Search Results (42)

Background: Pulmonary tuberculosis (TB) is a significant trigger of acute exacerbations of chronic obstructive pulmonary disease (AECOPD), so its timely and accurate diagnosis is essential. Also, the risk factors for TB occurrence in this population remain unclear. This study aimed to evaluate the performance of metagenomic next-generation sequencing (mNGS) for TB diagnosis in AECOPD patients, as well as to identify the associated risk factors. Methods: A retrospective observational cohort of 659 AECOPD patients with suspected pulmonary infection was enrolled. The microbial cell-free nucleic acids in bronchoalveolar lavage fluid samples were extracted and subjected to mNGS detection. The clinical data for each patient were collected from the hospital information system. The statistical analyses were performed with SPSS version 25.0. Results: A total of 170 cases, included for final analyses, were categorized into TB (n = 41), bacterial infection (n = 73), and non-infective control (n = 56) groups. Among these groups, the TB group had the highest intensive care unit (ICU) admission rate (46.34%) and longest median hospital stay (19.50 days) (p < 0.01). For TB diagnosis, mNGS demonstrated a greater sensitivity (86.00%), a lower specificity (93.30%), and a higher area under the curve (AUC, 0.877) than TB-DNA detection (70.21%, 100%, 0.848, respectively) and Xpert Mycobacterium tuberculosis/rifampicin (MTB/RIF) assay (63.83%, 100.00%, 0.870, respectively). Notably, mNGS identified the bacterial or viral co-infections in 18.00% of TB cases. Furthermore, the stringently mapped read number determined by mNGS showed a positive correlation with ICU admission rate (r = 0.76) and in-hospital mortality (r = 0.77). The lower body mass index (BMI) and reduced natural killer (NK) cell count were identified as the independent risk factors in the TB group (both p < 0.05). Conclusions: For the diagnosis of pulmonary TB in AECOPD patients, mNGS demonstrated comparable performance to TB-DNA detection and Xpert MTB/RIF assay, and also mNGS identified co-infections. In addition, a lower BMI and reduced NK cell count were identified as the independent risk factors for TB occurrence in this cohort. Full article

Type IV pili are widespread and multifunctional filamentous nanomachines that contribute to bacterial motility, adhesion, surface sensing, DNA uptake, biofilm formation, and, in some organisms, extracellular electron transfer. Owing to their small diameter, rapid dynamics, and sensitivity to experimental perturbation, type IV pili have historically been difficult to visualize in their native states. Recent advances in cryo-electron microscopy (cryo-EM), cryo-electron tomography (cryo-ET), fluorescence-based live-cell imaging, and label-free approaches such as interferometric scattering microscopy (iSCAT) have substantially expanded our ability to observe type IV pili across spatial and temporal scales. In this review, we summarize how these visualization strategies have reshaped current understanding of type IV pili, from conserved filament architecture and envelope-spanning assembly machineries to force-dependent behaviors and context-specific physiological functions. We further discuss how imaging-informed knowledge may support translational efforts, including antivirulence intervention, vaccine design, bioelectronic optimization, and microbial engineering, while emphasizing the current limitations of these applications. By integrating structural, dynamic, and functional perspectives, this review aims to provide a coherent framework for future studies of type IV pili in microbiology, infection biology, and biotechnology. Full article

