Photonic Diagnosis, Monitoring, Prevention, and Treatment of Infections and Inflammatory Diseases 2025

Infectious diseases represent a significant contributor to global mortality rates, underscoring the necessity for a comprehensive approach to pathogen diagnostics, antibiotic selection, and modulation of the host response. It is imperative that clinicians implement a stratification system and streamline the sampling process. Clinical research provides new information about the occurrence of illnesses and biomarkers, which in turn leads to the development of new diagnostic concepts. Photonic diagnostics have been demonstrated to be advantageous for time-critical decision-making and have the potential for bedside utilization. In vitro Raman-based diagnostics, commencing with laboratory-based tests, are being employed in sample-to-answer workflows to address clinically significant correlations between pathogen diagnosis and host response. The case of blood diagnostics provides an illustrative example of this phenomenon.

Conventional histology requires invasive acquisition of tissue samples via biopsy, manual tissue processing, slicing and staining by artificial markers to create high contrast in conventional white light microscopes. Multiphoton microscopy (MPM), on the other hand, can exploit the natural, optical contrast of autofluorescence (AF) from native proteins and second harmonic generation (SHG) from collagen fibers for label-free imaging of unstained tissue. Since both mechanisms are naturally confocal and since the infrared excitation in MPM has a greater penetration depth, MPM is ideal for optical sectioning and generating 3D image stacks. Previously, we used this technique to study different murine colitis models and identified distinct patterns of epithelial damage, immune infiltration or scar formation. Here, we used this database with several hundred image stacks to train a 3D convolutional neural network for the classification of disease activity and phenotype. In the future, the direct classification of label-free MPM image stacks could allow faster and potentially more accurate assessment of inflammatory bowel disease for preclinical research and for clinical bedside monitoring.

Botulinum neurotoxins (BoNTs) are the most potent toxins known in nature produced by Clostridium botulinum strains, which can cause life-threatening diseases in both humans and animals. The available in vivo mouse assay is inadequate for real-time and on-site assessment of outbreaks. Herein, we present a reflective-based approach for the detection of BoNT/C while estimating its activity. Two adjacent porous Si Fabry–a competitive immunoassay simultaneously utilizes Pérot interferometers to quantify minute BoNT/C concentrations and assess their endopeptidase activity. The reflectivity signals of each interferometer are amplified by biochemical reaction products infiltration into the scaffold or by peptide fragments detachment from the nanostructure. The optical assay is highly sensitive in compliance with the in vivo approach by presenting a detection limit of 4.24 pg mL–1. The specificity and selectivity of the designed platform are cross-validated against BoNT/B and BoNT/D, relevant to animal health. Finally, the analytical perform

Accurate diagnosis of respiratory virus co-infections is vital for effective treatment and management. This study introduces a label-free diagnostic platform combining surface-enhanced Raman scattering (SERS) with deep learning for the rapid detection and quantification of respiratory virus co-infections. Using silica-coated silver nanorod array SERS substrates, we collected spectra from 11 respiratory viruses, 9 2-virus mixtures， and 4 3-virus mixtures. Over 1.2 million SERS spectra were used to develop and optimize a deep learning model, MultiplexCR, which achieved 98.5% accuracy in virus classification and a mean absolute error (MAE) of 0.025 for concentration regression. A novel method for determining the limit of detection (LOD) was also introduced. In blind tests, the model maintained 95.2% accuracy, demonstrating its robustness. This platform shows great potential for rapid, point-of-care detection of virus co-infections, offering a powerful tool for managing complex infectious diseases.

In this study, we developed a novel SERS platform using filter paper with dendritic silver nanostructure/graphene oxide (GO) composite. By optimizing the morphology of the silver structures through reaction conditions and anchoring anti-IL-6 antibodies, we created a biosensor with enhanced IL-6 detection capabilities. The SERS-active substrates were prepared by immersing filter paper in GO suspension and drop casting synthesized dendritic silver nanoparticles. UV-Vis spectroscopy, TEM, FE-SEM, and AFM confirmed the substrate properties. This method achieved high sensitivity with a detection limit of 0.05 pM, demonstrating significant potential for IL-6 diagnosis.

