Enabling mRNA medicine for brain tumors

mRNA is a new class of drugs that has the potential to revolutionize the treatment of brain tumors. Thanks to the COVID-19 mRNA vaccines and numerous therapy-based clinical trials, it is now clear that lipid nanoparticles (LNPs) are a clinically viable means to deliver RNA therapeutics. However, LNP-mediated mRNA delivery to brain tumors remains elusive. Over the past decade, numerous studies have shown that tumor cells communicate with each other via small extracellular vesicles, which are around 100 nm in diameter and consist of lipid bilayer membrane similar to synthetic lipidbased nanocarriers. We hypothesized that rationally designed LNPs based on extracellular vesicle mimicry would enable efficient delivery of RNA therapeutics to brain tumors without undue toxicity. We synthesized LNPs using four components similar to the formulation used in the mRNA COVID19 vaccines (Moderna and Pfizer): ionizable lipid, cholesterol, helper lipid and polyethylene glycol (PEG)-lipid. For the in vitro screen, we tested ten classes of helper lipids based on their abundance in extracellular vesicle membranes, commercial availability, and large-scale production feasibility while keeping rest of the LNP components unchanged. The transfection kinetics of GFP mRNA encapsulated in LNPs and doped with 16 mol% of helper lipids was tested using GL261, U87 and SIM-A9 cell lines. Several LNP formations resulted in stable transfection (upto 5 days) of GFP mRNA in all the cell lines tested in vitro. The successful LNP candidates (enabling >80% transfection efficacy) were then tested in vivo to deliver luciferase mRNA to brain tumors via intrathecal administration in a syngeneic glioblastoma (GBM) mouse model, which confirmed luciferase expression in brain tumors in the cortex. LNPs were then tested to deliver Cre recombinase mRNA in syngeneic GBM mouse model genetically modified to express tdTomato under LoxP marker cassette that enabled identification of LNP targeted cells. mRNA was successfully delivered to tumor cells (70-80% transfected) and a range of different cells in the tumor microenvironment, including tumor-associated macrophages (80-90% transfected), neurons (31- 40% transfected), neural stem cells (39-62% transfected), oligodendrocytes (70-80% transfected) and astrocytes (44-76% transfected). Then, LNP formulations were assessed for delivering Cas9 mRNA and CD81 sgRNA (model protein) in murine syngeneic GBM model to enable gene editing in brain tumor cells. Sanger sequencing showed that CRISPR-Cas9 editing was successful in ~94% of brain tumor cells in vivo. In conclusion, we have developed a library of safe LNPs that can transfect GBM cells in vivo with high efficacy. This technology can potentially be used to develop novel mRNA therapies for GBM by delivering single or multiple mRNAs and holds great potential as a tool to study brain tumor biology.

Lipid nanoparticle library towards development next generation genomic medicines

Ionizable amino lipids are a major constituent of the lipid nanoparticles for delivering nucleic acid therapeutics (e.g., DLin-MC3-DMA in ONPATTRO , ALC-0315 in Comirnaty , SM-102 in Spikevax ). Scarcity of lipids that are suitable for cell therapy, vaccination, and gene therapies continue to be a problem in advancing many potential diagnostic/therapeutic/vaccine candidates to the clinic. Herein, we describe the development of novel ionizable lipids to be used as functional excipients for designing vehicles for nucleic acid therapeutics/vaccines in vivo or ex vivo use in cell therapy applications. We first studied the transfection efficiency (TE) of LNP-based mRNA formulations of these ionizable lipid candidates in primary human T cells and established a workflow for engineering of primary immune T cells. We then adapted this workflow towards bioengineering of CAR constructs to T cells towards non-viral CAR T therapy. Lipids were also tested in rodents for vaccine applications using self-amplifying RNA (saRNA) encoding various antigens. We have then evaluated various ionizable lipid candidates and their biodistribution along with the mRNA/DNA translation exploration using various LNP compositions. Further, using ionizable lipids from the library, we have shown gene editing of various targets in rodents. We believe that these studies will pave the path to the advancement in nucleic acid based therapeutics and vaccines, or cell gene therapy agents for early diagnosis and detection of cancer, and for targeted genomic medicines towards cancer treatment and diagnosis.

