Recently,the hepatitis of unknown etiology among children has shown a concentrated trend in many countries and regions,which has aroused concern from the World Health Organization. Based on the reports of existing cases,numerous hypotheses have been proposed,among which the hypothesis that adenovirus may be the cause of the hepatitis of unknown etiology among children is debated widely. According to the occurrence of the hepatitis of unknown etiology among children,the pathogenic characteristics of adenovirus and literatures on adenovirus hepatitis,this review analyzes the pathogens that may cause the hepatitis of unknown etiology among children and puts forward suggestions on the work to be carried out to clarify the pathogens. All of those details may provide a direction for timely and accurate diagnosis of related cases in China.
Objective To investigate the clinical characteristic and genetic variation analysis of 2 cases of neonatal butyrylcholinesterase deficiency（BCHED） caused by compound heterozygous mutation of BCHE gene. Methods Totally,the clinical characteristics of 2 cases of BCHED were analyzed,and trio-based whole-exome sequencing was performed. The harm of variants was evaluated by bioinformatics analysis,which were verified by Sanger sequencing. Results The 2 cases had decreased serum cholinesterase and increased bilirubin,without liver function injure. Case 1 had neonatal diarrhea symptom,and Case 2 had neonatal jaundice symptom. The results of genetic analysis showed that there were compound heterozygous mutations（c.401_c.402insA and c.221delC; c.401_c.402insA and c.127G>A） in BCHE gene in the 2 cases. Sanger sequencing confirmed the existence of mutation. c.221delC and c.127G>A were not reported in ClinVar and HGMD databases,which were new mutations. Conclusions The gene variation and phenotype spectra have further expanded through the discovery of different phenotypes and new variations of BCHE gene in the 2 neonatal cases of BCHED.
Objective To investigate how to obtain the appropriate evaluation standard based on the change or constant concentration value as performance standard through thrombelastogram proficiency verification data,and to improve continuously and effectively the determination level of thrombelastogram. Methods The data reported by laboratories participating in the 2018-2020 proficiency verification of Shanghai Center for Clinical Laboratory for thrombelastogram were collected. The relation of U-scores between the reported data of coagulation reaction time,coagulation formation time,angle and maximum amplitude of thrombus and the target value was evaluated,and the evaluation standard of each index based on the change or constant concentration value was obtained. Results The constant concentration values of coagulation reaction time,angle and maximum amplitude of thrombus could be used as evaluation criteria. Coagulation formation time can adopt the evaluation standard based on the change of concentration value. Conclusions The performance specifications of each index of thrombelastogram obtained in this study can be applied to the evaluation standards of proficiency verification and the requirements of allowable imprecision of internal quality control,which can evaluate the determination level of thrombelastogram in Shanghai objectively.
Objective To investigate the feasibility of replacing manual counting method with microscope in cell counts and white blood cell classification by automated hematology analyzer in body fluid mode. Methods A total of 287 serous cavity effusion specimens from inpatients were collected,and the manual counting method with microscope（manual method） and the instrumental determination method with automated hematology analyzer（instrumental method） were used for red blood cell and white blood cell counts and white blood cell classification. The manual method was used as the gold standard. The accuracy of instrumental method was evaluated. Results For the specimens whose white blood cell count 201-15 000 cells/μL or the red blood cell count ≥200 cells/μL,the instrumental method and manual method had good correlation in cell counts,and the 2 methods had a good correlation in white blood cell classification（P<0.01）. Conclusions The automated hematology analyzer has the advantages of rapidity,accuracy and reproducibility in cell counts and white blood cell classification of serous cavity effusion specimens. When the red blood cell and white blood cell counts are within certain linear ranges,instrumental method can replace the manual counting method with microscope.
Objective To establish and evaluate the performance and clinical application of plasma cfDNA determination in colorectal cancer patients based on next generation sequencing（NGS） platform. Methods Plasma circulating free DNA（cfDNA） determination system（cfDNA panel） for colorectal cancer patients had been established. The accuracy,precision,minimum sample amount and input of extraction,sensitivity and specificity of cfDNA panel were evaluated. The results were compared with both the results of Miseq Dx NGS platform and amplification refractory mutation system（ARMS）. Results The accuracy of cfDNA panel for colorectal cancer was 99.97%,and the determination rate of expected mutation was 100%. The minimum amount of plasma extraction was 1 mL,and the minimum amount of cfDNA input was 10 ng.The sensitivity was 0.2%. The 10 000 mg/L hemoglobin,500 mg/L bilirubin or 2% fat emulsion had no effet on the extraction of colorectal cancer cfDNA panel. Compared with the results of Miseq Dx NGS platform and ARMS,the consistency rates were 94.12% and 90.91%,respectively,and the positive and negative predictive rates of KRAS,NRAS,BRAF and PIK3CA were 100% and 99.60%,respectively. Conclusions The performance of cfDNA panel in colorectal cancer meets the needs of clinical application. The results of plasma cfDNA determination are highly consistent with the determination results of matched tumor tissue,which is a useful supplement for tissue biopsy.
The autoverification of reports is the future development trend of clinical laboratories,which can significantly shorten the laboratory sample turn-around time and improve work efficiency. The autoverification of coagulation tests is less used in clinical laboratories,due to many problems such as the identification of unqualified samples and the complex rules of autoverification. The basis for the implementation of autoverification is strict and standardized quality control of pre-analysis and interconnection of information systems. Based on the premise of accurate results,clinical laboratories can set appropriate autoverification rules according to its own conditions,continuously optimize the autoverification rules for coagulation test,and gradually increase the autoverification pass rate of clinical laboratory test.
