ObjectiveTo investigate the role of DEAD-box helicase 3 X-linked （DDX3X） in immune-mediated liver injury （ILI）， and to clarify its mechanism by regulating endoplasmic reticulum stress （ERS）-dependent apoptotic pathway and its association with the clinical progression of hepatitis B. MethodsMice were given injection of concanavalin A （ConA） via the caudal vein to establish a model of ILI， PBS （control group） and different concentrations of ConA were injected into the tail vein of hepatocyte-specific DDX3X-knockout mice （DDX3X^ΔHep and DDX3X-flox mice （DDX3X^fl/fl）， respectively.. The log-rank survival analysis， measurement of the serum levels of aspartate aminotransferase （AST） and alanine aminotransferase （ALT）， and HE staining of liver tissue were performed to assess liver injury， and qRT-PCR and Western Blot were used to measure the mRNA and protein expression levels of glucose-regulated protein 78 （GRP78）， CCAAT/enhancer-binding protein homologous protein （CHOP）， and DDX3X in liver tissue. Intraperitoneal injection of 4-phenylbutyric acid （4-PBA， 100 mg/kg） was performed to inhibit ERS. Serum samples （n=30） and liver tissue samples （n=6） were collected from healthy controls， chronic hepatitis B （CHB） patients， and hepatitis B virus-associated liver failure （HBV-LF） patients； ELISA was used to measure the serum level of DDX3X， and qRT-PCR/Western Blot was used to analyze the expression of targets in liver tissue. A one-way analysis of variance was used for comparison of continuous data between multiple groups， and the least significant difference t-test was used for further comparison between two groups. ResultsCompared with the control group of mice， the expression of DDX3X in the liver of mice induced by ConA was significantly increased after liver injury （P<0.05）， and hepatocyte-specific DDX3X knockout increased the 72-hour survival rate of mice by 55% （compared with 20% in the DDX3X^fl/fl group）， with significant reductions in the serum levels of ALT and AST （P<0.000 1） and the expression levels of the ERS markers GRP78 and CHOP （P<0.05）. After ERS was inhibited by 4-PBA， there was alleviation of liver injury （with reductions in ALT and AST， P <0.001） and a reduction in DDX3X expression （P<0.01）. The analysis of clinical samples showed that the mRNA and protein expression levels of liver DDX3X in CHB patients and HBV-LF patients were significantly higher than those in healthy controls （all P<0.01）， and there was a significant increase in the serum level of DDX3X in HBV-LF patients （P<0.000 1）. ConclusionDDX3X exacerbates ILI by regulating the ERS-dependent apoptotic pathway （GRP78/CHOP）， and its expression is associated with the progression of hepatitis B. Therefore， it can be used as a potential therapeutic target.

Sustained inflammatory responses are closely related to various severe diseases,and inhibiting the excessive activation of inflammasomes and pyroptosis has significant implications for clinical treatment.Natural products have garnered considerable concern for the treatment of inflammation.Huanglian-Wumei decoction(HLWMD)is a classic prescription used for treating inflammatory diseases,but the necessity of their combination and the exact underlying anti-inflammatory mechanism have not yet been elucidated.Inspired by the supramolecular self-assembly strategy and natural drug compatibility theory,we successfully obtained berberine(BBR)-chlorogenic acid(CGA)supramolecular(BCS),which is an herbal pair from HLWMD.Using a series of characterization methods,we confirmed the self-assembly mechanism of BCS.BBR and CGA were self-assembled and stacked into amphiphilic spherical supra-molecules in a 2:1 molar ratio,driven by electrostatic interactions,hydrophobic interactions,and π-πstacking;the hydrophilic fragments of CGA were outside,and the hydrophobic fragments of BBR were inside.This stacking pattern significantly improved the anti-inflammatory performance of BCS compared with that of single free molecules.Compared with free molecules,BCS significantly attenuated the release of multiple inflammatory mediators and lipopolysaccharide(LPS)-induced pyroptosis.Its anti-inflammatory mechanism is closely related to the inhibition of intracellular nuclear factor-kappaB(NF-κB)p65 phosphorylation and the noncanonical pyroptosis signalling pathway mediated by caspase-11.

