Objective To examine the changing prevalence and antimicrobial resistance profiles of Burkholderia cepacia in 52 hospitals across China from 2015 to 2021.Methods A total of 9 261 strains of B.cepacia were collected from 52 hospitals between January 1,2015 and December 31,2021.Antimicrobial susceptibility of the strains was tested using Kirby-Bauer method or automated antimicrobial susceptibility testing systems according to a unified protocol.The results were interpreted according to the breakpoints released in the Clinical & Laboratory Standards Institute(CLSI)guidelines(2023 edition).Results A total of 9 261 strains of B.cepacia were isolated from all age groups,especially elderly patients.The proportion was 11.1％(1 032 strains)in children,significantly lower than the proportion in adults.About half(46.5％,4 310/9 261)of the strains were isolated from patients at least 60 years old and 42.3％(3 919/9 261)of the strains were isolated from young adults.Most isolates(71.1％)were isolated from sputum and respiratory secretions,followed by urine(10.7％)and blood samples(8.1％).B.cepacia isolates were highly susceptible to the five antimicrobial agents recommended in the CLSI M100 document(33rd edition,2023).B.cepacia isolates showed relatively higher resistance rates to meropenem and levofloxacin.However,the resistance rates to ceftazidime,trimethoprim-sulfamethoxazole,and minocycline remained below 8.1％.The percentage of B.cepacia strains resistant to levofloxacin was the highest compared to other antibiotics in any of the three age groups(from 12.4％in the patients＜18 years old to 20.6％in the patients aged 60 years or older).Conclusions B.cepacia is one of the clinically important non-fermenting gram-negative bacteria.Accurate and timely reporting of antimicrobial susceptibility test results and ongoing antimicrobial resistance surveillance are helpful for rational prescription of antimicrobial agents and proper prevention and control of nosocomial infections.

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.

The amino acid sequences of the OmpA protein isolated from Escherichia coli QML2206-1(E.coli QML2206-1)in our laboratory were analyzed for homology with different strains of OmpA proteins using bioinformatics software,and the OmpA protein was analyzed for its physicochemical properties,transmembrane structure and signal peptide prediction,B-cell anti-genic epitope prediction,secondary and tertiary structure prediction.The OmpA gene fragment was ligated with pET-32a vector to construct a prokaryotic expression vector,which was purified by a nickel column affinity purification system after prokaryotic expression and optimization of ex-pression conditions in BL21(DE3).The purified recombinant protein was fully mixed with Freund's adjuvant to immunize mice,and the levels of mouse-specific IgG antibody and the expression levels of cytokines CD4,CD8 and IL-4 in mouse serum were detected by ELISA,and the immuno-protective effect was evaluated by mouse attack protection test.OmpA protein is a hydrophilic protein with no transmembrane structural domains and a secondary structure consisting mainly of irregular coils(47.98％)and α-helices(29.77％),with 12 antigenic epitopes that can bind to anti-bodies produced by B cells.The recombinant protein OmpA with a relative molecular mass of a-bout 55 kDa was successfully obtained by prokaryotic expression,and the highest expression was induced by IPTG concentration of 0.000 4 mmol/L for 6 h at 37 ℃.The serum-specific IgG anti-body potency of recombinant protein immunized mice was up to 1∶32 000;the expression levels of CD4,CD8 and IL-4 in the serum of immunized mice were elevated compared with those of the con-trol group.The survival rate of mice was 80％and 40％after attack with minimum lethal dose(MLD)and 2 times minimum lethal dose(2MLD),respectively.OmpA recombinant protein has good antigenicity and certain immunoprotective effects,and this study provides a technical basis for the next step in the development of a genetically engineered subunit vaccine against yak-appli-cable E.coli based on OmpA protein.

Objective:To investigate the clinical characteristics of a family with maturity-onset diabetes of the young type 3(MODY3) and assess its association with the novel HNF1A-c.47T>A variant.Methods:A genetic pedigree of the MODY3 family was constructed, and clinical data were collected. Whole-exome sequencing combined with Sanger sequencing was used, followed for familial co-segregation analysis. Bioinformatics tools, including multiple sequence alignment–based conservation analysis and protein structural prediction, were conducted to validate the association between the novel HNF1A-c.47T>A variant and MODY3.Results:Seven MODY3 patients were diagnosed in this family, all harboring the HNF1A-c.47T>A heterozygous variant. AlphaFold2 protein structure prediction indicated that the HNF1A-c.47T>A variant altered the conformation of the first pair of α-helices within the protein dimerization domain.Conclusion:Based on co-segregation analysis, sequence conservation assessment, protein structure prediction, and classification based on American College of Medical Genetics and Genomics(ACMG) Guidelines, the HNF1A-c.47T>A variant was determined to be pathogenic for MODY3. This study reports this novel pathogenic variant, expanding the mutational spectrum of MODY3. By revealing its disruptive effect on the protein dimerization domain, the findings provide a potential molecular basis for the diagnosis and management of patients carrying similar variants.

