Raman spectroscopy combined with machine learning offers a rapid, label-free approach for bacterial identification, but robust translation remains challenged by spectral variability, biological heterogeneity, and limited model interpretability. Here, we present an integrated evaluation of an optimized Spectral Transformer (ST) framework for Raman-based bacterial classification benchmarked against a systematically optimized one-dimensional convolutional neural network (1D-CNN). The comparison was performed using a curated 36-class dataset comprising 15 Gram-negative bacterial entries, 15 Gram-positive bacterial entries, one non-bacterial microorganism, and five background/reference classes, enabling evaluation of both species-level and fine-grained bacterial classification. Under 15 dB noise-augmented evaluation, the ST achieved 80.6% ± 0.3% accuracy and a Matthews correlation coefficient (MCC) of 0.801 ± 0.003, outperforming the 1D-CNN baseline with 72.9% ± 0.3% accuracy andanMCCof0.721±0.003. Integrated Gradients analysis combined with attention map visualization enabled multi-level model interpretation, revealing that the ST’s improved robustness correlates with more bounded attribution patterns during misclassification, whereas the 1D-CNN’s feature attribution becomes scattered under noise perturbation. Importantly, this interpretability-driven analysis identified model-specific failure modes in the baseline architecture, including an over-reliance on non-specific spectral regions under noise, which can inform future data collection strategies and guide refinements to experimental protocols. These results demonstrate that attention-based spectral modeling improves Raman-based bacterial classification under noise-perturbed conditions while enabling multi-level interpretability that bridges model understanding with actionable feedback on experimental design and data quality requirements.

Mice have been used as models of urinary tract infection (UTI) for decades and laid the basis for the fundamental understanding of UTI pathogenesis in humans. Here we report that the high urine osmolarity of mice impacts key aspects of Escherichia coli UTI pathogenesis and represents a confounder for the translation of results to humans.

Monitoring bacterial activity is essential for numerous scientific and industrial applications. However, current benchmark measurements, i.e., optical density (OD), exhibit a limited dynamic range and require transparent or translucent media. Conventional impedance spectroscopy involves direct electrode contact with the bacterial medium or biofilm, potentially perturbing the sample environment and compromising measurement fidelity. Moreover, many real-time methods rely on costly, specialized labware that limits scalability and versatility. Here, we introduce a non-contact impedance spectroscopy (NCIS) technique with customizable electrodes for off-the-shelf labware and show that the data collected from a KCl solution series agree well with the simplest electrolytic conductivity cell model solution, demonstrating the accuracy and simplicity of NCIS. As an example of bacterial activity monitoring, NCIS was performed in glass laboratory bottles and 24-well plates in which Staphylococcus epidermidis and Escherichia coli cultures were inoculated into Brain Heart Infusion media, maintained at 37 °C. Comparative OD measurements acquired intermittently from the same media exhibited a strong correlation between NCIS and OD data, confirming reliability and reproducibility. The bacterial culture was verified by Raman spectroscopy assisted by machine learning. NCIS eliminates the risks of contamination and sample alteration, minimizing costs and operational complexity and providing a scalable, versatile solution for biological and chemical research.

Objective: Abdominal aortic aneurysm (AAA) is a permanent local dilation of the abdominal aorta developed by chronic local inflammation and elastin degradation. Trigonella foenum-graecum (fenugreek) has been reported to have anti-inflammatory properties. We hypothesized that fenugreek supplementation can inhibit AAA growth. Materials and methods: AAA was induced in male Sprague-Dawley rats by intraluminal porcine pancreatic elastase infusion. The treatment by oral gavage was initiated post-operatively on day 1 and was administered daily (750 mg/kg/daily or 1500 mg/kg/daily or distilled water) for 27 days. AAA expansion was monitored weekly by ultrasound measurements in a blinded-to-treatment fashion; rats were euthanized 28 days after surgery. AAA cross-sections were examined histologically, where treatment allocation were blinded. Results: AAA developed in all three groups, yet there was no measured difference (p=0.104) in the maximal inner anterior-posterior abdominal aortic diameter on day 28 post-surgery between the control group (110% ± 70%, n=11), the low-dose fenugreek treatment group (LDF, 105% ± 68%, n=11), and the high-dose fenugreek treatment group (HDF, 153% ± 96%, n=13). Additionally, assessments of elastin structure in the AAA wall using Miller's stain revealed disorganized and ruptured fibers, but no significant differences in the severity of damage or elastin content among the groups were noted. No significant differences were observed in the presence of infiltrating neutrophils (as indicated by myeloperoxidase-positive cells), macrophage infiltration (% CD68-positive area), or area of vascular smooth muscle cells (a-SMA stained cells), or media thickness across control low-dose, and high-dose treatment groups. Conclusions: Daily fenugreek administrations did not halt AAA progression in either low-dose (750 mg/kg/daily) or high-dose (1500 mg/kg/daily) groups when compared to controls. These results did not show any beneficial effects of fenugreek supplementation in the aneurysm wall, and therefore, we can not recommend fenugreek supplementation as a treatment for patients with growing AAAs.

