Objective:To construct a recombinant bioluminescent bacteriophage (HT7) targeting Escherichia coli, and evaluate its ability to identify Escherichia coli. Methods:Initially, pCRISPR-sg (1-10) and PFN-1000 plasmid strains were constructed by genetic engineering, and the most efficient small guild RNA (sgRNA) were screened by bilayer plate. By the gene editing technique, which comprised homologous recombination and clustered regularly interspaced short palin dromic repeats (CRISPR)-Cas system, the Nanoluc luciferase gene was integrated into the downstream non-coding region of 10A gene of T7 phage, to constructe the bioluminescent phage HT7 successfully. The difference of biological characteristics between HT7 phage and T7 phage was evaluated by plaque assay and liquid amplification assay. In addition, 51 strains of Escherichia coli, 20 strains of Klebsiella pneumoniae, 14 strains of Staphylococcus aureus, 6 strains of Enterococcus faecium, 5 strains of Enterococcus faecalis, 3 strains of Acinetobacter baumannii and 1 strain of Pseudomonas aeruginosa were collected and isolated to evaluate the limit of detection and specificity of HT7 phage. Results:Among the 10 CRISPR-targeted cleavage systems constructed, sgRNA8 exhibited the highest cleavage efficiency, with a cleavage rate of 0.18. After three rounds of recombination screening using the pCas9/pCRISPR/PFN-1000 triple-plasmid system, PCR validation yielded recombinant phage bands at 2 798 bp, indicating the successful construction of the HT7 phage. The recombinant phage showed significant differences in biological characteristics in terms of lysis efficiency ( P<0.001), one-step growth curve ( P=0.001), and infection multiplicity ( P=0.031). Both lysis burst time and log growth node were extended by 10 min, with the optimal infection multiplicity being 0.1. Clinical sample testing identified lysis of 6 strains of Escherichia coli within 4.5 h, while other strains remained unaffected, with detection of pathogenic bacteria below 10 CFU/ml. Conclusions:The developed pCas9/pCRISPR/PFN-1000 triple-plasmid editing system efficiently edits the bacteriophage genome. The constructed HT7 fluorescent bacteriophage enables the detection of Escherichia coli below 10 CFU/ml within 4.5 hours, demonstrating low detection limits and high detection specificity.

Objective:To explore the clinical value of synovial fluid calprotectin for the diagnosis of periprosthetic joint infection (PJI).Methods:Based on prospective cohort study design, a total of 82 patients suspected of PJI after hip and knee arthroplasty in the First Medical Center of the PLA General Hospital from July 2021 to June 2022 were selected. Patients were divided into infection group (PJI, n=39) and non-infection group (non-PJI, n=43) according to the diagnostic criteria proposed by the Second International Consensus Conference in 2018. The matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was used for double-blind detection of calprotectin and internal reference standard (IRS) in synovial fluid of patients. The peaks of target protein and IRS were recorded for further analysis. Mann-Whitney U test was used to compare the concentrations of S100A8 and S100A9 between the two groups, and receiver operating characteristic curve (ROC) was used to analyze the diagnostic efficacy of S100A8 and S100A9 for PJI. Results:Calprotectin was detected as monomers S100A8 and S100A9. Synovial fluid S100A8 was significantly higher in the PJI group than that in the non-PJI group [1.57 (0.48, 4.17) vs 0.00 (0.00, 0.05), Z=?7.221, P<0.05]. Synovial fluid S100A9 was also significantly higher in the PJI group than that in the non-PJI group [0.74 (0.29, 1.70) vs 0.06 (0.00, 0.10), Z=?6.255, P<0.05]. When using S100A8 and S100A9 to diagnose PJI, the sensitivity were 97.4% and 87.2%, the specificity were 86.0% and 88.4%, and the area under the ROC were 0.964 (95% CI 0.929-0.998) and 0.902 (95% CI 0.924-0.996), respectively. Conclusion:The detection of synovial fluid S100A8 and S100A9 by MALDI-TOF MS can make a satisfactory diagnosis for PJI.

