Liver cancer is one of the most common malignant tumors globally, primarily including hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (ICC). To date, it lacks of an effective liquid biopsy for differential diagnosis. The direct test of tumor-derived small extracellular vesicles (sEVs) via sEV membrane proteins in serum can provide a rapid way to clinical diagnosis of liver cancer. However, the mainstream optical label techniques for quickly detecting serum protein biomarkers simultaneously target sEVs membrane proteins and homologous free proteins, which lowers diagnostic sensitivity and specificity considerably. In this work, we propose using meta-plasmonic biosensors to detect serum sEVs membrane proteins directly for liver cancer diagnosis, and evaluate the influences of homologous free proteins on testing results through a comprehensive biophysical modeling and experimental verifications. Our results show that the meta-plasmonic approach minimizes interference from homologous free proteins owing to its label-free optical readout, and demonstrates the exceptional sensitivity and specificity to distinguish the health control, benign liver disease, HCC, and ICC via the specific sEV membrane proteins of CD63 and GPC3. Our findings indicate that tumor-derived sEVs in serum can be detected directly by meta-plasmonic assays. Moreover, the proposed method for liver cancer detection could be also adaptable to other cancer diagnostics.

ABSTRACT Background Hyperammonemia is a central feature of hepatic encephalopathy (HE) and contributes to astrocytic dysfunction; however, the mechanisms underlying ammonia‐induced cytoskeletal remodeling remain unclear. This study aimed to identify key molecular mediators of ammonia‐induced astrocyte injury and to elucidate the role of ACTA2 in this process. Methods We established an in vitro HE model using the human astrocyte line SVG p12 treated with either ammonium chloride (NH 4 Cl) (5–20 mM) or serum from patients with HE. Cellular responses were evaluated by Cell Counting Kit‐8 assay and flow cytometry. RNA sequencing was performed to identify key regulatory genes, followed by molecular and functional validation of the lead candidate, ACTA2 , which encodes alpha‐smooth muscle actin ( α ‐SMA), using knockdown and overexpression strategies. Filamentous actin organization was assessed by phalloidin staining and fluorescence microscopy. The expressions of water channel (aquaporin‐4) and junction‐associated proteins (zonula occludens‐1, occludin, claudin‐5) were measured to characterize astrocytic junctional protein alterations. Results NH 4 Cl exposure effectively recapitulated the cytotoxic effects of serum from patients with HE, revealing a distinct biphasic response: low‐dose ammonia (5 mM) transiently boosted metabolic activity, whereas higher doses (10–20 mM) reduced viability, promoted apoptosis, and caused G2/M phase arrest. RNA‐seq, cytoskeletal analysis, and biochemical validation identified ACTA2 downregulation as a sentinel event in ammonia‐induced injury. Functional assays demonstrated that ACTA2 deficiency phenocopied NH 4 Cl‐induced cytotoxicity, while its restoration significantly mitigated apoptosis and cell cycle arrest. Ammonia treatment led to upregulation of aquaporin‐4 alongside altered expression of junctional proteins, marked by diminished zonula occludens‐1 and claudin‐5 expression and elevated occludin levels. Mechanistically, NH 4 Cl reduced the phosphorylation of FAK (Tyr397) and proto‐oncogene tyrosine‐protein kinase Src (c‐Src) (Tyr416), whereas ACTA2 overexpression in NH 4 Cl‐treated cells restored their activation, implicating suppression of the α ‐SMA‐FAK/c‐Src signaling axis in ammonia‐induced astrocytic injury. Conclusion This study demonstrates that NH 4 Cl disrupts cell cytoskeletal stability by dose‐dependently downregulating ACTA2 and suppressing the downstream FAK/c‐Src pathway, thereby affecting the viability and fate of SVG p12 cells and inducing molecular alterations associated with HE. This provides new mechanistic insights into understanding ammonia‐induced astrocytic dysfunction.