Determination of urinary or serum N-acetyl-β-d-glucosaminidase (NAG) activity as a tissue damage indicator is widely used in diagnosis of various pathologies, including diabetic nephropathy. Herein, we present a modified sensing approach for a rapid and reliable NAG activity determination in complex media using surface-enhanced Raman spectroscopy (SERS). Porous silicon (PSi) Fabry-Pérot interferometers were redesigned as sensitive SERS platforms utilizing the vast inherent surface area for silver (Ag) nanoparticles embedment. Interaction of the porous nanostructures with specific NAG-enzymatic products produces an indicative spectral fingerprint proportional in magnitude to its concentration. The sensitivity of Ag–PSi SERS substrates was evaluated in complex matrices presenting sufficient limits of detection compared with other advanced assays and techniques. The potential applicability of the suggested prototype for real-life scenarios was evaluated in vivo, in a model of insulin-dependent diabetes induced in sheep. The robust data confirm the application of SERS analysis for early diagnosis of pathology and for evaluation of clinical responses to pharmacological treatments.

Bovine mammary gland is susceptible to numerous bacterial infections resulting in inflammatory disease condition of bovine mastitis (BM) with a staggering economic impact on the dairy industry worldwide. Early BM detection is crucial for infection control within the dairy herd, which can be alleviated by antimicrobial therapy. N-acetyl-β-D-glucosaminidase (NAGase) is a prominent BM inflammatory biomarker secreted onto the blood circulation upon pathogenesis and then released into milk, capable of separating healthy quarters from subclinical and clinical BM cases. Herein, we report a sensitive, cost-effective and handy-to-use BM severity assay based on NAGase inherent content found in whole milk samples. Silver-coated porous Si (Ag-PSi) Fabry–Pérot interferometers were employed for quantifying the lysosomal activity in different milk qualities using surface-enhanced Raman spectroscopy(SERS). The optical response was proportional to the occurring pathogenesis induced by predominant bacteria found in dairy farms at different somatic cell levels. The optimized Ag-PSi SERS substrates were utilized for quantitative analysis of NAGase levels found at clinically relevant BM concentrations.

The overuse of antibiotics cause human health concerns due to antibiotic-resistant infections. For example, 2.8 million antibiotic-resistant infections including 35.000 deaths occur each year in US. Therapeutic drug monitoring (TDM) of antibiotics is an efficient approach to prevent and control the spread of antibiotic resistance. Recently, surface-enhanced Raman scattering (SERS) biosensors have demonstrated promising application in the field of TDM of antibiotics, because of their advantages of label-free, high sensitivity, rapid diagnostic, simple operation, and point-of-care potential. However, the practical application of SERS biosensors in clinical practice is still restricted by their sensing substrates, due to the challenges of poor-scaleup ability, poor reproducibility, and high-cost. In this work, we present an economic and scalable manufacturing process for paper-based SERS biosensors. The fast and quantitative Raman measurements of beta-lactam antibiotics in 1xPBS and urine solutions can be achieved using our SERS biosensors

Sepsis is a leading cause of death in infectious diseases, necessitating optimal antibiotic treatment with proper and quick monitoring of the drug dosage levels. Drug quantification in clinics is performed with time-consuming analytical methods. Therefore, introducing versatile techniques that quickly quantify antibiotic levels in patient's blood is essential. In this work, we develop an automated platform based on label-free surface-enhanced Raman spectroscopy (SERS) and a centrifugal microfluidics setting for quantifying meropenem (MER) in serum samples. We show the performance of this device for rapid, sensitive, and accurate quantification of meropenem in human patient samples. Therefore, showing the feasibility of SERS as a novel analytical method for point-of-care antibiotics in clinical settings.

Bacterial infections are a significant global problem. To reduce their implications innovative and rapid diagnostic tests are needed. Raman spectroscopy offers label-free, non-destructive and rapid diagnostic opportunities. The RamanBioAssay™, a flexible multiwell platform, provides rapid bacterial identification and antimicrobial susceptibility testing capabilities. Spectral fingerprints of bacteria are analyzed by means of statistical data analysis yielding bacterial identification and resistograms. The test turnaround time from bacterial sample to results is ≤ 3.5 hours which qualifies the RamanBioAssay™ as a rapid diagnostic platform.

Accurate and early diagnosis of infectious diseases is crucial for combating antibiotic resistance and improving patient outcomes. We developed an affordable and robust deep ultraviolet (DUV) Raman spectroscopy system using a mercury lamp at 253.65 nm, capable of resolving sub-1000 cm-1 Raman peaks. This system provides high-resolution spectral fingerprints for precise pathogen identification. Tested on various samples, including alcohols, foods, pharmaceuticals, saliva, soil, and plastics, it ensures precise identification and potential UV-C light disinfection. This dual-functionality system enhances infection diagnosis and control, offering a promising tool for clinical and field environments.