Objective:To investigate the clinical efficacy of proximal gastrectomy and total gastrectomy in the treatment of Siewert type Ⅱ and Ⅲ adenocarcinoma of esophagogastric junction (AEG).Methods:The retrospective cohort study was conducted. The clinicopathological data of 170 patients with Siewert type Ⅱ and Ⅲ AEG who were admitted to Guangdong Provincial People′s Hospital from January 2010 to December 2018 were collected. There were 125 males and 45 females, aged from 30 to 85 years, with a median age of 64 years. Of the 170 patients, 82 cases undergoing proximal gastrectomy were allocated into the proximal gastrectomy group and 88 cases undergoing total gastrectomy were allocated into the total gastrectomy group. Observation indica-tors: (1) surgical and postoperative situations; (2) follow-up and survival; (3) analysis of prognostic factors. Follow-up was conducted using telephone interview and outpatient examination to detect survival of patients up to December 2021. Measurement data with normal distribution were represented as Mean± SD, and comparison between groups was analyzed using the t test. Measure-ment data with skewed distribution were represented as M( Q1, Q3) or M(range), and comparison between groups was analyzed using the Mann-Whitney U test. Count data were described as absolute numbers or percentages, and comparison between groups was analyzed using the chi-square test or Fisher exact probability. Comparison of ordinal data was analyzed using the rank sum test. Kaplan-Meier method was used to draw survival curves, and Log-Rank test was used for survival analysis. COX proportional hazard model was used for univariate and multivariate analyses. Variables with P<0.1 in univariate analysis were included for multivariate analysis. Results:(1) Surgical and postoperative situations. Cases with surgical approach as transthoracic or thoraco-abdominal approach, transabdominal approach, the operation time, cases with volume of intra-operative blood loss ≤100 mL or >100 mL, cases with length of proximal margin ≤1.5 cm or >1.5 cm, cases with radical surgery outcome as R 0, R 1, R 2, the number of lymph nodes harvest, cases with anastomotic leakage, cases with anastomotic stricture, cases with incision infection, cases with pleural infection or effusion, cases with abdominal infection or ascites were 61, 21, (211±18)minutes, 46, 36, 44, 38, 73, 6, 3, 15(9,22), 5, 2, 2, 4, 2 in the proximal gastrectomy group, respec-tively. The above indicators were 12, 76, (263±15)minutes, 27, 61, 45, 43, 82, 4, 2, 23(18,32), 4, 1, 3, 1, 4 in the total gastrectomy group, respectively. There were significant differences in the surgical approach, operation time, volume of intraoperative blood loss and the number of lymph nodes harvest between the two groups ( χ2=63.94, t=-25.50, χ2=11.19, Z=-5.62, P<0.05). There was no significant difference in the length of proximal margin or radical surgery outcome between the two groups ( χ2=0.11, Z=-0.95, P>0.05) and there was no significant difference in the anastomotic leakage, anastomotic stricture, incision infection, pleural infection or effusion, abdominal infection or ascites between the two groups ( P>0.05). (2) Follow-up and survival. All the 170 patients were followed up for 89(64,106)months. Of the 170 patients, the 5-year overall survival rates were 43.8% and 35.5% of the Siewert type Ⅱ and Ⅲ AEG patients, respectively, showing no significant difference between them ( χ2=0.87, P>0.05). Of the patients with Siewert type Ⅱ AEG, the 5-year overall survival rates were 41.7% and 54.3% in the patients with proximal gastrectomy and the total gastrectomy, respectively, showing no significant difference between them ( χ2=1.05, P>0.05). Of the patients with Siewert type Ⅲ AEG, the 5-year overall survival rates were 31.3% and 37.5% in the patients with proximal gastrectomy and the total gastrectomy, respectively, showing no significant difference between them ( χ2=0.33, P>0.05). The 5-year overall survival rates were 39.0% and 44.2% in the proximal gastrectomy group and the total gastrectomy group, respectively, showing no significant difference between the two groups ( χ2=0.63, P>0.05). Of the patients in TNM stage Ⅰ, stage Ⅱ, stage Ⅲ, the 5-year overall survival rates were 65.3%, 36.3%, 27.1% in the proximal gastrectomy group, versus 83.3%, 48.0%, 39.7% in the total gastrectomy group, showing no signifi-cant difference between the two groups ( χ2=0.02, 1.50, 1.21, P>0.05). (3) Analysis of prognostic factors. Results of univariate analysis showed that pathological N staging, degree of tumor differen-tiation and radical surgery outcome were related factors influencing prognosis of AEG patients ( hazard ratio=1.71, 1.70, 2.85, 95% confidence interval as 1.16-2.60, 1.15-2.50, 1.58-5.14, P<0.05). Results of multivariate analysis showed that pathological N staging and radical surgery outcome were independent factors influencing prognosis of AEG patients ( hazard ratio=1.55, 2.18, 95% confidence interval as 1.05-2.31, 1.18-4.02, P<0.05). Conclusions:There is no significant difference in the prognosis of Siewert type Ⅱ and Ⅲ AEG patients undergoing proximal gastrectomy or total gastrectomy. Proximal gastrectomy can be used for the treatment of advanced Siewert type Ⅱ and Ⅲ AEG.