BACKGROUND:Identifying significant liver fibrosis (≥F2) is critical for initiating antiviral therapy in chronic hepatitis B (CHB). Chitinase-3-like protein 1 (CHI3L1) is a promising fibrosis biomarker, but its staging value remains unclear. We evaluated CHI3L1 (chemiluminescent immunoassay, CLIA) alone and with machine learning (ML) for differentiating mild (F0-F1) from significant (F2-F4) fibrosis. METHODS:We included 347 participants (303 primary, 44 external validation), most CHB patients with biopsy-confirmed METAVIR staging. Serum markers were measured, and 5 ML models were developed. RESULTS:CHI3L1 levels increased progressively with fibrosis severity and discriminated significant fibrosis (≥F2), with AUC 0.84 (95% CI 0.772-0.908) in the overall CHB cohort and 0.839 (95% CI 0.755-0.924) in CHB patients with normal ALT, outperforming conventional biomarkers. Incorporating CHI3L1 into ML further improved diagnostic performance; the Decision Tree model achieved optimal accuracy AUCs of 0.923 [95% CI 0.891-0.976] and 0.895 [95% CI 0.831-0.960], respectively. SHapley Additive exPlanations (SHAP) analysis identified CHI3L1 as the most influential feature (62.2%) in this model. The model performed consistently in the external validation cohort. CONCLUSIONS:CHI3L1 is a strong noninvasive biomarker for identifying clinically significant fibrosis in CHB. Integration with ML enhances fibrosis staging accuracy, supporting timely antiviral therapy.

Accurate subtyping of lung cancer is crucial for developing personalized treatment plans and improving patient outcomes. This study established machine learning models for lung cancer subtyping by integrating multidimensional hematological indicators, offering advantages such as non-invasiveness, repeatability, and the capability for dynamic disease monitoring. The study utilized data from 771 lung cancer patients for model construction and validation, and an additional 510 independent cases were collected to assess the clinical applicability of the models. Ten supervised learning algorithms were screened, ultimately identifying the XGBoost model as optimal for differentiating small cell lung cancer from non-small cell lung cancer, and the Random Forest model as optimal for distinguishing lung squamous cell carcinoma from lung adenocarcinoma. In the independent clinical validation cohorts from two centers, these two models achieved accuracies of 95% and 91%, respectively, demonstrating good clinical applicability and serving as a valuable complement to pathological biopsy.

Outer membrane vesicles (OMVs) are nanoscale structures naturally released by Gram-negative bacteria and play important roles in host-pathogen interactions. In Acinetobacter baumannii, a critical multidrug-resistant pathogen, OMVs contribute to virulence, immune modulation, and antibiotic resistance, while also representing promising vaccine candidates. In this review, we summarize current advances in the biogenesis, pathogenic functions, and vaccine applications of A. baumannii OMVs. We discuss the major mechanisms proposed for OMV biogenesis, factors regulating OMV production, and the diverse roles of OMVs in biofilm formation, resistance dissemination, and host immune responses. We further evaluate recent progress in native, detoxified, and engineered OMV-based vaccine platforms, with particular emphasis on endotoxin detoxification, antigen selection, and strategies for improving OMV yield and immunogenicity. We also compare the current status of A. baumannii OMV vaccines with the successful clinical development of meningococcal OMV vaccines and highlight the major challenges that remain for clinical translation. Collectively, this review provides an updated perspective on the opportunities and limitations of OMV-based interventions and outlines future directions for the development of safe, standardized, and broadly protective vaccines against A. baumannii.

BACKGROUND:Pancreatic ductal adenocarcinoma (PDAC) is difficult to detect early and highly aggressive, often leading to poor patient prognosis. Existing serum biomarkers like CA19-9 are limited in early diagnosis, failing to meet clinical needs. Machine learning (ML)/deep learning (DL) technologies have shown great potential in biomedicine. This study aims to establish PDAC differential diagnosis and prognosis assessment models using ML combined with serum biomarkers for early diagnosis, risk stratification, and personalized treatment recommendations, improving early diagnosis rates and patient survival. METHODS:The study included serum biomarker data and prognosis information from 117 PDAC patients. ML models (Random Forest (RF), Neural Network (NNET), Support Vector Machine (SVM), and Gradient Boosting Machine (GBM)) were used for differential diagnosis, evaluated by accuracy, Kappa test, ROC curve, sensitivity, and specificity. COX proportional hazards model and DeepSurv DL model predicted survival risk, compared by C-index and Log-rank test. Based on DeepSurv's risk predictions, personalized treatment recommendations were made and their effectiveness assessed. RESULTS:Effective PDAC diagnosis and prognosis models were built using ML. The validation set data shows that the accuracy of the RF, NNET, SVM, and GBM models are 84.21%, 84.21%, 76.97%, and 83.55%; the sensitivity are 91.26%, 90.29%, 89.32%, and 88.35%; and the specificity are 69.39%, 71.43%, 51.02%, and 73.47%. The Kappa values are 0.6266, 0.6307, 0.4336, and 0.6215; and the AUC are 0.889, 0.8488, 0.8488, and 0.8704, respectively. BCAT1, AMY, and CA12-5 were selected as modeling parameters for the prognosis model using COX regression. DeepSurv outperformed the COX model on both training and validation sets, with C-indexes of 0.738 and 0.724, respectively. The Kaplan-Meier survival curves indicate that personalized treatment recommendations based on DeepSurv can help patients achieve survival benefits. CONCLUSION:This study built efficient PDAC diagnosis and prognosis models using ML, improving early diagnosis rates and prognosis accuracy. The DeepSurv model excelled in prognosis prediction and successfully guided personalized treatment recommendations and supporting PDAC clinical management.