Invasive pulmonary mold infections (IPMIs) are critical complications in immunocompromised patients, contributing significantly to morbidity and mortality. Diagnosing pathogens like Aspergillus species (spp.) and the Mucorales remains challenging due to non-specific clinical presentations and the limitations of traditional culture methods. This review provides an up-to-date synopsis of IPMI diagnostic tools, focusing on their diagnostic performance, turnaround time (TAT), and cost-effectiveness. We conducted a narrative review of the current literature regarding clinical evaluation, radiographic findings, invasive diagnostics, and non-invasive assays, including next-generation sequencing (NGS) and volatile organic compounds (VOCs). Chest computerized tomography (CT) remains a vital first step, though classic signs like the “halo” or “reverse halo” are neither sensitive nor specific. Traditional diagnostics are limited by low sensitivity and delayed results. While plasma microbial cell-free DNA (mcfDNA) NGS offers rapid TAT (24–48 h) and high specificity, its suboptimal sensitivity for Aspergillus spp. (<50%) and high cost remain significant barriers. Investigational VOC “breath tests” show promising sensitivity (77–96%) but lack standardization. Future research must prioritize the standardization of non-invasive microbiologic testing modalities, particularly those with rapid TAT such as bedside “breath tests” and high-throughput mcfDNA NGS. Development of clinical algorithms that balance cost-effectiveness with timely pathogen diagnosis based on the patient’s degree of immunosuppression is essential to improve survival in high-risk populations. Full article

Background: Blood-derived circulating cell-free microbial DNA (cfmDNA) has emerged as a potential non-invasive biomarker source for cancer detection. However, low biomass and high susceptibility to analytical variability raise concerns regarding the stability and interpretability of inferred microbial signatures. This study aimed to evaluate how different taxonomic profiling strategies influence downstream machine learning-based classification and feature interpretation in lung cancer. Methods: cfDNA sequencing data from 168 individuals (80 lung cancer patients and 88 non-cancer controls) were analyzed using two taxonomic profiling workflows: a Bracken-based abundance estimation approach and a BLAST-refined alignment-based strategy. Microbial profiles derived from each pipeline were evaluated using supervised machine learning models within a nested cross-validation framework. Feature stability and fold-change trends were compared across profiling strategies. Results: A Random Forest model achieved robust classification performance under both workflows (AUC 0.852 for Bracken-derived data and 0.906 for BLAST-derived data). However, substantial pipeline-dependent variation was observed in feature selection patterns and quantitative fold-change directionality. Although 13 genera were consistently selected across cross-validation folds in both workflows, the magnitude and direction of abundance differences were not uniformly concordant. Conclusions: Blood-derived microbial DNA profiles can support machine learning-based lung cancer classification; however, feature-level interpretation remains sensitive to taxonomic assignment strategy. These findings underscore the importance of pipeline-aware interpretation and methodological transparency in low-biomass blood microbiome research. Full article

Background/Objectives: The gut–brain axis is increasingly recognized as a critical modulator of cognitive function. This study investigated the neurotoxic effects of combined exposure to bacterial lipopolysaccharide (LPS) and the antiretroviral drug zidovudine (ZDV) in a mouse model, and evaluated the protective potential of two probiotic interventions: Bacillus subtilis and a mixture of lactobacilli. Methods: Cognitive function was assessed using the Morris water maze (MWM). Gut microbiota composition was analyzed by 16S rRNA sequencing, and intestinal morphology was examined histologically. Gene expression of neuroinflammatory markers and mitophagy-related genes in brain tissue was quantified by RT-PCR. Plasma levels of cell-free mitochondrial DNA (cf-mtDNA) were measured as a marker of mitochondrial damage. Results: Combined LPS + ZDV exposure induced systemic inflammation, impaired spatial memory, damaged the intestinal mucosa, and caused dysbiosis characterized by an increase in pro-inflammatory Muribaculaceae. In the brain, LPS + ZDV significantly upregulated Tnfa expression, confirming neuroinflammation. Bacillus subtilis administration prevented cognitive deficits, maintained Tnfa at control levels, and significantly reduced Il1b and Il6 expression compared to the LPS + ZDV group. This was accompanied by activation of the PINK1/PTEN-dependent mitophagy pathway, prevention of cf-mtDNA release, and restoration of gut microbial diversity. In contrast, the Lactobacilli mixture not only failed to improve outcomes but was associated with exacerbated intestinal damage, more pronounced cognitive dysfunction, and no reduction in neuroinflammatory markers. Conclusions: Combined exposure to LPS and ZDV induces gut–brain axis dysfunction characterized by neuroinflammation, cognitive impairment, intestinal damage, and dysbiosis. Bacillus subtilis effectively preserves cognitive function through activation of PINK1/PTEN-dependent mitophagy and suppression of neuroinflammation, highlighting its potential as a therapeutic candidate for cognitive impairments associated with gut–brain axis dysfunction. The contrasting effects of the lactobacilli mixture underscore the critical importance of strain-specificity in probiotic interventions. Full article

