Objective Viral encephalitis is an infectious disease severely affecting human health.It is caused by a wide variety of viral pathogens,including herpes viruses,flaviviruses,enteroviruses,and other viruses.The laboratory diagnosis of viral encephalitis is a worldwide challenge.Recently,high-throughput sequencing technology has provided new tools for diagnosing central nervous system infections.Thus,In this study,we established a multipathogen detection platform for viral encephalitis based on amplicon sequencing. Methods We designed nine pairs of specific polymerase chain reaction(PCR)primers for the 12 viruses by reviewing the relevant literature.The detection ability of the primers was verified by software simulation and the detection of known positive samples.Amplicon sequencing was used to validate the samples,and consistency was compared with Sanger sequencing. Results The results showed that the target sequences of various pathogens were obtained at a coverage depth level greater than 20×,and the sequence lengths were consistent with the sizes of the predicted amplicons.The sequences were verified using the National Center for Biotechnology Information BLAST,and all results were consistent with the results of Sanger sequencing. Conclusion Amplicon-based high-throughput sequencing technology is feasible as a supplementary method for the pathogenic detection of viral encephalitis.It is also a useful tool for the high-volume screening of clinical samples.

Objective Genotypes(G)1,3,and 5 of the Japanese encephalitis virus(JEV)have been isolated in China,but the dominant genotype circulating in Chinese coastal areas remains unknown.We searched for G5 JEV-infected cases and attempted to elucidate which JEV genotype was most closely related to human Japanese encephalitis(JE)in the coastal provinces of China. Methods In this study,we collected serum specimens from patients with JE in three coastal provinces of China(Guangdong,Zhejiang,and Shandong)from 2018 to 2020 and conducted JEV cross-neutralization tests against G1,G3,and G5. Results Acute serum specimens from clinically reported JE cases were obtained for laboratory confirmation from hospitals in Shandong(92 patients),Zhejiang(192 patients),and Guangdong(77 patients),China,from 2018 to 2020.Seventy of the 361 serum specimens were laboratory-confirmed to be infected with JEV.Two cases were confirmed to be infected with G1 JEV,32 with G3 JEV,and two with G5 JEV. Conclusion G3 was the primary infection genotype among JE cases with a definite infection genotype,and the infection caused by G5 JEV was confirmed serologically in China.

Objective:To analyze the association between healthy lifestyle and mortality among Henan Province 35-74 years old individuals.Methods:Data from the programme of screening and intervention subjects with high-risk cardiovascular disease 99 133 adults were analyzed in a provincial cohort study of 16 counties. Four healthy lifestyle behaviors were assessed based on a questionnaire survey. Information on mortality endpoints was retrieved from the national death surveillance system. Cox proportional hazards regression models were used to estimate the associations between healthy lifestyles, mortality risk and population attributable fraction (PAF).Results:Out of the adult participants in Henan, 50.6% adhered to a healthy lifestyle, and only 0.1% adhered to 4 healthy lifestyle behaviours. During a mean of 4.5 years, 2 685 all-cause death and 1 283 cardiovascular deaths were documented. The decreased risk of mortality among individuals with non-smoking, moderate drinking, adequate exercise and healthy diet were 0.85 (95% CI: 0.77-0.94), 0.75 (95% CI: 0.63-0.89), 0.73 (95% CI: 0.67-0.79) and 0.86 (95% CI: 0.77-0.96), while the adjusted PAF for all-cause deaths were 5.2% (95% CI: 2.5%-7.9%), 24.0% (95% CI: 10.7%-36.4%), 19.4% (95% CI: 13.8%-24.8%) and 12.3% (95% CI: 3.4%-20.9%), respectively. A combined healthy lifestyle can bring more health benefits. Adherence to 4 healthy lifestyle behaviours could avoid 49.1% of all-cause death. Conclusion:Adherence to a healthy lifestyle can reduce the risk of death, and participants with a healthy lifestyle had a lower mortality risk.