Ricin toxin (RT), a highly potent plant-derived toxin, represents a critical threat due to its capacity to induce fatal acute lung injury (ALI) upon inhalation. While the epidermal growth factor receptor (EGFR), a receptor tyrosine kinase predominantly expressed on epithelial cells and fibroblasts, regulates cellular processes such as growth, proliferation, differentiation and inflammation, its involvement in RT-induced ALI remains unexplored. This study investigates this relationship using a mouse model of ALI induced by aerosolized RT at a dose of 2.0 × LD50 (approximately 0.01 mg kg −1). The results demonstrate that damage to alveolar epithelial type II (AT2) cells leads to the release of heparin-binding epidermal growth factor-like growth factor (HB-EGF), which activates EGFR on fibroblasts, exacerbating lung injury pathology and reducing survival. Mechanistically, EGFR activation in fibroblasts induces the early growth response protein 1 (EGR1), which subsequently enhances chemokine C-X-C motif ligand 1 (CXCL1) secretion 24 h post-exposure, promoting neutrophil infiltration in the lung. RNA sequencing analysis corroborates these findings. Notably, pharmacological inhibition of EGFR phosphorylation using Erlotinib (ERL) significantly mitigates the inflammatory response in RT-induced ALI. These results not only illuminate the immune response in lung tissue but also highlight EGFR signaling in fibroblasts as a pivotal mediator of RT-induced ALI. This study identifies a novel therapeutic strategy targeting EGFR signaling in fibroblasts for the treatment of inflammatory lung diseases.

Nanoplasmonic metasurface technology, known for its high sensitivity, has garnered significant attention in the field of cancer detection. However, its potential is currently hindered by the inefficient data processing and analysis of conventional biosensing approaches. Herein, a biosensing strategy based on the Kolmogorov-Arnold network (KAN)-enabled metasurface chip (metaEVchip) for ultrasensitive small extracellular vesicles (sEV) analysis in serum is proposed. By analyzing full-spectrum data from 600 pancreatic ductal adenocarcinoma (PDAC) patients and 1200 controls via KAN-powered deep learning nanoplasmonic biosensing, the strategy achieves an exceptional area under the curve (AUC) of 0.99 in an external validation set, outperforming traditional methods. Further exploration of this enhanced performance reveals KAN's mechanism for the simultaneous capture of multi-dimensional spectral features, an advantage that enables efficient data processing and accuracy. This advancement significantly expands the applicability of nanoplasmonic metasurfaces in biosensing and establishes a new paradigm for cancer screening and improved clinical management of multiple malignancies.