This study was conducted to examine the biochemical and morphological characteristics of biofilm-producing, colistin-resistant Pseudomonas aeruginosa using Raman spectroscopy and scanning electron microscopy (SEM). We have observed notable transitions in bacterial morphology and biochemical composition over time upon colistin treatment which leading to understand various aspects of bacterial response. Our findings demonstrate the potential of combining Spectroscopy based approach with multivariate analysis for monitoring and managing early detection of antibiotic resistance in hospital setups.

Invasive fungal infections are a major cause of global morbidity and mortality. The widespread use of antifungals and fungicides has resulted in increased antifungal resistance. Timely and accurate antifungal susceptibility testing (AFST) is essential to slow down the emergence of antifungal resistance. Here, we report a rapid testing procedure that determines the antifungal susceptibility of Candida species by stimulated Raman photothermal (SRP) metabolic imaging of deuterium oxide (D2O) metabolism at the single-cell level, based on a compact and robust fiber laser with rapid and wide tuning capability. The incorporation of D2O can be detected within 1 hour due to the high detection sensitivity and imaging speed of the SRP microscopy. The single-cell metabolism inactivation concentration (SC-MIC) value can be obtained within 3 hours, compared to the minimum 48 hours required by the Clinical and Laboratory Standards Institute (CLSI) AFST results. Our method does not rely on cell growth, making it a promising tool for rapid AFST. Additionally, it facilitates insights into resistance mechanisms and supports the precise administration of antifungal therapies.

Vibrational spectroscopy techniques are used to study leukocytes in full blood smear samples. Two modalities, fluorescence and optical photothermal spectroscopy, achieve this by first capturing high-resolution images of leukocytes and then collecting multiple spectra from each cell. Unsupervised hierarchical clustering analysis (HCA) is performed on the spectra's 2nd derivative to label leukocytes based on chemical signatures. This method correlates cluster data with actual leukocyte fractions in five specimens. Future work will use AI and machine learning to improve leukocyte classification by combining morphological and spectroscopic labels.

Raman spectroscopy (RS) is a powerful tool for infectious disease detection, enabling rapid discrimination of pathogens. Past works have primarily focused on using the fingerprint region (400-1800 cm 1) due to its high number of peaks representing different molecular bonds and have ignored the high-wavenumber region (2700-3600 cm-1) with the belief that its two major bands, representing Oxygen-Hydrogen and Carbon-Hydrogen bonds, yields minimal biochemical information. Here, we showcase that high-wavenumber RS is a feature-rich, low-background technique that can differentiate 16 microbial species with similar fidelity to fingerprint RS. Additionally, we show that the distinctive spectra of each class of biomolecule allow for the Carbon-Hydrogen band to be decomposed into a measure of the underlying molecular composition. When applied to the microbial spectra, this produces a characterization closely matching reported dry-mass compositions, indicating that high-wavenumber RS may be capable of non-destructively monitoring microbial composition over time.

Evaporative dry eye syndrome, often caused by meibomian gland dysfunction (MGD), results from a compromised lipid layer in the tear film, leading to increased tear evaporation. This study aimed to measure the lipid layer thickness by extracting white light interference patterns using the Unet++ deep learning algorithm for image segmentation, which showed superior performance. The pixel colors in the segmented interference images were mapped to thickness values with a custom table. The method successfully extracted lipid layer patterns with a 0.98 success rate. Significant differences in lipid layer thickness were found between subjects with and without dry eye disease, demonstrating the efficacy of this approach. The deep learning algorithm's ability to accurately segment and analyze lipid layer thickness could be applied to other ocular conditions, enhancing diagnostic precision and offering a scalable tool for routine eye health assessments, leading to early detection and personalized treatments.

Antimicrobial blue light (aBL) at 405 nm wavelength exhibits potential as a non-pharmacological approach to combat bacteria. However, in Staphylococcus aureus, staphyloxanthin, which acts as an antioxidant, renders this bacterium more tolerant to killing by aBL than others. In this study, we investigated the combination of 470 nm light, which can blench staphyloxanthin, with 405 nm aBL to improve the photo-killing of S. aureus. After an exposure of 360 J/cm2 at 470 nm and 216 to 360 J/cm2 exposures at 405 nm, > 4.5-log10 colony forming units (CFU) reduction was achieved in all six strains tested, including ATCC25923, ATCC33592, AR0570, AR0215, SH1000, and USA300. In summary, our preliminary results demonstrate the potential of this dual-wavelength strategy in potentiating the effectiveness of photo killing of S. aureus.