Patients with severe trauma require an extremely timely treatment and transfusion plays an irreplaceable role in the emergency treatment of such patients. An increasing number of evidence-based medicinal evidences and clinical practices suggest that patients with severe traumatic bleeding benefit from early transfusion of low-titer group O whole blood or hemostatic resuscitation with red blood cells, plasma and platelet of a balanced ratio. However, the current domestic mode of blood supply cannot fully meet the requirements of timely and effective blood transfusion for emergency treatment of patients with severe trauma in clinical practice. In order to solve the key problems in blood supply and blood transfusion strategies for emergency treatment of severe trauma, Branch of Clinical Transfusion Medicine of Chinese Medical Association, Group for Trauma Emergency Care and Multiple Injuries of Trauma Branch of Chinese Medical Association, Young Scholar Group of Disaster Medicine Branch of Chinese Medical Association organized domestic experts of blood transfusion medicine and trauma treatment to jointly formulate Chinese expert consensus on blood support mode and blood transfusion strategies for emergency treatment of severe trauma patients ( version 2024). Based on the evidence-based medical evidence and Delphi method of expert consultation and voting, 10 recommendations were put forward from two aspects of blood support mode and transfusion strategies, aiming to provide a reference for transfusion resuscitation in the emergency treatment of severe trauma and further improve the success rate of treatment of patients with severe trauma.

Objective To quantitatively analyze the impact of environmental hygiene on the occurrence of health-care-associated infections(HAI).Methods Monitoring data of HAI and environmental hygiene from a tertiary first-class hospital from January 2018 to December 2022 were collected,and the impact of environmental bacterial colony forming unit(CFU)on the occurrence of HAI was analyzed by a time-series generalized additive model.Results The single-contamination model showed a significant positive correlation between HAI and staff's hand bacterial CFU(β1=0.009,P=0.012).For an increase of 1 interquartile range(IQR)in the monthly mean CFU per dish(MCFU/Dish)of staffs'hand,the incidence of HAI increased by 13.28％(95％CI:2.82％-24.81％).Subgroup and lag effect analysis showed that when the monthly MCFU/Dish(after hand disinfection)of staffs'hand in-creased by one IQR,the excess risk(ER)of HAI for the month(lag0)was 16.26％(95％CI:15.45％-17.09％).In the multi-contamination model,the correlation between surface contamination and HAI was also statistically sig-nificant.Conclusion There is a significant correlation between hospital environmental hygiene and the occurrence of HAI.