In recent years Raman spectroscopy (RS) has become a promising tool for bacterial identification and antimicrobial resistance testing [1]. In this regard, multiple academic studies have shown excellent results, demonstrating high classification accuracy on multiple bacterial species [1]–[2]. These studies have typically applied complex, and time-consuming sample preparation, making their approaches unsuitable for clinical implementation. It has been a challenging task in realizing clinically relevant accuracies for bacteria identification using RS. Recent efforts involving machine-learning (ML) have demonstrated a large potential to reach high accuracies [1]–[2]. However, to bridge the gap between academic proof-of-concept and clinical application and for the purpose of introducing ML to data-limited domains. We propose a new ML transformer model, the spectral transformer (ST) [3]–[4]. We show that the ST can be used for fast and accurate classification of hyperspectral Raman images of bacteria. We Specifically demonstrate that our ST model can outperform state-of-the-art convolutional neural networks (CNNs) in scientific domains with limited availability of data [3]–[4]. We demonstrate the abilities of our ST ML model on Raman hyper-spectral images from 20 classes of reference bacteria as shown in figure 1. To further test the performance of the ST ML model in vivo we tested the concept on Raman data recorded on experimentally infected wounds in mice, with the aim to identify the species of the infecting organism.

OBJECTIVE:To define 'chronic' urinary tract infection (UTI) by reviewing current published research that employs this term. METHODS:We systematically searched Medline and Embase for studies covering all aspects of human UTI. For comparison, current urological and infectious disease guidelines were also reviewed. RESULTS:The electronic search yielded a total of 2175 articles, of which 154 were eventually included for data extraction. Of these, six studies presented a definition of chronic UTI within the text. The definitions were highly incongruent among studies and often identical to the current definition of 'recurrent' UTI. CONCLUSION:Chronic UTI is an undefined term, which has nevertheless been increasingly used in the past 15 years. Chronic UTI does not represent a distinct clinical or microbiological condition that justifies its use as medical diagnosis.

Bacterial enteric pathogens are major contributors to the global burden of diarrheal diseases and the associated consequences for human health including malnutrition, growth stunting, morbidity, and mortality. While mortality from diarrhea has decreased, incidence remains high, and better interventions for preventing disease are needed. Single-domain antibodies (i.e., VHHs), functioning as target-binding proteins in the gastrointestinal tract, have been proposed as a potential approach to mitigate bacterial pathogenesis. Here, we describe a mitigation strategy where precision binding of a bivalent VHH to the receptor-binding B-pentamer of heat-labile enterotoxin aggregates the AB5 toxin and impairs enterotoxigenic Escherichia coli colonization in a flow chamber model simulating the human intestine. The VHH construct also binds to the structurally similar cholera toxin and effectively abrogates its intestinal cell cytotoxicity in vitro. Based on these results, we believe that targeting virulence could emerge as a new strategy for the management of bacterial enteric pathogens, supporting gut health in at-risk populations alongside vaccination campaigns or in populations without access to vaccines.

