Objective:To investigate the composition of immune cells and fibroblastic reticular cells (FRCs) in the lymph nodes (LNs) follicles of human immunodeficiency virus (HIV)-infected individuals with varying immune statuses, and their association with HIV replication.Methods:Neck LNs samples were collected from 4 treatment-naive, newly diagnosed HIV-infected individuals with diverse immune statuses. RNA in situ sequencing was employed, with imaging achieved via rolling circle amplification and fluorescence labeling. By integrating cell segmentation and nuclear staining, single-cell data from up to one hundred thousand cells were generated per paraffin tissue section. Using lymphoid follicles as the unit of analysis, compositional changes in immune cells and FRCs were characterized, and their correlations with viral replication were evaluated. Results:The peripheral blood CD4 + T cell counts of samples LN_1, LN_2, LN_3, and LN_4 exhibited a sequential decrease. A total of 31, 15, 16, and 18 structurally intact follicles were identified in each sample, respectively. In the follicles of LN_1, the proportion of HIV-replicating cells positively correlated with cDCs abundance ( R2=0.2, P=0.011), and HIV RNA signals were spatially colocalized with cDCs and FRCs. In the follicles of LN_2, HIV RNA molecules showed preferential enrichment within FRCs. In sample LN_3, HIV RNA enrichment was observed in both cDCs and CD4 + T cells. In sample LN_4, the proportion of cells with HIV replication was positively correlated with the proportions of the following cells: cDCs ( R2=0.38, P=0.006 4), CD4 + T cells ( R2=0.28, P=0.025), and FRCs ( R2=0.26, P=0.029), and HIV RNA molecules were detected in cDCs, CD4 + T cells, and FRCs. LN_1 and LN_2 samples showed a trend toward negative correlation between HIV-replicating cell proportion and CD8 + T cells proportion. LN_4 sample demonstrated a significant positive correlation between HIV-replicating cell proportion and CD8 + T cells proportion ( R2=0.23, P=0.046). Conclusions:RNA in situ sequencing technology reveals unique distribution patterns of immune cells and viral replication in LNs follicles of HIV-infected individuals. The follicular immune microenvironment exhibits distinct characteristics associated with peripheral blood CD4 + T cell counts, providing novel insights into the spatial dynamics of HIV persistence and immune cell interactions during infection.

Objective To explore strategies for further subdivision of DRG in gastrointestinal surgery cases,providing references to enhance the differentiation of DRG subgroups.Methods A total of 5 108 gastrointestinal surgery cases were selected from a tertiary grade A hospital and a tertiary hospital in Hubei Province,spanning from January 2019 to June 30,2023,and another secondary hospital's data from 2020 and 2021.It employs single factor analysis and multiple linear regression analysis to identify factors influencing case costs.Additionally,the opinions of nine clinicans were gathered regarding factors affecting resource consumption in gastrointestinal surgery cases.The four selected case groups were further subdivided considers the peak characteristics of disease costs.It compares subdivided groups with the DRG Payment Subgroups Scheme(Version 2.0).Results Groups GB1,GB2,GC1,and GC2 were subdivided into 7,4,7,and 6 DRG groups,respectively.The coefficient of variation of each subdivided DRG were reduced,homogeneity was increased,and inter-group differentiation was increased.The results were consistent with the DRG Payment Subgroups Scheme(Version 2.0).Conclusion Based on DRG grouping,the DRG groups can be further subdivided according to the peak characteristics presented by case costs.This subdivision strategy is helpful to provide new ideas for case grouping of Medicare payment.