ABSTRACT To explore the influence of storage temperature and time on the stability of different concentrations of hepatitis C virus nucleic acid (HCV RNA) samples and to provide data reference for laboratory quality control. Serum samples of 10 patients with HCV RNA detection quantitation of 10 6 –10 8 IU/mL were collected. The samples of each patient were diluted into three concentrations: high, medium, and low. Then the samples of each concentration were divided into 21, which were divided into three groups according to the storage conditions of −20°C, 4°C, and 25°C, with seven samples in each group. The samples were selected from each group for quantitative detection of HCV RNA on day 0, day 1, day 3, day 5, day 7, day 14, and day 30. The results of each concentration and storage temperature sample remained stable within 5 days. Based on the mixed-effect linear model, the main effects of temperature, time, and concentration were statistically significant ( P < 0.01). There was an interaction effect between concentration and time ( P = 0.0448), and there was also an interaction effect between temperature and time ( P < 0.01). There was no interaction effect between concentration and temperature ( P = 0.11) or between concentration, temperature, and time ( P = 0.90). The results of serum samples with different concentrations of the HCV RNA remained stable within 5 days. The lower the initial concentration of HCV RNA serum sample, the worse the stability; the higher the storage temperature, the worse the stability. If conditions permit, the laboratory should store such samples at −20°C. IMPORTANCE Previously, there were few reports about the influence of different concentrations of sample nucleic acid on the stability of samples at various temperatures and times in various literatures. Therefore, it is necessary to analyze the influence of concentration factors on the stability of samples and test results at different storage times and temperatures. This study took the concentration of hepatitis C virus nucleic acid as the research object to further understand the stability of hepatitis C virus nucleic acid test samples under various storage conditions, to provide data reference for the treatment of hepatitis C virus nucleic acid and RNA test samples before clinical laboratory test, and provide guidance and help for the improvement of laboratory quality control.

Purpose: To investigate the genetic diversity of IncG plasmids, we have proposed a typing scheme based on replicon repA and performed comparative genomic analysis of five IncG plasmids from China. Methods: p30860-KPC, p116965-KPC, pA1705-KPC, pA1706-KPC and pNY5520-KPC total in five IncG plasmids from clinical isolates of Pseudomonas and Enterobacteriaceae, respectively, were fully sequenced and were compared with the previously collected Results: Based on phylogeny, IncG-type plasmids are divided into IncG-I to IncG-VIII, the five plasmids belong to IncG-VIII. A detailed sequence comparison was then presented that the IncG plasmid involved accessory region I (Tn5563a/b/c/d/e), accessory region II (ISpa19), and accessory region III (blaKPC-2-region). Expect for the pNY5520-KPC, the rest of the plasmids had the same backbone structure as the reference one. Within the plasmids, insertion sequences Tn5563d and Tn5563e were identified, a novel unknown insertion region was found in Tn5563b/c/d/e. In addition, Tn6376b and Tn6376c were newly designated in the study. Conclusion: The data presented here including a typing scheme and detailed genetic comparison which provide an insight into the diversification and evolution history of IncG plasmids.

Acinetobacter baumannii has been listed as one of the most critical pathogens in nosocomial infections; however, the key genes and mechanisms to adapt to the host microenvironment lack in-depth understanding. In this study, a total of 76 isolates (from 8 to 12 isolates per patient, spanning 128 to 188 days) were longitudinally collected from eight patients to investigate the within-host evolution of A. baumannii. A total of 70 within-host mutations were identified, 80% of which were nonsynonymous, indicating the important role of positive selection. Several evolutionary strategies of A. baumannii to increase its potential to adapt to the host microenvironment were identified, including hypermutation and recombination. Six genes were mutated in isolates from two or more patients, including two TonB-dependent receptor genes (bauA and BJAB07104_RS00665). In particular, the siderophore receptor gene bauA was mutated in multiple isolates from four patients with three MLST types, and all mutations were at amino acid 391 in ligand-binding sites. With 391T or 391A, BauA was more strongly bound to siderophores, which promoted the iron-absorption activity of A. baumannii at acidic or neutral pH, respectively. Through the A/T mutation at site 391 of BauA, A. baumannii displayed two reversible phases to adapt to distinct pH microenvironments. In conclusion, we demonstrated the comprehensive within-host evolutionary dynamics of A. baumannii, and discovered a key mutation of BauA site 391 as a genetic switch to adapt to different pH values, which may represent a model in the pathogen evolutionary adaption of the host microenvironment.

