Table of Content

30 June 2020, Volume 35 Issue 6

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Resistance mechanism and homology of ampicillin-resistant Haemophilus influenzae in children

WANG Jianglin, SUN Jie, YANG Huijian, WANG Bingjie, PAN Fen, ZHANG Hong

2020, 35(6):  513-518.  DOI: 10.3969/j.issn.1673-8640.2020.06.001

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Objective To study the resistance mechanism and homology of ampicillin-resistant Haemophilus influenzae（Hi） in children. Methods Totally,117 isolates of Hi were isolated. Antimicrobial susceptibility test was performed by microbroth dilution method. Beta-lactamase was determined by nitrocefin disk test. Beta-lactamase-positive ampicillin-resistance（BLPAR） genes,including TEM-1 and ROB-1,were amplified by polymerase chain reaction（PCR）,and ftsI gene was sequenced. Blast was compared with that of Hi RdKW20 to clarify the resistance mechanism of beta-lactamase-negative ampicillin-resistance（BLNAR）. The homology of Hi was determined by multiplex PCR,and the molecular epidemiological characteristics of Hi were clarified. Results Among the 117 isolates,the ampicillin resistance rate was 56.4%（66/117）,the beta-lactamase positive rate was 46.2%（54/117）,and TEM-1 gene was determined in all beta-lactamase-positive isolates. The determination rate of BLNAR was 10.3%（12/117）. The mutation rates of ftsI in BLPAR and BLNAR were 68.5%（37/54） and 75.0%（9/12）,and the genotype Ⅲ was the main type [73.0%（27/37） and 77.8%（7/9）]. The 51.4%（19/37） and 77.8%（7/9） of Asn526Lys replacement combined with Ser385Thr replacement occurred between BLPAR and BLNAR. The multiplex PCR typing of BLPAR was classified into 5 types,including a,b,c,d and e. The multiplex PCR typing of BLNAR was classified into 2 types,a and e,and these 2 types had clone-transmitted trends,and the proportions of type a in the 2 kinds of isolates were 63.0%（34/54） and 75%（9/12）,respectively. Conclusions The main resistance mechanism of beta-lactamase-positive Hi is to produce TEM-1. The main resistance mechanism of non-beta-lactamase Hi is caused by Asn526Lys replacement near KTG motif combined with Ser385Thr replacement near SSN motif. The ftsI genotype of BLPAR and BLNAR is mainly type Ⅲ. Both BLPAR and BLNAR have the trend of clone-transmission.

In vitro antimicrobial activity and biofilm inhibitory effect of imipenem combined with tigecycline against mucoid Pseudomonas aeruginosa

ZHONG Feng, FENG Yi, ZHANG Jingjing, XU Yan, FANG Yongmei, TAO Xiubin, WU Jing

2020, 35(6):  519-523.  DOI: 10.3969/j.issn.1673-8640.2020.06.002

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Objective To evaluate the inhibitory and biofilm inhibitory effect of imipenem combined with tigecycline on mucoid Pseudomonas aeruginosa（MPA） in vitro. Methods The minimum inhibitory concentration（MIC） of different concentrations of tigecycline and imipenem against 89 MPA isolates were determined by micro-broth dilution method. The checkerboard method was used to determine and classify the fractional inhibitory concentration index（FICI）,the time-killing curve was used to determine the combined effect,and the 96-well plate combined with crystal violet semi-quantitative staining was used to determine the biofilm inhibitory effect. Results The synergistic effect of tigecycline combined with imipenem on MPA was 46.07%（41 isolates）,partial synergistic effect was 14.6%（13 isolates）,and additive effect was 12.36%（11 isolates）,irrelevant effect was 26.97%（24 isolates）,and there was no antagonistic effect. At culturing for 24 h,the combined therapy showed >2 log₁₀ CFU/mL reduction in viable bacteria compared with tigecycline or imipenem alone. Compared with the use of tigecycline（1/4 MIC）,the combination of tigecycline with imipenem（1/4,1/2,1 MIC） reduced the counts of viable bacteria on each biofilm（P<0.05）. Compared with the use of imipenem（1/4 MIC）,the combination of imipenem with tigecycline（1/4,1/2,1 MIC） reduced the counts of viable bacteria on each biofilm（P<0.05）. Conclusions The combination of tigecycline with imipenem showed synergistic or partial synergistic effects on MPA. Combined therapy has higher antibacterial efficiency than that alone,and it has a certain enhancement effect on the inhibitory of biofilm.