MRSA infections pose an increasing risk in orthopaedic surgery. They develop resistance to antibiotics and form biofilms that protect them from therapeutic interventions. New treatment approaches are in need and antimicrobial photodynamic therapy (aPDT) is one of the most promising. This study was aimed at evaluating the effect of photons produced by laboratory induced bioluminescence and fluorescence MRSA. Using these observed effects can help identify the overestimation of antimicrobial PDT.

In the past two decades, many photosensitizers (PS) have been developed for photodynamic therapy (PDT) targeting cancers, microbial pathogens, and parasites in both clinical and pre-clinical settings. However, none of these PS are specific for bacteria. Given the growing threat of multidrug-resistant (MDR) bacteria, non-antibiotic approaches are actively sought after to battle these resilient microbes. To date, we will discuss bacteria-specific pro-PS that are activated exclusively in bacteria, but not in mammalian cells, distinguishing them from traditional PS that are active in both mammalian cells and bacteria. Two classes of pro-PS will be discussed: those that convert to PS from non-PS specifically in bacteria owing to higher levels of singlet oxygen production in bacteria upon light illumination and those utilizing porphyrin precursors in a bacteria-specific pathway to increase porphyrin biosynthesis. Because of the specificity, these pro-PS can be employed at high concentrations in conjunction and effectively eradicate a range of MDR bacteria, including their planktonic forms, mature biofilms, and persisters, irrespective of their antibiotic susceptibility.

The increasing prevalence of antibiotic resistance (AR) makes common bacterial infections difficult to treat, often leading to severe systemic infections. Recent studies have shown that photodynamic therapy (PDT), which uses photosensitizers (PS) activated by light, can disrupt bacterial AR mechanisms, enhancing antibiotic effectiveness. A major challenge is delivering PS and antibiotics simultaneously at the infection site. To overcome this, we developed a nanotechnology-based drug delivery system called photoactivatable multi-inhibitor liposome (PMIL). PMIL allows precise co-delivery of PS and antibiotics. Testing with S. aureus and E. coli models showed that PMIL impairs AR mechanisms and increases bacterial susceptibility to antibiotics, leading to more effective bacterial eradication. This innovative platform could transform clinical infection management by providing a new approach to treating antibiotic-resistant infections.

There is a high demand for innovative solutions to prevent infections in vulnerable settings, and thereby reduce the amount of antibiotics needed for treatment. In this context, antimicrobial photodynamic inactivation can serve as an effective means of disinfection for otherwise hard to treat components. In our study, we focus on its possible application on temporary polymer implants such as catheters or shunts. A stable covalent bond of the photosensitizer Rose Bengal to a silicon surface proved to be effective in killing sedentary bacteria by irradiation, preventing further colonization and biofilm formation. We present material processing protocols, dose-response relationships with free and bound RB, and investigations of the mode of action of the photosensitizer and its long-term stability. This work lays the foundation for stable, self-infecting polymer implants with the potential to significantly improve patient care.

Efflux pumps are membrane proteins that expel toxic substances, including antibiotics and photosensitizers. Efflux pump inhibitors (EPIs) block these pumps, enhancing antimicrobial treatments. Our study investigated reserpine, an EPI, with silver nanoparticles (Ag NPs) and methylene blue (MB) for pathogen photodeactivation. Results showed that reserpine, combined with Ag NPs and MB, significantly increased pathogen deactivation compared to Ag NPs and MB alone. Specifically, there was a 40% additional reduction in colony formation units (CFU) with reserpine and MB, and a 90% reduction with reserpine, Ag NPs, and MB. This suggests reserpine disrupts the cell's efflux mechanisms. Singlet oxygen production analysis indicated reserpine has no effect on it. Molecular docking revealed that reserpine binds more strongly to the AcrB efflux pump than MB, enhancing photodynamic therapy. Additionally, reserpine improved deactivation rates regardless of Ag NP size. Similar experiments with graphene quantum dot (GQD) nanoparticles will be presented.

Perforated appendicitis is associated with longer hospitalization and worse postoperative outcomes relative to uncomplicated appendicitis, due to the time required for intravenous antibiotics to control intra-abdominal infection. We propose intra-procedural photodynamic therapy (PDT) with methylene blue to rapidly disinfect the abdominal cavity following appendectomy. Towards this end, we have developed a rabbit model of perforated appendicitis to test PDT in vivo. In control animals that did not receive PDT, bacterial growth continued in the 24 hours following intervention. In animals receiving PDT, the bacterial burden was reduced by several orders of magnitude. These pilot results demonstrate the potential value of PDT in treating intra-abdominal infection secondary to perforated appendicitis.

