Ozone Eliminates SARS-CoV-2 from Difficult-to-Clean Office Supplies and Clinical Equipment.

(1) Background: Severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) continues to cause profound health, economic, and social problems worldwide. The management and disinfection of materials used daily in health centers and common working environments have prompted concerns about the control of coronavirus disease 2019 (COVID-19) infection risk. Ozone is a powerful oxidizing agent that has been widely used in disinfection processes for decades. The aim of this study was to assess the optimal conditions of ozone treatment for the elimination of heat-inactivated SARS-CoV-2 from office supplies (personal computer monitors, keyboards, and computer mice) and clinical equipment (continuous positive airway pressure tubes and personal protective equipment) that are difficult to clean. (2) Methods: The office supplies and clinical equipment were contaminated in an area of 1 cm2 with 1 × 104 viral units of a heat-inactivated SARS-CoV-2 strain, then treated with ozone using two different ozone devices: a specifically designed ozonation chamber (for low-medium ozone concentrations over large volumes) and a clinical ozone generator (for high ozone concentrations over small volumes). SARS-CoV-2 gene detection was carried out using quantitative real-time polymerase chain reaction (RT-qPCR). (3) Results: At high ozone concentrations over small surfaces, the ozone eliminated SARS-CoV-2 RNA in short time periods-i.e., 10 min (at 4000 ppm) or less. The optimum ozone concentration over large volumes was 90 ppm for 120 min in ambient conditions (24 °C and 60-75% relative humidity). (4) Conclusions: This study showed that the appropriate ozone concentration and exposure time eliminated heat-inactivated SARS-CoV-2 RNA from the surfaces of different widely used clinical and office supplies, decreasing their risk of transmission, and improving their reutilization. Ozone may provide an additional tool to control the spread of the COVID-19 pandemic.

Ozone treatment effectively eliminates SARS-CoV-2 from infected face masks.

The current COVID-19 pandemic is causing profound health, economic, and social problems worldwide. The global shortage of medical and personal protective equipment (PPE) in specialized centers during the outbreak demonstrated the need for efficient methods to disinfect and recycle them in times of emergency. We have previously described that high ozone concentrations destroyed viral RNA in an inactivated SARS-CoV-2 strain within a few minutes. However, the efficient ozone dosages for active SARS-CoV-2 are still unknown. The present study aimed to evaluate the systematic effects of ozone exposure on face masks from hospitalized patients infected with SARS-CoV-2. Face masks from COVID-19 patients were collected and treated with a clinical ozone generator at high ozone concentrations in small volumes for short periods. The study focused on SARS-CoV-2 gene detection (assessed by real-time quantitative polymerase chain reaction (RT-qPCR)) and on the virus inactivation by in vitro studies. We assessed the effects of different high ozone concentrations and exposure times on decontamination efficiency. We showed that high ozone concentrations (10,000, 2,000, and 4,000 ppm) and short exposure times (10, 10, and 2 minutes, respectively), inactivated both the original strain and the B.1.1.7 strain of SARS-CoV-2 from 24 contaminated face masks from COVID-19 patients. The validation results showed that the best condition for SARS-CoV-2 inactivation was a treatment of 4,000 ppm of ozone for 2 minutes. Further studies are in progress to advance the potential applications of these findings.

Effects of Ozone Treatment on Personal Protective Equipment Contaminated with SARS-CoV-2.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is causing profound health, economic, and social problems worldwide. Management of personal protective equipment (PPE) and its potential limited availability have created concerns about the increased risks for healthcare professionals at hospitals and nursing homes. Ozone is a powerful oxidant agent. The objectives of this study were to examine the effects of ozone treatment on PPE contaminated with SARS-CoV-2, and to explore whether relative humidity could modify those effects. METHODS: PPE contaminated by heat-inactivated SARS-CoV-2 were treated with different ozone concentrations, exposure times, and relative humidity conditions. SARS-CoV-2 gene amplification was assessed by real-time polymerase chain reaction. RESULTS: There was no amplification of SARS-CoV-2 in PPE after the following ozone exposures: 30 s at 10,000 ppm (20 g/m3), 5 min at 4000 ppm, and 10 min at 2000 ppm. At lower ozone concentrations, 4-12 ppm (0.008-0.024 g/m3), the effects were highly dependent on the relative humidity conditions. CONCLUSIONS: Oxidative stress induced by ozone exposure eliminated heat-inactivated SARS-CoV-2 in different PPE components under appropriate exposure times, ozone concentrations, and relative humidity conditions. These findings could have implications in decreasing the risk of contamination associated with personal protective equipment management and in increasing its availability. Further research in the original SARS-CoV-2 strain is guaranteed.