Objective To explore strategies for further subdivision of DRG in gastrointestinal surgery cases,providing references to enhance the differentiation of DRG subgroups.Methods A total of 5 108 gastrointestinal surgery cases were selected from a tertiary grade A hospital and a tertiary hospital in Hubei Province,spanning from January 2019 to June 30,2023,and another secondary hospital's data from 2020 and 2021.It employs single factor analysis and multiple linear regression analysis to identify factors influencing case costs.Additionally,the opinions of nine clinicans were gathered regarding factors affecting resource consumption in gastrointestinal surgery cases.The four selected case groups were further subdivided considers the peak characteristics of disease costs.It compares subdivided groups with the DRG Payment Subgroups Scheme(Version 2.0).Results Groups GB1,GB2,GC1,and GC2 were subdivided into 7,4,7,and 6 DRG groups,respectively.The coefficient of variation of each subdivided DRG were reduced,homogeneity was increased,and inter-group differentiation was increased.The results were consistent with the DRG Payment Subgroups Scheme(Version 2.0).Conclusion Based on DRG grouping,the DRG groups can be further subdivided according to the peak characteristics presented by case costs.This subdivision strategy is helpful to provide new ideas for case grouping of Medicare payment.

Objective:To investigate the composition of immune cells and fibroblastic reticular cells (FRCs) in the lymph nodes (LNs) follicles of human immunodeficiency virus (HIV)-infected individuals with varying immune statuses, and their association with HIV replication.Methods:Neck LNs samples were collected from 4 treatment-naive, newly diagnosed HIV-infected individuals with diverse immune statuses. RNA in situ sequencing was employed, with imaging achieved via rolling circle amplification and fluorescence labeling. By integrating cell segmentation and nuclear staining, single-cell data from up to one hundred thousand cells were generated per paraffin tissue section. Using lymphoid follicles as the unit of analysis, compositional changes in immune cells and FRCs were characterized, and their correlations with viral replication were evaluated. Results:The peripheral blood CD4 + T cell counts of samples LN_1, LN_2, LN_3, and LN_4 exhibited a sequential decrease. A total of 31, 15, 16, and 18 structurally intact follicles were identified in each sample, respectively. In the follicles of LN_1, the proportion of HIV-replicating cells positively correlated with cDCs abundance ( R2=0.2, P=0.011), and HIV RNA signals were spatially colocalized with cDCs and FRCs. In the follicles of LN_2, HIV RNA molecules showed preferential enrichment within FRCs. In sample LN_3, HIV RNA enrichment was observed in both cDCs and CD4 + T cells. In sample LN_4, the proportion of cells with HIV replication was positively correlated with the proportions of the following cells: cDCs ( R2=0.38, P=0.006 4), CD4 + T cells ( R2=0.28, P=0.025), and FRCs ( R2=0.26, P=0.029), and HIV RNA molecules were detected in cDCs, CD4 + T cells, and FRCs. LN_1 and LN_2 samples showed a trend toward negative correlation between HIV-replicating cell proportion and CD8 + T cells proportion. LN_4 sample demonstrated a significant positive correlation between HIV-replicating cell proportion and CD8 + T cells proportion ( R2=0.23, P=0.046). Conclusions:RNA in situ sequencing technology reveals unique distribution patterns of immune cells and viral replication in LNs follicles of HIV-infected individuals. The follicular immune microenvironment exhibits distinct characteristics associated with peripheral blood CD4 + T cell counts, providing novel insights into the spatial dynamics of HIV persistence and immune cell interactions during infection.

OBJECTIVE To establish the fingerprints of Kangfuyan capsules and carry out chemical pattern recognition analysis，and simultaneously determine the contents of five components so as to promote the quality standard of the drug. METHODS High performance liquid chromatography （HPLC）fingerprints of 11 batches of Kangfuyan capsules （S1-S11）were established by Similarity Evaluation System of TCM Chromatographic Fingerprint （2012 edition）；identification and attribution analysis of chromatographic peaks were carried out by comparison with the chromatograms of the reference substance and the decoction pieces of single ingredient. SPSS 26.0 and SIMCA 14.1 software were used for cluster analysis and principal component analysis. HPLC method was used to determine the contents of matrine ，phellodendrine chloride ，rutin，forsythoside A and berberine hydrochloride. RESULTS There were 29 common peaks in the fingerprints for 11 batches of samples ，and the similarity was higher than 0.99. A total of 5 chromatographic peaks were identified ，which are matrine （peak 3），phellodendron chloride （peak 14），rutin （peak 20），forsythiaside A （peak 22） and berberine hydrochloride （peak 28）. The results of cluster analysis and principal component analysis showed that S 1-S9 were clustered into one category ，and S 10 and S 11 were clustered into another category. The contents of above 5 components were 29.320 5-60.144 3，0.621 6-1.076 6，1.025 9-2.830 5，2.899 3-6.212 7 and 4.425 1-8.581 6 mg/g， respectively. CONCLUSIONS The established fingerprint and content determination method are stable and reliable ，and can provide reference for the quality control of the preparation in combination with chemical pattern recognition analysis.