Photosynthetic algae–microbial fuel cells (PAMFCs) are attractive for energy-positive wastewater treatment and carbon mitigation. However, PAMFC performance under continuous flow is often constrained by limited cathodic electron-acceptor supply and unstable photosynthetic biofilms, while the extent to which cathode interfacial engineering can stabilize diurnal power output and assimilative NH₄⁺–N removal remains unclear. In this study, the sponge-like and petal-like ZnO_0.2-NiO@rGO-modified carbon fibers (ZnO_0.2-NiO@rGO-pCFs and ZnO_0.2-NiO@rGO-pCFp) and pre-fabricated carbon felt (pCF) were used as cathode materials to construct three sets of PAMFC systems. Under light–dark cycling, the engineered cathodes reached steady operation within about 6.5 d and increased the steady-state voltage to approximately 0.35 V, compared with approximately 0.08 V for pCF. Under continuous-flow conditions, cathodic NH₄⁺–N removal exhibited a stable diurnal rhythm, with higher removal during illumination at about 43–51% than in the dark at about 29–30%, consistent with algal assimilation as the primary nitrogen sink, while cathode modification mainly improved the cathodic microenvironment and response stability. Compared with pCF, the ZnO_0.2–NiO@rGO cathode enriched a more even, Chlorophyta-dominated algal biofilm with an approximate relative abundance of 80%, indicating that its selective interfacial environment favors biofilm stabilization and sustains in situ oxygen production and cathodic electron-acceptor supply. Consequently, the composite cathode enhanced voltage output and stabilized light-enhanced, assimilative NH₄⁺–N removal under aeration-free operation, while establishing an interpretable link between electrochemical performance and 18S rDNA-derived community assembly features, thereby providing a low-cost cathode design basis for nitrogen removal in wastewater treatment. Full article

Bloodstream infections and sepsis necessitate rapid, sensitive, and clinically relevant diagnostics to minimize treatment delays and improve clinical outcomes. Next-generation sequencing enables culture-independent pathogen detection, antimicrobial resistance profiling, and genome-informed epidemiology. This narrative review integrates clinical evidence with practical workflows across three complementary approaches. We describe the use of plasma microbial cell-free DNA for broad organism detection and burden monitoring, as well as metagenomic next-generation sequencing of blood or plasma for unbiased pathogen discovery, including culture-negative and polymicrobial infections. Same-day Oxford Nanopore Technologies sequencing of positive blood culture broth is also discussed as a way to accelerate species identification, targeted resistance reporting, and infection-prevention decisions. We outline the sample-to-result steps, typical turnaround time (TAT), and stewardship-aligned decision points. Analytical reliability depends on effective reduction in human DNA background, stringent control of background and reagent-derived nucleic acids in low-biomass samples, and documented and validated bioinformatics workflows that are supported by curated taxonomic and resistance databases. Quantitative reports should adhere to validated thresholds and should be interpreted in the context of internal controls and clinical pretest probability. Ongoing challenges include variable correlation between genotype and phenotype for specific pathogen and antibiotic pairs, interpretation of low-level signals, and inconsistent regulatory and reimbursement environments. Advances in portable sequencing, faster laboratory and analytical workflows, and scaled liquid biopsy strategies may further reduce the TAT and expand access. Integrating these tools within One Health frameworks and global genomic surveillance programs could support early resistance detection and coordinated public health action, which could help to advance sepsis care toward more precise treatment and real-time infection control insights. Full article

