Background: Rapid molecular testing enhances pathogen detection in community-acquired pneumonia (CAP), yet its impact on clinical outcomes remains uncertain. We evaluated whether early precision treatment (PT) guided by evolving molecular diagnostics improves in-hospital mortality, and sought to identify specific patient subgroups that derive the greatest survival benefit. Methods: The multicentre retrospective CCAPP cohort study (June 2005–July 2024) included 2430 adults hospitalised with CAP across four general hospitals in Fujian Province, China, yielding 1904 patients with complete data for final analysis. Patients were categorized by diagnostic strategy: limited-target (LND; n=794; 11 bacterial and atypical pathogens) or diverse-target (DND; n=678; 33 bacterial, atypical, fungal, and viral pathogens) nucleic acid detection, or metagenomic next-generation sequencing (mNGS; n=432). On the basis of whether empirical anti-infective regimens administered within 3 days of admission covered clinically relevant pathogens, patients were classified as PT, non-precision treatment (NP), or no pathogen detected (NPD), and were further stratified into respiratory failure (RF) and non-respiratory failure (nRF) subgroups. Primary outcome was in-hospital mortality; secondary outcomes included length of stay and hospital costs. Confounding was addressed via 1:1:1 triad matching (PT:NP:NPD) in the DND (n=318) and mNGS (n=111) cohorts, analysed using conditional logistic regression and Kaplan-Meier methods. Findings: Pathogen positivity was 23·8% (LND), 60·0% (DND), and 32·6% (mNGS). PT was associated with significantly lower mortality compared with NP exclusively in the mNGS cohort, observed in both the overall population (3·4% vs 18·1%, p<0·001) and the RF subgroup (6·5% vs 34·1%, p=0·002). After matching, logistic regression in the DND cohort revealed that PT mortality benefit was strictly confined to the RF subgroup (OR 0·17, 95% CI 0·01–0·98; p=0·049; overall OR 0·43, p=0·220). Distinctly, PT in the mNGS cohort was associated with markedly lower mortality across both the overall population (OR 0·20, 95% CI 0·05–0·88; p=0·034) and the RF subgroup. Survival analyses corroborated this divergence: survival benefits for PT were consistent across the entire mNGS cohort (overall p=0·027; RF p=0·022), whereas DND-era benefits remained restricted to RF patients (p=0·040). PT did not significantly reduce length of stay or costs compared with non-PT. Interpretation: For hospitalised CAP, standard empirical therapy suffices for nRF patients, as PT yields no additional survival benefit, nor does it decrease length of stay or total costs. Conversely, RF patients derive substantial mortality reduction from PT guided by broad-spectrum diagnostics. While mNGS maximizes this protective effect, DND serves as a pragmatic alternative in resource-constrained settings.

Background The mechanisms underlying the early progression of lung adenocarcinoma (LUAD) remain unclear, and the interaction between circular RNAs (circRNAs) and the immune microenvironment lacks systematic investigation. While circHIF1A has been reported to possess oncogenic functions in colorectal and breast cancers, its specific role in LUAD and whether it regulates the tumor microenvironment via a competing endogenous RNA (ceRNA) mechanism remain to be elucidated. Methods Tumor and paired paratumorous normal tissues were collected from 80 LUAD patients. Using A549 and H1299 cell lines and the nude mouse subcutaneous xenograft model, we systematically analyzed the expression characteristics, biological functions, and molecular mechanisms of circHIF1A. Results circHIF1A was significantly upregulated in LUAD, and its expression level was positively correlated with advanced TNM stage and poor patient prognosis. Mechanistically, circHIF1A functions as the ceRNA to specifically sponge miR-486-5p, thereby relieving the transcriptional repression of its downstream target gene, GRHL2, forming the circHIF1A/miR-486-5p/GRHL2 regulatory axis. This axis not only directly enhances LUAD cell proliferation, stemness maintenance, migration, and invasion but also induces macrophage M2 polarization by promoting IL-10 secretion, thereby establishing an immunosuppressive microenvironment. In vivo experiments confirmed that intervening in the circHIF1A/miR-486-5p/GRHL2 axis significantly inhibits LUAD tumor growth. Conclusion circHIF1A regulates malignant phenotypes of LUAD cells and tumor microenvironment remodeling through a ceRNA network. The mediated circHIF1A/miR-486-5p/GRHL2 axis provides a novel molecular biomarker for LUAD prognostic assessment and represents a potential target for therapeutic intervention.