Soft Ionization: Improving Indoor Air Quality

In this paper we report two applications of a subcategory of air cleaning devices based on soft ionization that do not cause molecular fragmentation. A system that includes two unipolar ionizing modules has been used to simultaneously produce positive and negative ions in the air. In one set of experiments a large chamber (28 m3) was used to study the effect of ions on reducing PM1.0 particles produced by a research grade calibrated cigarette. The data presented in this paper were obtained using a carbon-brush-based bipolar ionizer and a MERV 10 filter with electret media in a recirculating HVAC system. Significant improvement in removal rate of fine and ultrafine particles was achieved when using the bipolar ionizer in conjunction with the MERV 10 filter. The second set of experiments were conducted using a 36 m3 chamber, following BSL-3 standards, to study the effect of ions on aerosolized SARS-CoV-2. Results of these investigations reveal the inactivation rate of SARS-CoV-2 are enhanced when ions are introduced in the air;inactivation rates were increased by more than 60%and 90%for ion densities of 10,000/cc and 18,000/cc. IEEE

Advances in approved nucleic acid drugs and lipid nanoparticle system

Nucleic acids, as a next generation of biotechnology drugs, not only can fundamentally treat diseases, but also own significant platform characteristics in view of technology and production. Therefore, nucleic acid-based drugs have broad clinical applications in biomedical fields. However, nucleic acids are degradable and unstable, and have very low intracellular delivery efficiency in vitro and in vivo, which greatly limits their applications. In recent years, ionizable lipid-based lipid nanoparticles have shown promising application potentials and have been successfully applied to COVID-19 (Coronavirus Disease 2019) vaccines in clinic. Lipid nanoparticles demonstrate high in vivo delivery efficiency and good safety profile due to their unique structural and physicochemical properties, which provides many possibilities for their clinical applications for nucleic acid delivery in the future. This review focused on the characteristics of nucleic acid drugs and their delivery barriers, and discussed the approved nucleic acid drugs to illustrate the key aspects of the success of their delivery carrier system. In addition, problems to be solved in the field were highlighted.Copyright © 2023, Chinese Pharmaceutical Association. All rights reserved.

Performance of in-duct bipolar ionization devices on pollutant removal and potential byproduct formation in indoor environments

The COVID-19 pandemic has highlighted the importance of indoor air quality (IAQ) since SARS-CoV-2 may be transmitted through virus-laden aerosols in poorly ventilated spaces. Multiple air cleaning technologies have been developed to mitigate airborne transmission risk and improve IAQ. In-duct bipolar ionization technology is an air cleaning technology that can generate ions for inactivating airborne pathogens and increasing particle deposition and removal while without significant byproducts generated. Many commercial in-duct ionization systems have been developed but their practical performance on pollutant removal and potential formation of byproducts have not been investigated comprehensively. The results in this study showed that the in-duct bipolar ionization technology can significantly improve the particle removal efficiency of the regular filter, while no significant ozone and ion were released to the indoor air. © 2022 17th International Conference on Indoor Air Quality and Climate, INDOOR AIR 2022. All rights reserved.

Analytical and biomedical applications of microfluidics in traditional Chinese medicine research

Traditional Chinese medicine (TCM) has significant benefits in the prevention and treatment of diseases due to its unique theoretical system and research techniques. However, there are still key issues to be resolved in the full interpretation and use of TCM, such as vague active compounds and mechanism of action. Therefore, it is promising to promote the research on TCM through innovative strategies and advanced cutting-edge technologies. Microfluidic chips have provided controllable unique platforms for biomedical applications in TCM research with flexible composition and large-scale integration. In this review, the analysis and biomedical applications of microfluidics in the field of TCM are highlighted, including quality control of Chinese herbal medicines (CHMs), delivery of CHMs, evaluation of pharmacological activity as well as disease diagnosis. Finally, potential challenges and prospects of existing microfluidic technologies in the inheritance and innovation of TCM are discussed.Copyright © 2022 Elsevier B.V.

A Review on Candida Auris, an Emerging Drug-resistant Fungal Pathogen.