Analysis on the technical key points of Salmonella isolation,screening and identification based on routine detection process

LIU Yue, ZHANG Hongzhi, GU Qifang, LIU Cheng, ZHU Yingying, XU Xuebin

2020, 35(6):  524-530.  DOI: 10.3969/j.issn.1673-8640.2020.06.003

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Objective To analyze and evaluate the technical key points for routine detection of Salmonella,using multi-dimensional composite process based on standard method. Method Process material included Salmonella selective agars and selective enrichment broth. Recovery ratios of 4 selective enrichment broths were detected by reference isolates,and the typical colony characteristics of 5 selective agars belonging to 3 kinds were verified. Suspected colony distinguishment,screening efficiency and sensitivity were compared using different selective agars and selective enrichment broth by actual samples. Results The recovery efficiency of Rappaport Vassiliadis broth（RV）,tetrathionate broth（TTB） and selenite brilliant green broth（SBG） was one dilution different from that of selenite cystine broth（SC）. The first 3 selective enrichment broths were effective for non-typhoidal Salmonella,and SC was effective for Salmonella typhi and Salmonellaparatyphoid. Screening method of double-sugar tubes accorded with the biochemical characteristics of confirmed reference isolates. Except CHORMagar Salmonella（CAS）,the others showed atypical colony characteristics,including H₂S negative isolates and small colonies. The 25 of 84 meat samples were positive,and 42 Salmonella isolates were isolated. The sensitivities of CAS and xylose lysine deoxycholate agar（XLD） combined with RV,TTB,SBG,SC were 88%,84%,72% and 0%,respectively. The sensitivities of RV with TTB,RV with SBG,TTB with SBG were 100%,92% and 88%,respectively. The sensitivities of RV,TTB,and SBG combined with CAS or XLD were 48.00% and 68.00%,respecitvely. The method of screening Salmonella like colonies had high specificity and positive predictive value by double-sugar tubes. Totally,42 Salmonella isolates were confirmed by biochemistry identifier,mass spectrometry and serotyping. Conclusions The effective combined processes could improve the sensitivities of Salmonella detection based on verification by double selective agar and double selective enrichment broths. The technical key points include the gradient selective enrichment,distinguish of the colonies and screening of biochemistry identifier by routine detection of Salmonella.

Diagnostic role of miRNA-21-5p for esophageal carcinoma and correlation of STAT3

WANG Yanzhong, DING Limin, CHEN Yirui, WANG Yongbin

2020, 35(6):  531-534.  DOI: 10.3969/j.issn.1673-8640.2020.06.004

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Objective To evaluate the diagnostic role of microRNA（miRNA）-21-5p for esophageal carcinoma and its mechanism. Methods A total of 78 patients with esophageal carcinoma from January 2016 to June 2018 in Xiasha Branch,Sir Run Run Shaw Hospital,Zhejiang University School of Medicine were enrolled as case group,and 69 healthy subjects were enrolled as control group. Blood and esophageal carcinoma and adjacent tissues were collected before treatment and during operation. The expression of miRNA-21-5p in serum was determined by real-time polymerase chain reaction（PCR）. The target gene of miRNA-21-5p was predicted by 3 databases,including picTar,TagetScan and Tarbase. The target gene intersection of the 3 databases was predicted using Draw venn diagram. Real-time PCR was used to determine the expression of signal transducer and activator of transcription 3（STAT3） mRNA in esophageal carcinoma and adjacent tissues in patients with esophageal carcinoma. Results The results of real-time PCR showed that miRNA-21-5p was highly expressed in serum of patients with esophageal carcinoma,which was higher than that in control group（P<0.01）. The high expression of STAT3 mRNA was also found in esophageal carcinoma tissues of patients with esophageal carcinoma,which was higher than that in adjacent tissues. Spearman correlation analysis showed that miRNA-21-5p and STAT3 were positively correlated. Conclusions The expression of miRNA-21-5p in serum of patients with esophageal carcinoma is high,and STAT3 mRNA is also highly expressed in esophageal carcinoma tissues. There is a positive correlation between miRNA-21-5p and STAT3.

Role of real-time fluorescence quantitative PCR-binding probe in the determination of MP and MP drug resistance

DU Jinlong, FENG Yan, LI Hengtao

2020, 35(6):  535-539.  DOI: 10.3969/j.issn.1673-8640.2020.06.005

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Objective To evaluate the role of real-time fluorescence quantitative polymerase chain reaction（PCR）-binding probe in the determination of Mycoplasma pneumoniae（MP）,to analyze the drug resistance mutation of MP,and to guide the clinical medication for children with MP infection. Methods A total of 70 patients with MP infection and 30 patients without MP infection in Fengcheng Hospital of Shanghai Fengxian District from September 2017 to August 2019 were enrolled,and throat swab specimens were collected from children. MP-DNA amplification and fluorescence probe method were used to determine the drug resistance mutation of MP and macrolide antibiotics. The data of children were collected,and the correlation with MP-DNA positive and drug resistance mutation was analyzed. Results The sensitivity and specificity of PCR-binding probe for determining MP were 80.0%（56/70） and 96.7%（29/30）,respectively. The MP-DNA positive determination rate of double serum MP-IgM antibody titer 4 times/4 times higher or lower group was higher than that of single serum titer ≥1:160 group. Sex,age,disease course and history of macrolide antibiotic treatment before admission had no effect on the positive rate of MP-DNA. The MP-DNA drug resistance mutation rate was 85.7%,and the rate of DNA drug resistance mutation in one course of treatment with macrolide antibiotics before admission was 90.5%,which was higher than those of untreated group（80.0%） and the group without finishing one course of treatment（50.0%）. Conclusions Real-time fluorescence quantitative PCR-binding probe can be used as an effective method for MP determination,providing a reference for the clinical use of antibiotics in MP infection children,which has a high clinical application value.