【Objective】 To compare the effect of different combinations of filter membranes on the removal of IgA by component Ⅱ solution. 【Methods】 The dissolved solution of component Ⅱ was filtered with different combinations of filter membranes, and the turbidity, IgA residue, molecular size distribution and purity were compared. 【Results】 After filtering through the combination of NA-16+ Millpore 0.5μ, NA-16+ EmphazeTMAEX HP, and ZetaPlus™ + Emphaze™AEX HP filter membranes, the turbidity of the filtrate decreased by 35.5%(22.4/63.1), 39.2%(24.8/63.3) and 59.8%(37.4/62.5), the IgA residue decreased by 5.8%(14.77/253.70), 20.0%(50.90/254.00) and 44.3%(112.13/253.33), respectively.Molecular size distribution was 99.60%, 99.85% and 100%, and the purity was 100%, 100% and 100%, respectively. 【Conclusion】 The combination of ZetaPlus™ + Emphaze™AEX HP filter membrane can not only effectively remove IgA in the dissolution solution of component Ⅱ with other quality indicators remaining high-level, which can be used in IVIG large-scale production process.

OBJECTIVE: To assess the effect of ergonomic factor load on work-related musculoskeletal disorders(WMSDs) in shipyard workers. METHODS: A total of 751 shipyard workers were selected as the research subjects using judgment sampling method. The exposure level of adverse ergonomic factors was assessed using the Quick Exposure Check method. The prevalence of WMSDs was investigated using the revised Musculoskeletal Disorders Investigating Questionnaire of our research group, and the relationship between them was analyzed. RESULTS: The proportions of high or very high level of work load exposure to the neck, back, shoulder and hand in shipyard workers from high to low were 66.4%, 63.5%, 59.8% and 43.7%(P<0.01) respectively. The proportions of occupational stress, driving, vibration and working rhythm at high or very high exposure level were 20.0%, 4.1%, 22.9% and 3.2%, respectively. The prevalence of WMSDs in four body parts of shipyard workers from high to low was the back, neck, hand and shoulder(the prevalence were 44.2%, 31.2%, 26.9% and 26.6%, respectively, P<0.01). After excluding the influence of confounding factors, multiple logistic regression analysis showed that the higher the labor load exposure level and longer of the vibration tool using, the higher the risk of shoulder WMSDs [odds ratio(OR) and 95% confidence interval(CI) were 1.25(1.04-1.51) and 1.33(1.05-1.69), respectively, P<0.05]. The higher the level of occupational stress, the higher the risk of back and neck WMSDs [OR(95%CI) was 1.29(1.05-1.58) and 1.42(1.15-1.77), respectively, P<0.05]. CONCLUSION: There was a dose-effect relationship between the exposure level of shoulder load, the time of using vibration tools and the shoulder WMSDs, and there was a dose-effect relationship between the occupational stress level and the WMSDs in the back and neck.