Candida auris, a multidrug-resistant yeast, can cause primary or secondary infections in a wide range of patients, including those diagnosed with the new coronavirus to even healthy individuals. The fungus has been reported in less than a decade on all six continents and in more than 45 countries. Ease of distribution, long shelf life, and resistance to several antifungal drugs have raised concerns about the prevention and management of patients with C. auris infection. Recent reports indicate serious challenges in identifying, understanding the mechanism of drug resistance, and preventing mortality from the infection with this microorganism. Given the prevalence of COVID-19 infection, it is important to identify patients colonized with C. auris correctly and at the early stages, to control and prevent a possible outbreak. In this article, the widespread occurrence of infections due to C. auris in the world and Iran, its clinical manifestations, risk factors, pathogenic mechanisms, diagnostic enhancements and challenges, drug resistance, treatment options, prevention, and control as well as concomitant C. auris infections in patients with COVID-19 virus, are reviewed.Copyright © 2022 Isfahan University of Medical Sciences(IUMS). All rights reserved.

Ventilator-associated lower respiratory tract infections: Etiology and diagnosis.

Objective. To review a literature published over the past 5 years and our own data on the etiology of lower respiratory tract infections (LRTI), antimicrobial resistance and its relationships between sepsis and choice of appropriate antibiotic therapy. Materials and methods. National Nosocomial Infections Surveillance (NNIS) criteria were used to diagnose LRTI. A review of the articles regarding LRTI from the Russian and international English language journals published over 6 years was performed. Identification of microorganisms was performed by culture over the period of 2003-2013;since 2014, MALDI-TOF MS method was used for this purpose. Results. Despite the ongoing policy to limit the use of antimicrobial therapy in the ICUs, there is an increase in carbapenemase-producing isolates in the ICUs from 2.2% (2018) to 11.7% (2020, 9 months). Along with the trend to increase in carbapenemase-producing pathogens causing LRTI, their variability is also increasing. In particular, it applies to strains producing carbapenemases OXA-48 or combination of OXA- 48 with KPC;with the trend to combined production of carbapenemase beginning at 2019. Conclusions. Carbapenemase producers are becoming more widespread in the ICU settings, including the lower respiratory tract in mechanically ventilated patients. Practitioners didn't get used to associate VAP with the Sepsis-3 criteria. The changes in etiology include the increased rate of carbapenem-resistant Enterobacterales and non-fermenting Gram-negative bacteria, primarily Acinetobacter spp., in Russia. It's due to improved quality of respiratory support and increased consumption of carbapenems, tigecycline and polymyxins. Significant increase of OXA-48-producing pathogens is likely to be associated with a poor compliance with temporary guidelines on COVID-19 with regard to antibiotic therapy.Copyright © 2021, Interregional Association for Clinical Microbiology and Antimicrobial Chemotherapy. All rights reserved.

Establishing Novel Antiretroviral Imaging for Hair to Elucidate Nonadherence: Protocol for a Single-Arm Cross-sectional Study.

BACKGROUND: Adherence to antiretroviral (ARV) therapy is critical for achieving HIV RNA suppression in people living with HIV and for preventing HIV infection in uninfected individuals using preexposure prophylaxis. However, a high level of adherence can be challenging to achieve for people living with HIV on lifelong ARVs and for HIV-negative individuals using daily preexposure prophylaxis who are not at daily risk for HIV infection. Current biological measures of adherence are invasive and use bioanalytical methods that do not allow for real-time feedback during a clinic visit. This study was designed to test the feasibility and acceptability of using MedViewer, a novel, minimally invasive, hair-based assay that measures longitudinal ARV drug adherence in real time and provides an output for provider-patient discussion. OBJECTIVE: The primary objectives were to investigate the feasibility of delivering the MedViewer results as planned, the acceptability of participation in a discussion of the MedViewer results, and the appropriateness of using MedViewer for adherence counseling. The secondary objectives were to investigate additional dimensions of feasibility, acceptability, and appropriateness of using the MedViewer test during a routine clinic visit for people with HIV. METHODS: The proposed study was a single-arm cross-sectional study among patients receiving HIV care and providers of HIV care in a southeastern infectious disease clinic. The study originally planned to implement the MedViewer test with 50 eligible patients who were living with HIV across 2 viral load strata (undetectable or detectable plasma HIV RNA over the previous 2 years), administer brief visit-specific questionnaires to all patient and provider participants, and conduct qualitative in-depth interviews and quantitative end-line questionnaires with a subsample of patient participants (n=30) and all provider participants. RESULTS: The Establishing Novel Antiretroviral Imaging for Hair to Elucidate Nonadherence study was funded by the National Institute of Allergy and Infectious Diseases and approved by the local institutional review board on November 4, 2019. Provider participant enrollment began on January 17, 2020, and patient participant enrollment began on January 22, 2020. Participant enrollment was halted on March 16, 2020, because of the COVID-19 pandemic (16 providers and 10 patients on study). Study activities resumed on February 2, 2021, with COVID-19 modifications approved by the local institutional review board. Participant enrollment closed on October 8, 2021, and data collection closed on November 15, 2021. In total, 36 unique patient participants, representing 37 samples, and 20 provider participants were enrolled. Data analysis and manuscript writing will take place throughout 2023. CONCLUSIONS: We anticipate that the data collected through this study will provide important insights regarding the feasibility, acceptability, and appropriateness of incorporating new real-time longitudinal, minimally invasive adherence tests into routine clinical care and identify potential barriers to medication adherence among patients. TRIAL REGISTRATION: ClinicalTrials.gov NCT04232540; https://clinicaltrials.gov/ct2/show/NCT04232540. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): RR1-10.2196/41188.