The regulatory mechanisms driving brain metastasis (BM) in non-small cell lung cancer (NSCLC) are complex, with Ceramide synthase 1 (Cers1) playing a critical role. However, the upstream factors controlling Cers1 expression remain unclear. Additionally, Kruppel-like factor 9 (KLF9) has been implicated as a potential tumor suppressor transcription factor (TF) in lung cancer. Our study aims to explore the physiological effects and molecular mechanisms of Cers1 upstream regulatory TFs in NSCLC BM, focusing on the link between KLF9 and Cers1. TFs regulating Cers1 expression were screened via GENECARD and UCSC databases. KLF9 expression and survival analysis in NSCLC were analyzed using TCGA and GEO data sets. Dual-luciferase assays were conducted to investigate the specific binding site of KLF9 on the Cers1 promoter. KLF9 genetic modulation impact on NSCLC BM was assessed through in vitro assays for proliferation, migration, and invasion, along with in vivo orthotopic xenograft models to evaluate tumor growth and metastasis. Our study revealed a significant downregulation of KLF9 in vitro, correlating with poor prognosis. KLF9 has a direct positive transcriptional regulation on Cers1, particularly on its promoter region (-711 nt/+22 nt). In vitro, KLF9 overexpression inhibited the ability of cells to penetrate a blood-brain barrier model. Moreover, in vivo experiments demonstrated a marked suppression of BM tumor formation upon KLF9 expression. This study reports KLF9's direct control over Cers1 expression by acting on its promoter. KLF9 demonstrates inhibitory effects on NSCLC BM, presenting a promising therapeutic avenue for NSCLC BM treatment.

Tumor mutational burden (TMB) is a recognized biomarker for predicting immunotherapy efficacy in non-small cell lung cancer (NSCLC). Its assessment requires whole-exome sequencing (WES), but the high cost and stringent sample requirements of WES limit its clinical application. This study aims to assess the predictive value of accessible systemic inflammation markers for identifying high TMB lung cancer populations. WES was performed on tumor samples and paired peripheral blood from 72 lung adenocarcinoma patients. Genomic analysis identified mutation patterns across different TMB groups. Systemic inflammatory markers, including the neutrophil-to-lymphocyte ratio (NLR), derived neutrophil-to-lymphocyte ratio (dNLR), lymphocyte-to-monocyte ratio (LMR), and platelet to lymphocyte ratio (PLR), were collected. Generalized linear models and restricted cubic spline (RCS) plots were used to explore the predictive value of these markers for TMB. The Xgboost model assessed the importance of each variable for TMB prediction. Among the 72 lung adenocarcinoma patients, missense mutations were the most common, with single nucleotide variants being the predominant mutation type. The most frequently mutated genes were EGFR (35

Brain metastasis (BM) is a significant factor contributing to the poor prognosis of patients with non-small cell lung cancer (NSCLC). Secreted phosphoprotein 1 (SPP1) is implicated in the progression and metastasis of several cancers. The role of SPP1 in NSCLC remains unclear, especially in NSCLC BM. This study aimed to identify genes associated with NSCLC BM and to investigate the involvement of SPP1 in NSCLC BM. Integrated genomic analysis was utilized to identify candidate genes in NSCLC. The expression levels of SPP1 were evaluated in NSCLC tissues and cell lines. In vitro and in vivo experiments were conducted to assess the effect of SPP1 on NSCLC cell behavior and BM. The potential mechanisms of SPP1 were demonstrated by CO-IP and liquid chromatography-mass spectrometry (LC-MS). The underlying mechanism involving the PI3K/AKT/mTOR pathway was explored. The results showed that SPP1 expression was upregulated in NSCLC tissues and cell lines. Depletion of SPP1 using shRNA inhibited cell proliferation, migration, and invasion in vitro and suppressed BM in vivo. Mechanistically, SPP1 facilitates the ubiquitination of P85α by interacting with the ubiquitin ligase RNF114, thus playing a role in regulating NSCLC BM through the PI3K/AKT/mTOR signaling pathway. Moreover, immunohistochemistry staining confirmed higher expression of SPP1 in NSCLC tissues with BM compared to those without BM. In summary, elevated SPP1 expression was associated with poor clinical outcomes in NSCLC patients. This study highlights the role of SPP1 as a regulator of cell metastasis and suggests its potential as a novel therapeutic target for BM in NSCLC.

PURPOSE:The objective of this trial was to evaluate the efficacy and safety of anti-IgE monoclonal antibody (CMAB007) in Chinese patients with inadequately controlled moderate or severe asthma with increased total IgE level despite medium or high dose inhaled corticosteroids (ICS)/long acting β₂-agonist (LABA) treatment. METHODS:This was a multicenter, randomized, placebo controlled, double blinded, phase 3 trial. Eligible patients with moderate or severe asthma with increased total IgE level (60-1,500 IU/mL) receiving optimal ICS-LABA were randomly assigned to receive CMAB007 or placebo treatment at a 2:1 ratio for 24 weeks. The primary efficacy endpoint was asthma exacerbation (AE) rate, the key second endpoints were asthma control test (ACT) score, pulmonary function and safety. RESULTS:A total of 392 patients were included in the efficacy analysis. AE rate was 0.45 with CMAB007 and 0.66 with placebo (hazard ratio, 0.68; 95% confidence interval, 0.49, 0.96; P = 0.030). The proportions of patients showing an increase of at least 3 points from baseline in ACT scores were greater in the CMAB007 group than in the placebo group at Week 8 (49.8% vs. 36.9%, P = 0.018), Week 16 (59.4% vs. 42.9%, P = 0.003) and Week 24 (60.2% vs. 47.0%, P = 0.019). Greater improvement of pre-bronchodialteor forced expiratory volume in 1 second was achieved in the CMAB007 group at 4 weeks (11.40% vs. 5.03%, P = 0.006), 8 weeks (16.27% vs. 6.57%, P = 0.003), 12 weeks (16.51% vs. 6.60%, P = 0.002) and 16 weeks (20.18% vs. 7.42%, P = 0.002). The incidence of adverse events was similar between the CMAB007 (77.2%) and placebo groups (75.2%). CONCLUSIONS:CMAB007 reduces AE and improves asthma control, lung function and quality of life without additional safety concern in Chinese patients having moderate to severe asthma with increased total IgE level. TRIAL REGISTRATION:ClinicalTrials.gov Identifier: NCT03468790.