Objective To summarize the changing prevalence of carbapenem resistance in Enterobacterales based on the data of CHINET Antimicrobial Resistance Surveillance Program from 2015 to 2021 for improving antimicrobial treatment in clinical practice.Methods Antimicrobial susceptibility testing was performed using a commercial automated susceptibility testing system according to the unified CHINET protocol.The results were interpreted according to the breakpoints of the Clinical & Laboratory Standards Institute(CLSI)M100 31st ed in 2021.Results Over the seven-year period(2015-2021),the overall prevalence of carbapenem-resistant Enterobacterales(CRE)was 9.43％(62 342/661 235).The prevalence of CRE strains in Klebsiella pneumoniae,Citrobacter freundii,and Enterobacter cloacae was 22.38％,9.73％,and 8.47％,respectively.The prevalence of CRE strains in Escherichia coli was 1.99％.A few CRE strains were also identified in Salmonella and Shigella.The CRE strains were mainly isolated from respiratory specimens(44.23±2.80)％,followed by blood(20.88±3.40)％and urine(18.40±3.45)％.Intensive care units(ICUs)were the major source of the CRE strains(27.43±5.20)％.CRE strains were resistant to all the β-lactam antibiotics tested and most non-β-lactam antimicrobial agents.The CRE strains were relatively susceptible to tigecycline and polymyxins with low resistance rates.Conclusions The prevalence of CRE strains was increasing from 2015 to 2021.CRE strains were highly resistant to most of the antibacterial drugs used in clinical practice.Clinicians should prescribe antimicrobial agents rationally.Hospitals should strengthen antibiotic stewardship in key clinical settings such as ICUs,and take effective infection control measures to curb CRE outbreak and epidemic in hospitals.

Objective To characterize the changing species distribution and antibiotic resistance profiles of respiratory isolates in hospitals participating in the CHINET Antimicrobial Resistance Surveillance Program from 2015 to 2021.Methods Commercial automated antimicrobial susceptibility testing systems and disk diffusion method were used to test the susceptibility of respiratory bacterial isolates to antimicrobial agents following the standardized technical protocol established by the CHINET program.Results A total of 589 746 respiratory isolates were collected from 2015 to 2021.Overall,82.6％of the isolates were Gram-negative bacteria and 17.4％were Gram-positive bacteria.The bacterial isolates from outpatients and inpatients accounted for(6.0±0.9)％and(94.0±0.1)％,respectively.The top microorganisms were Klebsiella spp.,Acinetobacter spp.,Pseudomonas aeruginosa,Staphylococcus aureus,Haemophilus spp.,Stenotrophomonas maltophilia,Escherichia coli,and Streptococcus pneumoniae.Each microorganism was isolated from significantly more males than from females(P＜0.05).The overall prevalence of methicillin-resistant S.aureus(MRSA)was 39.9％.The prevalence of penicillin-resistant S.pneumoniae was 1.4％.The prevalence of extended-spectrum β-lactamase(ESBL)-producing E.coli and K.pneumoniae was 67.8％and 41.3％,respectively.The overall prevalence of carbapenem-resistant E.coli,K.pneumoniae,Enterobacter cloacae,Pseudomonas aeruginosa,and Acinetobacter baumannii was 3.7％,20.8％,9.4％,29.8％,and 73.3％,respectively.The prevalence of β-lactamase was 96.1％in Moraxella catarrhalis and 60.0％in Haemophilus influenzae.The H.influenzae isolates from children(＜18 years)showed significantly higher resistance rates to β-lactam antibiotics than the isolates from adults(P＜0.05).Conclusions Gram-negative bacteria are still predominant in respiratory isolates associated with serious antibiotic resistance.Antimicrobial resistance surveillance should be strengthened in clinical practice to support accurate etiological diagnosis and appropriate antimicrobial therapy based on antimicrobial susceptibility testing results.