RESISTANCE FACTORS OF KLEBSIELLA PNEUMONIAE BACTERIA DURING COVID-19 PANDEMIC.

Multidrug-resistant K. pneumoniae bacterial strains producing extended range of beta-lactamases or carbapenemases are of serious clinical concern. The aim of the study was to determine the resistance factors of K. pneumoniae strains isolated from the lower respiratory tract of patients diagnosed with community-acquired pneumonia during the COVID-19 pandemic. Materials and methods. The study of resistance to antimicrobial drugs included 138 strains of K. pneumoniae isolated from the sputum of patients treated in infectious diseases monohospitals in the city of Tyumen and the Tyumen region within the period from May 2020 to June 2021. Among the strains examined, 51.4% of them were isolated from SARS-CoV-2 positive patients. The presence of resistance genes was determined by PCR in 71 strains of K. pneumoniae (34 strains from COVID-19-positive and 37 strains from COVID-19-negative patients). Identification of isolated bacterial strains was carried out according to the protein spectra by using a desktop time-of-flight mass spectrometer with matrix laser desorption MALDI-TOF MS (Bruker, Germany). The belonging of the strains to the hypermucoid phenotype was determined using the string test. Sensitivity to antimicrobial drugs was assessed in the disk diffusion method on Muller-Hinton medium. The sensitivity of culture strains to bacteriophage preparations was determined by the drop method (spot-test). In the study, we used "Polyvalent Sextaphage Pyobacteriophage" and "Purified Polyvalent Klebsiella Bacteriophage", JSC NPO Microgen, Russia. Detection of resistance genes to beta-lactam antibiotics by real-time PCR was carried out using the BakRezista kit (OOO DNA-technology, Russia). The results of the study evidence that K. pneumoniae bacteria isolated from COVID-19-positive and COVID-19-negative patients diagnosed with community-acquired pneumonia displayed a high resistance to antimicrobial drugs and commercial phage-containing drugs. Resistance of K. pneumoniae strains was recorded from 50% (to aminoglycosides and carbapenems) to 90% (to inhibitor-protected penicillins). Sensitivity to bacteriophages was noted on average in no more than 20% of strains. It is important to emphasize that strains isolated from COVID-19-positive patients more often showed a hypermucoid phenotype, suggesting a high bacterial virulence, and also showed greater resistance to all groups of antibacterial drugs examined in the study, which is confirmed by the presence of resistance genes of the ESBL group and carbapenemase. The results of the study suggest that the high level of resistance of K. pneumoniae strains isolated from COVID-19-positive patients is associated with immunosuppression provoked by the SARS-CoV-2 virus, which contributes to their colonization by more virulent strains.Copyright © 2022 Saint Petersburg Pasteur Institute. All rights reserved.

