Abstract: Managing safety and operational efficiency in large-scale events requires tools capable of capturing complex crowd dynamics while supporting rapid and informed decision-making. This paper presents a Generative AI-powered digital twin framework that integrates agent-based crowd simulation, an API-based execution pipeline, and a Large Language Model (LLM)-driven conversational interface within a unified system. The proposed approach enables dynamic configuration, execution, and analysis of crowd scenarios under varying operational conditions, including high-demand and emergency evacuation contexts. Experimental results demonstrate the system’s ability to reproduce nonlinear crowd dynamics, detect congestion patterns, and evaluate evacuation performance, providing actionable insights for planning and safety assessment. A key contribution lies in the introduction of an API-based execution paradigm that exposes the full simulation lifecycle (configuration, validation, execution, and output retrieval) through programmatic interfaces, enabling reproducible and scalable what-if analysis. Additionally, the integration of an LLM-based conversational interface allows non-technical users to interact with complex simulation models through natural language, significantly improving accessibility and usability. The framework is validated through a TRL-4 prototype, demonstrating robust performance, scalability, and interaction reliability. Overall, the proposed system advances digital twins from static analytical tools to executable, interactive, and user-centric platforms for decision support in complex urban environments.

Abstract: Nowadays, the organisational landscape aiming to provide value through their product and service offerings relies on having the infrastructure necessary to deliver at the expected service levels, as well as contributing to business continuity and organisational resilience in the face of modern organisational performance disruptions. This requires appropriate adaptation of existing frameworks, methods, and models to their business models which have generated consistent deliverables across time and industries. The same is applicable for the Romanian Information Technology (IT) organisations, which face increasing pressure to deliver within the expected quality, time, and budget parameters. Therefore, this paper aims to assess how stakeholder relationship management components, viewed through the Malcolm Baldrige National Quality Award (MBNQA) excellence framework, with impact on organisational quality and its contribution to business continuity and organisational resilience in Romanian IT organisations. This is a pilot-type study with a sample of N = 52 participants, to explore the applicability of the MBNQA framework within the Romanian IT sector. The results suggest that the four components of MBNQA focused on stakeholders (Leadership and Governance, Workforce, Customers and Markets, Community Engagement) may be suitable to be considered in assessment tools on the Romanian IT market. The “Workforce” variable emerges as the strongest area to focus on for achieving quality in stakeholder relationship management (SRM). Given its pilot delimitation, this study provides can be seen as providing an initial foundation for applying MBNQA in a specific regional IT context. While limited by sample size and geographic focus, the findings justify expanding the research to include broader population segments. Future research could transition from this correlational design to longitudinal frameworks to validate the associations across other multiple geographical markets.

Abstract: Previous studies on rats showed a deterioration of left ventricular (LV) function and myocardial injury characterized by oxidative/nitrosative stress, PARylation, and apoptosis in the heart after three days of hypoxia. In the present study on rats, we investigated whether a three-day recovery period in normoxia can reverse myocardial injury and dysfunction. Further, we studied the effects of norepinephrine (NE) administration as a model of strong sympathetic activation on hypoxia-induced LV dysfunction and myocardial damage, as well as their reversibility. Three days of normobaric hypoxia (10% O2) significantly decreased LV systolic function. Contrary to our expectations, NE infusion even aggravated the depression in LV function. These dysfunctions were completely reversed after three days of normoxic recovery. In contrast, nitrotyrosine as a marker of oxidative/nitrosative stress receded only partially, and poly-ADP-ribose (PAR) increased even further during the recovery period. Apoptosis-inducing factor receded at least partially indicating that PAR-related apoptosis (parthanatos) is not a major cause of hypoxia-induced LV dysfunction. Additional administration of NE mildly aggravated oxidative/nitrosative stress but did not significantly intensify PARylation and consequently, parthanatos. The findings demonstrate that hypoxia-induced LV dysfunction is reversible suggesting that subchronic hypoxia and subsequent reoxygenation has a better prognosis for the LV than classical ischemia/reperfusion injury.