Corona virus disease 2019（COVID-19） epidemic has seriously endangered human health and economic and social development. Accurate etiological determination in clinical laboratories is a prerequisite for epidemic prevention and control. At present,clinical laboratory determination technologies for severe acute respiratory syndrome coronavirus-2 mainly include nucleic acid,antibody and antigen determinations. Nucleic acid determination is a gold standard for the COVID-19 diagnosis with high sensitivity and specificity,but it has high requirements for personnel technical needs and experimental conditions. Antibody determination can be used as a supplement for nucleic acid determination to reduce missed determination and monitor the disease progress,while universal vaccination will affect the interpretation of antibody determination results,like false positivity. Antigen determination is convenient and fast,which can be operated at home,but its sensitivity is greatly affected by the disease process and viral load. In order to improve the efficiency of large-scale population screening,the National Health Commission has successively issued 5-in-1,10-in-1,and 20-in-1 mixed sampling technologies. However,the multiplication of preservation solution in the mixed sampling tube will result in virus dilution and increase the missed determination probability,and mixed sampling also aggravates the workload of epidemiological investigation and subsequent confirmation of the positive specimen in primary screening. With the progress of the COVID-19 epidemic,the SARS-CoV-2 continues to mutate,and the clinical laboratory determination technologies are gradually enriched and improved. Accurate analysis and judgment of the advantages and disadvantages of existing determination technologies and their application scenarios are conducive to the scientific prevention and control of the epidemic. This review will analyzes the following key clinical laboratory determination technical issues and put forward application suggestions,including the “mixed sampling” and “mixed determination” in nucleic acid determination,the impact of virus mutation on the performance of nucleic acid determination,the advantages and disadvantages of “endogenous” and “exogenous” internal standards in nucleic acid determination,the clinical significance and application scenarios of antigen determination and the value of antibody determination after vaccination.
Malignant tumors have evolved into a severe hazard to human health. Between China's tumor diagnosis and treatment and those of Europe,America and other developed countries,there is still a significant disparity. Early determination and accurate comprehensive diagnosis and treatment are essential for achieving the aim of a healthy China. However,there can be no proper therapy without an accurate diagnosis. Accurate diagnosis is based on a thorough understanding of tumor features and their evolution. The discovery of new tumor molecular features has changed tumor diagnostic and treatment concepts and strategies in the recent years,presenting a new opportunity and challenge in the area of tumor diagnosis and therapy. New tumor marker molecules are increasingly becoming more essential in clinical tumor diagnosis,recurrence,metastasis monitoring and prognosis.
Estrogen receptor（ER） belongs to the nuclear receptor superfamily,which mainly binds with the ligand estrogen to activate transcription of its target genes. The interaction of ER with estrogen could actively recruit co-regulator factors to influence transcription of ER-dependent genes. ER not only plays a role in promoting sexual organ maturation,parasexual development and maintaining sexual function,but also participates in the regulation of tumor cell growth,drug resistance,epithelial-mesenchymal transformation（EMT） and metastasis during the occurrence and development of tumors. ER is expressed at a low level in osteosarcoma. Regulating the expression level or activity of ER can effectively change the expression of estrogen/ER-dependent transcription factors and affect the invasion,metastasis,drug sensitivity and growth characteristics of tumors. This review focuses on the research progress of ER in the occurrence,development and treatment of osteosarcoma in the recent years,trying to provide new insights for clinic.
Objective To investigate the effects of antiestrogen（tamoxifen）on the proliferation and the expression level of CC chemokine ligand （CCL）2 of osteosarcoma cells. Methods The difference of estrogen receptor（ER） expression between osteosarcoma cells（143B,U-2OS） and osteoblast（hFOB1.19） was observed. CCK8 experiment and plate colony formation assay were used to observe the effects of tamoxifen on the proliferation of osteosarcoma cells. The relative expression levels of inflammatory factors,IL-6,IL-8,TGF-β,CCL2 and CCL5 in osteosarcoma cells were determined by fluorescence quantitative polymerase chain reaction（PCR）. The effects of tamoxifen on CCL2 expression of osteosarcoma cells in cell culture medium were determined by enzyme-linked immunosorbent assay（ELISA）. Results The expression level of ER mRNA in osteosarcoma cells（143B,U-2OS） was lower than that in osteoblast（hFOB1.19）. Tamoxifen could inhibit the proliferation of osteosarcoma cells. Tamoxifen could inhibit the expressions of inflammatory factors,IL-6,IL-8,TGF-β,CCL2 and CCL5,especially the chemokine CCL2 in osteosarcoma cells. Conclusions Tamoxifen could inhibit the proliferation of osteosarcoma cells and the expression of CCL2,suggesting that it might be used as a adjuvant drug for postoperative chemotherapy of osteosarcoma.
Objective To investigate the value of methylation determination of fecal adhesin syndecan 2（SDC2） gene in the auxiliary diagnosis of colorectal cancer. Methods Totally,51 patients with colorectal cancer（colorectal cancer group）,22 patients with adenoma（adenoma group） and 39 healthy subjects（healthy control group） were enrolled. The stool and serum samples of all the subjects were collected to determine fecal SDC2 gene methylation and serum carcinoembryonic antigen（CEA） and carbohydrate antigen 19-9（CA19-9）levels. Among them,26 fecal samples were pretreated by P12 automatic sample pretreatment instrument and manual method,and the correlation of the determination results after pretreatment by the 2 methods was compared. Pearson correlation analysis and kappa test were used to evaluate the correlation and consistency between the 2 methods. Results The methylation positive rate of fecal SDC2 gene in colorectal cancer group was higher than those in adenoma group and healthy control group（P<0.01）. In colorectal cancer group,the methylation positive rate of fecal SDC2 gene was higher than those of serum CEA and CA19-9（P<0.01）. There was no statistical significance in methylation positive rate of fecal SDC2 gene between patients with colorectal cancer stage Ⅰ-Ⅳ（P>0.05）. The methylation positive rate of fecal SDC2 gene in patients with colorectal cancer stage Ⅰ-Ⅲ was higher than those of CEA and CA19-9 in serum（P<0.01）. Receiver operating characteristic（ROC） curve analysis showed that the areas under curves（AUC） of the methylation of fecal SDC2 gene,serum CEA,serum CA19-9 and the combined determination for colorectal cancer diagnosis were 0.965,0.694,0.567 and 0.976,respectively. Serum CEA combined with the methylation of fecal SDC2 gene can increase the positive rate of colorectal cancer to 90.2%（46/51）. There was a positive correlation between the determination results of P12 automatic sample pretreatment instrument and manual method（r=0.994,P<0.001）,and the consistency was good（kappa=0.845,P<0.001）. Conclusions The methylation of fecal SDC2 gene has high sensitivity and specificity,which can be used as an auxiliary diagnosis of colorectal cancer,and it also has high clinical value in early colorectal cancer screening.