Objective:We conducted a real-world multi-center clinical study with a large sample size to comprehensively evaluate the performance of three commercial hepatitis C virus (HCV) core antigen assays. The study aimed to evaluate the performance for their use in HCV infection screening, and to provide clues for further improving the sensitivity and specificity of the assays.Methods:Key performance indicators including the lower limit of detection (LOD), diagnostic sensitivity, and specificity of three HCV antigen assays (the Architect, Laibo, and ChemClin HCV core antigen assays) were evaluated using commercial seroconversion panels reflecting early HCV infection and clinical routine serum samples of outpatients and inpatients from 3 tertiary hospitals from January 2018 to April 2022. Factors that affect the performance indicators were further investigated.Results:The window period for detecting HCV infection with the three antigen assays was equal to or slightly longer than that of the RNA assay, but all are shorter than that of the anti-HCV assay. There was a good linear positive correlation between HCV core antigen and HCV RNA levels in treatment naive patients with hepatitis C ( r=0.90, P<0.01). For the most common genotype 1b strain in China, the LOD of the three HCV assays were equivalent to 531 IU/ml (Architect), 3,698 IU/mL (Laibo), and 4,624 IU/mL (ChemClin) HCV RNA, respectively. Due to the skewed distribution of HCV RNA levels in treatment-naive hepatitis C patients, more than 95% of the patients had viral loads higher than 6 166 IU/ml. Therefore, the three HCV antigens assays still maintained a satisfactory diagnostic sensitivity (94.33%-99.40%). Among 54 immunodeficient patients (leukemia patients) with HCV infection, 9% (5/54) had negative anti-HCV results, while the HCV antigen assays found all these infectors. Through further experiments, we revealed the amino acid polymorphism in the core region of genotype 3 strain impaired the sensitivity of all three HCV antigen assays. In addition, the sensitivity of the two domestic assays was impaired by anti-HCV antibodies in the serum. The specificity of HCV antigen assays for diagnosing hepatitis C is 99.94% to 99.98%. The rheumatoid factors, autoantibodies, and other unknown interference substances can lead to a small number of low level, "false positive" antigen results. Conclusions:HCV core antigen assay may be used as a satisfactory approach of infection screening, especially for the immunodeficient patents. However, the sensitivity and specificity of the assays are influenced by multiple factors, which should be further improved.

目的 建立基质辅助激光解吸电离飞行时间质谱(MALDI-TOF MS)直接鉴定无菌体液病原菌快速、准确的方法.方法 2020年6-10月,连续收集解放军总医院第五医学中心检验科培养阳性的血液和腹水标本418份,其中血液标本220份、腹水标本198份.应用MALDI-TOF MS对阳性培养瓶进行前处理后直接快速鉴定,并与VITEK-2全自动微生物鉴定及药敏分析仪(简称VITEK-2)鉴定结果进行比对,两种方法鉴定结果不同的用分子生物学方法确认.结果 410份单菌株感染所致的阳性标本中,MALDI-TOF MS直接鉴定准确率为98.54％(404/410),VITEK-2鉴定的准确率为98.54％(404/410),两者比较差异无统计学意义(χ2=0.085,P>0.05).两种方法鉴定结果不同的单菌株共12株,经分子生物学方法确认,MALDI-TOF MS直接鉴定符合6株,准确率为50.00％(6/12).经MALDI-TOF MS直接鉴定:410株单菌株属、种水平鉴定率分别为93.17％(382/410)、82.68％(339/410);244株革兰阴性菌鉴定准确率为99.59％(243/244),属、种水平鉴定率分别为97.54％(238/244)、89.34％(218/244);149株革兰阳性菌鉴定准确率为97.32％(145/149),属、种水平鉴定率分别为87.92％(131/149)、74.50％(111/149);17株真菌鉴定准确率为94.12％(16/17),属、种水平鉴定率分别为76.47％(13/17)、58.82％(10/17);8株复合菌属、种水平鉴定率分别为75.00％(6/8)、25.00％(2/8).结论 应用MALDI-TOF MS技术对阳性培养瓶进行直接鉴定,可有效缩短检测时间,且结果准确率高,可作为脓毒症病原菌鉴定的重要手段,为临床快速、合理选用抗菌药物提供依据.