Abstract: The pronounced aero-hydrodynamic coupling effects of modern Wind-Assisted Propulsion System (WAPS) ships challenge the applicability of traditional stability frameworks, which are predicated on hydrostatic energy balance, in satisfying the dynamic constraints of the Second Generation Intact Stability Criteria (SGISC). This paper systematically reviews the methodological evolution of dynamic stability assessments for WAPS ships under extreme and damaged conditions. By introducing a "Hierarchy of Evidence" evaluation framework, this study delineates the applicability boundaries of aerodynamic Reduced-Order Models (ROM), extended 3/4-DOF maneuvering equations, and 6-DOF time-domain hybrid architectures, defining the role of high-fidelity CFD-VPP in establishing calibration benchmarks. The review also discusses the damping distortion mechanisms induced by multiphase flow sloshing under damaged conditions. Synthesized findings indicate that transitioning towards a 6-DOF time-domain coupled architecture provides clear advantages for capturing unsteady aerodynamic hysteresis and nonlinear interference. Meanwhile, surrogate models, such as Physics-Informed Neural Networks (PINNs), offer a potential pathway to mitigate the computational demands associated with long-term extreme value extrapolations. Ultimately, this review provides a methodological reference for the high-fidelity assessment of WAPS and the development of Digital Twin systems.

Abstract: Aging bridge infrastructure and limited inspection resources have created an urgent need for automated and reliable bridge condition assessment systems. Most existing deep learning-based inspection approaches detect damage types from images without considering the structural member on which the damage occurs, limiting their practical utility for maintenance decision-making. This study proposes a structure-aware deep learning framework for automated bridge inspection that integrates structural member segmentation, multiclass damage detection, and spatial damage-to-member association within a unified pipeline. A SegFormer-based semantic segmentation model was trained on a custom bridge inspection dataset comprising 1,339 images to identify three primary structural member classes — main girder, deck slab, and abutment — achieving a test mean Intersection over Union (mIoU) of 0.851. Boundary refinement using the Segment Anything Model (SAM) in mask-prompt mode was applied to improve mask precision during training data preparation. A YOLOv8s object detection model was trained on a custom bridge damage dataset of 6,531 images to detect two damage classes — crack and corrosion — achieving a mean Average Precision (mAP50) of 0.445 at a confidence threshold of 0.30. The framework associates detected damage with segmented structural members using a region-based spatial assignment strategy, enabling structure-aware outputs such as “crack on main girder” and “corrosion on deck slab.” Manual evaluation on 100 bridge inspection images demonstrated a damage detection accuracy of 70.0% fully correct and 84.0% fully or partially correct, and a member assignment accuracy of 62.0% fully correct and 87.0% fully or partially correct. The main girder class achieved the highest combined accuracy for both damage detection (90.9%) and member assignment (93.9%). These results demonstrate the potential of the proposed framework for practical automated bridge inspection and infrastructure monitoring applications.

Abstract: 1) Background: Coumestrol is a bioactive compound that inhibits HASPIN activity and prevents tau and H3 phosphorylation. Oral ingestion of CBSs increases blood testosterone levels, which decline with age causing late-onset hypogonadism. Oral ingestion of coumestrol-rich bean sprouts (CBSs) has been shown to suppress the onset of Alzheimer’s disease in 5xFAD mice and the onset of colon cancer in APC^min/+ mice. 2) Methods: We investigated the effect of oral ingestion of CBSs on the progression of aging in male senescence-accelerated prone 1 (SAMP1) mice allowed voluntary exercise or no exercise. The SAMP1 mice were divided into two groups fed either a standard diet or a diet including bean sprouts from 12 to 18 weeks of age. Each group was divided into two groups with voluntary exercise or no exercise. 3) Results: Voluntary exercise accelerated aging-related declines in blood testosterone levels, nerve growth factor levels, and spatial working memory, and oral ingestion of CBSs suppressed these age-related phenotypes, regardless of exercise. 4) Conclusion: Ingestion of CBSs prevented aging-related phenotypes in the experimental mice. A detailed analysis of the molecular mechanisms of coumestrol will be useful for understanding aging and preventing age-related diseases such as cancer, Alzheimer’s disease, and LOH.