Objective:To establish 30-day of age transcutaneous bilirubin (TcB) reference curves for healthy neonates, and to investigate regional variations in bilirubin dynamics.Methods:A multicenter retrospective cohort study was conducted. A total of 220 950 healthy neonates born at a gestational age of 35-<42 weeks, with a birth weight ≥2 000 g, who did not receive phototherapy within 60 h after birth were recruited. All of them underwent remote TcB monitoring using the Bilibaby remote jaundice monitoring system between August 1 st, 2020 and December 31 st, 2024 in 426 hospitals. TcB data were collected within the period from birth to 30-day of age. The P40, P75, and P95 of TcB values were calculated, and dynamic TcB curves for 30-day of age were constructed. Patterns of bilirubin change, rates of change, and transition outcomes were described. Regional comparisons between South and North were conducted using linear mixed-effects models for TcB trajectories and Pearson′s chi-square test for outcome differences. Results:A total of 220 950 neonates were included, of whom 101 711 (46.03%) were female. Gestational age at birth was (38.75±1.12) weeks, and birth weight was (3 272±417) g. TcB levels increased rapidly within 3-day of age, peaked at 4-6-day of age, with peak values at P40, P75, and P95 of 200.6, 239.7 and 275.4 μmol/L (11.8, 14.1 and 16.2 mg/dl), respectively. TcB levels gradually declined thereafter and stabilized after 13-day of age, with values at P40, P75, and P95 fluctuating between 147.9-159.8, 190.4-200.6, and 231.2-239.7 μmol/L (8.7-9.4, 11.2-11.8, 13.6-14.1 mg/dl), respectively. Notably, among neonates categorized as low-or low-intermediate-risk within 3-day of age, 6 700 (12.76%) progressed to intermediate-high or high risk between 4 and 30 days of age. Before 13-day of age, TcB levels in the southern regions were consistently higher than those in the northern regions ( P=0.039); from 14 to 30 days of age, the overall TcB levels had no statistically difference, but the temporal changes in TcB still showed regional differences (degrees of freedom=3, all interaction P<0.05). Among neonates classified as low-or low-intermediate risk within 3-day of age, 25 326 were from southern regions, of whom 4 254 (16.80%) progressed to intermediate-high or high risk between 4 and 30 days of age. In northern regions, 27 193 neonates were classified as low-or low-intermediate risk within 3-day of age, among whom 2 446 (8.99%) progressed to intermediate-high or high risk. The risk progression between the 2 regions had statistically difference ( χ2=716.49, P<0.001). Conclusions:A TcB percentile curve for neonates within 30-day of age was established, revealing that both the overall TcB level and its temporal trend were higher in southern than in northern newborns. These findings provide baseline data to support continuous management of neonatal jaundice.

Objective:To investigate the clinical characteristics of a family with maturity-onset diabetes of the young type 3(MODY3) and assess its association with the novel HNF1A-c.47T>A variant.Methods:A genetic pedigree of the MODY3 family was constructed, and clinical data were collected. Whole-exome sequencing combined with Sanger sequencing was used, followed for familial co-segregation analysis. Bioinformatics tools, including multiple sequence alignment–based conservation analysis and protein structural prediction, were conducted to validate the association between the novel HNF1A-c.47T>A variant and MODY3.Results:Seven MODY3 patients were diagnosed in this family, all harboring the HNF1A-c.47T>A heterozygous variant. AlphaFold2 protein structure prediction indicated that the HNF1A-c.47T>A variant altered the conformation of the first pair of α-helices within the protein dimerization domain.Conclusion:Based on co-segregation analysis, sequence conservation assessment, protein structure prediction, and classification based on American College of Medical Genetics and Genomics(ACMG) Guidelines, the HNF1A-c.47T>A variant was determined to be pathogenic for MODY3. This study reports this novel pathogenic variant, expanding the mutational spectrum of MODY3. By revealing its disruptive effect on the protein dimerization domain, the findings provide a potential molecular basis for the diagnosis and management of patients carrying similar variants.