Objectives: Two cases who underwent fetal pulmonary valvuloplasty (FPV) for pulmonary atresia with intact ventricular septum (PA-IVS) or critical pulmonary stenosis with intact ventricular septum (CPS-IVS) successfully were reported. The aim of the report was to explore the criteria for case selection, the technical essentials of FPV, and the postpartum outcome of the fetus. Methods: One case with PA-IVS and the other with CPS-IVS were enrolled in September 2016 and February 2017 in Guangdong General Hospital, and both cases were diagnosed with severe right ventricular dysplasia and tricuspid regurgitation by fetal echocardiogram. Parameters of right ventricle development and hemodynamics from echocardiography included tricuspid/mitral annulus (TV/MV), right ventricle/left ventricle long-axis (RV/LV), pulmonary/aortic annulus (PV/AV), tricuspid inflow duration/cardiac cycle, degree of tricuspid regurgitation (TR), blood flow direction of arterial duct and ductus venosus. Multidisciplinary team including the maternal-fetal cardiology, pediatric cardiology, cardiac surgery, obstetrics, neonatology and anesthesiology was summoned to discuss the indications and timing of PFV. Two cases underwent ultrasound-guiding trans-abdominal PFV at the 28 weeks of gestational age. Echocardiography was performed to observe the opening and closing of the pulmonary valve, and to evaluate the development of right ventricle and improvement in hemodynamics every 2-4 weeks until delivery. Results: From the technical perspective, pulmonary balloon valvuloplasty was successfully performed in these two cases. The opening of pulmonary valve improved in these two cases at 2-4 weeks after FPV. However, an obvious restenosis was detected in the first case at 5-8 weeks after FPV. In the first case, the echocardiography parameters including TV/MV, RV/LV, PV/AV and tricuspid inflow duration/cardiac cycle increased from 0.56, 0.42, 0.85,0.26 to 0.59, 0.51, 0.87, 0.32 at 5-8 weeks after FPV, respectively. However, the direction of blood flow through the arterial duct was still reverse. In the second case, TV/MV, RV/LV, PV/AV and tricuspid inflow duration/cardiac cycle ratio increased from 0.70, 0.63, 0.91,0.35 to 0.80, 0.80, 0.97, 0.42 at 5-8 weeks after FPV, respectively. The direction of blood flow through the arterial duct changed to bidirectional. Both fetuses were born alive. The first case underwent pulmonary valve commissurotomy and modified Blalock-Taussig shunt on the 8^th day after delivery and received follow-up for 6 months. The strategy for the next-step therapy was still pending. The second case underwent transcutaneous pulmonary balloon valvuloplasty on the 19^th day after delivery and received follow-up for 3 months. The opening of pulmonary valve improved obviously and the cardiac function was normal in the second case. Conclusions FPV is safe and effective for fetus during the second and third trimester of pregnancy, and FPV is beneficial for the development of fetal ventricle, valve and large artery. In addition, FPV may help to avoid the postnatal surgery for isolated single ventricle, improve fetal heart failure and prevent fetal death.

Objective: To survey the conduction and evaluate the effectiveness of extracorporeal membrane oxygenation (ECMO) therapy in pediatric intensive care unit (PICU) in China mainland. Methods: In a questionnaire-based survey, we retrospectively reviewed the application of ECMO in children's hospital and general hospital in China mainland to summarize and analyze the categories of diseases and prognosis of children treated with ECMO therapy. Results: By December 31, 2017, a total of 23 hospitals using ECMO, including 22 tertiary referral hospitals and 1 secondary hospital, among which 16 were children′s hospitals and 7 were general hospitals. Thirty-seven ECMO equipment was available. A total of 518 patients treated with ECMO, within whom 323 (62.4%) successfully weaned from ECMO and 262 (50.6%) survived to discharge. Among 375 pediatric patients, 233 (62.1%) were successfully weaned from ECMO and 186 (49.6%) survived to discharge. Among 143 newborn patients, 90 (62.9%) successfully weaned from ECMO, 76 (53.1%) survived to discharge. ECMO was applied in veno-arterial (VA) mode to 501 (96.7%) patients, veno-venous (VV) mode to 14 (2.7%) patients, and VV-VA conversion mode to 3 (0.6%) patients. Sixty-nine patients required extracorporeal cardiopulmonary resuscitation (ECPR), including 20 newborn patients (29.0%) and 38 pediatric patients (71.0%), who were all with cardiovascular disease. Neonatal respiratory distress syndrome (26/61), persistent pulmonary hypertension of the newborn (PPHN) (12/61), and meconium aspiration syndrome (MAS) (11/61) are the most common pulmonary diseases in newborn patients; among whom, infants with PPHN had highest survival rate (10/12), followed by MAS (9/11). Among newborn patients with cardiovascular diseases, those who admitted were after surgery for congenital cardiac disease were the most common (54/82), while those with septic shock had the highest survival rate (2/3). In pediatric pulmonary diseases, acute respiratory distress syndrome was the most common (42/93), while plastic bronchitis was with the highest survival rate (4/4), followed by viral pneumonia (13/16). Among pediatric cardiovascular diseases, congenital cardiac defect was the most common (124/282), while fulminant myocarditis had the highest survival rate (54/77). Conclusion The application of ECMO as a rescue therapy for children with severe cardiopulmonary failure has dramatically developed in China mainland.