Abstract: Life is a self-organizing and self-sustaining process that involves the transformation of energy, primarily regulated by the brain. The brain is made up of terminally differentiated postmitotic cells, and their proper functioning and longevity depend entirely on energy and glucose-fueled metabolism. Glucose metabolism is essential for maintaining normal neuronal function, homeostasis, and repair processes. Neurodegenerative changes begin to develop when glucose metabolism is insufficient. Brain involution often progresses in neuronal structures that have the highest levels of energy metabolism and activity. The primary age-related changes occur in the hypothalamus and in the GABA/glutamatergic systems of the cerebral cortex. These changes are closely linked to the age-related decline of nicotinamide adenine dinucleotide (NAD), which is a key mechanism that impairs glucose metabolism and leads to programmed neuronal death. Early warning signs typically include a combination of dysfunctions related to the affected areas. Research so far suggests that early supplementation with NAD precursors may slow the processes of aging and neurodegeneration.

Abstract: Background: Postural problems such as head forward posture, thoracic hyperkyphosis and lumbal hyperlordosis, when seen together, further complicate postural control, increasing the importance of comprehensive approaches in treatment. This study aims to examine the effect of 6 weeks telerehabilitation with web-based exercises and compare the home based exercises in individuals with postural problems. Trial Design: A Randomized Controlled Study. Methods: 34 volunteers with postural deformity among young adults were randomly divided into telerehabilitation (n=17) and control (n=1) groups. Craniovertebral, thoracic kyphosis, and lumbal lordosis angles of all individuals were evaluated with a smartphone application (Clinometer+ Bubble), hamstring, and pectoral muscle shortness with a goniometer, and trunk muscle endurance with endurance tests created by McGill and Sorenson. Whilst the tele-rehabilitation group was provided with a videobased exercise programme, the control group was advised to follow the same exercise programme at home. Exercises were performed 3 days a week for 6 weeks, 1 hour session. Participants in the telerehabilitation group were followed up with synconised video conference. Results: A significant difference was observed in the telerehabilitation group in muscle shortness and the endurance tests (p<0.05). Only a significant difference in left (p=0.03) and right (p=0.04) muscle shortness was observed in the home exercise group. Significant differences were observed in Craniovertebral and lumbal lordosis angles between groups (p<0.05), with the telerehabilitation group showing better outcomes. When examined kyphosis angle, muscle shortness, and endurance tests between groups were found to be similar (p>0.05). Conclusions: Six weeks of telerehabilitation can improve muscle shortness and trunk endurance in young adults with postural deformities. Both the exercise program using telerehabilitation and the home exercise program were beneficial for individuals with postural problems, with more favorable effects observed in the telerehabilitation group.

Abstract: Female rugby league performance is influenced by multiple interacting systems; however, the extent to which sensory and autonomic function differentiates playing level remains unclear. This study investigated whether visual, vestibular, somatosensory, and auto-nomic performance differ by playing level and positional group in female rugby league athletes. Elite and sub-elite athletes completed lower-limb proprioception testing using an Active Movement Extent Discrimination Assessment protocol, alongside visual–vestibular and autonomic measures obtained via a virtual-reality eye-tracking system. Bayesian hierarchical models were used to examine the effects of playing level, positional group (adjustables, backs, forwards), and their interaction, with posterior inference based on probability of direction and region of practical equivalence analyses. Interaction effects between level and position were observed for selected variables across somatosensory, vestibulo-oculomotor, and autonomic domains. Elite adjustables demonstrated higher ankle proprioceptive acuity than sub-elite adjustables (PD = 0.94), with additional interaction effects identified for vestibulo-oculomotor time on target (PD = 0.95) and autonomic dilation velocity (PD = 0.98). However, findings were not consistent across positional groups or outcome measures, and substantial within-group variability was evident. Overall, sensory and autonomic performance did not consistently differentiate playing level, suggesting limited utility for cross-sectional discrimination but potential value for longitudinal, individualised athlete monitoring.