Objective To investigate the clinical value of the SorCS1 gene promoter methylation determination in patients with colorectal cancer. Methods Cancer and adjacent tissues from 17 colorectal cancer patients were collected. The MassARRAY platform was used to determine the levels of gene promoter methylation in cancer tissues and adjacent tissues of patients with colorectal cancer,and the genes with different promoter methylation between cancer tissues and adjacent tissues were screened out and then compared with the Cancer Genome Atlasc（TCGA） database. Meanwhile,the methylation levels of SEPT9 gene were determined. The difference （Δ） determined by subtracting the determination value of gene promoter methylation in adjacent tissue from that in cancer tissue indicated the difference. Results A total of 2 genes（SorCS1 and CASR） were screened out. The levels of SorCS1,CASR and SEPT9 methylation in colorectal cancer tissues were higher than those in adjacent cancer tissues（P<0.001）. The comparison with TCGA database revealed that the expressions of SorCS1 gene were lower in patients with bladder urothelial carcinoma,cervical squamous cell carcinoma,colon adenocarcinoma,glioblastoma multiforme,rectum adenocarcinoma and stomach adenocarcinoma than that in healthy controls（P<0.05）,while the promoter methylation level of SorCS1 gene was higher in colon adenocarcinoma,rectum adenocarcinoma and lung adenocarcinoma than that in healthy controls（P<0.05）. The difference of CASR gene expression between colorectal cancer patients and healthy controls was not statistically significant（P>0.05）,and the data of CASR gene promoter methylation was not available in TCGA database. The differences between ΔSorCS1 and SEPT9 gene promoter methylation levels in colorectal cancer patients were associated with tumor stage,and those in progression stage（Ⅲ-Ⅳ） were higher than those in early stage（Ⅰ-Ⅱ）（P<0.05）. The gene promoter methylation differences were also associated with lymph node metastasis,and the higher level of ΔSorCS1 and SEPT9 gene promoter methylation levels appeared in the group of lymph nodes with metastasis than those without metastasis（P<0.05）,independent of sex,age,tumor site and maximum diameter of tumors（P>0.05）. However,there was no statistical significance in ΔCASR between different clinicopathological characteristics of patients（P>0.05）. The positive rate of SorCS1 gene methylation was 82.35%（14/17）,while that of SEPT9 gene methylation was 70.58%（12/17）,respectively. The positive rate of combined determination was 88.24%（15/17）. Conclusions SorCS1 gene promoter methylation determination has a high positive rate in colorectal cancer patients,and it is complementary to SEPT9 gene promoter methylation,which can be a potential new molecular marker for colorectal cancer development.
Objective To investigate the roles of serum human epidermal growth factor receptor-2（HER-2）and cluster of differentiation 44（CD44） in patients with colorectal cancer. Methods The levels of serum HER-2 and CD44 were determined by enzyme-linked immunosorbent assay in 38 colorectal cancer patients before and 2-3 d after operation,35 patients with benign colorectal diseases（benign disease group） and 36 healthy subjects（healthy control group）. The general data of all the subjects were collected. The correlation between serum HER-2 and CD44 levels in colorectal cancer patients was analyzed by Pearson correlation analysis. The efficiencies of each marker and combined determination in the auxiliary diagnosis of colorectal cancer were evaluated by receiver operating characteristic（ROC） curve. Results The preoperative serum HER-2 levels in colorectal cancer group were higher than those in benign colorectal disease and healthy control groups（P<0.05,P<0.000 1）. The preoperative serum CD44 levels in colorectal cancer group were higher than those in healthy control group（P<0.05）. Serum CD44 levels between colorectal cancer and benign colorectal disease group had no statistical significance（P>0.05）. The levels of serum HER-2 in colorectal cancer group were closely related to Ras mutation and peripheral nerve invasion（P<0.05）,and they were not related to serum CD44 levels（P>0.05）. Serum HER-2 and CD44 levels were not related to age,sex,tumor type,tumor size,TNM stage,lymph node metastasis and vascular invasion（P>0.05）. According to the results of ROC curve analysis,the areas under curves（AUC） of HER-2,CD44,carcinoembryonic antigen（CEA）,carbohydrate antigen 19-9（CA19-9） determinations and the combined determination in colorectal cancer diagnosis were 0.779,0.692,0.620,0.634 and 0.837,respectively. Postoperative levels of serum CD44 were reduced compared to preoperative levels in colorectal cancer group（P<0.000 1）,whereas no statistical significance was observed in serum HER-2 levels before and after operation（P>0.05）. Conclusions Serum HER-2 and CD44 levels are correlated in colorectal cancer patients. The combined determination of them may play a role in colorectal cancer auxiliary diagnosis and therapeutic monitoring.