Systemic Effects Induced by Hyperoxia in a Preclinical Model of Intra-abdominal Sepsis.

Supplemental oxygen is a supportive treatment in patients with sepsis to balance tissue oxygen delivery and demand in the tissues. However, hyperoxia may induce some pathological effects. We sought to assess organ damage associated with hyperoxia and its correlation with the production of reactive oxygen species (ROS) in a preclinical model of intra-abdominal sepsis. For this purpose, sepsis was induced in male, Sprague-Dawley rats by cecal ligation and puncture (CLP). We randomly assigned experimental animals to three groups: control (healthy animals), septic (CLP), and sham-septic (surgical intervention without CLP). At 18 h after CLP, septic (n = 39), sham-septic (n = 16), and healthy (n = 24) animals were placed within a sealed Plexiglas cage and randomly distributed into four groups for continuous treatment with 21%, 40%, 60%, or 100% oxygen for 24 h. At the end of the experimental period, we evaluated serum levels of cytokines, organ damage biomarkers, histological examination of brain and lung tissue, and ROS production in each surviving animal. We found that high oxygen concentrations increased IL-6 and biomarkers of organ damage levels in septic animals, although no relevant histopathological lung or brain damage was observed. Healthy rats had an increase in IL-6 and aspartate aminotransferase at high oxygen concentration. IL-6 levels, but not ROS levels, are correlated with markers of organ damage. In our study, the use of high oxygen concentrations in a clinically relevant model of intra-abdominal sepsis was associated with enhanced inflammation and organ damage. These findings were unrelated to ROS release into circulation. Hyperoxia could exacerbate sepsis-induced inflammation, and it could be by itself detrimental. Our study highlights the need of developing safer thresholds for oxygen therapy.

Differentiation at necropsy between in vivo gas embolism and putrefaction using a gas score.

Gas bubble lesions consistent with decompression sickness in marine mammals were described for the first time in beaked whales stranded in temporal and spatial association with military exercises. Putrefaction gas is a post-mortem artifact, which hinders the interpretation of gas found at necropsy. Gas analyses have been proven to help differentiating putrefaction gases from gases formed after hyperbaric exposures. Unfortunately, chemical analysis cannot always be performed. Post-mortem computed tomography is used to study gas collections, but many different logistical obstacles and obvious challenges, like the size of the animal or the transport of the animal from the stranding location to the scanner, limit its use in stranded marine mammals. In this study, we tested the diagnostic value of an index-based method for characterizing the amount and topography of gas found grossly during necropsies. For this purpose, putrefaction gases, intravenously infused atmospheric air, and gases produced by decompression were evaluated at necropsy with increased post-mortem time in New Zealand White Rabbits using a gas score index. Statistical differences (P<0.001) were found between the three experimental models immediately after death. Differences in gas score between in vivo gas embolism and putrefaction gases were found significant (P<0.05) throughout the 67h post-mortem. The gas score-index is a new and simple method that can be used by all stranding networks, which has been shown through this study to be a valid diagnostic tool to distinguish between fatal decompression, iatrogenic air embolism and putrefaction gases at autopsies.

Rectal pre-treatment with ozonized oxygen (O3) aggravates clinic status in septic rats treated with amoxicillin/clavulanate / El pretratamiento rectal con ozono (O3) empeora el estado clínico de ratas sépticas tratadas con amoxicilina/clavulánico

INTRODUCTION: Despite the advanced antibiotic therapies, sepsis continues being a clinical entity with high morbidity and mortality. The ozone/oxygen mixture (O3/O2) has been reported to exhibit positive effects on immunity. The aim of our study was to analyze whether (O3/O2) combined with amoxicillin/clavulanate has any influence on the morbidity and mortality of septic rats. METHODS: We used 48 Sprague-Dawley rats randomly allocated to 6 groups (n = 8): healthy (C), septic (I), healthy+ozone therapy (O3), septic+amoxicillin/clavulanate (AMC), septic+amoxicillin/clavulanate+ ozone therapy (AMC/O3 ) and septic + ozone therapy (I/O3 ). O3 /O2 was administered rectally at increasing O3 concentrations during 10 days prior to the onset of sepsis model (intraperitoneally injection of fecal material) or saline administration in healthy control rats. Later (post-inoculation), 3 days per week, O3 was also administered. Vital signs were recorded, and microbiological, hematological and histopathological studies were performed. RESULTS: The number of surviving animal/total was higher in AMC (8/8) than in AMC/O3 (4/8) p = 0.077. The percentage of surviving animals with pneumonia was higher in AMC/O3 than in AMC (100% vs 37.5%). In dead animals, AMC/O3 rats had a significantly higher percentage of lesions: Cardiac lesions, pulmonary hemorrhages and pleuritis (100%) and serositis/peritonitis (75%). Only Escherichia coli (2 different bio- types) was isolated from blood and/or peritoneal fluid from all infected groups. A significant decrease in the percentage of band neutrophils from the surviviors belonging to AMC/O3 vs AMC was observed (p < 0.05). CONCLUSION: Rectal pre-treatment with O3/O2 aggravates clinic status in septic rats treated with amoxi- cillin/clavulanate