Objective:Observing the effect of exosomes derived from hypoxic Bone marrow mesenchymal stem cells (BMSCs) on the function of chondrocytes, and exploring the role and mechanism of exosomal miR-196b-5p. Evaluating the application prospects of hypoxic BMSCs exosomes and miR-196b-5p for cartilage regeneration.Methods:Chondrocytes were cultured in the supernatant of BMSCs cultured under normoxia or hypoxia, respectively. The proliferation of chondrocytes was detected by CCK-8 assay and the expressions of Collagen type 2 (Col2), Col1, Aggrecan and SOX9 were detected by qPCR to evaluate the effect of hypoxic BMSCs paracrine on chondrocyte functions. Obtaining normoxic and hypoxic exosomes through ultracentrifugation, and testing their effects on the proliferation and anabolic-related genes of chondrocytes through CCK-8 assay and qPCR. Verifying the expression of miR-196b-5p in hypoxic exosomes based on exosomal miRNA array. Knocking out miR-196b-5p in hypoxic BMSCs, and detecting the effect of hypoxic exosomal miR-196b-5p on the functions of chondrocytes by loss-of-function assay. Predicting the downstream of miR-196b-5p through bioinformatics tools, and exploring the mechanism of hypoxic exosomal miR-196b-5p by gain-of-function assays. Hypoxic exosomes and miR-196b-5p-knockout hypoxic exosomes were loaded on silk fibroin hydrogel and subcutaneously into nude mice. After 4 weeks of culture, histological staining of saffron O, Masson and biochemical content of sGAG and collagen were performed to assess the application prospect of hypoxic exosomes and hypoxic exosomal miR-196b-5p on cartilage regeneration. Results:The results of CCK-8 assay and qPCR indicated that the supernatant of hypoxic BMSCs significantly promoted the proliferation of chondrocytes 1.20±0.07 and the expression of cartilage-related markers (Col2 2.95±0.17, Aggrecan 2.45±0.27, SOX9 2.92±0.29) compared to normoxic BMSCs (0.94±0.04, 1.89±0.09, 1.67±0.21, 1.76±0.16), the differences were statistically significant ( P<0.05). The result of CCK-8 assay showed that hypoxic exosomes (1.28±0.04) promoted the proliferation of chondrocytes compared to normoxic exosomes 1.05±0.06, the differences were statistically significant ( P<0.05). CCK-8 assay revealed that the down-regulation of miR-196b-5p in hypoxic exosomes 0.99±0.06 attenuated the proliferation of chondrocytes compared to control group 1.20±0.07, the differences were statistically significant ( P<0.05); the expression of Col2 0.56±0.04, Aggrecan 0.74±0.09, and SOX9 0.45±0.05 in chondrocytes was reduced in the miR-196b-5p knockdown group compared to the control group (1.00±0.09, 1.00±0.12, 1.00±0.07), the differences were statistically significant ( P<0.05). Co-transfection of pmirGLO-BACH1-WT reporter vector with miR-196b-5p mimics decreased the luciferase activity 0.73±0.06, the differences were statistically significant ( P<0.05). Co-transfection of pmirGLO-BACH1-MUT reporter vector with miR-196b-5p mimics showed no change in luciferase activity. BACH1 is the target of miR-196b-5p. Subcutaneous culture in nude mice showed that hypoxic exosomes significantly promoted the deposition of sGAG 383.2±21.54 and collagen 67.40±3.45, while reducing the expression of miR-196b-5p in hypoxic exosomes weakened the deposition of sGAG 258.4±19.50 and collagen 57.15±4.95, the differences were statistically significant ( P<0.05). Conclusion:Hypoxic exosomes promoted the functions of chondrocytes by inhibiting the expression of BACH1 through miR-196b-5p. Hypoxic exosomes can be applied in cartilage regeneration.

Methods: In this study, 267 matched pairs of AIS and controls were recruited. The participants underwent EMG measurements at their first presentation and did not receive any treatment for 6 months at which point they underwent EMG and radiographs. Early curve progression was defined as >5° in Cobb angle at 6 months. The root mean square of the EMG (rms-EMG) signal was recorded with the participants in sitting and back extension. The rms-EMG ratio at the upper end vertebrae, apical vertebrae (AV), and lower end vertebrae (LEV) of the major curve was calculated. Results: The rms-EMG ratio in the scoliosis cohort was high compared with that in the controls (sitting: 1.2±0.3 vs. 1.0±0.1, p<0.01; back extension: 1.1±0.2 vs. 1.0±0.1, p<0.01). An AV rms-EMG ratio in back extension, with a cutoff threshold of ≥1.5 in the major thoracic curve and ≥1.3 in the major lumbar curve, was a risk factor for early curve progression after 6 months without treatment (odds ratio, 4.1; 95% confidence interval, 2.8–5.9; p<0.01). Increases in side deviation (SD) (distance between the AV and the central sacral line) were related to a higher rms-EMG ratio in LEV of the major thoracic curve (baseline: rs=0.2, p=0.03; 6 months: rs=0.3, p<0.01). Conclusions An EMG discrepancy was detected in the scoliosis cohort, which was related to increases in SD in the major thoracic curve. The AV rms-EMG ratio in back extension was correlated with curve progression after 6 months of no treatment.