Introduction Lung adenocarcinoma (LUAD) is one of the leading causes of cancer-related deaths worldwide. While NCAPD2 has been implicated in promoting tumorigenesis across various cancer types, its specific role in LUAD remains underexplored. This study aims to elucidate the molecular mechanisms by which NCAPD2 contributes to LUAD progression, with a focus on its involvement in the AKTMDM2/E2F1 positive feedback loop.Materials and methods NCAPD2 expression in LUAD and normal tissues was analyzed using Western blotting and immunohistochemistry (IHC). Functional assays, including colony formation, wound healing, Transwell assays, and in vivo mouse models were conducted to evaluate the impact of NCAPD2 on LUAD cell proliferation, invasion, and metastasis. RNA sequencing and protein interaction experiments were used to investigate the role of NCAPD2 in the PI3K/AKT/MDM2 pathway and its interaction with E2F1.Results This study first identified that NCAPD2 expression is significantly upregulated in LUAD tissues, particularly in higher pathological stages. NCAPD2 overexpression promoted LUAD cell proliferation and metastasis, while its knockdown inhibited tumor growth and invasion. Mechanistically, NCAPD2 activated the PI3K/Akt pathway, facilitating the interaction between MDM2 and E2F1, reducing E2F1 ubiquitination, and increasing its expression. Furthermore, E2F1 enhanced NCAPD2 transcription, forming a positive feedback loop that drives LUAD progression.Conclusion This study reveals a novel role of NCAPD2 in promoting LUAD progression through the AKT/MDM2/E2F1 positive feedback loop. These findings provide new insights into the molecular pathogenesis of LUAD and suggest NCAPD2 as a potential therapeutic target for improving patient outcomes.

ObjectiveThis study aims to investigate the clinical application value of Metagenome Next-Generation Sequencing (mNGS) for pulmonary diffuse exudative lesions.MethodsFrom January 1, 2014, to November 31, 2021, 136 cases with chest radiologic presentations of pulmonary diffuse exudative lesions admitted to Fujian Provincial Hospital were included in the study; of those, 77 patients underwent mNGS pathogen detection. Based on the pathogen detection outcomes and clinical diagnoses, patients were categorized into an infection group (IG) and a non-infection group (NIG). A comparison was made between the diagnostic efficacy of the mNGS technique and traditional culture methods. Meanwhile, 59 patients clinically identified as having infectious pulmonary diffuse exudative lesions but who did not receive mNGS testing were designated as the non-NGS infection group (non-IG). A retrospective cohort study was conducted on patients in both the IG and non-IG, with a 30-day all-cause mortality endpoint used for follow-up.OutcomesWhen compared to conventional culture methods, mNGS demonstrated an approximate 35% increase in sensitivity (80.0% vs 45.5%, P<0.001), without significant disparity in specificity (77.3% vs 95.5%, P=0.185). Under antibiotic exposure, the positivity rate detected by mNGS was notably higher than that by traditional culture methods, indicating that mNGS is less affected by exposure to antibiotics (P<0.05). Within 30 days, the all-cause mortality rate for patients in the IG versus the non-IG was 14.55% and 37.29%, respectively (P<0.05). Following a COX regression analysis to adjust for confounding factors, the analysis revealed that a CURB-65 score ≥3 points (HR=3.348, P=0.001) and existing cardiovascular disease (HR=2.473, P=0.026) were independent risk factors for these patients. Conversely, mNGS testing (HR=0.368, P=0.017) proved to be an independent protective factor.ConclusionmNGS technology makes it easier to pinpoint the cause of pulmonary diffuse infectious exudative lesions without much interference from antibiotics, helping doctors spot and diagnose these issues early on, thereby playing a key role in helping them decide the best treatment approach for patients. Such conclusions may have a bias, as the performance of traditional methods might be underestimated due to the absence of complete results from other conventional diagnostic techniques like serological testing and PCR.