Abstract: Academic stress is a growing public health concern in higher education, particularly during examination periods, when increased anxiety and psychological distress negatively affect students’ wellbeing and academic performance. In this context, scalable and accessible interventions that promote emotional regulation and reduce stress are increasingly relevant within university health promotion strategies. This study examines the perceived benefits, conditions, and limitations of dog-assisted interventions in higher education, based on the implementation of the StressLess pro-gramme at the University of Costa Rica. A mixed exploratory design was used, combining qualitative narratives with quantitative content analysis. A total of 51 videoecorded testimonies were analysed, including university students (n = 22) and professionals from diverse disciplines (n = 29). Results indicate predominantly positive perceptions across all dimensions, particularly in emotional support and stress reduction. The only statistically significant difference be-tween groups was found in emotional support, with higher ratings among students. Professionals were more likely to identify implementation conditions, risks, and potential cognitive benefits. These findings support the integration of dog-assisted interventions as complementary strategies within university public health frameworks and contribute to the limited evidence available in Latin American contexts.

Abstract: Despite millennia of successful biological reproduction, the daily execution of child-rearing remains notoriously fraught and highly resistant to optimization. Societyfrequently attributes parental burnout and daily perceived failures in parenting tasks topsychological shortcomings, a lack of patience, or organizational failure. In this paper,we propose a mathematically rigorous defense of the exhausted parent by modelingroutine domestic tasks as formal decision problems. We demonstrate that the pursuit of“Optimal Parenting” (OP) is fundamentally intractable (assuming P ≠ NP). Byperforming polynomial-time reductions from classic NP-complete problems—specifically 0-1 Integer Linear Programming, Maximum Independent Set, and MAX-3-SAT—to simplified models of moral development, contradictory behavioral curricula,and developmental milestones, we prove that OP is strictly NP-hard. Consequently, weestablish that finding a perfect, conflict-free parenting strategy requires non-deterministic polynomial time, vastly exceeding the processing capabilities of anybiological parent. Our results mathematically absolve caregivers of domestic guilt andformally validate constraint relaxation (colloquially known as “lowering expectations”,or simply “doing one’s best”) as a necessary and optimal heuristic for surviving acomputationally hostile environment.

Abstract: Organic farming in the European Union is strongly shaped by Common Agricultural Policy (CAP) support, yet participation durability remains less examined than supported organic area or organ-ic–conventional comparisons. This study assesses whether the length of participation in CAP-supported organic farming is associated with the organizational, production, and economic out-comes of organic farms in Poland. It applies a two-level approach: CAP support trajectories based on ARMA data for 2008–2025 and organic production duration based on Polish FSDN data for 2008–2022. The results show that organic farming in Poland is highly CAP-dependent and follows an unstable trajectory, with expansion up to 2012–2013, subsequent decline, and renewed growth after 2019. Longer participation is associated with differences in land resources, supported organic UAA, ANCs conditions, production organization, and livestock presence, indicating both adapta-tion and structural selectivity. FSDN data show that fully organic farms have lower land and labor productivity than conventional farms, but persistent fully organic farms achieve similar income per hectare when subsidies are included; without subsidies, their income remains much weaker. The findings indicate that the evaluation of organic farming support should move beyond benefi-ciary counts and certified organic area to include participation durability, production-system co-herence, economic viability, and territorial embeddedness. More differentiated instruments are needed to strengthen durable, knowledge-intensive, and territorially embedded organic farming systems.