Objective To investigate the role of mitogen-activated protein kinase （MAPK）/extracellular regulated protein kinase （ERK） signaling pathway in reversing endocrine resistance of breast cancer. Methods Tamoxifen-resistant breast cancer cell lines,MCF-7/TAMR and T47D/TAMR,were established by human breast cancer cell lines,MCF-7 and T47D. The differences of cell proliferation and the expression of related MAPK/ERK signaling pathway molecules between MCF-7 and MCF-7/TAMR,T47D and T47D/TAMR were investigated. The suppressive effect of mitogen-activated protein kinase（MEK） inhibitor U0126 on MAPK/ERK signaling pathway in MCF-7/TAMR and T47D/TAMR was observed through the analysis of cell proliferation,cell cycle and transfactor expression. Results Compared with naïve MCF-7 or T47D cells,the proliferation and colony formation were increased （P<0.05,P<0.000 1）,and the expression of related MAPK/ERK signaling pathway molecules [phosphorylated extracellular regulated protein kinase（pERK）,ERK and c-MYC] was strongly increased. Suppressing MAPK/ERK signaling pathway by MEK inhibitor U0126,the growth of MCF7/TAMR or T47D/TAMR was decreased （P<0.000 1）,and cell cycle was arrested. The relative gene expression of transcription factors,steroid receptor coactivator-1（SRC-1）,E26 oncogene homolog-2（ETS-2） and c-JUN,and the expressions of their proteins were decreased by MEK inhibitor U0126（P<0.01）. Conclusions The inhibition of MAPK/ERK signaling pathway could reverse the endocrine resistance of breast cancer cells,which can provide a reference for the treatment of patients with endocrine resistant breast cancer.
Objective To investigate the interaction between interleukin-8（IL-8） and microRNA-182（miR-182） in non-small cell lung cancer（NSCLC） and the mechanism of promoting the proliferation and migration of NSCLC. Methods A total of 40 patients with NSCLC（NSCLC group） were enrolled to collect preoperative serum samples and intraoperative cancer tissues and adjacent tissues. Another 40 healthy subjects（healthy control group） were enrolled as well. Serum IL-8 level and the expression level of miR-182 in cancer and adjacent tissues were determined. Cytological experiments were performed to analyze the effects of miR-182 and IL-8 on proliferation and migration of human NSCLC cell line A549,the regulatory effect of IL-8 on miR-182 and the signaling pathway of miR-182 affecting IL-8 expression. Results Serum IL-8 level in NSCLC group was higher than that in healthy control group（P<0.01）. The relative expression of miR-182 in cancer tissues of NSCLC patients was higher than that in adjacent tissues（P<0.01）. IL-8 promoted the proliferation and migration of A549 cells through miR-182. IL-8 up-regulated the expression of miR-182 through signal transducer and activator of transcription 3（STAT3）. MiR-182 up-regulated IL-8 expression by regulating nuclear factor-κB（NF-κB） signaling pathway. Conclusions Both IL-8 and miR-182 are highly expressed in NSCLC patients,and the positive feedback regulatory axis of IL-8/STAT3/miR-182/NF-κB/IL-8 promotes the occurrence and development of NSCLC.
Objective To investigate the application value of next generation sequencing technology in preimplantation genetic testing（PGT） of autosomal recessive polycystic kidney disease（ARPKD） family. Methods A case of ARPKD family was selected,and the mutation of polycystic kidney and hepatic disease 1（PKHD1） gene in family members was investigated by Sanger sequencing. The coding region of PKHD1 gene was selected as target region,120 high-density closely linked single nucleotide polymorphisms（SNP） were selected as the genetic linkage markers in the upstream and downstream 2 M regions of the gene. The SNP haplotypes of family members were constructed by selecting effective SNP sites by multiplex polymerase chain reaction（PCR） and next generation sequencing,and the risk chromosomes of gene mutation were determined. After the whole genome amplification of the trophoblast cells obtained by blastocyst biopsy,the mutation sites of embryonic PKHD1 gene were sequenced directly,and the embryonic SNP haplotypes were constructed for linkage analysis by next generation sequencing. Sanger sequencing was used to verify the next generation sequencing results of embryonic PKHD1 gene mutation sites. Low depth chromosome aneuploidy screening was carried out both in the normal and the heterozygous embryos. Results The father of the family members carried PKHD1 gene c.5935G>A,which was heterozygous. The mother carries PKHD1 gene c.10058T>G,which was heterozygous. The proband carried double heterozygous mutations of PKHD1 gene c.5935G>A and c.10058T> G. Among the 5 embryos used for biopsy,2 cases were undetected mutations,and 3 cases carried heterozygous mutations. Low depth chromosome aneuploidy screening showed that the 3 of 5 embryos were euploidy,and 2 embryos were aneuploidy. A healthy baby was delivered at full term after a well-developed euploid embryo with no mutation,which was implanted into the mother's uterus. Conclusions The application of next generation sequencing in PGT of ARPKD can block the risk of recurrence of this single gene disease in the family,and avoid the abortion caused by the selection of aneuploid embryos.
Objective To investigate the application of morphological analysis of circulating tumor cells（CTC） in tumor clinical examination. Methods Peripheral blood samples of 212 patients with tumors were collected and analyzed by CTCBIOPSY detection system. The morphological analysis of CTC was performed by microscopy. The clinical applicability of morphological criteria was further verified by immunohistochemistry with CD45 and CD31 antibodies. The relationship between CTC count and detection rate and clinical factors,such as tumor type and clinical stage,was analyzed. Results The detection rate of CTC was 41.51%,and the specificity was 100%. Comparing the CTC-positive group with the CTC-negative group,there was statistical significance in age（P=0.036） and TNM stage（P<0.001）. The detection rate of CTC and CTC count were different in different types of tumors and different stages of the same type of tumor. The detection rate of CTC was the highest in patients with advanced lung cancer and prostate cancer,and the difference of CTC count in patients with early and advanced tumors had statistical significance（P<0.001）. Conclusions CTC morphological analysis can preserve the integrity of cells,and it can judge rapidly the CTC of various tumors,with good specificity and general sensitivity.