PurposeAvailable research indicates that the mammalian target of rapamycin complex 1 (mTORC1) signaling pathway is significantly correlated with lung cancer brain metastasis (BM). This study established a clinical predictive model for assessing the risk of BM based on the mTORC1-related single nucleotide polymorphisms (SNPs).MethodsIn this single-center retrospective study, 395 patients with non-small cell lung cancer were included. Clinical, pathological, imaging, and mTORC1-related single nucleotide polymorphism data were collected. Lasso regression was used to identify variables related to the risk of BM in lung cancer, and a nomogram was constructed. Internal validation was performed using 1,000 bootstrap samples. We plotted the receiver operating characteristic (ROC) curve and calculated the area under the curve (AUC). The calibration of the model was assessed using calibration curves and the Hosmer-Lemeshow goodness-of-fit test, and decision curve analysis (DCA) was plotted to evaluate the net clinical benefit.ResultsThe nomogram's predictive factors included lung cancer histology, clinical N stage, CEA, neutrophil to lymphocyte ratio (NLR), lymphocyte to monocyte ratio (LMR), RPTOR: rs1062935, and RPTOR: rs3751934. The AUC of the model in the training set and internal validation were 0.849 and 0.801, respectively. The calibration curves and Hosmer-Lemeshow test both indicated a good fit.ConclusionThe nomogram has practicality and efficacy in predicting the high risk of BM in lung cancer patients, confirming that single nucleotide polymorphisms in the mTORC1 pathway genes may be good predictors in clinical prediction models.

Third-generation EGFR-TKIs can be used to treat advanced non-small cell lung cancer patients with T790M resistance mutation induced by first- or second-generation EGFR-TKIs. However, it will also result in drug resistance, and the resistance mechanisms of third-generation EGFR-TKIs are complex. Here we reported a patient diagnosed with advanced lung adenocarcinoma and EGFR positive in September 2016. Following first-line targeted therapy with gefitinib, genetic testing showed EGFR T790M positive, which resulted in a change to osimertinib targeted therapy. In May 2021, troponin and creatinine levels were elevated, and the tumor hyperprogressed to severe lung cancer. Repeated genetic testing revealed that EGFR genotype converted to a non-classical mutation and EGFR T790M turned negative, which caused third-generation EGFR-TKI resistance. As a result, afatinib combined with anlotinib was selected to stabilize the patient’s condition. We were inspired by the case that it reflects the significance and necessity of exploring the resistance mechanism and dynamically detecting genetic status throughout the course of treatment, which may help realize individualized precision therapy, and maximize the potential of patient.

BACKGROUND:Tuberculosis is a chronic infectious disease and an important public health problem. Despite progress in controlling tuberculosis, the incidence of tuberculosis in China is still very high, with 895000 new cases annually. This case report describes the investigation of a case of severe disseminated tuberculosis in a young adult with normal immune function, conducted to ascertain why a Mycobacterium tuberculosis (M. tuberculosis) strain caused such severe disease. CASE SUMMARY:A previously healthy 28-year-old woman presented to our hospital with a 1-month history of fever and fatigue. She was diagnosed with severe disseminated pulmonary tuberculosis, spinal tuberculosis with paravertebral abscesses, and tuberculous meningitis. M. tuberculosis was isolated from bronchoalveolar lavage fluid. She was treated with standard antituberculous therapy and underwent debridement, bone graft, and internal fixation surgery for spinal tuberculosis. She responded to therapy and regained her ability to walk following the surgery. We analysed the whole-genome sequence of the strain and designated it BLM-A21. Additional M. tuberculosis genomes were selected from the Virulence Factor Database (http://www.mgc.ac.cn/cgi-bin/VFs/genus.cgi?Genus=Mycobacterium) for comparison. An evolutionary tree of the BLM-A21 strain was built using PhyML maximum likelihood software. Further gene analysis revealed that, except for the pks1 gene, BLM-A21 had similar virulence genes to the CDC 1551 and H37Rv strains, which have lower dissemination. CONCLUSION:We speculate that the pks1 virulence gene in BLM-A21 may be the key virulence gene responsible for the widespread dissemination of M. tuberculosis infection in this previously healthy adult with normal immune function.

OBJECTIVE:To compare the diagnostic efficacy of metagenomic next generation sequencing (mNGS) with traditional fungal culture, (1,3)-β-D glucan (G) test, and galactomannan (GM) test in diagnosing invasive pulmonary aspergillosis (IPA) and to explore the advantages and disadvantages of mNGS for IPA diagnosis. METHODS:A retrospective analysis was conducted on 136 patients admitted to the Department of Respiratory and Critical Care Medicine of Affiliated Hospital of Putian University from March 2018 to March 2020. Among them, there were 66 patients with IPA (IPA group) and 70 without (non-IPA group). Baseline data, inflammatory factors, cytokines, and specimens such as bronchoalveolar lavage fluid (BALF) and blood of these patients were collected. Fungal culture test, G test, GM test and mNGS test were performed. Information included for analysis encompassed patients' host factors, clinical features, chest scanning images, laboratory test results, and treatment outcome. RESULTS:There was no statistical difference in the baseline data or inflammatory factors in patients between the IPA group and the non-IPA group. Further analysis showed that the sensitivity of mNGS in diagnosing IPA was 53.03%, which was higher than that of traditional fungal culture test (27.27%), G test (31.82%), and GM test (34.85%). Notably, when combining fungal culture, G test, GM test, and mNGS, the sensitivity increased to 69.70%, with a specificity of 97.14%. The sensitivity of the combined test was higher than that any of the tests alone for diagnosing IPA. CONCLUSION:mNGS test offers superior diagnostic performance for IPA in comparison to traditional tests, particularly for testing samples like bronchoalveolar lavage fluid and bronchial secretions. The test result remains valuable even after aspergillus treatment. In addition, the use of mNGS in conjunction with other traditional tests, such as fungal culture test, G test, and GM test, can enhance the diagnostic efficacy for IPA.