Abstract: Biodegradable polymer systems based on poly(3-hydroxybutyrate) (PHB) and poly(butylene adipate-co-terephthalate) (PBAT) have attracted significant attention for fused deposition modeling (FDM)-based orthopedic applications due to their biodegradability, tunable mechanical behavior, and potential to reduce stress-shielding effects associated with metallic implants. However, the immiscibility of PHB/PBAT blends, limited melt stability, and poor balance between stiffness and ductility restrict their processability and functional performance. In this study, rheology was employed as the central design parameter to establish the relationship between reactive compatibilization, melt structure evolution, filament formation, printability, mechanical response, and degradation behavior in PHB/PBAT-based systems. PHB/PBAT blends containing varying Joncryl® ADR and MgO nanoparticle contents were prepared through reactive melt blending, followed by filament extrusion and FDM processing. FTIR analysis confirmed epoxy-mediated reactions between Joncryl and polyester chain ends, indicating chain extension, branching, and enhanced interfacial interactions. Rheological analysis demonstrated that reactive compatibilization significantly increased storage modulus, complex viscosity, melt elasticity, and relaxation times, particularly at low frequencies, indicating the formation of a more interconnected viscoelastic network favorable for stable filament extrusion and shape retention during FDM processing. Stress relaxation measurements further confirmed delayed stress dissipation and enhanced melt structural recovery in compatibilized systems. In contrast, MgO incorporation introduced rheological heterogeneity and altered relaxation dynamics through polymer-filler interactions and localized chain confinement. Mechanical characterization revealed a transition from brittle PHB behavior to ductile PBAT-rich systems. Among the investigated formulations, PHB/PBAT/J0.3 exhibited the most favorable balance between tensile strength, elongation, toughness, and filament stability, while excessive MgO loading reduced ductility and impact resistance despite modest stiffness enhancement. SEM observations demonstrated improved phase morphology and interfacial adhesion after reactive compatibilization, whereas MgO-containing systems exhibited increased structural heterogeneity. Thermal analysis showed that compatibilization modified crystallization behavior through chain branching and reduced crystallinity, while MgO influenced crystallization efficiency and degradation pathways. In vitro degradation in phosphate-buffered saline (PBS) solution at 37 °C demonstrated controlled degradation behavior and gradual pH evolution over 42 days. The results demonstrate that reactive compatibilization governs the viscoelastic state required for stable FDM processing and balanced mechanical performance, while MgO provides secondary control over stiffness and degradation behavior. The developed biodegradable PHB/PBAT-based systems show promising potential for additively manufactured orthopedic and biomedical applications where controlled degradation, flexibility, and processability are required.

Abstract: Background/Objectives: The semi-solid enteral nutrition method, in which semi-solid nutrients are administered through a gastrostoma rather than liquid nutrients, is a medical technology for nutritional management developed in Japan. This nutritional method has spread rapidly, and in 2018, the Home Semi-solid Enteral Nutrition Guidance and Management fee was newly established in the payment system for medical services. However, there are few studies evaluating this nutrition method in actual clinical practice. Methods: Using the DeSC medical claims database, we investigated patient characteristics, the number of fee calculations, and the length of hospital stay for all hospitalizations and for pneumonia during and after fee calculation. Results: Home semi-solid enteral nutrition guidance and management was received at least once by 3105 patients (mean age: 79.4 years). Cerebrovascular disease, nervous system disease, dementia, and pneumonia were the most common diseases at the start of guidance and management, and the mean number of sessions received was 7.7. The mean length of stay for any hospitalization was significantly shorter during the guidance and management period than after the last guidance and management session (8.2 vs. 29.1 days). Notably, the mean length of hospital stay for pneumonia was also significantly shorter during the guidance and management period than after the last guidance and management session (3.1 vs. 10.2 days). Conclusions: This study clarified the clinical status and outcomes of patients who received Home Semi-solid Enteral Nutrition Guidance and Management. The results suggested that application of the semi-solid enteral nutrition method can contribute to stable patient home care.