Objective To investigate the interference effect of hemolysis,icterus and lipemia on the determination of prealbumin（PA） by immunoscattering turbidimetry. Methods A fresh serum sample was used as a basic sample,interfering substances [hemoglobin（Hb）,bilirubin and lipid] were added,and the PA level was determined by immunoscattering turbidimetry. According to the EP07-A3 document of the Clinical and Laboratory Standards Institute（CLSI）,the paired difference experiment and dose-effect experiment were performed. Results PA had no interference with 10 g/L Hb（hemolysis index of 1 000）and 200 mg/L（340 μmol/L） bilirubin（icterus index of 20）. The lipid turbidity interfering substance with a lipid turbidity index of 987 had a negative interference on the determination of PA. The degree of interference had a linear relationship with the lipid turbidity index. For samples with low values of PA,when the lipid turbidity index≤120,there was no interference to PA determination. The linear equation between lipid turbidity index and interference rate was Y=-0.077X+0.979（r 2=0.999）. For samples with high values of PA,when the lipid turbidity index≤203,there was no interference to PA determination. The linear equation between lipid turbidity index and interference rate was Y=-0.040X-0.168（r2=0.992）. Conclusions Generally clinical hemolytic and icteric samples will not interfere with the PA determination results by immunoscattering turbidimetry. The samples with lipid turbidity index>120 will cause negative interference to PA determination results.
Objective To investigate the relationship between procalcitonin（PCT） level and all-cause death in patients undergoing maintenance hemodialysis（MHD）. Methods A total of 160 patients undergoing MHD were enrolled,and their clinical data,including sex,age,body mass index（BMI）,smoking history,primary disease,comorbidities,dialysis duration,New York Heart Association（NYHA） cardiac function classification and laboratory indicators,were collected. Receiver operating characteristic（ROC） curve was used to determine the optimal cut-off value of death and survival in MHD patients. The patients with PCT> optimal cut-off value were taken as high PCT group,and those with PCT≤optimal cut-off value were taken as low PCT group. The influencing factors of high PCT were analyzed by binary Logistic regression. Kaplan-Meier survival curve was used to analyze the survival rate of MHD patients. Cox proportional risk regression analysis was used to evaluate the relationship between PCT and all-cause death in MHD patients. Results The dialysis duration and CRP levels in high PCT group were higher than those in low PCT group（P<0.01）,but there was no statistical significance for the other indicators（P>0.05）. Binary Logistic regression analysis showed that increased CRP level was a risk factor for high PCT [odds ratio（OR）=1.182,95% confidence interval（CI）1.043-1.339,P=0.009]. Kaplan-Meier survival curve analysis showed that the survival rate of high PCT group was lower than that of low PCT group（Log-rank χ2=6.707,P=0.01;Breslow test value was 6.828,P=0.009）. Cox proportional risk regression analysis showed that high triglyceride（TG） level was a protective factor for all-cause death in MHD patients [hazard ratio（HR）=0.166,95%CI 0.071-0.387]. High PCT level was a risk factor for all-cause death in MHD patients（HR=4.409,95%CI 1.757-11.064）. Conclusions High PCT level may be an independent risk factor for all-cause death in MHD patients.
Objective To evaluate the feasibility of ampicillin to predict the imipenem susceptibilities of Enterococcus faecalis and Enterococcus faecium. Methods Totally,127 non-duplicated isolates of Enterococcus faecalis and 124 isolates of Enterococcus faecium were collected from 23 hospitals. The susceptibilities of Enterococcus faecalis and Enterococcus faecium to penicillin,ampicillin and imipenem were determined by broth microdilution and disk diffusion methods. Results For Enterococcus faecalis,which were both sensitive or resistant to penicillin and ampicillin,the categorical agreement（CA） of ampicillin-imipenem by broth microdilution was 100.0%,and there was no very major error（VME） and major error（ME）. For Enterococcus faecalis,which were resistant to both penicillin and ampicillin,the CA of ampicillin-imipenem by disk diffusion was 77.8%,and the ME accounted for 22.2%. For penicillin-resistant but ampicillin-sensitive Enterococcus faecalis,the CA of ampicillin-imipenem were 57.1% and 81.8% by broth microdilution and disk diffusion. Conclusions Ampicillin is better than penicillin to predict the susceptibilities of Enterococcus faecalis and Enterococcus faecium to imipenem. The susceptibilities to ampicillin can not be used to predict the susceptibilities of penicillin-resistant but ampicillin-sensitive Enterococcus faecalis and Enterococcus faecium to imipenem. Enterococcus faecalis and Enterococcus faecium isolates being sensitive to penicillin are predictably susceptible to ampicillin.
Objective To establish and validate a method for the simultaneous determination of testosterone（T）,dihydrotestosterone（DHT）,androstenedione（AD）,dehydroepiandrosterone（DHEA） and 17alpha-hydroxyprogesterone（17OHP） in human serum by ultra-performance liquid chromatography-tandem mass spectrometry（UPLC-MS/MS）,and to establish the reference intervals of 5 steroid hormones in healthy adults. Methods The serum samples were purified by SOLAμ HRP solid-phase extraction,and they were separated by Poroshell 120 EC-C18 analytical column. Mobile phase was constituted of 30% acetonitrile solution and 90% acetonitrile solution containing 0.05% formic acid with gradient elution. The electrospray positive ion multi-reaction monitoring mode was used for determination,and the isotope internal standard method was used to quantify the concentration of steroid hormones. The performance of the method was verified comprehensively,and the reference intervals suitable for the local population in Shanghai were established. Results The minimum limits of quantification of the 5 steroid hormones in human serum were 0.01-0.10 ng/mL. The standard curve range covered 1 000 times （r2≥0.999 5） and showed good linearity,and the sample after dilution can be reported to cover the range of 5 000 times. The average accuracy of 5 steroid hormones ranged from 95.0% to 105.6%,with the precision of <8%. None of the 8 structural analogs interfered with the target analytes,high triglyceride blood did not affect the determination results,and hemolysis resulted in low DHEA concentration. The extraction recoveries of the analytes were 60.5%-93.3%,and there was no obvious matrix effect after calibration of internal standard. Analytes demonstrated good stability at room temperature for 4 h,at -25--15 ℃ for 30 d or at -90--70 ℃ for 100 d. The method was used and participated in a survey of endocrine accuracy verification supplied by the National Center for Clinical Laboratories in 2019. The determination deviations of T and 17OHP were -5.00%-0.38%. The results of 367 physical examination subjects were analyzed and showed that there was statistical significance in the concentration of 5 steroid hormones between females and males（P<0.05）. There was statistical significance between the 2 age subgroups（aged 16-50 years and aged>50 years） of female groups（P<0.05）. Conclusions A UPLC-MS/MS has been established for the determination of 5 steroid hormones in human serum,and the reference intervals suitable for the Central Laboratory of Shanghai Xuhui Central Hospital have been verified.