Abstract Background Ceramide metabolism is crucial in the progress of brain metastasis (BM). However, it remains unexplored whether targeting ceramide metabolism may arrest BM. Methods RNA sequencing was applied to screen different genes in primary and metastatic foci and whole-exome sequencing (WES) to seek crucial abnormal pathway in BM + and BM-patients. Cellular arrays were applied to analyze the permeability of blood–brain barrier (BBB) and the activation or inhibition of pathway. Database and Co-Immunoprecipitation (Co-IP) assay were adopted to verify the protein–protein interaction. Xenograft and zebrafish model were further employed to verify the cellular results. Results RNA sequencing and WES reported the involvement of RPTOR and ceramide metabolism in BM progress. RPTOR was significantly upregulated in BM foci and increased the permeability of BBB, while RPTOR deficiency attenuated the cell invasiveness and protected extracellular matrix. Exogenous RPTOR boosted the SPHK2/S1P/STAT3 cascades by binding YY1, in which YY1 bound to the regions of SPHK2 promoter (at -353 ~ -365 nt), further promoting the expression of SPHK2. The latter was rescued by YY1 RNAi. Xenograft and zebrafish model showed that RPTOR blockade suppressed BM of non-small cell lung cancer (NSCLC) and impaired the SPHK2/S1P/STAT3 pathway. Conclusion RPTOR is a key driver gene in the brain metastasis of lung cancer, which signifies that RPTOR blockade may serve as a promising therapeutic candidate for clinical application.

OBJECTIVE To investigate the clinical characteristics of non-tuberculous Mycobacteria pulmonary disease（NTMPD）, identify the species of non-tuberculous Mycobacteria（NTM） with metagenomic sequencing and perform the cluster analysis. METHODS A total of 79 patients who were diagnosed with NTMPD in Fujian Provincial Hospital from Jan 2015 to Dec 2020 were enrolled in the study, the related clinical data were collected, the whole genome sequencing was carried out for 29 clinical NTM isolates by means of metagenomic next generation sequencing（mNGS）, the average nucleotide identity（ANI） analysis was performed for identification of the strains, and the cluster heat map of the ANI value was drawn for the clinical NTM isolates. RESULTS The number of confirmed cases of NTMPD was increased year by year. Among the 79 patients with NTMPD, there were 40 male cases and 39 female cases, the average age was（60.78±12.04） years old, and about 55.70%（44/79） had bronchiectasia. Cough（86.08%） and expectoration（75.95%） were the major clinical manifestations of the NTMPD patients; bronchiectasis（55.70%, 44/79）, tree-in-bud signs（43.04%, 34/79） and nodules（35.44%, 28/79） were the common CT imaging features. Alveolar lavage fluid specimens were dominant among the specimens cultured positive for NTM, accounting for 60.76%（48/79）. Totally 10 species of NTM were identified among the 29 strains of NTM, 58.62% of which were Mycobacterium avium complex, and 24.14% were Mycobacterium abscess. Among the M.avium complex strains, M. intracellulare, M.paraintracellulare and M. chimera had close ties of consanguinity, and the fast-growing M. abscess was distantly related to slow-growing NTM strains. CONCLUSION The patients with NTMPD lack typical clinical symptoms. It is necessary to remain vigilant for the possibility of NTM infection among the patients with CT imaging manifested as bronchiectasis complicated with nodules and tree-in-bud signs. There is certain association between the genotypes and the phenotypes of the NTM strains.

Brain metastasis (BM) is common in patients with non-small cell lung cancer (NSCLC) and is associated with a poor prognosis. Ceramide synthase 1 (CERS1) participates in malignancy development, but its potential role in NSCLC BM remains unclear. This study aimed to explore the physiological effects and molecular mechanism of CERS1 in NSCLC BM. CERS1 expression was evaluated in NSCLC tissues and cell lines, and its physiological roles were subsequently explored in vivo and in vitro. Mass spectrometry and co-immunoprecipitation were performed to explore CERS1-interacting proteins. The associated signaling pathways of CERS1 in NSCLC BM were further investigated using bioinformatics analysis and molecular biotechnology. We demonstrated that CERS1 was significantly downregulated in NSCLC cell lines and BM tissues, and its upregulation was associated with better prognoses. In vitro, CERS1 overexpression inhibited cell migration, invasion, and the ability to penetrate the blood-brain barrier. Moreover, CERS1 interacted with ubiquitin-specific protease 14 (USP14) and inhibited BM progression by downregulating the PI3K/AKT/mTOR signaling pathway. Further, CERS1 expression substantially suppressed BM tumor formation in vivo. This study demonstrated that CERS1 plays a suppressor role in NSCLC BM by interacting with USP14 and downregulating the PI3K/AKT/mTOR signaling pathway, thereby serving as a novel therapeutic target for NSCLC BM.