Objective To investigate the distribution and antimicrobial resistance profiles of common pathogens isolated from cerebrospinal fluid(CSF)in CHINET program from 2015 to 2021.Methods The bacterial strains isolated from CSF were identified in accordance with clinical microbiology practice standards.Antimicrobial susceptibility test was conducted using Kirby-Bauer method and automated systems per the unified CHINET protocol.Results A total of 14 014 bacterial strains were isolated from CSF samples from 2015 to 2021,including the strains isolated from inpatients(95.3％)and from outpatient and emergency care patients(4.7％).Overall,19.6％of the isolates were from children and 80.4％were from adults.Gram-positive and Gram-negative bacteria accounted for 68.0％and 32.0％,respectively.Coagulase negative Staphylococcus accounted for 73.0％of the total Gram-positive bacterial isolates.The prevalence of MRSA was 38.2％in children and 45.6％in adults.The prevalence of MRCNS was 67.6％in adults and 69.5％in children.A small number of vancomycin-resistant Enterococcus faecium(2.2％)and linezolid-resistant Enterococcus faecalis(3.1％)were isolated from adult patients.The resistance rates of Escherichia coli and Klebsiella pneumoniae to ceftriaxone were 52.2％and 76.4％in children,70.5％and 63.5％in adults.The prevalence of carbapenem-resistant E.coli and K.pneumoniae(CRKP)was 1.3％and 47.7％in children,6.4％and 47.9％in adults.The prevalence of carbapenem-resistant Acinetobacter baumannii(CRAB)and Pseudomonas aeruginosa(CRPA)was 74.0％and 37.1％in children,81.7％and 39.9％in adults.Conclusions The data derived from antimicrobial resistance surveillance are crucial for clinicians to make evidence-based decisions regarding antibiotic therapy.Attention should be paid to the Gram-negative bacteria,especially CRKP and CRAB in central nervous system(CNS)infections.Ongoing antimicrobial resistance surveillance is helpful for optimizing antibiotic use in CNS infections.

Objective To investigate the antimicrobial resistance of clinical isolates from elderly patients(≥65 years)in major medical institutions across China.Methods Bacterial strains were isolated from elderly patients in 52 hospitals participating in the CHINET Antimicrobial Resistance Surveillance Program during the period from 2015 to 2021.Antimicrobial susceptibility test was carried out by disk diffusion method and automated systems according to the same CHINET protocol.The data were interpreted in accordance with the breakpoints recommended by the Clinical and Laboratory Standards Institute(CLSI)in 2021.Results A total of 514 715 nonduplicate clinical isolates were collected from elderly patients in 52 hospitals from January 1,2015 to December 31,2021.The number of isolates accounted for 34.3％of the total number of clinical isolates from all patients.Overall,21.8％of the 514 715 strains were gram-positive bacteria,and 78.2％were gram-negative bacteria.Majority(90.9％)of the strains were isolated from inpatients.About 42.9％of the strains were isolated from respiratory specimens,and 22.9％were isolated from urine.More than half(60.7％)of the strains were isolated from male patients,and 39.3％isolated from females.About 51.1％of the strains were isolated from patients aged 65-＜75 years.The prevalence of methicillin-resistant strains(MRSA)was 38.8％in 32 190 strains of Staphylococcus aureus.No vancomycin-or linezolid-resistant strains were found.The resistance rate of E.faecalis to most antibiotics was significantly lower than that of Enterococcus faecium,but a few vancomycin-resistant strains(0.2％,1.5％)and linezolid-resistant strains(3.4％,0.3％)were found in E.faecalis and E.faecium.The prevalence of penicillin-susceptible S.pneumoniae(PSSP),penicillin-intermediate S.pneumoniae(PISP),and penicillin-resistant S.pneumoniae(PRSP)was 94.3％,4.0％,and 1.7％in nonmeningitis S.pneumoniae isolates.The resistance rates of Klebsiella spp.(Klebsiella pneumoniae 93.2％)to imipenem and meropenem were 20.9％and 22.3％,respectively.Other Enterobacterales species were highly sensitive to carbapenem antibiotics.Only 1.7％-7.8％of other Enterobacterales strains were resistant to carbapenems.The resistance rates of Acinetobacter spp.(Acinetobacter baumannii 90.6％)to imipenem and meropenem were 68.4％and 70.6％respectively,while 28.5％and 24.3％of P.aeruginosa strains were resistant to imipenem and meropenem,respectively.Conclusions The number of clinical isolates from elderly patients is increasing year by year,especially in the 65-＜75 age group.Respiratory tract isolates were more prevalent in male elderly patients,and urinary tract isolates were more prevalent in female elderly patients.Klebsiella isolates were increasingly resistant to multiple antimicrobial agents,especially carbapenems.Antimicrobial resistance surveillance is helpful for accurate empirical antimicrobial therapy in elderly patients.