Objective To investigate plasma exosome-derived circular RNA（exo-circRNA） with diagnostic potential for hepatocellular carcinoma（HCC）. Methods A total of 256 patients with HCC who received curative resection（25 cases belonged to discovery cohort,126 cases belonged to training cohort and 105 cases belonged to validation cohort）,100 patients with liver cirrhosis（50 cases belonged to training cohort and 50 cases belonged to validation cohort） and 125 healthy subjects（healthy control group,25 individuals belonged to discovery cohort,50 individuals belonged to training cohort and 50 individuals belonged to validation cohort）were enrolled. The plasma exosome was isolated by precipitation,and the expression level of exo-circRNA was determined by real-time quantitative polymerase chain reaction（RT-qPCR）. The new exo-circRNA model for HCC diagnosis was constructed using Logistic regression analysis,and the diagnostic performance of this exo-circRNA model was evaluated by receiver operating characteristic（ROC） curve. Results Compared with normal liver cell line L02,102 exo-circRNA expression in liver cancer cell line Hep3B were increased（P<0.05）. After conducting integrative analysis with exoRBase database,4 circRNA with increased expression in HCC patients were identified（circ_0000690,circ_0001359,circ_0000396 and circ_0000722）. The expressions of circ_0000690,circ_0001359 and circ_0000396 were increased in HCC group compared with those in healthy control group（P<0.05）. There was no statistical significance in the expression of circ_0000722 between the 2 groups（P>0.05）. The expression of circ_0000690 was higher in HCC group compared to those in liver cirrhosis group and healthy control group in both training and validation cohorts（P<0.001）. The expression of circ_0001359 showed escalating pattern in healthy control group,liver cirrhosis group and HCC group（P<0.05）. There was no statistical significance in the expression of circ_0000396 between HCC group and liver cirrhosis group（P>0.05）,but they were still higher than those in healthy control group（P<0.05）. The established diagnostic model was Logit（P）=1.110×circ_0000690+0.822×circ_0001359+0.622×circ_0000396-4.153. ROC curve analysis results showed that,in the training cohort,the areas under curves（AUC）for diagnosing HCC of circ_ 0000690,circ_ 0001359,circ_0000396 and integrated model were 0.802,0.726,0.621 and 0.859,respectively. Meanwhile,the AUC for diagnosing early HCC（defined as Barcelona stage 0+A） of circ_ 0000690,circ_ 0001359,circ_0000396 and integrated model were 0.767,0.698,0.611 and 0.847,respectively. The AUC for diagnosing alpha-fetoprotein（AFP）-negative（<20 ng/mL） HCC were 0.810,0.695,0.588 and 0.894,respectively. In the validation cohort,the AUC for diagnosing HCC of circ_ 0000690,circ_0001359,circ_0000396 and integrated model were 0.752,0.663,0.615 and 0.847,respectively. The AUC for diagnosing early HCC of circ_ 0000690,circ_ 0001359,circ_0000396 and integrated model were 0.763,0.673,0.591 and 0.845,respectively. Moreover,the AUC for diagnosing AFP-negative HCC of circ_ 0000690,circ_ 0001359,circ_0000396 and integrated model were 0.702,0.670,0.641 and 0.840,respectively. Conclusions The plasma exo-circRNA model established in this study has potential diagnostic value for HCC,especially AFP-negative and early HCC.
Objective To evaluate the performance of a method for the detection of hepatitis B virus（HBV） RNA,to compare serum HBV RNA levels in patients with different results of serological markers of HBV infection,and to evaluate the relationship between HBV RNA and HBV DNA. Methods The performance of the method for HBV RNA detection was evaluated,which included precision,sensitivity and linearity. The sera of 81 patients with hepatitis B surface antigen（HBsAg）-positive chronic hepatitis B were collected,and the levels of HBV RNA,HBV DNA and serological markers were detected simultaneously. The patients were classified according to the different results of hepatitis B e antigen（HBeAg）,serological markers of HBV infection and medication. Results The intra-assay precisions of high and low HBV RNA values were 0.56% and 2.30%,and the inter-assay precisions were 3.13% and 6.03%,respectively,which all met the clinical requirements. The sensitivity of the assay was achieved by repeating the assay 20 times for samples close to the lower limit of detection stated by the manufacturer,and the detection rate was 100%,which met the clinical requirement. It was linear in the range of HBV RNA 1.0×102-1.0×107 copies/mL. Serum HBV RNA and HBV DNA loads in HBeAg-positive group were higher than those in HBeAg-negative group（P<0.01,P<0.05）,and the HBeAg-positive and hepatitis B e antibody（HBeAb）-negative group had the highest HBV RNA loads among all the groups with different results of serological markers of HBV infection（P<0.01）. Serum HBV DNA and HBV RNA loads were higher in untreated group than those in treated group（P<0.01,P<0.05）. In 81 patients with chronic hepatitis B,HBV RNA loads were positively correlated with HBV DNA load and HBeAg positive titer（r=0.348 and 0.544,respectively,P<0.05） and without HBsAg titer（r=0.04,P>0.05）. In HBeAg-positive patients with chronic hepatitis B,serum HBV RNA loads were positively correlated with serum HBV DNA load（r=0.338,P<0.05）. In HBeAg-negative patients with chronic hepatitis B,serum HBV RNA levels did not correlate with serum HBV DNA load（r=0.14,P>0.05）. Conclusions Serum HBV RNA level is a potential new marker for monitoring the viral activity and therapeutic efficacy of HBV infection.