Objective:To establish 30-day of age transcutaneous bilirubin (TcB) reference curves for healthy neonates, and to investigate regional variations in bilirubin dynamics.Methods:A multicenter retrospective cohort study was conducted. A total of 220 950 healthy neonates born at a gestational age of 35-<42 weeks, with a birth weight ≥2 000 g, who did not receive phototherapy within 60 h after birth were recruited. All of them underwent remote TcB monitoring using the Bilibaby remote jaundice monitoring system between August 1 st, 2020 and December 31 st, 2024 in 426 hospitals. TcB data were collected within the period from birth to 30-day of age. The P40, P75, and P95 of TcB values were calculated, and dynamic TcB curves for 30-day of age were constructed. Patterns of bilirubin change, rates of change, and transition outcomes were described. Regional comparisons between South and North were conducted using linear mixed-effects models for TcB trajectories and Pearson′s chi-square test for outcome differences. Results:A total of 220 950 neonates were included, of whom 101 711 (46.03%) were female. Gestational age at birth was (38.75±1.12) weeks, and birth weight was (3 272±417) g. TcB levels increased rapidly within 3-day of age, peaked at 4-6-day of age, with peak values at P40, P75, and P95 of 200.6, 239.7 and 275.4 μmol/L (11.8, 14.1 and 16.2 mg/dl), respectively. TcB levels gradually declined thereafter and stabilized after 13-day of age, with values at P40, P75, and P95 fluctuating between 147.9-159.8, 190.4-200.6, and 231.2-239.7 μmol/L (8.7-9.4, 11.2-11.8, 13.6-14.1 mg/dl), respectively. Notably, among neonates categorized as low-or low-intermediate-risk within 3-day of age, 6 700 (12.76%) progressed to intermediate-high or high risk between 4 and 30 days of age. Before 13-day of age, TcB levels in the southern regions were consistently higher than those in the northern regions ( P=0.039); from 14 to 30 days of age, the overall TcB levels had no statistically difference, but the temporal changes in TcB still showed regional differences (degrees of freedom=3, all interaction P<0.05). Among neonates classified as low-or low-intermediate risk within 3-day of age, 25 326 were from southern regions, of whom 4 254 (16.80%) progressed to intermediate-high or high risk between 4 and 30 days of age. In northern regions, 27 193 neonates were classified as low-or low-intermediate risk within 3-day of age, among whom 2 446 (8.99%) progressed to intermediate-high or high risk. The risk progression between the 2 regions had statistically difference ( χ2=716.49, P<0.001). Conclusions:A TcB percentile curve for neonates within 30-day of age was established, revealing that both the overall TcB level and its temporal trend were higher in southern than in northern newborns. These findings provide baseline data to support continuous management of neonatal jaundice.

The amino acid sequences of the OmpA protein isolated from Escherichia coli QML2206-1(E.coli QML2206-1)in our laboratory were analyzed for homology with different strains of OmpA proteins using bioinformatics software,and the OmpA protein was analyzed for its physicochemical properties,transmembrane structure and signal peptide prediction,B-cell anti-genic epitope prediction,secondary and tertiary structure prediction.The OmpA gene fragment was ligated with pET-32a vector to construct a prokaryotic expression vector,which was purified by a nickel column affinity purification system after prokaryotic expression and optimization of ex-pression conditions in BL21(DE3).The purified recombinant protein was fully mixed with Freund's adjuvant to immunize mice,and the levels of mouse-specific IgG antibody and the expression levels of cytokines CD4,CD8 and IL-4 in mouse serum were detected by ELISA,and the immuno-protective effect was evaluated by mouse attack protection test.OmpA protein is a hydrophilic protein with no transmembrane structural domains and a secondary structure consisting mainly of irregular coils(47.98％)and α-helices(29.77％),with 12 antigenic epitopes that can bind to anti-bodies produced by B cells.The recombinant protein OmpA with a relative molecular mass of a-bout 55 kDa was successfully obtained by prokaryotic expression,and the highest expression was induced by IPTG concentration of 0.000 4 mmol/L for 6 h at 37 ℃.The serum-specific IgG anti-body potency of recombinant protein immunized mice was up to 1∶32 000;the expression levels of CD4,CD8 and IL-4 in the serum of immunized mice were elevated compared with those of the con-trol group.The survival rate of mice was 80％and 40％after attack with minimum lethal dose(MLD)and 2 times minimum lethal dose(2MLD),respectively.OmpA recombinant protein has good antigenicity and certain immunoprotective effects,and this study provides a technical basis for the next step in the development of a genetically engineered subunit vaccine against yak-appli-cable E.coli based on OmpA protein.

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.