Abstract: The research paper explores the intersections of colonialism, apartheid, gender, sexuality, and nationalism through the narratives of women from Mpumalanga, in South Africa. I critically examine how these factors shape the identities and experiences of Black women in South Africa, especially in the wake of the country’s transition to democracy. I employ qualitative methods, engaging with women’s narratives alongside existing literature, to frame my analysis. This approach allows for an understanding of women’s embodied experiences and how these relate to broader socio-political contexts. By utilising personal narratives, this research highlights the lived realities of Black women, illuminating the ongoing effects of socio-political and cultural legacies on their sexualities and identities. The theoretical frameworks guiding my analysis are rooted in decolonial and African feminist scholarship. These frameworks challenge conventional narratives on gender and sexuality, emphasising the need to contextualise African experiences within their unique historical and socio-cultural landscapes. I draw on diverse scholarly contributions that examine the effects of the colonial gaze, gender roles imposed by colonial ideologies, and the historical narrative of Black women under surveillance. To conclude, through women’s personal narratives and Focus Group Discussions, my research aims to critically interrogate the historical constructions of gender and sexuality, foregrounding women's voices to deepen understanding of power dynamics in post-apartheid South Africa. Through this, I assert that the legacies of racial and sexual disparity continue to influence contemporary discourses and women’s lived experiences today.

Abstract: Background: Pumped-storage hydropower (PSH) plants remain a cornerstone of grid stability and renewable-energy integration in Europe, yet a large share of the European fleet is ageing and was built under design codes that predate modern geotechnical standards. The Dychów pumped-storage power plant (ESP Dychów, 88 MW, Lubusz Voivodeship, western Poland), in service since the 1930s, is classified as part of the national critical energy infrastructure. Its earthen retaining structures have a documented history of surface mass movements – including the 1997 landslide on the frontal dam, which damaged around 60 m of the crest road and the powerhouse – and require recurrent safety assessment in complex glaciotectonic settings. Methods: Electrical resistivity tomography (ERT) performed in a gradient array was combined with hydrogeological observations and finite-element slope-stability analyses in ZSoil. Two cross-sections were examined – the earthen frontal dam of the upper reservoir and the embankment of the derivation channel feeding the reservoir – each with four calculation variants and two groundwater scenarios. The shear-strength reduction technique was used to obtain the “global” safety factor SF. Results: SF equals 1.75 for the frontal dam and ranges from 1.80 to 2.10 for the channel embankment. Parametric reduction of the internal friction angle of saturated medium sand yields limit values of φ = 12.3° (dam) and φ = 20.3° (embankment), clearly below realistic in-situ values. Conclusions: Both structures comply with the safety threshold F ≥ 1.50 specified for Class I hydraulic engineering structures. The presented non-invasive ERT–FEM workflow offers a cost-effective tool for the periodic reassessment of ageing PSH infrastructure, supporting its continued role in balancing variable renewable generation in decarbonising power systems.

Abstract: One of the most important traditional dairy products in Kosovo is cheese. On the other hand, there are few data regarding the characterization and classification of cheese in Kosovo. The purpose of this paper is to identify and classify the varieties of cheese in Kosovo, depending on their physicochemical characteristics and properties. This paper collects samples of cheese from various regions in Kosovo and analyzes them regarding some of the most important physicochemical properties. The results showed that there are differences in their composition, which range from 42.5 to 74.5% in the moisture content, average 24.8% for the fat content of the cheese, and average 18.6% for the protein content of the cheese. There are differences about the kinds of milk used to produce these kinds of cheese, which are dominated by cow milk, followed by sheep, goat, and mixed milk. Cluster analysis grouped the cheeses into distinct categories corresponding to soft, semi-soft, and semi-hard varieties. As can be seen from the information provided above, one can notice that various kinds of cheese are available in Kosovo. Information provided above can be considered to serve as the scientific basis for standardization of traditional cheese products. Information provided above can be of major importance for quality control of cheese products in Kosovo.