Clustered regularly interspaced short palindromic repeats（CRISPR）-CRISPR-associated protein（Cas） system is a powerful tool for gene editing. Compared with Cas9,Cas13a targeted multiple gene transcription products and regulated gene function expression,filling Cas9 with defects such as editing and missing effect only limited to DNA level. Moreover,using the characteristics of Cas13a targeting RNA,the system has been successfully transformed into the next generation of nucleic acid diagnostic tools. In this review,the latest research progress of CRISPR-Cas13 system in the field of gene editing and molecular diagnosis is summarized,and the application prospect of this system has prospected.
C-reactive protein（CRP）,which can be determined by serum as well as whole blood,has been traditionally utilized as a marker of infection. Many brands of whole blood CRP determination equipments have been widely used in medical institutions,especially in women's and children's hospitals. However,the standard for the performance evaluation of whole blood CRP determination system is not established. According to the data of 26 women's and children's hospitals,this consensus aims to provide the performance status of whole blood CRP determination system,and further put forward the performance evaluation requirements of whole blood CRP determination system in combination with clinical needs,biological variation and industry standards,so as to ensure the determination quality.
Objective To investigate the optimal cut-off values of mean corpuscular volume（MCV） and mean corpuscular hemoglobin（MCH） for screening thalassemia in adults in Dehong,Yunnan. Methods Totally,9 562 adults undergoing outpatient examination,physical examination,premarital examination and genetic counseling in the People's Hospital of Dehong Dai and Jingpo Autonomous Prefecture were enrolled,and they were classified into thalassemia group（2 662 cases） and healthy control group（6 900 cases） through the determination of common gene mutations of thalassemia. Receiver operating characteristic（ROC） curve was drawn based on the MCV and MCH results obtained from routine blood test in order to determine the optimal screening cut-off values and efficiencies. Results The optimal cut-off value of MCV was 83.95 fL,the screening sensitivity was 82.7%,and the specificity was 89.7%. The optimal cut-off value of MCH was 28.15 pg,the screening sensitivity was 88.7%,and the specificity was 89.2%. Compared with the cut-off values widely used in China（MCV<80 fL and MCH<27 pg）,the screening cut-off values in this study may reduce the missed diagnosis rate,and the missed diagnosis rate of screening with MCV and MCH combinedly was the lowest. Conclusions Epidemic areas with unique mutation profiles may establish their screening cut-off values,and the combined screening of MCV or MCH may reduce the missed diagnosis rate and improve screening efficiency.
Objective To compare and evaluate the ability to detect pathogens between blood culture bottles of new antibiotic adsorption materials and those of activated carbon adsorption materials. Methods According to the common bacterial distribution of Sun Yat-sen University Cancer Center,18 selected isolates（clinical isolates and ATCC standard isolates） were chosen to simulate patients' samples with non-antibiotic treatment and antibiotic treatment including vancomycin,daptomycin,imipenem and piperacillin-tazobactam. Those samples were added into blood culture bottles of 2 different types of antibiotic adsorption materials to evaluate their positive rates and detection times in BacT/ALERT system. Results Blood cultures of new antibiotic adsorption material and activated carbon adsorption material showed 100% recovery on 18 pathogens,like Gram-positive and Gram-negative bacteria and yeast,except for Staphylococcus haemolyticus. The time to detection of 13 species of pathogens was faster in new antibiotic adsorption material when compared with activitied carbon adsorption material（P<0.05）. The average time to detection in new antibiotic adsorption aerobic material was 0.72-68.24 h,which was faster than that in activated carbon adsorption aerobic material,while the time to detection of new antibiotic adsorption anaerobic material showed 0.48-14.48 h shorter than that of activated carbon adsorption anaerobic material in detecting pathogens. When specific antibiotic added,both new antibiotic adsorption materials showed 100% recovery to 3 pathogens,while the recovery in activated carbon adsorption materials was 33%. The average time to detection was ranged from 12.72 h to 18.08 h in new antibiotic adsorption aerobic material and from 11.76 h to 28.16 h in new antibiotic adsorption anaerobic material. Conclusions In the absence of antibiotic interference,blood cultures with different antibiotic adsorption materials show the same positive rate of target pathogen,and those of new antibiotic adsorption material have a shorter time to detection. In the presence of antibiotic interference,compared with the activated carbon adsorption material,the blood culture bottles of the new antibiotic adsorption material have a higher detection ability of pathogens.
Low-density lipoprotein receptor（LDLR） is one type of cell surface glycoprotein,which plays a role in maintaining cholesterol metabolism balance in vivo. LDLR gene mutation can cause the decreasing,even the absence of LDLR on the cell surface,reduce the body's ability to metabolize cholesterol,lead to elevated plasma low-density lipoprotein cholesterol（LDL-C） level and deposition in different tissues and organs,and cause a variety of lipid metabolism disorders. Accurate and rapid determination of LDLR gene mutations and analysis of changes in LDLR function are essential for the diagnosis and differential diagnosis of familial hypercholesterolemia（FH）. This review focuses on the progress in the methodology and clinical application of LDLR gene mutation and function determination,in order to raise awareness and concern of its importance in early diagnosis and intervention of FH patients.
