Objective:To establish and optimize a novel method, focus forming assay (FFA), for quantitation of influenza A virus (FluA) and compare its application performance with traditional plague forming assay (PFA).Methods:The foci chromogenic effects of three peroxidase substrates in immunostaining were compared. The PFA and FFA methods were used to explore FluA incubation times and plaque morphology on 12-well plates, and to determine optimal incubation times and virus adsorption volumes for different FluA subtypes on 96-well plates. The correlation between FFA and PFA was evaluated, and the optimized FFA was applied to the in vitro antiviral efficacy analysis of Favipiravir and neutralization test against different subtypes of FluA. Results:TRUEBLUE substrate was identified as the optimal substrate for foci visualization. Compared with the PFA, the FFA showed improved sensitivity and reduced detection time in FluA titration, and good correlation was shown between the two methods′ results. By replacing the 96-well plate with the 12-well plate for FFA titration of different subtypes of FluA, the detection time was shortened, and the amount of serum samples used could be further reduced by optimizing the virus adsorption volume. The half-maximal effective concentration of favipiravir against influenza viruses assessed by the FFA and PFA methods showed no significant difference, and was consistent with the results obtained from quantitative PCR. Additionally, the focus reduction neutralization test and hemagglutination inhibition assays demonstrated strong correlation in determining antibody titers against FluA in serum neutralization assays.Conclusions:The improved FFA method developed here provides a more efficient experimental tool for FluA titration, antiviral drug screening and broad-spectrum vaccine evaluation.

Objective: To explore the experience of self growth among caregivers of adolescents with non suicidal self injury (NSSI), so as to provide practical reference for improving the family support system of adolescents with NSSI. Methods: From August to November 2023, a purposive sampling method was used to select 21 caregivers of adolescents with NSSI who were treated in a tertiary grade A psychiatric hospital in Xinxiang City, Henan Province for semi structured in depth interviews. The Colaizzi s seven step qualitative analysis method was applied to analyze the data and extract themes related to caregiver growth Results: A total of 5 themes and 12 sub themes of benefit finding were extracted from caregivers of adolescents with NSSI: personal growth (increased psychological resilience, enhanced awareness of independent learning, improved self reflection ability, better understanding and acceptance of NSSI behaviors in adolescents), improvement of family relationships (improved parent-child relationship, improved relationships among family members), perceived social support (more peer support, support from close friends), improvement of caregiving ability (creating a favorable rehabilitation environment, improved caregiving skills), and changes in life and education attitudes (adopting a healthier lifestyle, reasonably adjusting expectations for children). Conclusions During the process of caring for adolescents with NSSI, caregivers from positive experiences in multiple aspects such as personal growth, family relationships, social support, and life education attitudes. Improving the mental health level of caregivers and optimize the family rehabilitation environment, will help reduce the occurrence of NSSI behaviors among adolescents.

Objective This study aims to explore the correlation between the grade of testicular tissue damage and clinical features which is used to evaluate the viability of testicular tissue after the orchidopexy for testicular torsion.Methods We conducted a retrospective,multicenter analysis of pathological specimens obtained from patients who under-went unilateral orchiectomy due to testicular torsion.Tissue viability was reassessed by the Mikuz grading scale.The corre-lations among tissue viability,duration of torsion symptoms,the angle of torsion,preoperative Doppler ultrasound image and intraoperative testicular resuscitation methods were analyzed.Results Eighty-five pathological specimens were en-rolled from 4 medical centers.The age range was 18 years(IQR 14-33),with a median torsion angle of 480°(IQR 180°-1080°).The interval from the torsion onset to surgical intervention was 18 hours(IQR 9-84).In the subgroup analysis of different torsion durations,the distribution of Grade I injuries was as follows.The subgroup within 12 hours ac-counted for 37.5％.The subgroup between 12 to 24 hours accounted for 14.28％.And the subgroup between 24 to 48 hours ac-counted for 6.45％.The subgroup exceeding 48 hours only accounted for 3.13％.Four patients received orchiectomy due to pre-operative ultrasound image indicating necrosis,correlating with Mikuz grade 3 tissue damage.The decision was made based on the failure of color recovery of affected testis in 57 cases,revealing 12.28％with grade 1 and 35.89％with grade Ⅱ damage.Twenty-four cases were identified as an Arda score of 3,with 4.16％showing grade 1 and 20.83％showing grade Ⅱ Mikuz damage.The method of Arda's grading could be used to determine the vitality of testicular tissue more accurately compared to the post-resusci-tation color observation method(P=0.032).Conclusion Pathological reassessment provides a precise determination of tis-sue viability in the affected testis.In some instances,the testicular tissue damage can be found potentially reversible.