目的 探讨我院肺癌患者合并下呼吸道感染的病原菌分布特征及药物敏感性情况,为临床合理应用抗菌药物提供参考依据.方法 从我院病历管理系统中收集2020～2021年肺癌合并下呼吸道感染患者的临床病历资料,对患者临床特征、病理分型、病原菌特征、药物敏感性及临床抗感染治疗等情况进行了回顾性分析.结果 肺癌合并下呼吸道感染的76例患者中,肺部CT检查结果显示肺部炎症占89.5％;病理分型以非小细胞肺癌为主;共分离出病原菌120株,其中革兰阴性菌91株,占75.8％,革兰阳性菌20株,占16.7％,真菌9株,占7.5％;76例患者中9例为混合感染,发生率为11.8％,以真菌混合革兰阴性菌感染为主.克雷伯菌属和铜绿假单胞菌对亚胺培南、美罗培南、哌拉西林/他唑巴坦、阿米卡星的敏感率均在93.0％以上,对左氧氟沙星和环丙沙星的敏感70.0％～93.3％;主要革兰阳性菌对奎奴普丁/达福普汀、利福平、万古霉素、利奈唑胺的敏感率均为100％;有69.7％的患者经抗感染治疗得到缓解或控制,根据药敏试验结果调整经验用药进行针对性治疗的患者占30.3％.结论 我院肺癌合并下呼吸道感染的病原菌以革兰阴性菌为主,以克雷伯菌属和铜绿假单胞菌最为常见.不同病原菌对不同抗菌药物敏感性不同,临床可选用哌拉西林/他唑巴坦等药物对革兰阴性菌进行经验性治疗,可选用奎奴普丁/达福普汀、利福平、万古霉素、利奈唑胺等药物对革兰阳性菌进行经验性治疗.临床医师应结合患者情况综合评定是否经验性使用碳青霉烯类药物,并根据药物敏感性试验结果进行针对性治疗,及时控制感染,改善患者预后,提高肺癌患者的救治成功率.

Owing to their diagnostic and therapeutic potential, extracellular vesicles (EVs) derived from tumour cells have recently garnered great interest. The presence of different glycosylation sites at the EV surface supports the need for efficient glycosylated EV isolation. Here, we developed a GlyExo-Capture technique for robustly capturing fucosylated EVs from sera and cell supernatants. Lens culinaris lectin (LCA)-immobilized magnetic complexes were found to capture approximately 60% of the total EVs from HepG2 cells. The capture efficiency was reduced to less than 40% in nontumorigenic MIHA cells. Notably, the cellular uptake pattern of highly fucosylated EVs was markedly different from that of EVs with low fucosylation. The unearthing of enriched fucosylated EV miRNA cargos by next-generation deep sequencing (NGS) revealed 75 differentially expressed miRNAs (DEMs) in hepatocellular carcinoma (HCC). Among them, a 4-miRNA panel was chosen and yielded an area under the ROC curve (AUC) of 0.86 and 0.84 for the detection of HCC from non-HCC controls in testing samples and independent validation samples, respectively. The 4-miRNA signature was independent of alpha-fetoprotein (AFP), and a combined model with AFP yielded an increased AUC of 0.92. In conclusion, we developed a high-throughput method for capturing fucosylated EVs efficiently and shed light on the use of fucosylated EVs as potential sources of miRNAs for cancer biomarker detection. ### Competing Interest Statement The authors have declared no competing interest.

目的:分析解放军总医院血液病住院患者血流感染病原菌的分布特点和耐药性,为临床合理选用抗菌药物提供参考依据.方法:对2019年至2021年血液病和非血液病住院患者送检的血培养及药敏结果进行回顾性分析,用VITEK-2微生物分析仪进行微生物鉴定和药敏试验.结果:血液病患者血培养阳性标本病原菌以革兰阴性杆菌为主(71.2％),其次是革兰阳性菌(26.8％)、真菌(2.0％);主要病原菌有大肠埃希菌、肺炎克雷伯菌、凝固酶阴性葡萄球菌、铜绿假单胞菌等;主要革兰阴性菌对阿米卡星、亚胺培南、美罗培南敏感率较高;主要革兰阳性菌对万古霉素、利奈唑胺敏感率较高;葡萄球菌属未发现万古霉素、替加环素和利奈唑胺耐药株;分离的真菌对二性霉素B和伏立康唑的敏感率高.对血液病和非血液病患者血流感染的病原菌进行比较,发现血液病患者铜绿假单胞菌和嗜麦芽窄食单胞菌构成比高于非血液病患者,凝固酶阴性葡萄球菌和白色假丝酵母菌明显低于非血液病患者(P＜0.05).血液病患者大肠埃希菌对碳青霉烯类药物、喹诺酮类药物等的耐药率高于非血液病患者,差异有统计学意义(P＜0.05);肺炎克雷伯菌和铜绿假单胞菌对碳青霉烯类、喹诺酮类等抗菌药物的耐药率均低于非血液病患者,差异有统计学意义(P＜0.05).结论:革兰阴性菌是引起本院血液病住院患者血流感染最主要的病原菌,以大肠埃希菌最为常见,与本院非血液病患者病原菌分布结果大致相同.革兰阴性菌对阿米卡星、亚胺培南、美罗培南、哌拉西林/他唑巴坦敏感率高,革兰阳性菌对万古霉素、利奈唑胺敏感率高.本研究发现血液病患者大肠埃希菌对碳青霉烯类药物、喹诺酮类药物等的耐药率高于非血液病患者,在临床诊治中应加强对碳青霉烯类药物和喹诺酮类药物的关注和监控,及时进行病原菌和耐药性监测、正确选用抗菌药物、及早控制血流感染,对预后非常重要.