Relevant factors on short-term critical events in patients with corona virus disease 2019

WANG Bin, YUAN Xu, HAN Feng, TANG Ning

2020, 35(6):  540-545.  DOI: 10.3969/j.issn.1673-8640.2020.06.006

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Objective To investigate the factors for short-term critical events in patients with corona virus disease 2019（COVID-19）. Methods A total of 73 patients with COVID-19 were enrolled from January 13,2020 to February 15,2020. According to whether these patients used high flow nasal catheter oxygen therapy,non-invasive or invasive respiratory aids,the patients were classified into critical group and non-critical group. The clinical data were collected. The determination results of 14 routine indicators [prothrombin time（PT）,activated partial thromboplastin time（APTT）,fibrinogen（Fg）,thrombin time（TT）,D-dimer（DD）,antithrombin activity（AT）,white blood cell（WBC） count,lymphocyte（Ly） count,platelet（PLT） count,C-reactive protein（CRP）,alanine aminotransferase（ALT）,aspartate aminotransferase（AST）,blood urea nitrogen（BUN） and serum creatinine（Cr）] at admission,>1-≤4 d and >4-≤8 d and clinical outcome data（observed until March 6,2020） were collected. The differences in the 2 groups were compared. The risk factors of short-term critical events in COVID-19 patients were analyzed by receiver operating characteristic（ROC） curve analysis. Results The age,number of diabetes mellitus cases and death rate in critical group were higher than those in non-critical group（P<0.05）. The levels of PT,DD,WBC count,CRP,AST and BUN in critical group at the 3 stages were higher than those in non-critical group（P<0.05）. For different courses,APTT,Fg,TT,AT,Ly count,PLT count and Cr had statistical significance between critical group and non-critical group（P<0.05）. The level of ALT had no statistical significance at the 3 stages between the 2 groups（P>0.05）. DD and PLT count were independent risk factors and independent protective factors for short-term critical events. ROC curve analysis indicated that the areas under curves（AUC） of DD,PLT count and DD+PLT count were 0.820 [95% confidence interval（CI） 0.739-0.902],0.764（95%CI 0.652-0.876） and 0.809（95%CI 0.704-0.915）,respectively. The sensitivities of DD,PLT count and DD+PLT count were 67.4%,85.8% and 67.9%,and the specificities were 93.7%,60.7% and 86.8%,respectively. Conclusions DD and PLT count have relations with the occurrence of short-term critical events in patients with COVID-19.

Role of severe acute respiratory syndrome coronavirus 2 antibody detections in the diagnosis of corona virus disease 2019

WANG Jianru, YANG Jinbo, ZHANG Chi

2020, 35(6):  546-550.  DOI: 10.3969/j.issn.1673-8640.2020.06.007

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Objective To investigate the role of severe acute respiratory syndrome coronavirus 2（SARS-CoV-2） IgM and IgG antibody detections in the diagnosis of corona virus disease 2019（COVID-19）. Methods A total of 173 patients diagnosed with COVID-19 were enrolled. There were 140 cases of being positive for the first time of SARS-CoV-2 nucleic acid test and 33 cases of being eventually diagnosed for several times of SARS-CoV-2 nucleic acid test. Totally,101 patients without COVID-19 were enrolled as control group. Colloidal gold immunochromatography assay（CGIA） was used to detect SARS-CoV-2 IgM and IgG antibodies,and fluorescence quantitative polymerase chain reaction（PCR） was used to detect SARS-CoV-2 nucleic acid. Taking clinical diagnosis as gold standard,the efficiency of SARS-CoV-2 IgM and IgG antibody detections in the diagnosis of COVID-19 was analyzed. Results The sensitivities of SARS-CoV-2 IgM and IgG antibody detections were 76.9% and 74.6%,the specificities were 94.0% and 95.1%,and the consistency rates were 83.2% and 82.1%,which were well consistent with the clinical diagnosis（kappa values were 0.663 and 0.644）,respectively. Receiver operating characteristic（ROC） curve analysis showed that the areas under curves（AUC） of SARS-CoV-2 IgM and IgG antibody detections for the diagnosis of COVID-19 were 0.85. Conclusions The detections of SARS-CoV-2 antibodies have high sensitivities,specificities and consistency rates,which can be used for the auxiliary diagnosis of COVID-19.