Objective:To report the nationwide surveillance results of pathogenic profiles and antimicrobial resistance patterns of Gram-positive bloodstream infections in China in 2023.Methods:The clinical isolates of Gram-posttive bacteria from blood cultures were collected in member hospitals of National Bloodstream Infection Bacterial Resistant Investigation Collaborative System（BRICS）during January to December 2023. Antimicrobial susceptibility testing was performed using the dilution method recommended by the Clinical and Laboratory Standards Institute（CLSI）. Statistical analyses were conducted using WHONET 5.6 and SPSS 25.0 software.Results:A total of 4 385 Gram-positive bacterial isolates were obtained from 60 participating center. The top five pathogens were Staphylococcus aureus（ n=1 544，35.2%），coagulase-negative Staphylococci（ n=1 441，32.9%）， Enterococcus faecium（ n=574，13.1%）， Enterococcus faecalis（ n=385，8.8%），and α-hemolytic Streptococci（ n=187，4.3%）. The prevalence of methicillin-resistant Staphylococcus aureus（MRSA）and methicillin-resistant coagulase-negative Staphylococci（MRCNS）was 26.2%（405/1 544）and 69.8%（1 006/1 441），respectively. Notably，all Staphylococci remained susceptible to glycopeptide or daptomycin. Staphylococcus aureus demonstrated excellent susceptibility（>97.0%）to cephalobiol，rifampicin，trimethoprim-sulfamethoxazole，linezolid，minocycline，tigecycline，and eravacycline. No Enterococcus exhibiting resistance to linezolid were detected. Glycopeptide resistance was uncommon but more frequent in Enterococcus faecium（resistance to vancomycin and teicoplanin：both 1.7%）compared to Enterococcus faecalis（both 0.3%）. The detection rates of MRSA and MRCNS exhibited significant regional variations across the country（ χ2=17.674 and 148.650，respectively，both P<0.001）. No vancomycin-resistant Enterococci were detected in central China. Institutional comparison demonstrated higher prevalence of MRSA（ χ2=14.111， P<0.001）and MRCNS（ χ2=4.828， P=0.028）in provincial hospitals than that in municipal hospitals. Socioeconomic analysis identified elevated detection rates of both MRSA（ χ2=18.986， P<0.001）and MRCNS（ χ2=4.477， P=0.034）in less developed regions（per capita GDP