IntroductionThis study explored the differences in clinical characteristics between the 2009 pandemic influenza A (H1N1) and SARS-CoV-2 BA.2 variant (Omicron) infections in patients younger than age 65 years, to improve identification of these diseases and better respond to the current epidemic. MethodsData from 127 patients with the 2009 pandemic influenza A (H1N1) diagnosed between May and July of 2009 and 3,265 patients with Omicron diagnosed between March and May of 2022 were collected. Using a 1:2 match based on age (difference <2 years), sex, and underlying diseases, data from 115 patients with the 2009 pandemic influenza A (H1N1) infection (H1N1 group) and 230 patients with SARS-CoV-2 Omicron BA.2 infection (Omicron group) were analyzed. The clinical manifestations were compared between the groups, logistic regression was performed to identify possible independent risk factors for each group, and multiple linear regression was used to analyze the factors predicting time for nucleic acid negativization (NAN). ResultsThe median [interquartile range] age of the two groups was 21 [11, 26] years. Compared with the H1N1 group, the Omicron group had: lower white blood cell counts and C-reactive protein levels; less fever, nasal congestion, sore throat, cough, sputum, and headache; and more olfactory loss, muscle soreness, and lactate dehydrogenase (LDH) abnormalities. Patients in the Omicron group used fewer antibiotics and antiviral drugs, and the time for NAN was longer (17 [14,20] VS 4 [3,5] days, P<0.001). Logistic regression showed that fever, cough, headache, and increased white blood cell count were more strongly correlated with the H1N1 group, while muscle soreness and LDH abnormalities were more strongly correlated with the Omicron group. Fever (B 1.529, 95% confidence interval [0.149,2.909], P=0.030) significantly predicted a longer time for NAN in patients with Omicron. DiscussionThere are significant differences in clinical characteristics between SARS-CoV-2 Omicron infection and the 2009 pandemic influenza A (H1N1) infection. Recognition of these differences has important implications for clinical practice.

BACKGROUND:This study aimed to compare the clinical characteristics of infections caused by different pathogens and then establish a viral/bacterial infection prediction model to guide early clinical identification of pathogens in inpatients with community-acquired pneumonia(CAP). METHODS:A total of 687 patients who were diagnosed with CAP in our hospital between March 2012 and December 2018 were studied.Basic data,clinical symptoms,laboratory examinations,and imaging examinations of patients were collected,and a virus/bacteria prediction equation was established.In the prediction model,the relevant variables were screened according to a univariate logistic regression analysis,and then,a multivariate logistic regression analysis was performed to establish the prediction equation. RESULTS:The proportions of patients with muscle soreness and headaches were significantly higher in the viral infection group than in the bacterial infection group.Procalcitonin(PCT)concentrations,the erythrocyte sedimentation rate(ESR),and the neutrophil alkaline phosphatase(NAP)score were significantly higher in the bacterial infection group than in the viral infection group.Creatine kinase concentrations were significantly higher in the viral infection group than in the bacterial infection group(P＜0.05).A higher proportion of patients had lung degeneration in the atypical pathogen infection group than in other groups(P=0.005).Patchy shadows were more common in the viral infection group than in the other groups.A binary logistic regression equation was obtained that could predict the probability of viral infection(sensitivity:57.5％,specificity:67.7％,and area under the receiver operating characteristics curve:0.651). CONCLUSIONS:Adult patients with CAP and viral infection are more likely to have headaches and muscle soreness than those with bacterial infection.An elevated PCT concentration,NAP score,and ESR indicate a high possibility of bacterial infection.We successfully established a viral and bacterial infection prediction model.