Background/Objectives: In this study, three antimicrobial peptides (1–3) were conjugated onto bare aluminum nanoparticles (NP) to produce peptide-conjugated nanoparticles (NP1–NP3) in order to evaluate their biological effects. Methods: The peptide-functionalized Al₂O₃ nanoparticles were characterized and subsequently analyzed for their antimicrobial activity against selected bacterial strains. The findings were compared with those of bare Al₂O₃ nanoparticles and free antimicrobial peptides. Through this comparison, the enhanced impact of combining nanoparticles with peptides in addressing antimicrobial resistance was demonstrated. Additionally, biofilm inhibition, microbial cell viability inhibition, DNA cleavage, antioxidant, and amylolytic activity assays were performed to comprehensively evaluate the biological functionality of the synthesized nanoparticles. Results: Although all tested samples exhibited significant antimicrobial activity, peptide-conjugated nanoparticles NP1, NP2, and NP3 provided superior activity with an MIC value of 16 mg/L. The highest biofilm inhibition activities were observed for NP2 as 53% and 70% against S. aureus and P. aeruginosa, respectively. Additionally, NP1–NP3 inhibited microbial cell viability by 100% at a concentration of 6.25 mg/L and free peptide 3 displayed E. coli inhibition as 100% at a concentration of 12.5 mg/L. Furthermore, we evaluated the biological potential of antimicrobial peptide-functionalized Al₂O₃ nanoparticles through antibiofilm, antioxidant, antidiabetic activities, and DNA cleavage assays. Peptide-conjugated nanoparticles NP1, NP2, and NP3 exhibited the highest antioxidant activities as 43.70%, 45.22%, and 59.57%, respectively. Except for NP3, the compounds were observed to act as α-amylase enzyme activators. NP and NP1–NP3 completely degraded the supercoiled circular form into small pieces. Conclusions: Our findings suggest that peptide–aluminum nanoparticle conjugation may be a promising formulation for enhancing biological activity. Further in vitro and in vivo tests may help clarify the therapeutic potential of this novel nanoformulation. Full article

Malaria remains a considerable challenge to international health, especially in returning travelers from endemic regions where exposure risk may be downplayed. Prompt and accurate diagnosis is crucial, especially when conventional diagnostic techniques are insufficient. This case report presents a 59-year-old man who developed fever, rash, and myalgia after returning from the Amazon rainforest. Initial laboratory tests demonstrated leukopenia, thrombocytopenia, transaminitis, and hyperbilirubinemia. Despite these abnormal results and a clinically suspicious presentation, malaria smears were negative. Since the symptoms did not resolve, a Karius test—a plasma-based microbial cell-free DNA sequencing assay—successfully detected the presence of Plasmodium vivax, thus establishing the diagnosis. The patient needed several treatment regimens for the recurrent attacks, including chloroquine and primaquine, artemether-lumefantrine, and eventually a combination of quinine and doxycycline together with a prolonged course of primaquine. His symptoms resolved completely after the last treatment regimen, along with the normalization of the blood counts and liver function tests. This case demonstrates the limitations of smear microscopy diagnosis in P. vivax infections, highlights the role of molecular diagnostics like the Karius test, and stresses the importance of preventing relapses with adequate hypnozoite clearance. It further highlights the importance of clinician awareness and diligent follow-up in cases of travel-related Malaria, especially those with unusual presentations or recurrent symptoms. Full article