Correlation of laboratory index and clinical classification among 342 corona virus disease 2019 patients in Ezhou

LIU Sijia, CHENG Feng, YANG Xiaoyu, HE Jie, LI Hui, ZHANG Wei, ZHANG Jianbo, GONG Guofu, DUAN Xiuqun

2020, 35(6):  551-556.  DOI: 10.3969/j.issn.1673-8640.2020.06.008

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Objctive To investigate the correlation of laboratory index and clinical classification in corona virus disease 2019（COVID-19） patients. Methods A total of 342 patients with COVID-19 in Ezhou Central Hospital from January 23,2020 to February 12,2020 were enrolled. There were 196 cases of general group,97 cases of severe group and 49 cases of critical group. Their age,sex,basic diseases,co-infection and blood indexes were compared. Receiver operating characteristic（ROC） curve was used to evaluate the roles of blood indexes in the diagnosis of COVID-19. Results The age and prevalence of basic diseases among the 3 groups had statistical significance（P<0.05）. The proportion of males in critical group was higher than that of females（P<0.05）. The infection rate of co-infection with other respiratory pathogens had statistical significance among the 3 groups（P<0.05）. The infection rates of Mycoplasma pneumoniae and influenza B virus were the highest. In severe group,lymphocyte percentage（LYMPH%）,total protein（TP） and albumin（Alb） were lower than those in general group（P<0.05）,while D-dimer（DD）,fibrinogen（Fg）,erythrocyte sedimentation rate（ESR） and C-reactive protein（CRP） were higher（P<0.05）. In critical group,LYMPH%,TP and Alb were lower than those in severe group（P<0.05）,while white blood cell（WBC） count,DD and procalcitonin（PCT） were higher（P<0.05）. According to ROC curve analysis,ESR,Alb and LYMPH% were the most valuable blood indexes to distinguish general and severe groups,and PCT,LYMPH% and Alb were the most valuable blood indexes to distinguish severe and critical groups. Conclusions ESR,Alb,LYMPH% and PCT determinations are helpful to diagnose the conditions.

Study on early laboratory warning of severe COVID-19

ZHA Qiongfang, FENG Bo, LI Xiaoning, ZHOU Donghua, KANG Yu, QIN Hui

2020, 35(6):  557-560.  DOI: 10.3969/j.issn.1673-8640.2020.06.009

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Objective To evaluate the predictive role of early laboratory indicators in the severity of corona virus disease 2019（COVID-19）. Methods Retrospective analysis was used to analyze the laboratory results in 85 patients with COVID-19 at the time of admission,and the results included blood routine test,liver function,kidney function,blood coagulation function,interleukin 6（IL-6）,ferritin,C-reactive protein（CRP）,erythrocyte sedimentation rate（ESR）,procalcitonin（PCT）,myoglobin,troponin and amyloid A. Results In severe group,the levels of white blood cell（WBC） count,the absolute value of neutrophil（NEUT#）,neutrophil-lymphocyte ratio（NLR）,lactate dehydrogenase（LDH）,CRP,D-dimer（DD）,ferritin and creatinine were higher than those in mild group（P<0.05）. WBC count,NEUT#,NLR,LDH,myoglobin,CRP and ferritin were predictive risk factors of COVID-19. NLR was an independent predictor for the severity of COVID-19. When the cut-off value of NLR was 5.61,the sensitivity and specificity were 68.75% and 78.38%,respectively. Conclusions The increasing of WBC count,NEUT#,NLR,LDH,myoglobin,CRP and ferritin suggests the risk of severity,and NLR is an independent prediction for the severity of COVID-19.

Predictive role of serum fibronectin level in the second trimester of pregnancy for patients with gestational diabetes mellitus complicated with preeclampsia

XU Zhen, MA Lijuan, WU Siyi, WANG Yan, JIA Keke

2020, 35(6):  561-565.  DOI: 10.3969/j.issn.1673-8640.2020.06.010

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Objective To investigate the predictive role of serum fibronectin（Fn） level in the second trimester of pregnancy for patients with gestational diabetes mellitus（GDM） complicated with preeclampsia（PE）. Methods A prospective case-control study was conducted to select 458 serum samples from pregnant women with GDM from 24 to 28 weeks of gestational weeks to observe their pregnancy outcomes. Totally,30 patients developed PE as PE-GDM group,10 patients were diagnosed as gestational hypertension（GH） as GH-GDM group,and 40 cases of GDM with full-term normal delivery and no complication were enrolled from 458 subjects（GDM group）. Immunofluorescence assay was used to determine the levels of Fn and related clinical biochemical indicators in the 3 groups. Results The level of Fn in PE-GDM group was 221.72（202.33,264.52）mg/L,those in GH-GDM group and GDM group were 185.13（174.30,216.21）mg/L and 198.77（185.65,208.37）mg/L,respectively（P<0.05）. There was no statistical significance in Fn levels between GH-GDM group and GDM group（P>0.05）. The levels of triglyceride（TG） and pre-pregnancy body mass index（BMI） in PE-GDM group were 3.60（2.96,4.62）mmol/L and （25.10±3.20）kg/m²,respectively,and they were higher than those in GH-GDM group [2.32 （2.00,2.81）mmol/L and （23.97±3.36）kg/m²] and GDM group [2.52 （2.13,2.94）mmol/L and （23.38±3.61）kg/m²]（P<0.05）. Receiver operating characteristic（ROC） curve analysis showed that the areas under curves（AUC） of Fn,TG and pre-pregnancy BMI were 0.778,0.794 and 0.654,respectively,and the 95% confidence intervals（CI） were 0.665-0.890,0.681-0.907 and 0.526-0.782,respectively. The AUC of the combined determination of the 3 indicators was 0.818,and the 95% CI was 0.714-0.921. Conclusions Serum Fn levels in the second trimester have a certain predictive value for GDM patients with PE. The combined determination of Fn,TG and pre-pregnancy BMI in the second trimester of pregnancy is important in predicting GDM with PE.