Staphylococcus lugdunensis (S. lugdunensis) is a coagulase-negative Staphylococcus comprising the normal skin microbiota, primarily colonizing the lower abdomen and extremities.[1]S. lugdunensis has attracted substantial attention in recent years since the discovery of lugdunin by Zipperer et al[2] in 2016. Lugdunin is a secondary metabolite synthesized by non-ribosomal peptide synthetase (NRPS) that inhibits the growth of various gram-positive bacteria, including methicillin-resistant S. aureus (MRSA),[2] suggesting its potential as an antibiotic. To date, 28 complete genome sequences and 11 partial-assembly genome sequences of S. lugdunensis have been published and uploaded to the GenBank database (https://www.ncbi.nlm.nih.gov/genome/browse/#!/prokaryotes/2548/, January 18, 2022). However, there are limited data on the genome sequences of S. lugdunensis from mainland China, and the effects of geographical origin on genetic variations in S. lugdunensis remain unknown. We analyzed six clinical strains of S. lugdunensis isolated from Beijing, Wuhan, and Fujian, China [Supplementary Table 1, https://links.lww.com/CM9/B273]. The minimum inhibitory concentrations (MICs) of antibiotics were evaluated using the VITEK2 Compact system (bioMerieux, Inc., Durham, USA). Cefoxitin disc dilution (cefoxitin DD), broth microdilution for oxacillin, and oxacillin salt agar methods were used as reference methods for assessing methicillin resistance. The complete genomes of the six S. lugdunensis strains were obtained by next-generation sequencing and Oxford Nanopore sequencing, and then compared with genomes available in the GenBank database. We further characterized the resistome and secondary metabolism gene clusters of these staphylococcal cassette chromosome mec (strains using computational approaches. We identified SCCmec) elements of methicillin-resistant S. lugdunensis (MRSL) and performed collinearity analysis of these strains.[3] Comparisons were performed for the obtained nucleotide sequences of the lug operon with those in the GenBank database using Blastx and Blastp tools against the proteins of the operon. We further explored the relationships of genetic variations of the lug operon with geographical origins and the clonal complex (CC). The detailed methods are reported in the Supplementary Materials, https://links.lww.com/CM9/B273. The genome sizes of the six strains ranged from 2.59 to 2.71 Mbp, with a GC content of 33.7% to 33.9% [Supplementary Figure 1, https://links.lww.com/CM9/B273]. All strains contained 2405 to 2523 coding sequences, with 60 to 61 tRNAs, 16 to 19 rRNAs, and 51 to 53 sRNAs. Two to three plasmids were identified in five strains (except RMLUG5). Multilocus sequence typing revealed sequence type (ST)3 (RMLUG1, RMLUG3, RMLUG6), ST27 (RMLUG2), ST34 (RMLUG4), and ST6 (RMLUG5). In antimicrobial susceptibility tests by VITEK2, three of the six strains were resistant to cefoxitin and oxacillin, as confirmed by the oxacillin broth microdilution method and oxacillin salt agar screening. Cefoxitin DD tests showed that strains RMLUG1 and RMLUG6 were susceptible to cefoxitin, although near the breakpoint [Supplementary Table 2, https://links.lww.com/CM9/B273]. The three MRSL strains were susceptible to antibacterial agents showing activity against MRSA, such as linezolid, vancomycin, and teicoplanin [Supplementary Table 3, https://links.lww.com/CM9/B273]). Notably, the methicillin resistance gene mecA was identified in the three MRSL strains (RMLUG1, RMLUG3, and RMLUG6) and in one methicillin-susceptible S. lugdunensis (MSSL) strain (RMLUG2). The SCCmec elements were V(5C2) (RMLUG1, RMLUG2, and RMLUG6) and IVi (2B) (RMLUG3). Only the ccr class 9 (ccrC2 allele 1) gene was identified in the MSSL strain RMLUG4, which is a composite of the SCCmec element. Supplementary Figure 2, https://links.lww.com/CM9/B273 illustrates the SCCmec structure of the six strains. Strains RMLUG1 and RMLUG6 were consistent with SCCmec type V from S. aureus strain WIS [Supplementary Figure 2A, https://links.lww.com/CM9/B273]. Compared with SCCmec type IVi in S. aureus strain JCSC6668, strain RMLUG3 contained a 1014 bp deletion of the gene encoding ISSep1-like transposase [Supplementary Figure 2B, https://links.lww.com/CM9/B273]. The ST27 strain RMLUG2 contained a complete SCCmec type V, including a type mecA class C2 complex and ccrC1 complex, but in a SCCmec type VII-like order: orfX-J3-ccr-J2-mec-J1. Moreover, the IS431 upstream of mecA had the opposite orientation to that downstream, in contrast with the structure of SCCmec type VII, suggesting a novel SCCmec type V variant [Supplementary Figure 2C, https://links.lww.com/CM9/B273]. The SCCmec elements of RMLUG4 did not carry the mecA gene but contained ccr class 9 (ccrC2) and determinants of resistance to heavy metals such as arsenic and copper. According to SCCmec classification,[4] this element is described as an SCC element, which was designated SCCRMLUG4 [Supplementary Figure 2D, https://links.lww.com/CM9/B273]. MIC tests and genomic analysis indicated that strain RMLUG2 carried the mecA gene but did not exhibit oxacillin resistance. No mutation of the mecA gene was observed among the MRSL strains. C to T substitution was identified at position –33 (i.e., 33 bp upstream of the start codon) within the mecA promoter in strain RMLUG2, which also occurred in RMLUG1 and RMLUG6. A comparison of the sequences of FemXAB family genes between strain RMLUG2 and the MRSL strain JICS135 revealed several mutations in femX (eg, R176K