INTRODUCCIÓN: A pesar de los avances en terapia antibiótica, la sepsis sigue siendo una entidad clínica con alta morbimortalidad. Se ha publicado que la mezcla ozono/oxígeno (O3/O2) presenta efectos beneficiosos sobre el sistema inmunológico. El objetivo de este estudio es analizar si (O3 /O2 ) combinado con amoxicilina/clavulánico tiene efectos en la morbimortalidad de ratas sépticas. MÉTODOS: Utilizamos 48 ratas Sprague-Dawley distribuidas aleatoriamente en 6 grupos (n = 8): sanas (C), sépticas (I), sanas+ozonoterapia (O3), sépticas+amoxicilina/clavulánico (AMC), sépticas+amoxicilina/ clavulánico + ozonoterapia (AMC/O3 ) y sépticas + ozonoterapia (I/O3 ). (O3 /O2 ) se administró por vía rectal a concentraciones crecientes de O3 los 10 días previos a la instauración del modelo de sepsis (inyección intraperitoneal de material fecal) o de la administración de solución salina, en las ratas control. Posteriormente (postinoculación) se continuó administrando (O3 /O2 ), 3 días por semana. Registramos los signos vitales y realizamos estudios microbiológicos, histopatológicos y hematológicos. RESULTADOS: El número de supervivientes/total fue mayor en AMC (8/8) que en AMC/O3 (4/8), p = 0,077. El porcentaje de supervivientes con neumonía fue mayor en AMC/O3 que en AMC (100% vs 37,5%). Entre los fallecidos, AMC/O3 tenía un porcentaje mayor de lesiones: cardiacas, hemorragias pulmonares y pleuritis (100%) y serositis/peritonitis (75%). A partir de la sangre y/o líquido peritoneal de los grupos infectados se aislaron exclusivamente Escherichia coli (2 biotipos diferentes). Observamos una disminución significativa en el porcentaje de neutrófilos en banda en las supervivientes pertenecientes a AMC/O3 vs AMC (p < 0,05). CONCLUSIÓN: El tratamiento rectal previo con (O3 /O2 ) agrava el estado clínico en ratas sépticas tratadas con amoxicilina/clavulánico

Altered Profile of Circulating Endothelial-Derived Microparticles in Ventilator-Induced Lung Injury.

OBJECTIVES: Pulmonary endothelial cell injury is central to the pathophysiology of acute lung injury. Mechanical ventilation can cause endothelial disruption and injury, even in the absence of preexisting inflammation. Platelet-endothelial cell adhesion molecule-1 is a transmembrane protein connecting adjacent endothelial cells. We hypothesized that injurious mechanical ventilation will increase circulating lung endothelial-derived microparticles, defined as microparticles positive for platelet-endothelial cell adhesion molecule-1, which could serve as potential biomarkers and mediators of ventilator-induced lung injury. DESIGN: Prospective randomized, controlled, animal investigation. SETTING: A hospital preclinical animal laboratory. SUBJECTS: Forty-eight Sprague-Dawley rats. INTERVENTIONS: Animals were randomly allocated to one of the three following ventilatory protocols for 4 hours: spontaneous breathing (control group), mechanical ventilation with low tidal volume (6 mL/kg), and mechanical ventilation with high tidal volume (20 mL/kg). In both mechanical ventilation groups, positive end-expiratory pressure of 2 cm H2O was applied. MEASUREMENTS AND MAIN RESULTS: We analyzed histologic lung damage, gas exchange, wet-to-dry lung weight ratio, serum cytokines levels, circulating endothelial-derived microparticles, platelet-endothelial cell adhesion molecule-1 lung protein content, and immunohistochemistry. When compared with low-tidal volume mechanical ventilation, high-tidal volume ventilation increased lung edema score and caused gas-exchange deterioration. These changes were associated with a marked increased of circulating endothelial-derived microparticles and a reduction of platelet-endothelial cell adhesion molecule-1 protein levels in the high-tidal volume lungs (p < 0.0001). CONCLUSIONS: There is an endothelial-derived microparticle profile associated with disease-specific features of ventilator-induced lung injury. This profile could serve both as a biomarker of acute lung injury and, potentially, as a mediator of systemic propagation of pulmonary inflammatory response.