Objective:To report the results of bacterial resistant investigation collaborative system（BRICS）on the distribution and antimicrobial resistance profile of clinical Gram-negative bacteria isolates from bloodstream infections in China in 2023，and provide reference for clinical tretment of bloodstream infections and prevention and control of bacterial resistance.Methods:The clinical isolates of Gram-negative bacteria from blood cultures in member hospitals of BRICS were collected during January 2023 to December 2023. Antibiotic susceptibility tests were conducted by agar dilution or broth dilution methods recommended by Clinical and Laboratory Standards Institute（CLSI）. WHONET 5.6 and SPSS 25.0 were used to analyze the data.Results:During the study period，11 492 strains of Gram-negative bacteria were collected from 60 hospitals，of which 10 098（87.9%）were Enterobacterales and 1 394（12.1%）were non-fermentative bacteria. The top 5 bacterial species were Escherichia coli（50.0%）， Klebsiella pneumoniae（26.1%）， Pseudomonas aeruginosa（5.1%）， Acinetobacter baumannii complex（5.0%）and Enterobacter cloacae complex（4.1%）. The ESBL-producing rates in Escherichia coli， Klebsiella pneumoniae and Proteus mirablilis were 46.8%（2 685/5 741），18.3%（549/2 999）and 44.0%（77/175），respectively. The prevalence of carbapenem-resistant Escherichia coli（CREC）and carbapenem-resistant Klebsiella pneumoniae（CRKP）were 1.3%（76/5 741）and 15.0%（450/2 999）；32.9%（25/76）and 78.0%（351/450）of CREC and CRKP were sensitive to ceftazidime/avibactam combination，respectively. 94.7%（72/76）and 90.2%（406/450）of CREC and CRKP were sensitive to aztreonam/avibactam combination. Furthermore，57.9%（44/76）and 79.1%（356/450）were sensitive to imipenem/relebactam combination. The prevalence of carbapenem-resistant Acinetobacter baumannii（CRAB）complex was 64.6%（370/573），while more than 80.0% of CRAB complex was sensitive to tigecycline，eravacycline and polymyxin B. The prevalence of carbapenem-resistant Pseudomonas aeruginosa（CRPA）was 17.0%（99/581）. There were differences in the composition ratio of Gram-negative bacteria in bloodstream infections and the prevalence of important Gram-negative bacteria resistance among different regions in China，with statistically significant differences in the prevalence of CREC，CRKP，CRPA and CRAB complex（ χ2=10.6，28.6，10.8 and 19.3， P<0.05）. The prevalence of ESBL-producing Escherichia coli， CREC，CRAB complex and CRKP were higher in provincial hospitals than those in municipal hospitals（ χ2=12.5，9.8，12.7 and 57.8，all P<0.01）. Conclusions:Gram-negative bacteria are the main pathogens causing bloodstream infections in China，and Escherichia coli is ranked in the top，while the trend of Klebsiella pneumoniae increases continuously with time. CRKP infection shows a slow upward trend，CREC infecton maintains a low prevalence level，and CRAB complex infection continues to exhibit a high prevalence rate. The composition and resistance patterns of pathogens causing bloodstream infections vary to some extent across different regions and levels of hospitals in China.

Objective To construct a uricase-deficient mouse model with stable inheritance using the CRISPR/Cas9 system,and evaluate its ability to simulate the disease characteristics of patients with hyperuricemia.Methods Double single guide RNAs(sgRNAs)were designed on both sides of exon 2～4 of the Uox gene.sgRNA and Cas9 mRNA for gene knockout were microinjected into the fertilized eggs of mice.After culture for 2～4 h,the embryos were transferred to surrogate mother mice to produce an F0 generation.Uox-knockout mice were identified by polymerase chain reaction and sequencing analysis.Positive mice were then mated with wild-type(WT)mice to produce an F1 generation,and heterozygous female and male F1 mice were then selected to obtain homozygous F2 mice.Serum and urine levels of uric acid,creatinine,and urea,and serum alanine aminotransferase(ALT)and aspartate aminotransferase(AST)levels were detected and compared between homozygous and wild-type mice.Pathological changes in kidney and liver tissues were observed by hematoxylin and eosin and Masson staining.Results Urine levels of serum uric acid(male:(4116.8±1928.1)μmol/L,P＜0.001;female:(2998.0±547.7)μmol/L,P＜0.01)and serum levels of uric acid(male:(478.4±114.6)μmol/L,P＜0.001;female:(507.7±129.6)μmol/L,P＜0.001),creatinine((91.8±55.6)μmol/L,P＜0.001),urea((28.6±13.9)mmol/L,P＜0.05),ALT((53.3±23.3)U/L,P＜0.01),and AST((203.3±70.3)U/L,P＜0.001)were significantly increased in Uox-/-mice compared with WT mice.Histopathological examination showed moderate hepatocyte degeneration in the liver,moderate-to-severe tubular cystic dilation,degeneration,and fibrosis in the kidney,glomerular hypertrophy and hyperplasia,small-vessel dilation and congestion,and infiltration of stromal monocytes and lymphocytes in Uox-/-mice.Conclusions We successfully established a homozygous uricase-deficient mouse strain using CRISPR/Cas9 technology,as a suitable animal model for research in the field of hyperuricemia.

