic breast cancer (MBC).Methods:A total of 58 patients with MBC were enrolled into this study from Jul. 2009 to Jan. 2014. All patients received albumin-bound paclitaxel-based chemotherapy. The adverse reactions were evaluated every cycle, and the short-term response was evaluated every two cycles. The patients were followed-up, and the survival was analyzed.Results:58 patients with refractory MBC were evaluable for response, 67.2% of patients received multiple line (≥3 lines) chemotherapy, 32.8% of patients with first and second line of chemotherapy were involved metastasis within one year after adjuvant chemotherapy, 84.5% of patients with visceral metastasis and 93.1% with prior taxane treatment. The objective response rate (ORR) was 13.8%, and disease control rate (DCR) was 60.3%, the median progression free survival (PFS) was 4.0 months, and the overall survival (OS) was 10.1 months. For 23 patients with triple negative breast cancer, ORR was 13.0% and DCR was 56.5%, the median PFS was 4.1 months, and OS was 6.6 months. The main toxicity was myelosuppression (grades 3 and 4 neutropenia, anemia and thombocytopenia were seen in 34.5%, 12.1% and 6.9% of patients, respectively), gastrointestinal reactions, sensory neuropathy, myodynia/arthragia, fatigue, alopecia and so on.Conclusion:The albumin-bound paclitaxel-based chemotherapy can be used in aggressive and refractory MBC. It also showed antitumor activity in taxanes-resistance patients and triple negative patients with good safety and tolerance.
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Objective:To observe whether murine bone marrow mesenchymal stem cells(MSCs)implantation improves the survival of hepatocellular carcinoma(HCC)-bearing mice,and to investigate whether MSCs can differentiate to hepatocytes in HCC microenvironment in a mouse model of orthotopic HCC and its effects on tumor cells.Methods:Murine bone morrow MSCs were obtained through adherent culture method combined with magnetic cell sorting CD45-,CD11b-cells,labeled with CFSE in vitro.Phenotypes of MSCs were analyzed by flow cytometry.A murine model of orthotopic HCC was induced by intrahepatic injection of 5?105 murine H22 hepatoma cells in a BALB/c mouse.In this experiment,twelve BALB/c mice were randomly divided into MSCs implantation and saline administration groups on the 7th day after establishment of orthotropic HCC.CFSE labeled MSCs were injected into tumor and/or normal liver tissue in MSCs implantation group.The life span of HCC-bearing mice was observed.After three weeks,albumin was determined by immunohistochemistry.The livers of HCC-bearing mice were observed by light microscopy.Results:In MSCs group,the mean survival time was 25 days(95%Confidence interval:22-28 d),while,in the control group,mean survival time was 21 days(95%CI:20-23 d).However,the statistical difference was not obvious between the two groups(P=0.0713).It showed that the CFSE labeled cells mainly localized in the border of tumor,also a few in the tumor bed,and expressed albumin.Interestingly,there was a larger area of necrosis in the tumor bed,compared with that in the controls.Conclusion:It can be concluded that MSCs not only could engraft in livers of carcinoma-bearing BALB/c mice,but also could differentiate to hepatocyte-like cell.At the same time,MSCs might induce tumor cells necrosis.Further mechanism need to be studied.