Urinary tract infections (UTIs) are the main complication associated with clean intermittent catheterization (CIC) and are facilitated by post-void residual urine and trauma to the mucosa during voiding. The risk of UTI may be diminished by reducing the residual volumes and preventing microtrauma caused by mucosal suction through the eyelets of conventional eyelet catheters (CEC). A new micro-hole zone catheter (MHZC) was developed and tested in an ex vivo porcine lower urinary tract model and in vivo , in pigs, against a CEC. It was shown that, irrespective of the micro-hole diameter, the new catheter ensured increased flowrates and significantly lower residual volumes at the first flow-stop. Furthermore, with a micro-hole diameter of 0.4 mm, mucosal suction was virtually eliminated, regardless of the insertion depth or simulated intra-abdominal pressure mimicking sitting or standing humans. Pressure profile experiments and endoscopy studies indicated that the bladder gradually folds against the drainage tip of the new catheter, without blocking the flow, and, unlike with the CEC, sharp pressure variations and flow-stops did not occur during voiding. The MHZC outperformed the CEC in all tested scenarios and decreased residual volumes, thus potentially decreasing the risk of UTIs.

Herein, we report the hit-to-lead identification of a drug-like pleuromutilin conjugate 16, based on a triaromatic hit reported in 2020. The lead arose as the clear candidate from a hit-optimization campaign in which Gram-positive antibacterial activity, solubility, and P-gp affinity were optimized. Conjugate 16 was extensively evaluated for its in vitro ADMET performance which, apart from solubility, was overall on par with lefamulin. This evaluation included Caco-2 cell permeability, plasma protein binding, hERG inhibition, cytotoxicity, metabolism in microsomes and CYP3A4, resistance induction, and time-kill kinetics. Intravenous pharmacokinetics of 16 proved satisfactory in both mice and pigs; however, oral bioavailability was limited likely due to insufficient solubility. The in vivo efficacy was evaluated in mice, systemically infected with Staphylococcus aureus, where 16 showed rapid reduction in blood bacteriaemia. Through our comprehensive studies, lead 16 has emerged as a highly promising and safe antibiotic candidate for the treatment of Gram-positive bacterial infections.

Immunocompromised patients infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can have a longer duration of viral shedding and persistence of symptoms. The optimal treatment strategy for these patients remains to be established. This case describes a male in his late sixties with follicular lymphoma and persistent symptoms of infection with SARS-CoV-2 variant BA.2 who was treated with remdesivir five times over a period of six months. The clinical effect of remdesivir treatment decreased over time, and further viral sequencing revealed the emergence of mutations across the SARS-CoV-2 genome. Due to the lack of other treatment options, the patient was treated with a combination of remdesivir and molnupiravir for 10 days, and epcoritamab was discontinued, which led to the cessation of symptoms. This case illustrates the risk of a diminished effect of remdesivir with prolonged use and the need for treatment guidelines for immunocompromised patients with persistent COVID-19.

AbstractRaman spectroscopy provides non-destructive, label-free quantitative studies of chemical compositions at the microscale as used on NASA’s Perseverance rover on Mars. Such capabilities come at the cost of high requirements for instrumentation. Here we present a centimeter-scale miniaturization of a Raman spectrometer using cheap non-stabilized laser diodes, densely packed optics, and non-cooled small sensors. The performance is comparable with expensive bulky research-grade Raman systems. It has excellent sensitivity, low power consumption, perfect wavenumber, intensity calibration, and 7 cm−1 resolution within the 400–4000 cm−1 range using a built-in reference. High performance and versatility are demonstrated in use cases including quantification of methanol in beverages, in-vivo Raman measurements of human skin, fermentation monitoring, chemical Raman mapping at sub-micrometer resolution, quantitative SERS mapping of the anti-cancer drug methotrexate and in-vitro bacteria identification. We foresee that the miniaturization will allow realization of super-compact Raman spectrometers for integration in smartphones and medical devices, democratizing Raman technology.