Objective To construct a uricase-deficient mouse model with stable inheritance using the CRISPR/Cas9 system,and evaluate its ability to simulate the disease characteristics of patients with hyperuricemia.Methods Double single guide RNAs(sgRNAs)were designed on both sides of exon 2～4 of the Uox gene.sgRNA and Cas9 mRNA for gene knockout were microinjected into the fertilized eggs of mice.After culture for 2～4 h,the embryos were transferred to surrogate mother mice to produce an F0 generation.Uox-knockout mice were identified by polymerase chain reaction and sequencing analysis.Positive mice were then mated with wild-type(WT)mice to produce an F1 generation,and heterozygous female and male F1 mice were then selected to obtain homozygous F2 mice.Serum and urine levels of uric acid,creatinine,and urea,and serum alanine aminotransferase(ALT)and aspartate aminotransferase(AST)levels were detected and compared between homozygous and wild-type mice.Pathological changes in kidney and liver tissues were observed by hematoxylin and eosin and Masson staining.Results Urine levels of serum uric acid(male:(4116.8±1928.1)μmol/L,P＜0.001;female:(2998.0±547.7)μmol/L,P＜0.01)and serum levels of uric acid(male:(478.4±114.6)μmol/L,P＜0.001;female:(507.7±129.6)μmol/L,P＜0.001),creatinine((91.8±55.6)μmol/L,P＜0.001),urea((28.6±13.9)mmol/L,P＜0.05),ALT((53.3±23.3)U/L,P＜0.01),and AST((203.3±70.3)U/L,P＜0.001)were significantly increased in Uox-/-mice compared with WT mice.Histopathological examination showed moderate hepatocyte degeneration in the liver,moderate-to-severe tubular cystic dilation,degeneration,and fibrosis in the kidney,glomerular hypertrophy and hyperplasia,small-vessel dilation and congestion,and infiltration of stromal monocytes and lymphocytes in Uox-/-mice.Conclusions We successfully established a homozygous uricase-deficient mouse strain using CRISPR/Cas9 technology,as a suitable animal model for research in the field of hyperuricemia.

Objective This study aims to explore the correlation between the grade of testicular tissue damage and clinical features which is used to evaluate the viability of testicular tissue after the orchidopexy for testicular torsion.Methods We conducted a retrospective,multicenter analysis of pathological specimens obtained from patients who under-went unilateral orchiectomy due to testicular torsion.Tissue viability was reassessed by the Mikuz grading scale.The corre-lations among tissue viability,duration of torsion symptoms,the angle of torsion,preoperative Doppler ultrasound image and intraoperative testicular resuscitation methods were analyzed.Results Eighty-five pathological specimens were en-rolled from 4 medical centers.The age range was 18 years(IQR 14-33),with a median torsion angle of 480°(IQR 180°-1080°).The interval from the torsion onset to surgical intervention was 18 hours(IQR 9-84).In the subgroup analysis of different torsion durations,the distribution of Grade I injuries was as follows.The subgroup within 12 hours ac-counted for 37.5％.The subgroup between 12 to 24 hours accounted for 14.28％.And the subgroup between 24 to 48 hours ac-counted for 6.45％.The subgroup exceeding 48 hours only accounted for 3.13％.Four patients received orchiectomy due to pre-operative ultrasound image indicating necrosis,correlating with Mikuz grade 3 tissue damage.The decision was made based on the failure of color recovery of affected testis in 57 cases,revealing 12.28％with grade 1 and 35.89％with grade Ⅱ damage.Twenty-four cases were identified as an Arda score of 3,with 4.16％showing grade 1 and 20.83％showing grade Ⅱ Mikuz damage.The method of Arda's grading could be used to determine the vitality of testicular tissue more accurately compared to the post-resusci-tation color observation method(P=0.032).Conclusion Pathological reassessment provides a precise determination of tis-sue viability in the affected testis.In some instances,the testicular tissue damage can be found potentially reversible.