Abstract As a new type of pollutant, microplastics have attracted extensive attention. Children in a critical stage of growth and development are vulnerable to microplastics. Summarzing the relevant laws and regulations and the source of microplastics, the paper demonstrates the ways of microplastics entering human body, some toxic effects of microplastics found in recent experimental studies and their potential hazards to children s health are introduced in detail.

Objective:To analyze the distribution of clinically isolated fungal strains and their resistance to common antifungal drugs in Shandong province.Methods:Through the Shandong Children’s Bacterial & Fungal Drug Resistance Surveillance and Research Collaborative Network, a total of 1 030 fungi were collected in 46 hospitals of Shandong province from January 1 to December 31, 2018. The source and type of strains were analyzed, and antifungal drug sensitivity tests were performed by using the micro-dilution method. Whonet 5.6 and SPSS 22.0 were applied to analyze the data.Results:The overall main strains were Candida albicans (38.74%, 399/1 030), Candida tropicalis (16.99%, 175/1 030) and Candida parapsilosis (16.41%, 169/1 030); the main fungi strains in child patients were C. albicans (52.50%, 63/120), C. parapsilosis (12.50%, 15/120) and C. tropicalis (9.17%, 11/120); the main fungi strains in adult patients were C. albicans (36.37%, 331/910), C. tropicalis (17.03%, 155/910) and C. parapsilosis (15.27%, 139/910). The isolation rate of main Candida strains from January to March and August to December was much higher than that of other months. The drug resistance rates of C. albicans to fluconazole and voriconazole were 7.14% and 7.43%, respectively, and the drug resistance rates to itraconazole were 50.44%. The resistance rates of C. tropicalis to fluconazole, voriconazole and itraconazole were 29.05%, 23.29% and 48.65%, respectively. The sensitivity rates of C. parapsilosi to fluconazole, voriconazole and itraconazole were 93.06%, 93.75% and 94.44%, respectively. Candida glabrata showed a dose-dependent sensitivity rate of 2.33% to fluconazole. Analysis of 244 blood fungi strains showed that non-candida albicans bacteremia accounted for 70.08%. In the pathogen spectrum covering 92.22%, fluconazole was sensitive to 64.65% of the pathogens, voriconazole was 68.88%, and amphotericin B was 88.75%. After quantification, the effective rates of fluconazole, voriconazole and amphotericin B in the clinical treatment of fungal bacteremia were 70.10%, 74.69% and 96.23%, respectively. Among them, the sensitivity rate of voriconazole to C. tropicalis was lower than that of fluconazole. Conclusions:Candida is the main clinical fungus isolates in hospitals of Shandong province. The resistance rate of C. tropicalis to azole antifungal drugs is on the rise, and the sensitivity of other Candida species to clinically used antifungal drugs is basically stable.

Objective:To explore the characteristics of long-lived memory CD8 + T cells and its relationship with disease progression in HIV-1 infected patients. Methods:Twenty-six treatment-na?ve and 26 antiretroviral therapy (ART)-treated HIV-1 infected patients, as well as 11 healthy controls were recruited in this study. Peripheral blood mononuclear cells were isolated and analyzed by flow cytometry to detect the frequency and function of long-lived memory CD8 + T cells. Results:The frequency of long-lived memory CD8 + T cells decreased significantly in treatment-na?ve patients compared with that in healthy controls, and was not fully restored in ART-treated group. In addition, the frequency of long-lived memory CD8 + T cells is positively correlated with CD4 + T cell count and CD4/CD8 ratio, while negatively correlated with HIV-1 viral load. Conclusions:The disease progression in chronic HIV-1 infected individuals is accompanied by continuous damage to long-lived memory CD8 + T cell.