Objective This study aimed to investigate the antimicrobial resistance profiles of bacterial strains isolated from pediatric intensive care units(PICU)in China for better antimicrobial therapy.Methods Clinical isolates were collected from 17 institutions,including tertiary care children's hospitals and pediatric department of tertiary general hospitals in China from January 1,2020 to December 31,2022.Antimicrobial susceptibility testing was carried out according to a unified protocol using Kirby-Bauer method or automated systems.Results were interpreted according to the breakpoints released by the Clinical and Laboratory Standards Institute(CLSI)in 2020.Results A total of 10 688 isolates were collected,including gram-positive organisms(39.2％)and gram-negative organisms(60.8％).The top three organisms were S.aureus(13.6％,1 453/10 688),A.baumannii(10.0％,1 067/10 688),and coagulase-negative Staphylococcus(9.9％,1 058/10 688).Multi-drug resistant organisms(MDROs)were very common in children.The prevalence of methicillin-resistant Staphylococcus aureus(MRSA),carbapenem-resistant Enterobacterales(CRE),carbapenem-resistant E.coli,carbapenem-resistant K.pneumoniae(CRKP),carbapenem-resistant A.baumannii(CRAB),and carbapenem-resistant P.aeruginosa(CRPA)was 41.1％,19.4％,8.8％,30.9％,67.4％,and 28.8％,respectively.Overall,more than 50％of Enterobacteriales isolates were resistant to cephalosporins,while nearly 25％of Enterobacteriales isolates were resistant to carbapenems.MDROs were highly resistant to commonly used antibiotics.More than 80％of CRE and CRAB strains were resistant to all beta-lactam antibiotics.CRE and CRAB showed low resistance rates to tigecycline and polymyxin.CRPA showed lower resistance rates to piperacillin,beta-lactamase inhibitor combinations than the resistance rates to third and fourth generation cephalosporins.All of the Staphylococcus and Enterococcus isolates were susceptible to vancomycin and tigecycline.None of PRSP strains isolated from meningitis and nonmeningitis samples were resistant to rifampicin,vancomycin,or linezolid.The prevalence of β-lactamase-negative ampicillin-resistant(BLNAR)strains was 43.3％in Haemophilus influenzae.Conclusions MDROs were prevalent in PICU.It is necessary to establish an effective multidisciplinary team(MDT)to control the antimicrobial resistance.