COVID-19: From the Molecular Mechanisms to Treatment

The 2019 novel coronavirus (SARS-CoV-2) causes severe pneumonia called COVID-19 and leads to severe acute respiratory syndrome with a high mortality rate. The SARS-CoV-2 virus in the human body leads to jumpstarting immune reactions and multi-organ inflammation, which has poorer outcomes in the presence of predisposing conditions, including hypertension, dyslipidemia, dysglycemia, abnormal adiposity, and even endothelial dysfunction via biomolecular mechanisms. In addition, leucopenia, hypoxemia, and high levels of both cytokines and chemokines in the acute phase of this disease, as well as some abnormalities in chest CT images, were reported in most patients. The spike protein in SARS-CoV-2, the primary cell surface protein, helps the virus anchor and enter the human host cells. Additionally, new mutations have mainly happened for spike protein, which has promoted the infection's transmissibility and severity, which may influence manufactured vaccines' efficacy. The exact mechanisms of the pathogenesis, besides molecular aspects of COVID-19 related to the disease stages, are not well known. The altered molecular functions in the case of immune responses, including T CD4+, CD8+, and NK cells, besides the overactivity in other components and outstanding factors in cytokines like interleukin-2, were involved in severe cases of SARS-CoV-2. Accordingly, it is highly needed to identify the SARS-CoV-2 bio-molecular characteristics to help identify the pathogenesis of COVID-19. This study aimed to investigate the bio-molecular aspects of SARS-CoV-2 infection, focusing on novel SARS-CoV-2 variants and their effects on vaccine efficacy.Copyright © 2022 NRITLD, National Research Institute of Tuberculosis and Lung Disease, Iran.

mRNA expression and delivery efficacy of lipid nanoparticles in the cells breast tumor microenvironment: in vitro and in vivo evaluation

During the COVID-19 pandemics we have all witnessed the clinical importance of mRNA as current vaccines and future therapeutics. mRNA therapies have a potential to revolutionize cancer treatment. Delivery of mRNA requires lipid nanoparticles (LNP) to protect the cargo from degradation. mRNA has a negative charge and depends on positively charged lipids to be encapsulated in LNP. These lipids can be either ionizable at certain pH or constantly cationic. Even though previous studies had evaluated the formulation properties of ionizable and cationic LNP systems, there is the need to understand their specificity in terms of mRNA delivery and protein expression in breast cancer tumor microenvironment. The objective of this work was to assess the kinetics of LNP cellular uptake and mRNA expression inv breast cancer (BC) cells and fibroblasts, the most frequent cell type in the tumor microenvironment cells, while studying the mechanisms involved in differential behaviors of LNP formulated with cationic and ionizable lipids. To achieve this goal mRNA-LNP containing ionizable lipids (LNP-A) and cationic lipids (LNP-B) were designed and formulated using Nanoassemblr Benchtop microfluidics mixer (Precision NanoSystems). mRNA-LNP were characterized for size, zeta potential using dynamic light scattering (DLS) and mRNA encapsulation efficiency using RiboGreen assay. LNP were tagged with rhodamine lipid to investigate the uptake kinetic and a reporter GFP mRNA to evaluate mRNA expression in murine 4T1 and human MCF7, MDA-231, SUM-159 and T47D breast cancer cells and BJ fibroblasts. Live fluorescence microscopy imaging, IncuCyte S3, was used to determine the LNP uptake and GFP mRNA expression. In vitro biocompatibility was assessed with WST-1 assay. Additionally, expression of mRNA delivered from LNP in tumor microenvironment was evaluated in vivo in a syngeneic 4T1 breast cancer model using mRNA luciferase and IVIS imaging. mRNA-LNPs possessed an average diameter of 77 - 107 nm, narrow size distribution, neutral zeta potential and high mRNA encapsulation efficiency (>94%). Our results demonstrated that mRNA expression was higher in breast cancer cells when delivered from LNP-A formulation and in BJ fibroblasts when delivered from LNP-B. LNP-A, the ionizable LNP, was tested in the breast cancer cells to confirm the efficacy of the delivery. The highest transfection efficacy, from high to low, T-47D, MCF7, SUM-159, 4T1 and MDA-231.We have further investigated the cellular uptake mechanisms of LNP using uptake pathway inhibitors for caveolae endocytosis, clathrin endocytosis, and phagocytosis. Our data confirm that there are differences in mechanisms that govern the uptake of mRNA LNP in breast cancer cells and fibroblasts. Clathrin-mediated endocytosis was active in 4T1 breast cancer cells for ionizable and cationic LNP. Interestingly, despite in vitro differences in uptake and mRNA expression, in vivo results show that both formulations efficiently delivered luciferasemRNA in the tumor microenvironment. Histology results demonstrated similar luciferase expression for both LNP in tumors. Additionally, we were able to confirm the prominent presence of fibroblast and similar distribution in the 4T1 subcutaneous model which could explain the similar efficacy of cationic and ionizable LNP. Understanding uptake and mRNA expression of different LNP formulations in the tumor microenvironment can help in achieving the necessary protein expression for breast cancer therapies. Furthermore, determining the most efficient carrier in early stages may reduce the time required for clinical translation. Acknowledgement: This research was supported in part by CPRIT Core for RNA Therapeutics and Research.