Objective:To evaluate the positive rate of nucleic acid detections in throat swab and stool specimens of patients, thereby to provide a reference to the clinical laboratory diagnosis.Methods:Forty-one confirmed patients were enrolled in this study, their throat swab and stool specimens were collected for detections. All the samples were determined by a fluorescence quantitative PCR method . The positive rate and concordance were evaluated for these two types of samples.Results:Among those 41 patients, 9 of them had positive result in throat swab samples, accounted for 30%. Five patients had positive result in stool samples, accounted for 12.2%. Only two patients had concordant positive result . Kappa tests showed that the concordance was poor, kappa index was 0.153. Conclusions:The nucleic acid detection in stool specimen is a complemental method for respiratory tract samples and therefore may provide. additional information for infectiousness of patients.

Objective:In this article, we analyzed and discussed the clinical characteristics and S region gene sequencing of hepatitis B virus in HBsAg anti-HBs coexistent patients.Methods:Data of 5 serologic markers of hepatitis B and quantitative result, liver function and HBV DNA load of HBsAg positive patients were collected, and their basic clinical information were recorded. According to the positive and negative result of Anti-HBs, the clinical and virological characteristics of these two groups were analyzed. At the same time, among 17 320 patients with HBsAg positive HBV infection, 994 cases were tested by gene sequencing. The S region amino acid mutation, site mutation detection rate and genotype of 994 HBV infected patients with gene sequencing were statistically analyzed.Results:The positive rate of HBsAg and Anti-HBs was 4.36% (756/17 320). HBV-related cirrhosis in HBsAg+ /Anti-HBs+ group (19.71%) was significantly higher than that in HBsAg+ /Anti-HBs-group (15.94%), while chronic hepatitis B (62.04%) was significantly lower than that in HBsAg+ /Anti-HBs-group (67.06%). At the same time, the positive rates of HBsAg-quantification (QN) and ALT in HBsAg+ /Anti-HBs+ group were significantly lower than those in HBsAg+ /Anti-HBs-group, the positive rate of HBeAg was significantly higher than that in HBsAg+ /Anti-HBs-group, and the HBV DNA was higher than that in HBsAg+ /Anti-HBs-group, but the difference was no statistical significance. Gene sequencing was performed in 994 HBV patients. Genotype C (81.79%) had the highest proportion, genotype B (17.40%) was the second, and genotype D (0.80%) was the least in two groups. In genotype C HBV infected patients, the detection rate of sP120Q/T/A/S mutant in HBsAg+ /Anti-HBs+ group was significantly higher than that in HBsAg+ /Anti-HBs-group. Meanwhile, regardless of genotype B or C or overall comparison, the detection rate of sG145A/E/K/R mutant of HBV infected patients in HBsAg+ /Anti-HBs+ group was significantly higher than that in HBsAg+ /Anti-HBs-group, these differences were all statistically significant.Conclusions:The hepatitis B patients with coexistence of HBsAg and Anti-HBs were more likely to develop cirrhosis, and the hepatitis B patients with HBV gene sequencing results were mainly type C2. The drug resistance variation of S-region sP120Q/T/A/S and sG145A/E/K/R mutants of patients with HBV infection is an important reason for the coexistence of HBsAg and Anti-HBS.