Objective To explore the effect and preliminary mechanism of the plant-derived immunostimulatory molecule,ophiopogonin,on enhancing the immune response of a subunit vaccine with the receptor-binding domain(RBD)of coronavirus spike protein as the antigen.Methods CCK-8 assay was used to determine the cytotoxicity of ophiopogonin D'(OPD')on bone marrow-derived dendritic cells(BMDCs).Female Balb/c mice were randomly divided into RBD,RBD/OPD',RBD/Alum,and control groups.The immunization dose was 5 μg of antigen per mouse and 100 μg of adjuvant per mouse,and immunization was carried out according to the intramuscular injection immunization procedure on days 0,21,and 42.The titers of specific IgG and its subtype antibodies were detected by ELISA.The cytokine levels in the supernatant of splenocytes were detected using ELISA.The number of splenocytes secreting IFN-γ was detected by ELISpot.Laser confocal microscopy was employed to observe the uptake of antigen by BMDCs.The phagocytic ability of BMDCs for antigen was quantitatively analyzed by flow cytometry.The mechanism of its enhanced immune effect was preliminarily explored using transcriptomics technology combined with bioinformatics research.Results When the concentration of OPD'was less than 5 μg/mL,the survival rate of BMDCs was 100%.After a single intramuscular injection in mice,except for a slight decrease in body weight,the other biochemical indicators were within corresponding normal ranges.After intramuscular injection immunization of the vaccine,the titers of serum-specific IgG,IgG1,and IgG2a in the RBD/OPD'group were significantly higher than those in the RBD group(P<0.05).Compared with the RBD group,the RBD/OPD'group induced a high-level Th1 cell immune response of IL-1β,TNF-α,and IFN-γ(P<0.01)and had more lymphocytes secreting IFN-γ(P<0.001).Laser confocal microscopy displayed that BMDCs took up more antigens after OPD'treatment,which was further confirmed with flow cytometry in quantitative analysis on antigen uptake rate(P<0.01).Transcriptomics results indicated that there was more significant enrichment of the PPAR signaling pathway in the RBD/OPD'group than the RBD group,suggesting that OPD'may activate the PPAR signaling pathway to exert its adjuvant effect.Conclusion OPD'effectively enhances the immune response of the RBD subunit vaccine,and its action mechanism may be related to the activation of the PPAR signaling pathway.

Objective To establish a mouse model of infection with the minimum lethal dose of Vibrio vulnificus(V.vulnificus)and to evaluate the protective efficacy of inactivated whole-cell(IWC)vaccine using this model.Methods A mouse model of lethal-dose infection was established by intraperitoneal injection of different doses of V.vulnificus.Bacterial colonization in the organs was detected with tissue homogenate plating,and pathological changes in the organs were observed after tissue section staining.Flow cytometry was used to detect immune cell responses after liver tissues were digested into single-cell suspension.IWC vaccine of V.vulnificus was prepared,and the mice were immunized through different routes to observe the protective efficacy of the vaccine.Results A mouse model of infection with the minimum lethal dose at 1×106 CFU of V.vulnificus was successfully established.After infection,the bacteria were mainly colonized in the liver of mice and caused severe pathological damages.Compared with the uninfected mice,the proportion of neutrophils in the liver was significantly increased in the infected mice,whereas the proportions of B cells and T cells were correspondingly decreased(P<0.05).A single intramuscular or intraperitoneal injection of the IWC vaccine could protect the mice effectively against lethal infection of V.vulnificus in 7 d later(P<0.01),although the level of serum IgG having no significant increase.Conclusion A mouse model of lethal-dose infection with V.vulnificus is successfully established,with histopathological characteristics.The IWC vaccine of V.vulnificus rapidly mediates immune protection in this model probably independent of IgG.

In this work,near infrared carbon quantum dots(NIR-CDs)were synthesized by hydrothermal method using biomass material Clausena lansium leaves.The synthesized NIR-CDs emitted maximum fluorescence signal at 677 nm,which was independent of excitation wavelength.The characterization results showed that there were abundant groups on the surface of NIR-CDs.Pd2+could form non-fluorescent compounds with the surface groups of NIR-CDs,resulting in fluorescence quenching(Fluorescence signal was denoted as F0).Because chlorpromazine hydrochloride(CPZ)parent nucleus contained unoxidized S atom,CPZ could form stable colored complex with Pd2+under acidic conditions.In the presence of CPZ,Pd2+dissociated from the surface of NIR-CDs and bonded with CPZ,so that the fluorescence signal could be restored(Fluorescence signal was denoted as F).An"on-off-on"fluorescence sensor was thus constructed.The fluorescence signal recovery value of NIR-CDs(△F=F-F0)showed a good linear relationship with the concentration of CPZ in the range of 5.68-28.43 μg/mL,and the detection limit(3σ)was 0.078 μg/mL.The sensor was applied to determination of CPZ in pharmaceutical preparations,and the recoveries were 94％-106％.The developed fluorescence sensor was expected to be used in quality control of actual pharmaceutical preparations.