Role of CD55,CD59 and FLAER combined determination for the diagnosis of PNH

LU Jiacai, HUANG Ying, MO Yang, ZHOU Xiaofen, YAO Xin

2020, 35(6):  566-569.  DOI: 10.3969/j.issn.1673-8640.2020.06.011

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Objective To determine the expressions of CD55,CD59 and fluorescent aerolysin（FLAER） of anemia patients,and to evaluate the role of CD55,CD59 and FLAER combined determination for the diagnosis of paroxysmal nocturnal hemoglobinuria（PNH）. Methods A total of 157 inpatients,including 38 patients with iron deficiency anemia（IDA）,23 patients with megaloblastic anemia（MA）,27 patients with myelodysplastic syndrome（MDS）,25 patients with aplastic anemia（AA）,31 patients with autoimmune hemolytic anemia（AIAH） and 13 patients with PNH,were enrolled. Totally,20 healthy subjects were enrolled as control group. Erythrocyte and granulocyte CD55,CD59 and granulocyte and monocyte FLAER were determined. Results Erythrocyte and granulocyte CD55^- and CD59^- percentages in PNH group had statistical significance compared with control group（P<0.05）. In PNH group,granulocyte and monocyte FLAER^- percentages were >50%,which were higher than erythrocyte and granulocyte CD55^- and CD59^- percentages（P<0.05）. The consistency rate of CD55,CD59 and FLAER determinations was 100%. There were 3 AA patients and 2 MDS patients with FLAER deletion and normal CD55 and CD59 expressions. There were 2 AIHA patients with partial erythrocyte CD55 and CD59 deletion and normal granulocyte CD55 and CD59 and FLAER expression. Conclusions CD55 and CD59 expression defects are the main characteristics of PNH,and FLAER determination combined with CD55 and CD59 plays a role in the diagnosis and differential diagnosis of PNH.

Changes of neutrophil oxidative phagocytosis and related receptors in sputum samples of COPD patients

YAN Peiyi, ZHANG Ji, TU Huanping, ZHANG Li, QIU Yue, ZHANG Fengying, HU Yiwen, JIN Shu

2020, 35(6):  570-577.  DOI: 10.3969/j.issn.1673-8640.2020.06.012

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Objective To investigate the roles of neutrophil oxidative phagocytosis and related receptors [chemokine C-C receptor -1（CCR1）,Toll-like receptor（TLR） 2 and TLR4] in patients with chronic obstructive pulmonary disease（COPD）. Methods A flow cytometry method was established to determine neutrophil oxidative phagocytosis and related receptors in sputum samples of COPD patients. By flow cytometry,neutrophil oxidative phagocytosis in peripheral blood and sputum samples of 30 COPD patients（COPD group）,30 pneumonia patients（pneumonia group） and 30 healthy subjects（healthy control group） was determined. Neutrophil oxidative phagocytosis and the expression levels of CCR1,TLR2 and TLR4 on the surface were determined. Results The neutrophil oxidative phagocytosis of peripheral blood in pneumonia group and COPD group was lower than that in healthy control group（P<0.05）,and there was no statistical significance between pneumonia and COPD groups（P>0.05）. There was statistical significance in the expression of CCR1 on the surface of peripheral blood neutrophils among COPD,pneumonia and healthy control groups（P<0.05）,but there was no statistical significance for TLR2 and TLR4 expression levels（P>0.05）. The neutrophil oxidative phagocytosis in sputum samples in COPD group was lower than that in pneumonia group（P<0.05）,the expression level of CCR1 was higher than that in pneumonia group（P<0.05）,and there was no statistical significance for the expression levels of TLR2 and TLR4（P>0.05）. Pearson correlation analysis showed that the positive rate of neutrophil oxidative phagocytosis in sputum samples in COPD group was negatively correlated with the expression level of CCR1（r=-0.548,P<0.01）,and it had no correlation with the expression levels of TLR2 and TLR4（r=-0.206 and -0.219,P>0.05）. Conclusions Neutrophil oxidative phagocytosis in sputum samples of COPD patients is decreased,which can increase CCR1 expression level on the surface,suggesting that COPD patients'respiratory neutrophil oxidative phagocytosis injury may be one of the mechanisms of COPD pathogenesis.