Objective To investigate the clinical value of whole blood zinc in children and adolescents with neurally-mediated syncope（NMS）. Methods Totally,102 NMS children（NMS group） who underwent head-up tilt test（HUTT） and tested trace elements and vitamins（Vit） were enrolled. According to the results of HUTT,83 cases were classified into HUTT positive group,and 19 cases were classified into HUTT negative group. The HUTT positive group included 32 cases of vasovagal syncope（VVS） and 53 cases of postural orthostatic tachycardia syndrome（POTS）. Two patients were diagnosed with both VVS and POTS. A total of 100 healthy age-matched children who tested trace elements and 100 healthy age-matched children who tested Vit were enrolled as control group A and control group B,respectively. The levels of zinc,calcium,copper,magnesium and iron in whole blood and the levels of Vit A,Vit B1,Vit B2,Vit B6,Vit B9,Vit C,Vit D and Vit E in serum were determined. Binary Logistic regression analysis was used to assess risk factors of NMS. Receiver operating characteristic（ROC） curve was used to evaluate the predictive value in diagnosing NMS and the predictive value in differentiating VVS and POTS. Results Compared with control group A,the differences of zinc,calcium and iron had statistical significance in NMS group（P<0.05）. There was no statistical significance in the other items between NMS group and control groups（P>0.05）. The levels of zinc,calcium and iron in HUTT positive group were lower than those in control group A（P<0.05）. There was no statistical significance in trace element levels between HUTT negative group and HUTT positive group,control group A（P>0.05）. The level of zinc in VVS group was lower than that in control group A（P<0.05）. The levels of zinc,calcium,magnesium and iron in POTS group were lower than those in control group A（P<0.05）. The level of magnesium in VVS group was higher than that in POTS group（P<0.05）. Binary Logistic regression analysis showed that the decrease of zinc [odds ratio（OR）=0.726,95% confidence interval（CI） 0.652-0.807] increased the risk of NMS,and the decrease of calcium（OR=1.772,95%CI 0.639-4.909） and iron（OR=0.472,95%CI 0.090-2.476） had no statistical significance on the occurrence of NMS. When the cut-off value of zinc was 80.09 μmol/L,the area under curve（AUC）was 0.800,and the sensitivity and specificity for the diagnosis of NMS were 74.5% and 80.0%,respectively. When the cut-off value of magnesium was 1.33 mmol/L,the AUC was 0.631,and the sensitivity and specificity for the differential diagnosis between VVS and POTS were 67.9% and 59.4%,respectively. Conclusions Zinc deficiency may be related to children and adolescents with NMS. Magnesium may have a clinical value in the differential diagnosis of VVS and POTS.
Recently,hypervirulent Klebsiella pneumoniae has emerged as a concerning pathogen. In contrast to nosocomial infections in immuno-compromised patients with classic Klebsiella pneumoniae,hypervirulent Klebsiella pneumoniae usually causes community acquired infections in healthy individuals and is prone to metastasize to distant sites,including most commonly the eye,lung and central nervous system,and often requiring source control. The genetic determinants of hypervirulent Klebsiella pneumoniae are often found on large virulence plasmids as well as chromosomal mobile genetic elements which can be used as biomarkers to distinguish hypervirulent Klebsiella pneumoniae from common Klebsiella pneumoniae clinical isolates. These distinct virulence determinants of hypervirulent Klebsiella pneumoniae include up to 4 siderophore systems for iron acquisition,increased capsule production,K1 and K2 capsule types and colibactin toxin. Alarmingly,multidrug-resistant hypervirulent isolates have emerged,creating a new challenge in combating this already dangerous pathogen. In order to better understand hypervirulent Klebsiella pneumoniae,the characteristics,epidemiology,virulence related factors and drug resistance with hypervirulent Klebsiella pneumoniae will be introduced in detail.
Clinical laboratory examinations for parasitic diseases play an important role in the diagnosis of parasitic diseases. Clinical laboratory examinations for parasitic diseases include pathogenic examinations,immunological examinations and molecular biological examinations. Although the traditional parasitological examination is intuitive and accurate,it has some limitations. With the rapid development of medical technologies,parasite laboratory examination technology has also made great progress. Immunological examinations and molecular biological examinations have good sensitivities,specificities and repeatabilities. This review summarizes and introduces clinical laboratory examination technologies for parasitic diseases.
Objective To analyze the clinical symptom and laboratory examinations of a case of green neutrophil and monocyte inclusions with malaria pigment retrospectively,and to improve the understanding of this case. Methods Through the clinical and pathologic findings,including complete blood counts,C-reactive protein,biochemical examination,malaria parasite antigen test,malaria parasite morphological examination and oil red O staining,the clinical symptom and laboratory examinations' characteristics were analyzed combined with literatures. Results The patient presented with high fever and joint pain as the initial clinical symptoms. Platelet count was 12×109/L,the percentage of neutrophil was 82.7%,and C-reactive protein was >200 mg/L,accompanied by liver and renal dysfunction. Plasmodium falciparum was detected on peripheral blood smear. Green inclusions and malaria pigment were found in neutrophils and monocytes at the later stage of treatment,and the oil red O staining of green inclusions was positive. Conclusions The presence of green inclusions and malaria pigment is an important indication for the prognosis of severe malaria,which should be warned to clinicians.
China has made remarkable achievements in the prevention and control of human parasitic diseases,becoming the first country in the elimination of lymphatic filariasis. Malaria has been eliminated in China,which is approved by World Health Organization in 2021. Schistosomiasis in many provinces reaches the standard of transmission interruption or elimination. The infection rate of soil-transmitted nematode drops to a new historical low. However,the population of its infection is still tens of millions. Foodborne parasitic diseases,such as clonorchiasis sinensis,still pose a great threat to food safety in China. Although the vector-borne and zoonotic parasitic diseases have been controlled to some extent,the epidemic continues and relapses. The incidence of opportunistic parasitic diseases has increased in the population with immunodeficiency,and the cases of imported parasitic diseases have also increased. The review focuses on the results of parasitic diseases' prevention and control in China,understanding the epidemic status of important human parasitic diseases,analyzing the transmission risks and challenges of human parasitic diseases' prevention and control,so as to provide a reference for the health administrative department formulating the strategies and measures of parasitic diseases' prevention and control in the new period,and to provide epidemiological datum reference for the diagnosis and treatment of clinical parasitic diseases.
