Sedimentation field-flow fractionation for rapid phenotypic antimicrobial susceptibility testing: a pilot study.

BACKGROUND: The increase in antibiotic resistance is a major public health issue. The development of rapid antimicrobial susceptibility testing (AST) methods is becoming a priority to ensure early and appropriate antibiotic therapy. OBJECTIVES: To evaluate sedimentation field-flow fractionation (SdFFF) as a method for performing AST in less than 3 h. METHODS: SdFFF is based on the detection of early biophysical changes in bacteria, using a chromatographic-type technology. One hundred clinical Escherichia coli strains were studied. A calibrated bacterial suspension was incubated for 2 h at 37°C in the absence (untreated) or presence (treated) of five antibiotics used at EUCAST breakpoint concentrations. Bacterial suspensions were then injected into the SdFFF machine. For each E. coli isolate, retention times and elution profiles of antibiotic-treated bacteria were compared with retention times and elution profiles of untreated bacteria. Algorithms comparing retention times and elution profiles were used to determine if the strain was susceptible or resistant. Performance evaluation was done according to CLSI and the ISO standard 20776-2:2021 with broth microdilution used as the reference method. RESULTS: AST results from SdFFF were obtained in less than 3 h. SdFFF showed high categorical agreement (99.8%), sensitivity (99.5%) and specificity (100.0%) with broth microdilution. Results for each antimicrobial were also in agreement with the ISO 20776-2 recommendations, with sensitivity and specificity of ≥95.0%. CONCLUSIONS: This study showed that SdFFF can be used as a rapid, accurate and reliable phenotypic AST method with a turnaround time of less than 3 h.

Sedimentation Field-Flow Fractionation: A Diagnostic Tool for Rapid Antimicrobial Susceptibility Testing.

Conventional antimicrobial susceptibility testing (AST) methods require 24-48 h to provide results, creating the need for a probabilistic antibiotic therapy that increases the risk of antibiotic resistance emergence. Consequently, the development of rapid AST methods has become a priority. Over the past decades, sedimentation field-flow fractionation (SdFFF) has demonstrated high sensitivity in early monitoring of induced biological events in eukaryotic cell populations. This proof-of-concept study aimed at investigating SdFFF for the rapid assessment of bacterial susceptibility to antibiotics. Three bacterial species were included (Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa) with two panels of antibiotics tailored to each bacterial species. The results demonstrate that SdFFF, when used in "Hyperlayer" elution mode, enables monitoring of antibiotic-induced morphological changes. The percentage variation of the retention factor (PΔR) was used to quantify the biological effect of antibiotics on bacteria with the establishment of a threshold value of 16.8% to differentiate susceptible and resistant strains. The results obtained with SdFFF were compared to that of the AST reference method, and a categorical agreement of 100% was observed. Overall, this study demonstrates the potential of SdFFF as a rapid method for the determination of antibiotic susceptibility or resistance since it is able to provide results within a shorter time frame than that needed for conventional methods (3-4 h vs 16-24 h, respectively), enabling earlier targeted antibiotic therapy. Further research and validation are necessary to establish the effectiveness and reliability of SdFFF in clinical settings.

Canadian laboratory incidents with human pathogens and toxins: An overview of reports, 2016-2022.

Background: When the Public Health Agency of Canada's Human Pathogens and Toxins Act and Human Pathogens and Toxins Regulations came into force, the reporting of laboratory incidents to the Laboratory Incident Notification Canada (LINC) surveillance system became mandatory. This report summarizes the laboratory exposure and non-exposure data reported from 2016 to 2022, with a particular focus on factors that are not typically presented in LINC's annual report. Methods: Reported laboratory incidents from 2016 to 2022 were analyzed. Exposures were analyzed by severity, occurrence and root cause, and affected individuals were analyzed by disease outcome, role and applied interventions. Non-exposures were analyzed by incident type. Exposure and non-exposure incident rates were calculated. Results: Events reported to LINC totalled 928. Of those, 355 were confirmed non-exposures, 361 were confirmed exposures, and 111 were other events. Both exposure and non-exposure incident rates per 100 active licences peaked in 2018 (9.44 and 7.11, respectively). Most exposures were rated as minor or negligible severity. The most cited exposure occurrence types were sharps-related and procedure-related (23% each), and standard operating procedure-related root causes were most cited (24%). While 781 individuals were affected in the exposure incidents, most did not develop a laboratory-acquired infection (n=753; 96%) and received at least one form of treatment post-exposure (n=717; 92%). Inadvertent possession/production cases were the most common non-exposure incidents reported. Conclusion: Exposure and non-exposure incident rates have decreased since 2018. Among exposure incidents, sharps-related and procedure-related occurrences were the most common, and the root cause was usually a standard operating procedure. Non-exposure incidents were mostly inadvertent possession/production cases. Exposure and illness outcome severity was mostly minor.