Rectal pre-treatment with ozonized oxygen (O3) aggravates clinic status in septic rats treated with amoxicillin/clavulanate.

INTRODUCTION: Despite the advanced antibiotic therapies, sepsis continues being a clinical entity with high morbidity and mortality. The ozone/oxygen mixture (O3/O2) has been reported to exhibit positive effects on immunity. The aim of our study was to analyze whether (O3/O2) combined with amoxicillin/clavulanate has any influence on the morbidity and mortality of septic rats. METHODS: We used 48 Sprague-Dawley rats randomly allocated to 6 groups (n=8): healthy (C), septic (I), healthy+ozone therapy (O3), septic+amoxicillin/clavulanate (AMC), septic+amoxicillin/clavulanate+ozone therapy (AMC/O3) and septic+ozone therapy (I/O3). O3/O2 was administered rectally at increasing O3 concentrations during 10 days prior to the onset of sepsis model (intraperitoneally injection of fecal material) or saline administration in healthy control rats. Later (post-inoculation), 3 days per week, O3 was also administered. Vital signs were recorded, and microbiological, hematological and histopathological studies were performed. RESULTS: The number of surviving animal/total was higher in AMC (8/8) than in AMC/O3 (4/8) p=0.077. The percentage of surviving animals with pneumonia was higher in AMC/O3 than in AMC (100% vs 37.5%). In dead animals, AMC/O3 rats had a significantly higher percentage of lesions: Cardiac lesions, pulmonary hemorrhages and pleuritis (100%) and serositis/peritonitis (75%). Only Escherichia coli (2 different biotypes) was isolated from blood and/or peritoneal fluid from all infected groups. A significant decrease in the percentage of band neutrophils from the surviviors belonging to AMC/O3vs AMC was observed (p<0.05). CONCLUSION: Rectal pre-treatment with O3/O2 aggravates clinic status in septic rats treated with amoxicillin/clavulanate.

Soluble platelet-endothelial cell adhesion molecule-1, a biomarker of ventilator-induced lung injury.

INTRODUCTION: Endothelial cell injury is an important component of acute lung injury. Platelet-endothelial cell adhesion molecule-1 (PECAM1) is a transmembrane protein that connects endothelial cells to one another and can be detected as a soluble, truncated protein (sPECAM1) in serum. We hypothesized that injurious mechanical ventilation (MV) leads to shedding of PECAM1 from lung endothelial cells resulting in increasing sPECAM1 levels in the systemic circulation. METHODS: We studied 36 Sprague-Dawley rats in two prospective, randomized, controlled studies (healthy and septic) using established animal models of ventilator-induced lung injury. Animals (n = 6 in each group) were randomized to spontaneous breathing or two MV strategies: low tidal volume (VT) (6 ml/kg) and high-VT (20 ml/kg) on 2 cmH2O of positive end-expiratory pressure (PEEP). In low-VT septic animals, 10 cmH2O of PEEP was applied. We performed pulmonary histological and physiological evaluation and measured lung PECAM1 protein content and serum sPECAM1 levels after four hours ventilation period. RESULTS: High-VT MV caused severe lung injury in healthy and septic animals, and decreased lung PECAM1 protein content (P < 0.001). Animals on high-VT had a four- to six-fold increase of mean sPECAM1 serum levels than the unventilated counterpart (35.4 ± 10.4 versus 5.6 ± 1.7 ng/ml in healthy rats; 156.8 ± 47.6 versus 35.6 ± 12.6 ng/ml in septic rats) (P < 0.0001). Low-VT MV prevented these changes. Levels of sPECAM1 in healthy animals on high-VT MV paralleled the sPECAM1 levels of non-ventilated septic animals. CONCLUSIONS: Our findings suggest that circulating sPECAM1 may represent a promising biomarker for the detection and monitoring of ventilator-induced lung injury.