and D341E) and femB (eg, D245N and D259Y). The femX and femB genes of strains RMLUG1, RMLUG3, and RMLUG6 matched those of strain JICS135. A comparison of the genomes of the six strains to 14 lug operon genes (lugJ to lugM; Supplementary Table 4, https://links.lww.com/CM9/B273) identified the complete lug operon in all strains except RMLUG2, which only contained lugM. There was a nonsense mutation (g.308T > A) in lugM in strain RMLUG2. Six frameshift mutations were identified in all strains in the genes encoding NRPS enzymes. All frameshift mutations resulted from the deletion of one or two nucleotides, leading to premature termination codons. Additionally, an 18 bp deletion was identified at the beginning of the putative regulator lugR gene in our five strains. An insertion segment, g.67_68insATTTTATACAGGAAGAAG, was identified in lugZ of strains RMLUG4 and RMLUG5. Among all strains, 34 of the 39 genomes in the GenBank database contained 14 complete lug genes, two of which belonged to the same type of strain (NCTC12217) despite different assembly levels from different laboratories. Similar to strain RMLUG2, strains VCU150, VISLISI_25, and C_33 harbored only lugM, but had missense mutations instead of the nonsense mutation. Further analysis of the 38 strains indicated missense mutations in 12 of 14 genes (excluding lugI and lugD), making this the most common mutation type. Missense mutations were detected in lugJ (g.394G > A) in all CC1 isolates. Interestingly, missense mutations were found in lugE (g.331A > G) in 32 of the 38 S. lugdunensis strains, but were not present in any ST1 strain, except strain HKU09-01. Missense mutations in lugG (g.551C > T) were present in all CC6 isolates. In addition, genetic variations in the lug operon were independent of geographical origin. In the current study, the MIC tests and genomic analysis indicated discordance between genotype and phenotype, similar to the findings of Kao et al[5] There are several possible reasons for this discrepancy. First, the heterogeneity of the mecA gene can suppress methicillin resistance. Gargis et al[6] reported that a frameshift mutation in mecA and lack of the full mecA promoter sequence can cause susceptibility to oxacillin and cefoxitin, respectively. Additionally, Chen et al[7] reported that the –33C to T substitution within the mecA promoter can lower the oxacillin MIC, whereas this mutation was identified in strains RMLUG2(MSSL), RMLUG1(MRSL), and RMLUG6(MRSL). Second, mutations in auxiliary genes may contribute to decreased resistance to methicillin. Giannouli et al[8] reported that the accumulation of amino acid changes in FemXAB family proteins may affect cell wall synthesis, leading to atypical oxacillin responsiveness. Several mutations were detected in the femX, femA, and femB genes between strain RMLUG2 and S. lugdunensis JICS135, which might account for the lowered MIC. Moreover, we identified CC-dependent genetic variations of the 14 lug genes, indicating that the lug operon may not be conserved in the S. lugdunensis genome. Lebeurre et al[9] detected significant genetic variations independent of CCs in the lug locus. However, they only searched against genes encoding four NRPS enzymes (lugA, lugB, lugC, and lugD) and one regulator gene (lugR), with the results being somewhat consistent with our findings. Our results indicated that not all S. lugdunensis isolates harbor the lug operon. Four strains containing only lugM varied in clinical source and geographical location; thus, no link was evident between S. lugdunensis lug polymorphisms and geographical origin. There were some limitations to this study. First, we only described the phenotypes and genotypes of the six strains, which could not establish a causal relation. Functional studies are needed to investigate the activity of resistance genes and the lug operon. Second, the GenBank data may be biased because some sequences were obtained only by third-generation sequencing without assembling sequence fragments obtained by other methods. Thus, the accuracy of the sequences should be taken into account. In particular, some nucleotides need to be corrected to confirm the mutations of the lug operon in the future. Overall, our findings revealed variations in the genomes of S. lugdunensis strains isolated from different cities in China. We report novel SCCmec and SCCRMLUG4 elements, suggesting the need for additional studies on this species. Comparative analysis of the lug operon for all available genomes demonstrated CC-dependent genetic variations; however, further research is needed to elucidate the relationships between genotypes and phenotypes. Data availability The complete genome sequences have been deposited at GenBank under the accessions CP084480-CP084483 (RMLUG1), CP084434-CP084436 (RMLUG2), CP084437-CP084439 (RMLUG3), CP084440-CP084442 (RMLUG4), CP084443 (RMLUG5), and CP084444-CP084446 (RMLUG6). Acknowledgments The authors thank Pengcheng Du (Beijing YuanShengKangTai (ProtoDNA) Genetech Co Ltd.) for the technical help in Oxford Nanopore sequencing. We would like to thank Jing Wu for the collection of patient characteristics. We would like to thank Dr. Ronghua Liu (Microbiology Laboratory of Linfen Central Hospital, Linfen, Shaanxi, China) for antibiotic susceptibility test. Funding This work was supported by grants from the National Science and Technology Major Project of China (No. 2017ZX10103004-006) and the National Key Research and Development Programme of China (No. 2016YFC0903800). Conflicts of interest None.