Surveillance of laboratory exposures to human pathogens and toxins, Canada, 2022.

Background: The Laboratory Incident Notification Canada (LINC) surveillance system was launched in 2015 to monitor the mandated national reporting of laboratory incidents. This report describes the laboratory exposures reported in 2022. Methods: Exposure incidents were analyzed by activity, occurrence, sector, root cause and pathogens/toxins implicated, while affected individuals were analyzed by education, exposure route, role and years of laboratory experience. An analysis of the median number of exposures per month was conducted, and time between the exposure incident date and the date the incident was reported to LINC was examined. Results: Forty confirmed laboratory exposure incident reports were received, with two suspected laboratory-acquired infections. The exposure incident rate per 100 active licences was 3.8, and the number of exposure incidents was highest in September. The majority of exposure incidents involved risk group 2 pathogens (n=27; 63%) and non-security sensitive biological agents (n=36; 84%). Microbiology was the most cited activity occurring during the exposure event (n=20; 50%), and sharps and procedure-related issues were the most common occurrences (n=15; 24.2% each). Most incidents were reported by the academic sector (n=16; 40%). Human interaction was the most common root cause (n=20; 23.8%) and most affected individuals were technicians/technologists (n=68; 73.1%). The median time delay between the incident date and reporting date was 5.5 days. Conclusion: The exposure incident rate was lower in 2022 than in 2021. Incidents related to sharps and standard operating procedures remained the most common occurrence types. The most cited root cause of exposure incidents involved human interaction.

Improvement of Standardization of Molecular Analyses in Hematology: The 10-year GBMHM French Experience.

Molecular tests have become an indispensable tool for the diagnosis and prognosis of hematological malignancies and are subject to accreditation according to the International Standard ISO 15189. National standardization of these techniques is essential to ensure that patients throughout France benefit from the same care. We report here on the experience of the GBMHM (Groupe des Biologistes Moléculaires des Hémopathies Malignes). By organizing External Evaluation of Quality (EEQ) programs and training meetings, the GBMHM has contributed to improvement and standardization of molecular tests in 64 French laboratories. A retrospective analysis of the quality-control results of 11 national campaigns spanning 10 years was performed for the 3 most frequently prescribed tests: BCR-ABL1, JAK2 V617F, and lymphoid clonality. For each test, particular attention was placed on comparing methodologies and their evolution throughout the period. The establishment of the BCR-ABL1, JAK2 V617F, and lymphoid clonality EEQ programs and the associated training meetings have initiated a process of collective standardization concerning the methods of implementation (JAK2 V617F) and the interpretation and formulation of results (lymphoid clonality). In addition, it resulted in objective improvement in technical performance (BCR-ABL1). Our evaluation of the impact of these EEQ programs demonstrates that it is possible to obtain reproducible values across different laboratories in France by applying national recommendations. To our knowledge, this is the first publication that evaluates the impact of a national quality assurance program on improving molecular results in hematology.

Comparison of SimoaTM and EllaTM to assess serum neurofilament-light chain in multiple sclerosis.

We compared SimoaTM and EllaTM immunoassays to assess serum neurofilament-light chain levels in 203 multiple sclerosis patients from the OFSEP HD study. There was a strong correlation (ρ = 0.86, p < 0.0001) between both platforms. The EllaTM instrument overestimated values by 17%, but as the data were linear (p = 0.57), it was possible to apply a correction factor to EllaTM results. As for SimoaTM , serum neurofilament-light chain levels measured by EllaTM were correlated with age and EDSS and were significantly higher in active multiple sclerosis, suggesting that these assays are equivalent and can be used in routine clinical practice.

Supplementation with a Bioactive Melon Concentrate in Humans and Animals: Prevention of Oxidative Damages and Fatigue in the Context of a Moderate or Eccentric Physical Activity.

Exercise is recognized to provide both physical and psychological health benefits. However, oxidative stress can occur and induce muscular damages. SOD B® M is a melon concentrate, well known to counteract oxidative stress and prevent its side effects. The present study aimed to evaluate the potential of the melon concentrate in the context of both a strong and isolated effort associated with deleterious effects, and a moderate and regular physical activity considered as beneficial. First, a preclinical study was set up on rats to evaluate its potential on the prevention of damages induced by an eccentric exercise. Secondly, the combined effect of the melon concentrate and a regular standardized physical training was studied on the overall physical condition of healthy subjects in a randomized, double-blind, placebo-controlled trial. Repeated measures Analysis of Variance (ANOVA), student's t test and Mann-Whitney test were used for statistical analyses. Melon concentrate helped to prevent gastrocnemius damages induced by the eccentric exercise. It allowed a reduction of fibrosis by approximately 38% and a reduction of Tumor Necrosis Factor- α (TNF-α) plasma level by 28%. This supplementation also induced a rearrangement of myosin fibers and an increase in PGC-1α plasma level. In the clinical study, melon concentrate was able to decrease oxidative stress and C-Reactive protein (CRP) plasma level. Besides, magnesium (Mg) plasma level was higher in the context of a regular training performed by healthy subjects supplemented with the melon concentrate. Therefore, the melon concentrate allowed a better adaptation to effort linked to PGC-1α activation: a regulator of energy metabolism. The antioxidant properties of the melon concentrate and its ability to mobilize magnesium also suggest that the supplementation could induce a better resistance to fatigue and recovery during regular physical activity.