BackgroundReliable animal models are critical for preclinical research and should closely mimic the disease. With respect to route of infection, pathogenic agent, disease progression, clinical signs, and histopathological changes. Sheep have similar bone micro- and macrostructure as well as comparable biomechanical characteristics to humans. Their use in bone research is established, however their use in bone infection research is limited. This systematic review will summarise the key features of the available bone infection models using sheep, providing a reference for further development, validation, and application.MethodThis systematic review was designed according to the PRISMA guidelines and registered with PROSPERO. Quality was assessed using SYRICLE's risk of bias tool adapted for animal studies. PubMed, MEDLINE, Web of Science and EMBASE were searched until March 2022.1921 articles were screened by two independent reviewers, and 25 were included for analysis.ResultsModels have been developed in nine different breeds. Staphylococcus aureus was used in the majority of models, typically inoculating 108 colony forming units in tibial or femoral cortical defects. Infection was established with either planktonic or biofilm adherent bacteria, with or without foreign material implanted. Most studies used both radiological and microbiological analyses to confirm osteomyelitis.ConclusionsThere is convincing evidence supporting the use of sheep in bone infection models of clinical disease. The majority of sheep studied demonstrated convincing osteomyelitis and tolerated the infection with minimal complications. Furthermore, the advantages of comparable biology and biomechanics may increase the success for translating in vivo results to successful therapies.The Translational potential of this articleIn the realm of preclinical research, the translation to viable clinical therapies is often perilous, and the quest for reliable and representative animal models remains paramount. This systematic review accentuates the largely untapped potential of sheep as large animal models, especially in bone infection research. The anatomical and biomechanical parallels between sheep and human bone structures position sheep as an invaluable asset for studying osteomyelitis and periprosthetic joint infection. This comprehensive exploration of the literature demonstrates the robustness and translational promise of these models. Furthermore, this article underscores the potential applicability for sheep in developing effective therapeutic strategies for human bone infections.

Uropathogenic Escherichia coli (UPEC) can undergo extensive filamentation in the host during acute urinary tract infections (UTIs). It has been hypothesised that this morphological plasticity allows bacteria to avoid host immune responses such as macrophage engulfment. However, it is still unclear what properties of filaments are important in macrophage-bacteria interactions. The aim of this work was to investigate the contribution of bacterial biophysical parameters, such as cell size and shape, and physiological parameters, such as cell surface and the environment, to macrophage engulfment efficiency. Viable, reversible filaments of known lengths and volumes were produced in the UPEC strain UTI89 using a variety of methods, including exposure to cell-wall targeting antibiotics, genetic manipulation and isolation from an in vitro human bladder cell model. Quantification of the engulfment ability of macrophages using gentamicin-protection assays and fluorescence microscopy demonstrated that the ability of filaments to avoid macrophage engulfment is dependent on a combination of size (length and volume), shape, cell surface and external environmental factors. UTI89 filamentation and macrophage engulfment efficiency was also found to occur independently of the SOS-inducible filamentation genes, sulA and ymfM in both in vivo and in vitro models of infection. Compared to filaments formed via antibiotic inhibition of division, the infection-derived filaments were preferentially targeted by macrophages. With several strains of UPEC now resistant to current antibiotics, our work identifies the importance of bacterial physiological and morphological states during infection.

SARS-CoV-2 virus infects cells by engaging with ACE2 requiring protease TMPRSS2. ACE2 is highly expressed in kidneys. Predictors for severe disease are high age and male sex. We hypothesized that ACE2 and TMPRSS2 proteins are more abundant (1) in males and with increasing age in kidney and (2) in urine and extracellular vesicles (EVs) from male patients with COVID-19 and (3) SARS-CoV-2 is present in urine and EVs during infection. Kidney cortex samples from patients subjected to cancer nephrectomy (male/female; < 50 years/˃75 years, n = 24; ˃80 years, n = 15) were analyzed for ACE2 and TMPRSS2 protein levels. Urine from patients hospitalized with SARS-CoV-2 infection was analyzed for ACE2 and TMPRSS2. uEVs were used for immunoblotting and SARS-CoV-2 mRNA and antigen detection. Tissue ACE2 and TMPRSS2 protein levels did not change with age. ACE2 was not more abundant in male kidneys in any age group. ACE2 protein was associated with proximal tubule apical membranes in cortex. TMPRSS2 was observed predominantly in the medulla. ACE2 was elevated significantly in uEVs and urine from patients with COVID-19 with no sex difference compared with urine from controls w/wo albuminuria. TMPRSS2 was elevated in uEVs from males compared to female. ACE2 and TMPRSS2 did not co-localize in uEVs/apical membranes. SARS-CoV-2 nucleoprotein and mRNA were not detected in urine. Higher kidney ACE2 protein abundance is unlikely to explain higher susceptibility to SARS-CoV-2 infection in males. Kidney tubular cells appear not highly susceptible to SARS-CoV-2 infection. Loss of ACE2 into urine in COVID could impact susceptibility and angiotensin metabolism.