Objective:To understand the characteristics of serological detection indicators of patients with hepatitis B virus (HBV) infection in three hospitals in Beijing from 2018 to 2021.Methods:The five markers of hepatitis B, liver function tests, HBV DNA load, AFP and PT test results and basic clinical information of HBsAg positive HBV infected patients in three hospitals in Beijing from 2018 to 2021 were collected. Then the diagnosis of HBV infection, the positive patterns of serological indicators for five markers of hepatitis B and the clinical characteristics of hepatitis B patients were analyzed by SAS 9.4 statistical software.Results:Among the 1 026 604 patients who were tested for the five markers of hepatitis B or hepatitis B surface antigen quantification (HBsAg-QN) in three hospitals in Beijing from 2018 to 2021, the positive detection rate of HBsAg was 53.50%. The annual positive detection rate of HBsAg was 57.22%, 55.05%, 53.64% and 47.69% successively, showing a downward trend year by year. 111 709 hepatitis B patients were divided into 1-30, 31-60 and>60 years old groups according to their age. The main diseases of the three groups of HBV infected patients was chronic hepatitis B (CHB), and the proportion of patients with acute hepatitis B (AHB) and CHB decreased with age, while the proportion of patients with HBV-related liver cirrhosis, HBV-related liver cancer, liver cancer surgery and liver transplantation increased with age, the difference of which was statistically significant (all P<0.05). In this research, a total of 24 positive patterns of the five markers of hepatitis B were detected, including 7 common patterns (the main pattern was 145), 14 rare patterns (the main pattern was 1345), and 3 unusual patterns (the main pattern was 12345). The age, male ratio, HBeAg positive detection rate, HBV DNA positive detection rate and load, TBIL, ALT, AFP and PT results in the HBsAg positive group (90 011cases) were higher than those in the HBsAg negative group (21 698 cases), and the above results of the two groups of hepatitis B patients were higher than those of the healthy control group (20 623 cases). The albumin (ALB) results were the lowest in the HBsAg positive group and the highest in the healthy control group. And the differences were statistically significant (all P<0.05). Conclusions:From 2018 to 2021, the positive rate of HBsAg among the patients who received the five markers of hepatitis B or HBsAg-QN test in three hospitals in Beijing decreased year by year. Age was associated with disease progression in patients with hepatitis B. The positive patterns of five markers of hepatitis B in HBV infected people showed diversity.

Objective: To construct the lentivirus overexpression vector with two label genes fused with CopGFP and PuroR and to detect the emission of green fluorescence as well as resistance to puromycin in liver cancer cells infected with lentivirus packaged with the above vector. Methods: Firstly, two fragments containing copGFP and PuroR coding sequences were amplified from pCDH-CMV-MCS-copGFP and pLKO.1 respectively; secondly, the two amplified regions were fused with each other by recombinant PCR; thirdly, the fusion DNA fragment was cut and inserted into pCDH-CMV-MCS-copGFP vector, which was linearized with the same restriction endonuclease as used to digest fusion DNA fragment: BamH Ⅰ and Sal Ⅰ. The fusion region in the constructed vector was confirmed by DNA sequencing. The checked vector was co-transfected with package assistant plasmids, namely PLP1, PLP2 and VSVG into in 293T cells and the culture supernatant was subjected to centrifuge and infect liver cancer MHCC97H cells, which were then used to detect their resistance to puromycin (infected cells were treated with 1 mg/ml puromycin for 7 days after infection) and to observe green fluorescence emission in microscope. To determine its efficiency in expressing foreign target protein, the Sp1 coding region was inserted into the MCS sites of the vector, and Sp1 mRNA and protein expression levels were compared with the vehicle vector by RT-qPCR and Western blot. Results: The lentivirus overexpression vector with two label genes fused with CopGFP and PuroR was successfully constructed, and the liver cancer cells infected with lentivirus packaged with the vector expressing two labeling genes fused with CopGFP and PuroRshowed both emission of green fluorescence and resistance to puromycin simultaneously, while cells containing with the vector inserted with Sp1 coding region improved Sp1 mRNA level with 3.3 fold and protein level with 2.2 fold higher in comparison with cells containing the vehicle vector (P＜0.01). Conclusion: The fused label genes consisting of copGFP and PuroR are correctly cloned into the lentivirus vector and confer cells with the ability to emission of green fluorescence and resistance to puromycin, besides, the vector may promote the expression of the target gene with long coding sequence.