An Investigation into the Presence of Pathogens on Inanimate Surfaces in the Hospital Environment

Background The hospital environment is understood to play an important role in the transmission of nosocomial pathogens, with inanimate surfaces facilitating pathogen movement and persistence in the environment. The majority of studies of surface contamination have been carried out in outbreak conditions or on high-dependency wards. Current surface cleaning guidance only requires surfaces to be visually clean. Microbiological standards for cleanliness have been proposed however they are not widely adopted and little implementation guidance is available. Evidence-based surface sampling protocols are required for the transmission risk surfaces pose to be adequately quantified and addressed. Methods Environmental surface samples will be collected in a number of in- and outpatient settings staff and public areas, such as waiting rooms. This will be done before and after cleaning. Pathogens associated with nosocomial infection (e.g., ESKAPE pathogens Clostridioides difficile and Candida auris) will be identified through culture methods and MALDI-ToF mass spectrometry. Antimicrobial susceptibility profiles of isolates will be determined following EUCAST protocols. Realtime PCR will be utilized to identify viral pathogens (including norovirus adenovirus influenza and SARS-CoV-2) present. Following this samples will be collected for community composition sequencing allowing for non-culturable microorganisms to be identified. Whole genome sequencing will be performed on any pathogens of interest isolated during this investigation. Results and Conclusion Fewer microorganisms are expected to be isolated after cleaning than prior and samples from inpatient environments will have higher proportions of pathogens. The hospital microbiome has rarely been investigated outside of outbreak conditions. This study presents a novel, systematic approach to assess the microorganisms present in the hospital environment and how they are impacted by current cleaning measures. This will build a comprehensive picture of the hospital microflora and provide an evidence base for the development of surface sampling protocols helping inform clinical risk assessments and subsequently improving patient outcomes.

Development of respirator design for children using bamboo-based activated carbon filter and bipolar ionization

This paper proposes a respirator for children integrated with bamboo-based activated carbon filters and bipolar ionization as the sterilizer to increase protection from Covid-19 and other pathogens. The respirator, named as Bion-Kids, was designed based on the children's head-and-face anthropometry from direct measurement. The duration of sterilization process is controlled based on the activities, which are classified using the Recurrent Neural Network (RNN) model with dataset acquired from the accelerometer. Manufacturing process of the main body prototype comprises heat bending and 3D printing. The bamboo-based activated carbon filter was synthesized by applying heat and pressure at the strewed activated carbon particles on the N95 filter. Observation using scanning electron microscope showed the ability of the bamboo-based activated carbon filter to adsorb and trap the particles. Result of the qualitative fit test and questionnaire survey indicated that Bion-Kids is suitable and comfortable for the children's face. The result of filtration and bipolar ionization sterilization system analysis have also met the ISO14698 standard by not showing any infiltration of microorganisms on the blood agar media. This device may become an early-stage personal protective equipment against the transmission of Covid-19 and other pathogens. © 2022 THE AUTHORS

Carbon nanotube substrates enhance SARS-CoV-2 spike protein ion yields in matrix-assisted laser desorption-ionization mass spectrometry

Nanostructured surfaces enhance ion yields in matrix-assisted laser desorption-ionization mass spectrometry (MALDI-MS). The spike protein complex, S1, is one fingerprint signature of Sars-CoV-2 with a mass of 75 kDa. Here, we show that MALDI-MS yields of Sars-CoV-2 spike protein ions in the 100 kDa range are enhanced 50-fold when the matrix-analyte solution is placed on substrates that are coated with a dense forest of multi-walled carbon nanotubes, compared to yields from uncoated substrates. Nanostructured substrates can support the development of mass spectrometry techniques for sensitive pathogen detection and environmental monitoring. © 2023 Author(s).

Confirmation of radiant catalytic ionization efficacy for airborne SARS-CoV-2 elimination indoors using "COVID19 traps".