Objective:To analyze the molecular characteristics and antibiotic susceptibility of two strains of Nocardia farcinica isolated from patients with joint infection using whole genome sequencing. Methods:Two strains of Nocardia farcinica causing knee-joint infections in two elderly patients were collected in January 2020. Whole genome sequencing was used to determine the nocardia species. Drug sensitivity test was performed using the micro-broth dilution and E-test method according to CLSI M24 guideline. ABRicate was used to analyze drug resistance and virulence genes. Snippy and other bioinformatic tools were used for genomic comparison, and to construct SNP homologous tree. Results:The clinical isolates in this study were both Nocardia farcinica. Antimicrobial susceptibility test showed the isolates were resistant to ceftriaxone, cefepime, cefotaxime and trimethoprim/sulfamethoxazole (TMP/SMX). Imipenem, linezolid and amoxicillin-clavulanic acid showed good activity. Four antibiotic resistance genes including class A β-lactamase gene far-1, RNA polymerase binding protein gene RbpA, multi-drug resistance efflux pump transcription activator gene MtrA and regulatory transcription factor gene vanR-O were identified in the Nocardia farcinica genomes, which conferred resistance to beta-lactams, rifampicin, macrolides and vancomycin respectively. No acquired TMP/SMX resistance genes were identified. There are multiple missense mutations in the dihydrofolate reductase family genes. Four virulence genes of icl, mbtH, phoP,and relAthat are homologous to Mycobacterium tuberculosis were found. SNP homologous tree analysis showed the two Nocardia strains were closely related, and there were only ten SNP sites, six compound substitutions and one deletion mutation between them. Conclusions:Whole genome sequencing technology is helpful to explore the molecular characteristics and resistance mechanisms of Nocardia species. Nocardia farcinica has a trend of spreading in China. Resistance to TMP/SMX is worthy of attention. The mutation of genes involved in the metabolic pathway of dihydrofolate might be one of multiple TMP/SMX resistance mechanisms.

Objective:To analyze the molecular characteristics and antibiotic susceptibility of two strains of Nocardia farcinica isolated from patients with joint infection using whole genome sequencing. Methods:Two strains of Nocardia farcinica causing knee-joint infections in two elderly patients were collected in January 2020. Whole genome sequencing was used to determine the nocardia species. Drug sensitivity test was performed using the micro-broth dilution and E-test method according to CLSI M24 guideline. ABRicate was used to analyze drug resistance and virulence genes. Snippy and other bioinformatic tools were used for genomic comparison, and to construct SNP homologous tree. Results:The clinical isolates in this study were both Nocardia farcinica. Antimicrobial susceptibility test showed the isolates were resistant to ceftriaxone, cefepime, cefotaxime and trimethoprim/sulfamethoxazole (TMP/SMX). Imipenem, linezolid and amoxicillin-clavulanic acid showed good activity. Four antibiotic resistance genes including class A β-lactamase gene far-1, RNA polymerase binding protein gene RbpA, multi-drug resistance efflux pump transcription activator gene MtrA and regulatory transcription factor gene vanR-O were identified in the Nocardia farcinica genomes, which conferred resistance to beta-lactams, rifampicin, macrolides and vancomycin respectively. No acquired TMP/SMX resistance genes were identified. There are multiple missense mutations in the dihydrofolate reductase family genes. Four virulence genes of icl, mbtH, phoP,and relAthat are homologous to Mycobacterium tuberculosis were found. SNP homologous tree analysis showed the two Nocardia strains were closely related, and there were only ten SNP sites, six compound substitutions and one deletion mutation between them. Conclusions:Whole genome sequencing technology is helpful to explore the molecular characteristics and resistance mechanisms of Nocardia species. Nocardia farcinica has a trend of spreading in China. Resistance to TMP/SMX is worthy of attention. The mutation of genes involved in the metabolic pathway of dihydrofolate might be one of multiple TMP/SMX resistance mechanisms.