Background/Objectives: 16S ribosomal RNA sequencing has, for several years, been the main means of identifying bacterial and archaeal species. Low-throughput Sanger sequencing is often used for the detection and identification of microbial species, but this technique has several limitations. The use of high-throughput sequencers may be a good alternative to improve patient identification, especially for polyclonal infections and management. Kraken 2 and KrakenUniq are free, high-throughput tools providing a very rapid and accurate classification for metagenomic analyses. However, Kraken 2 can present false-positive results relative to KrakenUniq, which can be limiting in hospital settings requiring high levels of accuracy. The aim of this study was to establish an alternative next-generation sequencing technique to replace Sanger sequencing and to confirm that KrakenUniq is an excellent analysis tool that does not present false results relative to Kraken 2. Methods: DNA was extracted from reference bacterial samples for Laboratory Quality Controls (QCMDs) and the V2-V3 and V3-V4 regions of the 16S ribosomal gene were amplified. Amplified products were sequenced with the Illumina 16S Metagenomic Sequencing protocol with minor modifications to adapt and sequence an Illumina 16S library with a small 500-cycle nano-flow cell. The raw files (Fastq) were analyzed on a commercial Smartgene platform for comparison with Kraken 2 and KrakenUniq results. KrakenUniq was used with a standard bacterial database and with the 16S-specific Silva138, RDP11.5, and Greengenes 13.5 databases. Results: Seven of the eight (87.5%) QCMDs were correctly sequenced and identified by Sanger sequencing. The remaining QCMD, QCMD6, could not be identified through Sanger sequencing. All QCMDs were correctly sequenced and identified by MiSeq with the commercial Smartgene analysis platform. QCMD6 contained two bacteria, Acinetobacter and Klebsiella. KrakenUniq identification results were identical to those of Smartgene, whereas Kraken 2 yielded 25% false-positive results. Conclusions: If Sanger identification fails, MiSeq with a small nano-flow cell is a very good alternative for the identification of bacterial species. KrakenUniq is a free, fast, and easy-to-use tool for identifying and classifying bacterial infections. Full article

Background: The Karius Test (KT), a microbial cell-free DNA next-generation sequencing assay, is increasingly utilized in challenging infectious syndromes. However, its real-world clinical utility and cost-effectiveness remain uncertain. Methods: We conducted a retrospective review of 88 KT results from adult patients at UC San Diego Health between July 2017 and April 2024. Each case was evaluated for clinical impact using standardized criteria. We analyzed diagnostic yield, turnaround time, number needed to test (NNT), and institutional billing data for reimbursement and cost implications. Results: Of 88 unique tests, forty-nine (55.7%) identified at least one pathogen. Eleven (12.5%) had a positive clinical impact—eight provided the only microbiologic diagnosis, and three were faster than conventional methods. Vascular/graft infections showed the highest yield. Twenty-one tests had a neutral impact; fifty-six showed no clinical benefit. The Median turnaround time was 3 days. The NNT was 6.1 or 2.75 including neutral cases. Cost analysis revealed a substantial financial burden without transparent reimbursement mechanisms in inpatient settings. Conclusion: The KT demonstrates modest clinical utility with noteworthy benefits in select scenarios. Given its high cost and variable impact, we advocate for diagnostic stewardship led by infectious disease specialists to optimize test use and minimize unnecessary expense. Full article

Background/Objectives: The main objective of the study is to assess the clinical utility of microbial cell-free DNA (mcfDNA) in neutropenic patients diagnosed with acute myeloid leukemia (AML) undergoing chemotherapy in the outpatient setting. Neutropenia is a common complication in this patient cohort and enhances the risk of fatal opportunistic bacterial and fungal infections. Accurate and timely diagnosis of these infections in outpatient asymptomatic individuals is critical. Methods: Fourteen patients were studied in this prospective observational case series. Traditional blood cultures (BCs) were obtained when clinically indicated and blood samples were collected for plasma mcfDNA metagenomic sequencing up to two times a week at outpatient oncology appointments. Results were compared in identifying potential infectious agents. Results: BCs identified pathogens in only two patients, despite several cases where infection was suspected. In contrast, mcfDNA testing detected pathogens in 11 of the 14 patients, including bacteria, such as Staphylococcus aureus, and invasive fungi, such as Candida and Aspergillus species, and Pneumocystis jirovecii. Conclusions: In the outpatient setting, mcfDNA surveillance offers a more reliable method for detecting pathogens. This approach identified actionable microbiologic results in immunocompromised individuals who did not meet standard clinical criteria for suspicion of infection. Further research is required to confirm the potential of mcfDNA surveillance in an outpatient setting to guide more accurate treatment decisions, reduce extensive clinical investigations, and improve neutropenic patient outcomes. Full article