Abstract: The 12-lead electrocardiogram is essential for cardiovascular diagnosis but limited by inter-observer variability, low sensitivity for subclinical disease, and labor-intensive telemonitoring analysis. Artificial intelligence (AI), particularly deep learning, addresses these constraints by extracting high-dimensional patterns that correlate with arrhythmias, structural abnormalities, and systemic conditions. This review synthesizes recent AI-enabled ECG advances, covering technical foundations—including foundation models and validation strategies—and clinical applications such as arrhythmia detection, structural heart disease identification, and digital biomarker derivation. We discuss emerging trends like self-supervised learning, multimodal integration, generative models, and explainability techniques. Furthermore, we address critical challenges regarding generalizability, algorithmic bias, privacy, and regulatory frameworks. Finally, we outline research priorities, including curated open datasets, personalized continuous-learning systems, and deployment in resource-limited settings. With rigorous validation, transparent governance, and human-centered design, AI-ECG has the potential to democratize cardiovascular diagnostics and improve clinical outcomes across diverse environments.

Abstract: Background. The main protease (Mpro/3CLpro) of coronaviruses (CoVs) is an essential enzyme involved in viral replication and represents an attractive target for antiviral drug discovery. Based on the similar binding pocket residues within the Mpro of different CoVs, the study aimed to identify potential inhibitors of Mpro from PDB ID 6M2N, using inte-grated computational approaches. Methods. Structure-based pharmacophore modeling, virtual screening, molecular docking, MM-GBSA binding energy calculation, and molecu-lar dynamics (MD) simulation were performed using BIOVIA Discovery Studio. The vali-dated pharmacophore model was utilized to screen the ZINC database, followed by dock-ing and 100 ns MD simulation analyses of the top-ranked compounds. Results. The pharmacophore model 01 demonstrated favourable predictive performance (AUC = 0.781). Virtual screening identified 483 compounds, from which 21 compounds were selected for docking studies. Among them, ZINC95473654 (Lig-1), ZINC95473725 (Lig-2), and ZINC08792368 (Lig-3) exhibited strong binding affinity toward Mpro. Lig-1 demonstrated the best docking score and binding free energy along with stable interactions with key cat-alytic residues HIS41, CYS145, and GLU166. MD simulation analyses further confirmed that Lig-1 and Lig-2 maintained stable conformations and persistent intermolecular inter-actions throughout the 100 ns simulation period. Conclusion. The findings suggest that Lig-1, followed by Lig-2, may serve as promising CoVs Mpro inhibitors and warrant fur-ther experimental validation. Further experimental validation are required to consolidate the identified compounds as universal inhibitors of the CoVs-Mpro enzyme.

Abstract: Effective canine gastrointestinal health depends on suppression of enteric pathogens and maintenance of epithelial barrier integrity. Limosilactobacillus reuteri ATCC PTA 6127 (Lr6127) is a dog‑derived probiotic, but evidence supporting its protective capacity remains limited. Here, we evaluated the antimicrobial and epithelial‑supportive effects of Lr6127 using a canine epithelial cell model. Cell‑free supernatant (CFS) from Lr6127 significantly inhibited the growth of canine‑relevant pathogens, including Escherichia coli (52.0 ± 1.3%), Clostridium perfringens (54.0 ± 2.7%), and Salmonella typhimurium (48.6 ± 1.2%), compared with the medium control (P <  0.0001). Pathogen inhibition increased in a dose‑dependent manner with increasing CFS concentrations. Untargeted metabolomic analysis revealed enrichment of multiple antimicrobial‑associated metabolites, indicating multi‑component pathogen suppression, with genomic analysis supporting the aromatic amino acid–derived metabolite findings. In addition, viable Lr6127 significantly reduced epithelial adhesion of all tested pathogens (P <  0.01). Beyond direct antimicrobial effects, Lr6127 CFS promoted epithelial wound healing at later time points, accompanied by coordinated modulation of proteins associated with cytoskeletal remodeling and barrier repair. Collectively, these findings suggest that Lr6127 is associated with pathogen suppression, reduced adhesion, and epithelial wound repair mechanisms.