Radiant catalytic ionization (RCI) is a novel technology that uses the appropriate wavelength (240-260 nm) and the phenomenon of photo-oxidation leading to permanent removal of viruses, bacteria, and fungi. Here, two analyses were performed. The first of them was a complete analysis of environmental biosecurity in a hospital environment. The second one was a longitudinal study with 40 patients with confirmed COVID19 and high viral load to assess the efficacy of RCI technology eliminating airborne SARS-CoV-2 indoors. A significant decrease in the number of bacteria and fungi colony-forming units (CFUs) was found in rooms with RCI when compared with rooms without it (p=0.03 for both of them). In the second part of the study, 16 samples out of 40 (40%) were positives when RCI technology was absent; whereas, these samples were negative when the equipment was on. Incidence rates (IR) with their Poisson 95% Confidence Intervals (CI) were calculated as the number of positive tests with the purifier or without it, showing an IR difference of 48.5% [CI(15.9-81), p=0.004]. Furthermore, the IR ratio was calculated obtaining a value of 3.3, confirming that RCI diminished more than 3-fold the presence of the SARS-CoV-2 in the air of the patients' rooms, thus laying the first stone in the fight for prevention of SARS-CoV-2 dissemination indoors.

Nanokit coupled electrospray ionization mass spectrometry for analysis of angiotensin converting enzyme 2 activity in single living cell.

Angiotensin-converting enzyme 2 (ACE2) is not only an enzyme but also a functional receptor on cell membrane for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Here, the activity of ACE2 in single living cell is firstly determined using a nanokit coupled electrospray ionization mass spectrometry (nanokit-ESI-MS). Upon the insertion of a micro-capillary into the living hACE2-CHO cell and the electrochemical sorting of the cytosol, the target ACE2 enzyme hydrolyses angiotensin II inside the capillary to generate angiotensin 1-7. After the electrospray of the mixture at the tip of the capillary, the product is differentiated from the substrate in molecular weight to achieve the detection of ACE2 activity in single cells. The further measurement illustrates that the inflammatory state of cells does not lead to the significant change of ACE2 catalytic activity, which elucidates the relationship between intracellular ACE2 activity and inflammation at single cell level. The established strategy will provide a specific analytical method for further studying the role of ACE2 in the process of virus infection, and extend the application of nanokit based single cell analysis.

Massive empyema due to Lactococcus garvieae

Introduction Lactococcus garvieae, a zoonotic pathogen, may rarely infect humans through the consumption of fish. Documented manifestations of L. garvieae infection in humans include infective endocarditis, prosthetic joint infections, liver abscesses, peritoneal dialysis-associated peritonitis, osteomyelitis, meningitis, infective spondylodiscitis, acalculous cholecystitis, and urinary tract infection. Case report An 87-year-old female was hospitalized for coffee-ground emesis secondary to acute gastritis after eating cooked fish. One week after her discharge, she developed new-onset confusion and was returned to the hospital. Chest computed tomography revealed total consolidation of the left lung and a multiloculated left pleural effusion. The patient required intubation and direct admission to the intensive care unit. Pleural fluid and blood cultures grew L. garvieae, which was susceptible to ceftriaxone, penicillin, and vancomycin. Despite intensive antibiotic therapy and supportive care for thirteen days, the patient remained in irreversibl e shock, and the family opted for comfort care. Conclusions Heretofore unreported, this case demonstrates that L. garvieae can cause bronchopneumonia and empyema. Copyright © GERMS 2022.

How to dissect viral infections and their interplay with the host-proteome by immunoaffinity and mass spectrometry: A tutorial

[Display omitted] • Immunohistochemistry with magnetic core nanoparticles to isolate viruses. • The use of MALDI-MS for rapid virus detection is explained in detail. • The use of ESI-MS/MS to pinpoint host-patient crosstalk is explained in detail. • The absolute quantitative MS is explained for large-scale protein quantitation. The capabilities of bioanalytical mass spectrometry to (i) detect and differentiate viruses at the peptide level whilst maintaining high sample throughput and (ii) to provide diagnosis and prognosis for infected patients are presented as a tutorial in this work to aid analytical chemists and physicians to gain insights into the possibilities offered by current high-resolution mass spectrometry technology and bioinformatics. From (i) sampling to sample treatment;(ii) Matrix-Assisted Laser Desorption Ionization- to Electrospray Ionization -based mass spectrometry;and (iii) from clustering to peptide sequencing;a detailed step-by-step guide is provided and exemplified using SARS-CoV-2 Spike Y839 variant and the variant of concern SARS-CoV-2 Alpha (B.1.1.7 lineage), Influenza B, and Influenza A subtypes AH1N1pdm09 and AH3N2. [ FROM AUTHOR]