Objective:To establish and optimize a novel method, focus forming assay (FFA), for quantitation of influenza A virus (FluA) and compare its application performance with traditional plague forming assay (PFA).Methods:The foci chromogenic effects of three peroxidase substrates in immunostaining were compared. The PFA and FFA methods were used to explore FluA incubation times and plaque morphology on 12-well plates, and to determine optimal incubation times and virus adsorption volumes for different FluA subtypes on 96-well plates. The correlation between FFA and PFA was evaluated, and the optimized FFA was applied to the in vitro antiviral efficacy analysis of Favipiravir and neutralization test against different subtypes of FluA. Results:TRUEBLUE substrate was identified as the optimal substrate for foci visualization. Compared with the PFA, the FFA showed improved sensitivity and reduced detection time in FluA titration, and good correlation was shown between the two methods′ results. By replacing the 96-well plate with the 12-well plate for FFA titration of different subtypes of FluA, the detection time was shortened, and the amount of serum samples used could be further reduced by optimizing the virus adsorption volume. The half-maximal effective concentration of favipiravir against influenza viruses assessed by the FFA and PFA methods showed no significant difference, and was consistent with the results obtained from quantitative PCR. Additionally, the focus reduction neutralization test and hemagglutination inhibition assays demonstrated strong correlation in determining antibody titers against FluA in serum neutralization assays.Conclusions:The improved FFA method developed here provides a more efficient experimental tool for FluA titration, antiviral drug screening and broad-spectrum vaccine evaluation.

Objective:To report the nationwide surveillance results of pathogenic profiles and antimicrobial resistance patterns of Gram-positive bloodstream infections in China in 2023.Methods:The clinical isolates of Gram-posttive bacteria from blood cultures were collected in member hospitals of National Bloodstream Infection Bacterial Resistant Investigation Collaborative System（BRICS）during January to December 2023. Antimicrobial susceptibility testing was performed using the dilution method recommended by the Clinical and Laboratory Standards Institute（CLSI）. Statistical analyses were conducted using WHONET 5.6 and SPSS 25.0 software.Results:A total of 4 385 Gram-positive bacterial isolates were obtained from 60 participating center. The top five pathogens were Staphylococcus aureus（ n=1 544，35.2%），coagulase-negative Staphylococci（ n=1 441，32.9%）， Enterococcus faecium（ n=574，13.1%）， Enterococcus faecalis（ n=385，8.8%），and α-hemolytic Streptococci（ n=187，4.3%）. The prevalence of methicillin-resistant Staphylococcus aureus（MRSA）and methicillin-resistant coagulase-negative Staphylococci（MRCNS）was 26.2%（405/1 544）and 69.8%（1 006/1 441），respectively. Notably，all Staphylococci remained susceptible to glycopeptide or daptomycin. Staphylococcus aureus demonstrated excellent susceptibility（>97.0%）to cephalobiol，rifampicin，trimethoprim-sulfamethoxazole，linezolid，minocycline，tigecycline，and eravacycline. No Enterococcus exhibiting resistance to linezolid were detected. Glycopeptide resistance was uncommon but more frequent in Enterococcus faecium（resistance to vancomycin and teicoplanin：both 1.7%）compared to Enterococcus faecalis（both 0.3%）. The detection rates of MRSA and MRCNS exhibited significant regional variations across the country（ χ2=17.674 and 148.650，respectively，both P<0.001）. No vancomycin-resistant Enterococci were detected in central China. Institutional comparison demonstrated higher prevalence of MRSA（ χ2=14.111， P<0.001）and MRCNS（ χ2=4.828， P=0.028）in provincial hospitals than that in municipal hospitals. Socioeconomic analysis identified elevated detection rates of both MRSA（ χ2=18.986， P<0.001）and MRCNS（ χ2=4.477， P=0.034）in less developed regions（per capita GDP