Traumatic cerebrospinal fluid leakage commonly presents in traumatic brain injury patients, and it may lead to complications such as meningitis, ventriculitis, brain abscess, subdural hematoma or tension pneumocephalus. When misdiagnosed or inappropriately treated, traumatic cerebrospinal fluid leakage may result in severe complications and may be life-threatening. Some traumatic cerebrospinal fluid leakage has concealed manifestations and is prone to misdiagnosis. Due to different sites and mechanisms of trauma and degree of cerebrospinal fluid leak, treatments for traumatic cerebrospinal fluid leakage varies greatly. Hence, the Craniocerebral Trauma Professional Group of Neurosurgery Branch of Chinese Medical Association and the Neurological Injury Professional Group of Trauma Branch of Chinese Medical Association organized relevant experts to formulate the " Chinese expert consensus on the diagnosis and treatment of traumatic cerebrospinal fluid leakage in adults ( version 2023)" based on existing clinical evidence and experience. The consensus consisted of 16 recommendations, covering the leakage diagnosis, localization, treatments, and intracranial infection prevention, so as to standardize the diagnosis and treatment of traumatic cerebrospinal fluid leakage and improve the overall prognosis of the patients.

Abstract： Objective　To elucidate the antigenic antibody reaction of recombinant expression of non-structural protein 1 (NS1) of Japanese encephalitis (JE) virus with various mosquito-borne flaviviruses, including JE virus, and the antigenic antibody reaction of serum samples of patients infected with JE virus in acute stage. Methods　In this study, Escherichia coli prokaryotic expression vector (pET) system was used to recombinant express Japanese encephalitis virus NS1 gene. Western Blot assay was performed to detect the antibody responses of the recombinantly expressed protein against a variety of mosquito-transmitted flaviviruses, including JE virus, as well as antigen-antibody reactions of serum from patients with acute JE virus infection. Results　The NS1 gene expression product of JE virus (P3 strain) was in the form of an inclusion body, and the denatured and renatured expression product was displayed as a single band in the denatured gel (polyacrylamide gel electrophoresis, PAGE), with a molecular weight of about 45 000. The results of further antigen-antibody analysis showed that the antigen/antibody hybridization reaction of the expression product with polyclonal or monoclonal antibody of JE virus (mosquito isolates, encephalitis isolates) and serum samples of patients with acute JE virus infection could be completely consistent. The recombinant product showed negative antigen/antibody hybridization reactions with mosquito-transmitted flaviviruses, such as dengue virus and yellow fever virus polyclonal antibodies, but positive reactions with polyclonal antibodies to West Nile virus and Murray Valley encephalitis virus. Conclusions　In this study, the recombinant expression of the NS1 protein of JE virus was successfully obtained, and the antigen/antibody reaction between the recombinant protein and samples of patients infected with mosquito-borne flavivirus and JE virus was analyzed. The study results provide important basic data for elucidating the antigen-antibody reaction between the NS1 protein of JE virus and mosquito-borne flavivirus. The recombinant expression protein obtained in this study provides an important material basis for further research on the function of JE virus NS1 protein.

Objective:To determine the evaluation indexes of national standard strains of genotypes 1, 3 and 5 of Japanese encephalitis virus (JEV) and evaluate the national standard JEV strains.Methods:According to the national standard strain evaluation technical standards of pathogenic microbial bacteria (virus) species, based on the application of Japanese encephalitis virus research, and according to the morphological characteristics, biological characteristics, molecular biological characteristics and other research data to identify the characteristics of G1, G3 and G5 genotypes of JEV.Results:Spherical virus particles with a diameter of about 60 nm were visible under electron microscope of the three Japanese encephalitis virus strains. The cytopathic effect was mainly characterized by cell shrinkage and exfoliation in BHK-21 and Vero cell lines, cell fusion and exfoliation were shown after infection with C6/36 cell line; the virus titer was 10 5-10 7 PFU/ml, and the plaque size was different by genotype. The median lethal dose of intrabitoneal challenge in G1, G3 and G5 JEV in three weeks-old mice was 50.51 PFU, 6.98 PFU, and 8.13 PFU, and the median lethal dose of intracranial challenge in five weeks mice was 3 PFU, 0.3 PFU, 1.35 PFU. The whole genome length of G1, G3 and G5 JEV was 10 967 bp, 10 976 bp and 10 983 bp, respectively. Conclusions:Three genotypic national standard strains of JE V were identified and evaluated by electron microscopy, cell, animal and genome laboratory indexes, which provided reference for the identification and evaluation of other national standard strains of JEV.