Photobiomodulation boosts bacterial metabolism, leading to an increase in ATP and ROS, thereby enhancing the effectiveness of ABL in MRSA biofilms

Antimicrobial resistance poses a critical public health threat worldwide, making it urgent for the development of new strategies to address the problem. While aPDT has proven to be effective against multidrug-resistant bacteria, challenges in delivering the PS to a targeted site can limit its efficacy. Superhydrophobic (SH) antimicrobial photodynamic therapy (SH-aPDT) is an attractive new technology that can address these challenges by isolating the PS into an SH bandage, with the bandage emitting airborne singlet oxygen for aPDT. This unique “contact-free” technique is expected to improve aPDT efficiency, hence addressing current aPDT limitations.

Wound chronicity is mainly attributed to a persistent tissue level of bacteria in the wound site. Despite the availability of a variety of antimicrobial strategies, the presence of drug-resistant bacteria makes conventional conservative treatment less effective and even frustrates the confidence of patients. The advantages of PDT as a modality for infected wounds are receiving increasing attention, and the development of photosensitizers can promote the continuous advancement of this technology. As a new chlorine derivative with improved chlorine structure, STBF possesses better hydrophilicity and its mediated PDT is believed to be promising for infected wounds. STBF-PDT had not received much attention in the treatment of resistant bacterial infections and infected wounds so far. Our frontier investigation elucidated the powerful antimicrobial utility of STBF-PDT on MRSA and efficacy on MRSA-infected mouse wounds. Furthermore, our clinical study proved that STBF-PDT could effectively and safely treat MRSA-infected wounds and improved the wound-QoL of patients.

Bacterial otitis media (OM) is a prevalent disease occurring in 90%+ of children before age 5. For treatment, a course of broad-spectrum antibiotics is often effective, though may have long-term side effects on the microbiome and increase risks of antibiotic resistance. Plasma medicine harnesses technology used in semiconductor manufacturing as promising therapeutic approaches for many infectious diseases. Here, results are presented from a new cold microplasma (CMP)-based treatment platform for OM. The presentation will include experimental validation of a protocol to deliver treatment noninvasively through the eardrum and into the middle ear cavity. This project is the first application and demonstration of CMP to successfully clear bacterial OM in vivo in a preclinical animal model. Minor side-effects and other points for refinement will be reviewed, in addition to current limitations and future work. Plasma medicine as a non-pharmacological therapy for OM may significantly impact the management of this common disease.

Photosensitization using endogenous photosensitizers and blue light showed an effective and safe alternative antimicrobial therapy for skin infections. Incorporating nanotechnology in the field of photosensitization could have a good impact on overcoming some limitations of photosensitizers and enhancing antimicrobial photodynamic therapy. Nanoparticles reduces the probability of aggregation of photosensitizers and subsequently enhance the efficacy of photosensitization.

Antimicrobial Blue Light (aBL) promotes effective antimicrobial treatment through the production of radical oxygen species (ROS) by photoexcitation of endogenous chromophores. Tetracyclines used as potential photosensitizers concurrently with aBL amplified antimicrobial activity against MRSA and P. aeruginosa in planktonic culture and biofilm. We compared the effects of Minocycline, Doxycycline, Omadacycline, and Tigecycline with aBL therapy. We found significant differences between the older tetracyclines and the newer, extended-spectrum tetracyclines.

As with conventional fibre optics, light will propagate within a laminar flow of transparent fluid. Hence ‘laminar flow Photodynamic Therapy’ is explored using a photosensitizer solution to deliver light for activation directly into a wound. By adding a delivery mechanism of irradiation via wound irrigation, we develop a means of optical activation of a photoreactive solute inside a wound to destroy bacteria within a deep and occluded wound site without recourse to antibiotics. The project quantified the design parameters necessary for an operationally practical approach for clinical use. PDT via laminar flow fibre-optic delivery for infection treatment during irrigation could improve treatment efficacy and provide a novel tool combating growing AMR threats.

This study investigated the effects of laser irradiation on the microscopic structural and diffusion properties of articular cartilage. Following 1550 nm laser treatment of bovine cartilage samples, significant structural changes revealing formation of pronounced radiation induced channels were observed. 27%-154% increase in sample permeability was also recorded.