Osteoporosis is a systemic bone disorder characterized by decreased bone mass and deteriorated microarchitecture, leading to an increased risk of fractures. Recent studies have revealed that its pathogenesis involves complex biological processes beyond bone remodeling, including oxidative stress, chronic inflammation, cellular senescence, osteoimmunology, gut microbiota alterations, and epigenetic modifications. Oxidative stress disrupts bone homeostasis by promoting excessive free radical production and osteoclast activity. Chronic inflammation and the accumulation of senescent cells impair skeletal repair mechanisms. Advances in osteoimmunology have highlighted the critical role of immune–bone crosstalk in regulating bone resorption and formation. Moreover, the gut–bone axis, mediated by microbial metabolites, influences bone metabolism through immune and endocrine pathways. Epigenetic changes, such as DNA methylation and histone modification, contribute to gene–environment interactions, affecting disease progression. Multi-omics approaches (genomics, proteomics, and metabolomics) systematically identify molecular networks and comorbid links with diabetes/cardiovascular diseases, revealing pathological feedback loops that exacerbate bone loss. In conclusion, osteoporosis pathogenesis extends beyond bone remodeling to encompass systemic inflammation, immunometabolic dysregulation, and gut microbiota–host interactions. Future research should focus on integrating multi-omics biomarkers with targeted therapies to advance precision medicine strategies for osteoporosis prevention and treatment. Full article

Infective endocarditis (IE) remains a diagnostic challenge, particularly in cases where microbiological diagnosis is not established. Advances in molecular diagnostics have expanded the ability to identify causative pathogens beyond traditional culture-based methods. This review explores the role of molecular assays, including pathogen-specific PCR, multiplex PCR, broad-range PCR, and shotgun metagenomic sequencing, in diagnosing IE. These molecular techniques enhance pathogen detection, especially in patients with prior antibiotic exposure, and improve diagnostic accuracy in culture-negative IE. Broad-range PCR assays and metagenomic sequencing offer the untargeted detection of a wide spectrum of organisms. Despite their advantages, limitations such as availability, interpretation challenges, and a lack of antimicrobial susceptibility testing remain. A multimodal approach integrating molecular diagnostics with conventional methods is essential to optimize patient management. Further research is needed to refine diagnostic algorithms and improve cost-effectiveness in clinical practice. Full article

The presence and biological role of extracellular DNA (eDNA) have been explored in diverse microbial environments. Nonetheless, it has not been studied in the chicken gut microbiome. This study aims to investigate eDNA in the chicken cecum, analyzing cecal samples from broiler chickens using three preparation methods: Whole Cecal Suspension (WCS), Washed Cell Pellets (WCP), and Cell-Free Supernatant (CFS). The 16S rRNA gene-based microbiota analysis revealed distinct microbial communities in CFS compared to WCS and WCP (p = 0.001). Notably, specific taxa, including Anaerofilum, Anaerotruncus, Oscillospira, Syntrophomonas, and Delftia, were enriched in CFS. Confocal fluorescence microscopy, employing stains such as Propidium Iodide (PI), GelGreen, and SYTO 9, confirmed the presence of eDNA with filaments observed in WCS and CFS. Colocalization of PI and GelGreen™ validated the extracellular nature of eDNA, while DNase I treatment selectively degraded eDNA, further confirming its extracellular nature. Our findings in this study highlight the presence of eDNA in the chicken cecal microbiome, and the presence of eDNA associated with specific taxonomic groups suggest that it might play a specific role in the biological function of the cecal microbiome, which warrants further investigation in the future. Full article