Objective To explore the clinical value of flow cytometry( FCM) and DNA automated cell image analyzer ( AICM) in determine the character of ascites and pleural effusion.Methods This was a cross-sectional study.203 ascites and pleural effusionsamples were random selected from PLA hospital inpatients between August 2013 to June 2014 .The DNA content of sediment cells were detectedthrough the FCM and AICM respectively benign and malignant disease were differentiated according the counts and proportion of aneuploid cells.The sensitivity, specificitywere calculated byROC curves.Results The sensitivity, specificity and accuracy of flow cytometry cell in detectingtumor cells were 78.6%,80.0% and 79.2%%, while the sensitivity, specificity and accuracy of image analyzer were 83.5%,78.6% and 81. 3%respectively.When FCM and AICMwere combined ,the sensitivity, specificity and accuracyincreased to 92.2%, 86.3% and 89.6%.Conclusions Compared toconventional cytology test, the sensitivity and specificity were significantly high when the two methods were combined .Therefore, the combination method can be used to assist in clinical identification of the nature of ascites and pleural effusion and to help the diagnosis of disease.

Objective To develop an up-converting phosphor technology-based lateral-flow ( UPT-LF) assay for rapid detection of Listeria monocytogenes, named LM -UPT-LF.Methods Monoclonal antibodies against p 60, which was the specific virulence factor of L.monocytogenes,were prepared and covalently conjugated with up-converting phosphor nanopar-ticles (UCP-NPs) as bio-label.Then, LM-UPT-LF was established with double-antibody sandwich mode-based LF assay. Detection performance , including sensitivity and specificity , was evaluated .Results The samples with absolute contamina-ted amount of L.monocytogenes cells <10, 10-99, and 100-1000 cfu could be significantly detected as positive after in-cubation at 20 h, 18 h, and 16 h, respectively.Other 13 kinds of food-borne pathogens with concentration of 109 cfu/ml did not caused any non-specific reaction .Conclusion The established LM-UPT-LF assay could detect L.monocytogenes with high sensitivity , specificity and simplicity and provides an alternative method for food safety control .

Objective To evaluate the association between interleukin-10(IL-10 )polymorphisms and endometriosis suscepti-bility by meta-analysis.Methods Computer retrieval Pubmed,Embase,Chinese HowNet,Chinese biomedical and Wanfang database.Published data relating to case-control studies reporting the link between IL-10 polymorph-isms and endometriosis were retrieved.Meta-analysis was conducted to determine whether the IL-10 gene polymorphisms were associated with endo-metriosis.Results Six studies were finally accepted for analysis.There were five studies focused on-819T/C polymorphism and six studies focused on-592A/C polymorphism.The odds ratio of each genetype for endometriosis:TT/CC,TC/CC,AA/CC and AC/CC were 0.90(0.81~0.99),P<0.05;0.92(0.82~1.02),P>0.05;0.92(0.79~1.07),P>0.05;0.98(0.90~1.06),P>0.05.No significant publication bias was found regarding TT/CC,TC/CC,AA/CC and AC/CC genotypes(P=0.81,0.81,0.71,0.26).Conclusion IL-10 gene-819TT genotypes may be a protetive factor of endometriosis.There was no related to show that other genotypes in position-819 and-592 were associated with endometriosis.