Clinical role of serum cathepsin D determination in non-alcoholic fatty liver disease

WANG Qianqing, HONG Juan, DU Chao, CHU Qinghua

2020, 35(6):  578-582.  DOI: 10.3969/j.issn.1673-8640.2020.06.013

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Objective To investigate the clinical role of serum cathepsin D（CTSD） in non-alcoholic fatty liver disease（NAFLD）. Methods A total of 356 NAFLD patients were enrolled and classified into 3 groups according to ultrasound diagnosis,which were mild group（146 patients）,moderate group（113 patients） and advanced group（97 patients）. Totally,300 healthy subjects were enrolled as control group. Serum levels of CTSD,alanine aminotransferase（ALT）,aspartate aminotransferase（AST）,gamma-glutamyltransferase（GGT）,triglyceride（TG）,total cholesterol（TC） and low-density lipoprotein cholesterol（LDL-C） were determined. Pearson correlation analysis and Spearman correlation analysis were used to assess the correlation between each indicator and the severity of NAFLD. Receiver operating characteristic（ROC） curve was used to evaluate the diagnosis value of these indicators. Results Serum levels of ALT,TC,TG,LDL-C and CTSD in NAFLD group were higher than those in control group（P<0.01）,which had statistical significance among NAFLD groups（P<0.01）. Compared with control group,serum levels of AST and GGT were increased in NAFLD group （P<0.01）,and those in advanced group were higher than those in moderate group（P<0.01）,while there was no statistical significance between control and mild groups（P>0.05）. Serum CTSD was positively correlated with the severity of NAFLD （r_s=0.800 5,P<0.000 1）,ALT（r=0.541,P<0.01）,TC（r=0.662,P<0.01） and LDL-C（r=0.571,P<0.01）. ROC curve analysis showed that the areas under curves（AUC） of CTSD,ALT and GGT in the diagnosis of NAFLD were 0.940,0.822 and 0.627,respectively. The optimal cut-off values were 12.67 pg/mL,38.00 U/L and 53.00 U/L,respectively. The sensitivities were 76.97%,51.69% and 34.55%,and the specificities were 91.33%,93.00% and 92.33%,respectively. The AUC of CTSD combined with ALT determination in the diagnosis of NAFLD was 0.960,the sensitivity was 88.48%,and the specificity was 90.00%. Conclusions Serum level of CTSD is correlated with the severity of NAFLD,and it can be used as a biomarker for the early diagnosis and staging of NAFLD.

Correlation between interleukin and high-sensitivity C-reactive protein with primary angle-closure glaucoma

YAO Tianyue, LI Jian, SONG Yunxiao, YUAN Wenhua, ZHAO Zhiyun, GE Wen

2020, 35(6):  583-586.  DOI: 10.3969/j.issn.1673-8640.2020.06.014

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Objective To investigate the clinical roles of interleukin（IL）-1,IL-2,IL-6,IL-8,IL-10 and high-sensitivity C-reactive protein（hs-CRP） levels in primary angle-closure glaucoma（PACG）. Methods A total of 72 patients with PACG and 55 healthy subjects were enolled. General information and eye parameters [intraocular pressure,central corneal thickness（CCT）,vertical cup-disc ratio（VCDR）,axial length（AL）,anterior chamber depth（ACD）,visual field mean deviation（MD） and visual field mean sensitivity（MS）] were collected and recorded in detail. Binary Logistic regression analysis was used to assess the risk factors of PACG. Spearman correlation analysis was used to assess the relationship between IL and eye parameters. Results The levels of IL-6,IL-8 and hs-CRP in PACG group were higher than those in control group（P<0.05）. There was no statistical significance in IL-1,IL-2 and IL-10 between PACG and control groups（P>0.05）. There was a positive correlation of IL-6 level with MD and VCDR in PACG patients（r=0.431 and 0.270,P<0.05）. There was a negative correlation between CCT and IL-8（r=-0.279,P=0.023）,and there were positive correlations of IL-8 with MD and VCDR（r=0.322 and 0.330,P<0.05）. Multivariate binary Logistic regression analysis showed that IL-6 [odds ratio（OR）=1.296,95% confidence interval（CI）1.009-1.664] and IL-8（OR=1.106,95% CI 1.044-1.172） were correlated with PACG. Conclusions IL-6,IL-8 and hs-CRP may be related to PACG.