Background Urinary tract infection (UTI) is a common disease with a significant risk of relapse. Deliberate bladder colonization with asymptomatic Escherichia coli is being explored as a potential strategy to fend off invading uropathogens, thereby mitigating the risk of symptomatic UTI. Currently, one major obstacle is the low success rates for achieving persistent bladder colonization with asymptomatic bacteria and experimental challenge studies are lacking. Here, we assessed the influence of an indwelling bladder catheter on the ability of asymptomatic E. coli to colonize the bladder and to assess the protective efficacy of such colonization against experimental UTI with uropathogenic E. coli.Methods Pigs with or without indwelling bladder catheters were experimentally inoculated with the asymptomatic E. coli strain 83972 and subsequently challenged by inoculation with a uropathogenic E. coli isolate, UTI89. The animals were monitored with regular urine and blood samples and bladders and kidneys were harvested at termination.Results All pigs with indwelling catheters were colonized by E. coli 83972 in response to inoculation, compared to pigs without catheters in which only 1 of 8 animals were colonized. When removing the catheter, E. coli 83972 were spontaneously cleared. Colonization with E. coli 83972 prevented experimental infection in 50% of animals, whereas all control animals became infected.Conclusions The presence of indwelling bladder catheters strongly facilitates the colonization of E. coli 83972, indicating that individuals with catheters may be particularly suited for receiving this treatment. The research supports prophylactic colonization with E. coli 83972 as a potential strategy to reduce the risk of UTIs. The presence of an indwelling urinary catheter significantly increases bladder colonization with the asymptomatic Escherichia coli bacteriuria strain 83972. Colonization with E. coli 83972 prevents 50% of experimental urinary tract infections.

BackgroundArteriovenous (AV) grafts often develop severe complications of stenosis due to neointimal proliferation that occurs either at the venous anastomosis site or at the outflow receiving vein. This study compares primary patency during 12 months of follow up for a new experimental Biomodics© interpenetrating polymer network (IPN) drug-eluting graft prototype with state-of-the-art GORE® ACUSEAL (ACUSEAL) in an AV graft model in sheep.Methods and resultsAn end-to-end bypass from the common carotid artery to the jugularis vein was performed bilaterally in 12 sheep. The usage of ACUSEAL or the IPN, both 6.0 mm in diameter, was determined via randomization. The sheep were followed up every 4 weeks with ultrasonic duplex scanning to determine patency; the experienced observer was blinded to the randomization. One sheep died after 11 days, and the final sample accordingly consisted of 11 animals. When comparing neointimal hyperplasia after 12 months in the two grafts, Fisher's exact test showed a significant difference with none out of 11 in the IPN grafts and 9 out of 11 in the ACUSEAL graft (p < 0.001). However, the Biomodics© IPN exhibited severe deterioration over time.ConclusionsAlmost all of the grafts occluded during the 12 months of follow up. Although the zwitterion-bounded interpenetrating drug eluting polymer network showed signs to impair neointimal hyperplasia and thrombosis, age-related degeneration hindered demonstrating a potential improvement in patency.

Fucoidans are a heterogenous class of fucose-rich sulfated carbohydrates which have attracted increasing attention in cancer and inflammation research due to their bioactive properties. There are reports that fucoidans may have direct antibacterial effects and synergy with antibiotics. However, the literature is conflicting, potentially due to the limited reporting of origin, characteristics, and extraction methods of the fucoidans tested. Here we report the results of 18 defined fucoidans screened for direct, indirect, and synergistic antibacterial effects. 15 distinct fucoidan fractions, isolated from Laminaria hyperborea using a solvent-free extraction process, were characterised for molecular weight, pH, viscosity, and sulfur content. These, together with three commercially available crude fractions, were assessed at concentrations from 0.03125-24 mg mL-1 for minimum inhibitory concentration against Staphylococcus aureus, Streptococcus mutans and Streptococcus sanguinis. Furthermore, we tested a selection of fucoidans for antibacterial synergy with vancomycin and indirect antibacterial effects in whole blood survival assays. Reverse-transcription quantitative polymerase chain reaction (RT-qPCR) was performed to assess the stress response in fucoidan-treated S. aureus cultures. We have identified one fucoidan fraction with bactericidal activity against diverse bacteria. This effect is dose-, fucoidan fraction- and bacteria-specific, and furthermore, not related to osmotic stress. No synergistic effects were observed with fucoidan in combination vancomycin. Fucoidans have exciting potential as antimicrobial agents. Further analysis is required to establish the precise molecular characteristics responsible for their potent bactericidal activity.