Comparison of 2 high-throughput nucleic acid determination methods for determining multiple respiratory pathogens

JING Hongbo, LI Zhan, HE Mu, WANG Yanbo, LI Wen, MA Hongmei

2020, 35(6):  590-594.  DOI: 10.3969/j.issn.1673-8640.2020.06.016

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Objective To compare the efficiency of multiplex fluorescence quantitation polymerase chain reaction（PCR） and TaqMan array card（TAC）for determining multiple respiratory pathogens. Methods The clinical respiratory specimens were collected,and the nucleic acid was determined by multiplex fluorescence quantitation PCR and TAC. The sensitivities and specificities of the 2 methods were compared. Results The positive rate,sensitivity and specificity of multiplex fluorescence quantitation PCR were 30.1%,93.4% and 98.9%. The positive rate,sensitivity and specificity of TAC were 27.9%,87.4% and 98.9%. The consistency of the 2 methods was 92.8%,and there was no statistical significance in the sensitivities and specificities of the 2 methods（P>0.05）. Conclusions The sensitivities and specificities of the 2 methods are close. The spectrum of respiratory pathogens determined by TAC is broader,and it can be used as an efficient method for rapid screening of multiple respiratory pathogens in emergency circumstances.

A novel electrochemical biosensor for the detection of serum CYFRA21-1

WANG Jianfu, LI Xiaoping, CHEN Wenhu, FAN Kai, QIAN Miaomiao

2020, 35(6):  595-600.  DOI: 10.3969/j.issn.1673-8640.2020.06.017

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Objective To construct a highly sensitive electrochemical biosensor for the detection of cytokeratin 19-fragment（CYFRA21-1）. Methods Three-dimensional graphene @gold nanoparticles（3D-G@Au）nanocomposite was modified on glassy carbon electrode surface to enhance the conductivity of electrochemical biosensor. Anti-CYFRA21-1 was captured and fixed on the modified 3D-G@Au glassy carbon electrode through the cross-linking of chitosan and glutaraldehyde. The 3D-G@Au electrochemical biosensor had been completed as to determine CYFRA21-1. The electrochemical and determination performance of successfully constructed 3D-G@Au electrochemical biosensor was evaluated preliminarily. Results The cyclic voltammetry showed that the redox peaks of bare glassy carbon electrode,3D-G modified glassy carbon electrode and 3D-G@Au modified glassy carbon electrode increased gradually,indicating that the 3D-G@Au electrochemical biosensor was successfully constructed. The optimal incubation time of antigen-antibody interaction was 60 min,and the optimal incubation temperature was 35 ℃. CYFRA21-1 was detected by 3D-G@Au electrochemical biosensor,the linear range was 0.1-300.0 ng/mL,and the determination limit was 0.1 ng/mL（signal-to-noise ratio=3）. 10 mmol/L Vit C,10 mmol/L DA and 10 mmol/L UA had no interference to the determination,and the reproducibility was good [relative standard deviations（RSD） were 1.08% and 3.11%], compared with that of chemiluminescence method (RSD was 19.6%-4.78%). The 3D-G@Au electrochemical biosensor can be stable for 30 d at 4 ℃. Conclusions It is potential for the determination of serum CYFRA21-1 with 3D-G@Au electrochemical biosensor.

Clinical evaluation of homemade Precil C3510 automatic coagulation analyzer for routine coagulation parameters

ZHANG Qiang, LIN Jing, ZONG Xiaolong, SUN Guang, LIU Junfeng

2020, 35(6):  601-608.  DOI: 10.3969/j.issn.1673-8640.2020.06.018

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Objective To evaluate the clinical performance of homemade Precil C3510 automatic coagulation analyzer for routine coagulation parameters [prothrombin time（PT）,activated partial thromboplastin time（APTT）,fibrinogen（Fib） and D-dimer（DD）] and its application in monitoring the efficiency of warfarin and unfractionated heparin anticoagulant therapy. Methods According to the Clinical and Laboratory Standards Institute（CLSI） guidelines,the imprecision,interference,linearity and reference intervals of Precil C3510 for the determinations of PT,APTT,Fib and DD were validated. The consistency between Precil C3510 and STA-Compact automatic coagulation analyzer（STA-Compact） was evaluated by Spearman correlation,Passing-Bablok regression and Bland-Altman plot with STA-Compact as a reference system. The consistency of the international normalized ratio（INR） between Precil C3510 and STA-Compact for patients receiving warfarin therapy was evaluated. APTT monitoring range for Precil C3510 based on the standard of 0.3-0.7 IU/mL of anti-Xa activity determined by STA-R Evolution was investigated. Results The imprecision,linearity and reference intervals of Precil C3510 met the manufacturers' requirements. Triglyceride with 3.2 mg/mL and more higher concentrations had a negative effect on DD. The 2 instruments had good consistency in determining PT and Fib. There was no systematic,proportional and random differences,and the deviations were within clinical acceptance interval. APTT determined by the 2 instruments had systematic difference [regression equation intercept was -18.121 6,95% confidence interval（CI） -26.003 9--12.522 1],proportional difference （slope was 1.261 8,95%CI 1.123 4-1.435 6） and random difference（the bias within ±1.96s accounting for 92.31%）,and the average bias （-18.9%,95%CI -22.9%--14.9%）was not within the clinical acceptance interval（-7.5%-7.5%）. The results of the 2 instruments had a consistency in judging APTT based on their respective reference intervals（Kappa value was 0.87,95%CI 0.74-0.99）. The APTT-based unfractionated heparin anticoagulant therapeutic range of Precil C3510 was 62-108.1 s. The INR of 35 patients（87.5%）receiving warfarin therapy showed a INR difference<0.5,indicating a good correlation between the 2 instruments. Conclusions Precil C3510 has good analytical performance for routine coagulation parameters,and it can be used to monitor the anticoagulation effect of warfarin and unfractionated heparin for routine analysis.

Evaluation on the differential ability of visual APTT mixed correction test

PENG Zheng, TANG Ning, ZOU Zhiping, XIAO Fang

2020, 35(6):  609-613.  DOI: 10.3969/j.issn.1673-8640.2020.06.019

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Objective To evaluate the differential ability of visual activated partial thromboplastin time（APTT） mixed correction test for coagulation factor deficiency and coagulation inhibitor. Methods A total of 62 specimens with prolonging APTT were collected,including 20 cases of coagulation factor deficiency,22 cases of coagulation inhibitor Ⅷ positive and 20 cases of lupus anticoagulant（LA） positive. Visual APTT mixed correction test and Rosner index method were used. Results The sensitivities of Rosner index method and visual APTT mixed correction test for judging coagulation factor deficiency were 100.0%,and the sensitivities for judging LA were 95.0%. The sensitivities for judging coagulation inhibitor were 95.5% by visual APTT mixed correction test and 50% difference method. Conclusions Using the visual APTT mixed correction test,the differential ability of coagulation factor deficiency and coagulation inhibitor is comparable to Rosner index method,and visual APTT mixed correction test is feasible for interpreting results.

Role of FadD9 recombinant protein for the diagnosis of active tuberculosis

ZHANG Junli, LIN Chen, WANG Yuchen, LIU Jun, YUAN Li, PAN Zhifen, ZHANG Lu

2020, 35(6):  614-619.  DOI: 10.3969/j.issn.1673-8640.2020.06.020

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Objective To investigate the role of FadD9 recombinant protein of Mycobacterium tuberculosis（MTB） for the diagnosis of active tuberculosis. Methods FadD9 recombinant protein was prepared and identified. C57BL/6 mouse model was used to evaluate its immunogenicity. Using Ag85A protein as positive reference antigen,the specific antibody levels of FadD9 recombinant protein in serum samples of 145 active tuberculosis patients（active tuberculosis group）,38 close-contact tuberculosis patients（MTB close-contact group） and 101 healthy subjects（healthy control group） were determined. The FadD9 recombinant protein was incubated with blood samples of 58 tuberculosis patients and 34 non-tuberculosis patients at 37 ℃ for 20-24 h,and then was determined by interferon-gamma release assay（IGRA）. The results were compared with the standard tube in IGRA kit,and the sensitivity and specificity of FadD9 recombinant protein in the diagnosis of tuberculosis were analyzed by receiver operating characteristic（ROC）curve. Results FadD9 recombinant protein could induce a high level of humoral immune response in mice,which was close to that of Ag85A. Serum anti-FadD9 antibody level in active tuberculosis group was higher than those in healthy control group and MTB close-contact group（P<0.000 1）. There was no statistical significance between MTB close-contact group and healthy control group（P>0.05）. There was no statistical significance among the 3 groups in the level of anti-Ag85A antibody（P>0.05）. For the diagnosis of tuberculosis,the area under curve（AUC） of FadD9 recombinant protein was 0.682,the optimal cut-off value was 0.343,and the sensitivity and specificity were 96.8% and 37.5%,respectively. Conclusions FadD9 recombinant protein is potential in the diagnosis of active tuberculosis.

Research progress on neutrophil extracellular traps in ulcerative colitis

LUO Ting, PAN Xiujun

2020, 35(6):  626-630.  DOI: 10.3969/j.issn.1673-8640.2020.06.023

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Ulcerative colitis（UC） is an autoimmune disease associated with chronic bowel inflammation,the pathogenesis of UC is unclear,but its etiology is multifactorial,and the incidence of UC is increasing. Neutrophil granulocytes play roles in the immune response of humans and are the first cells to migrate to the site of inflammation. Traditionally,neutrophils defend the host against pathogens,the important functions of neutrophils include chemotaxis,phagocytosis and degranulation. Recent massive data support that neutrophils were found to have a specific mechanism of immune defense by forming neutrophil extracellular traps（NET）. NET not only play a defense reaction but also are related to the pathogenesis of various kinds of diseases,including autoimmune diseases. Investigating the relationship of NET in the development of autoimmune diseases will further help with understanding the etiology and pathogenesis of autoimmune diseases. This review focuses on the role of NET in the pathogenesis of UC,in order to provide a reference for the therapy and diagnosis of UC.