A Specific Melon Concentrate Exhibits Photoprotective Effects from Antioxidant Activity in Healthy Adults.

Skin is the largest body organ and the first barrier to exogenous threats. This organ is constantly exposed to external factors such as ultraviolet radiation, which induces many adverse effects including sunburn, depigmentation, photo aging, photo immune suppression, and even skin cancer. Antioxidants seem to be good candidates in order to reduce ultraviolet-mediated damages and to prevent the health consequences of ultraviolet exposure. The present investigation aims to further characterize the potential skin photoprotective effects of a food supplementation and a topical administration of a melon concentrate alone or in combination. A clinical study assessing the Minimal Erythema Dose (MED) was first set up to evaluate photoprotection. Afterward, an independent in vitro study was performed on human skin explants from a donor to evaluate the effect of the melon concentrate at different levels including on the sunburn cells formation and on the endogenous antioxidant enzymes and its influence on melanin. Clinical study results demonstrate that melon concentrate application and/or supplementation increased MED. It also increased the endogenous antioxidant enzymes and reduced sunburn cells and melanin level on irradiated skin explants. Therefore, it is suggested that melon concentrate administration (oral and/or topical) could be a useful strategy for photoprotection due to its antioxidant properties.

Placebo-controlled double-blind clomipramine trial for the treatment of anxiety or fear in beagles during ground transport.

The purpose of this explorative study was, first, to document changes in physiological parameters and behavior observed in dogs following ground transport and, second, to measure the effects on the above variables of a short-term administration of clomipramine, anecdotally already prescribed in private veterinary practice to reduce fear, anxiety, or both. Twenty-four beagles were randomly allocated to either clomipramine (2 mg/kg, q12h for 7 d) or placebo treatment, and then transported 3 times in a truck for 1 hour. Physiological parameters (cortisol, neutrophil:lymphocyte (N:L) ratio, heart rate) and behavior were recorded and analyzed. Clomipramine significantly reduced plasma cortisol (P < 0.05) following transport and tended (P = 0.07) to reduce N:L ratio. Clomipramine tended to only reduce "moving and panting" and drooling. Short-term administration of clomipramine appears to slightly reduce fear, anxiety, or both during transport. More research is needed to confirm the efficacy of this treatment and the appropriate dosage.

Relaxin and atrial natriuretic peptide pathways participate in the anti-fibrotic effect of a melon concentrate in spontaneously hypertensive rats.

BACKGROUND: In spontaneously hypertensive rats (SHR), a model of human essential hypertension, oxidative stress is involved in the development of cardiac hypertrophy and fibrosis associated with hypertension. Dietary supplementation with agents exhibiting antioxidant properties could have a beneficial effect in remodeling of the heart. We previously demonstrated a potent anti-hypertrophic effect of a specific melon (Cucumis melo L.) concentrate with antioxidant properties in spontaneously hypertensive rats. Relaxin and atrial natriuretic peptide (ANP) were reported to reduce collagen deposition and fibrosis progression in various experimental models. OBJECTIVE: The aim of the present investigation was to test the hypothesis that, beside reduction in oxidative stress, the melon concentrate may act through relaxin, its receptor (relaxin/insulin-like family peptide receptor 1, RXFP1), and ANP in SHR. DESIGN AND RESULTS: The melon concentrate, given orally during 4 days, reduced cardiomyocyte size (by 25%) and totally reversed cardiac collagen content (Sirius red staining) in SHR but not in their normotensive controls. Treatment with the melon concentrate lowered cardiac nitrotyrosine-stained area (by 45%) and increased by 17-19% the cardiac expression (Western blot) of superoxide dismutase (SOD) and glutathione peroxidase. In addition, plasma relaxin concentration was normalized while cardiac relaxin (Western blot) was lowered in treated SHR. Cardiac relaxin receptor level determined by immunohistochemical analysis increased only in treated SHR. Similarly, the melon concentrate reversed the reduction of plasma ANP concentration and lowered its cardiac expression. CONCLUSIONS: The present results demonstrate that reversal of cardiac fibrosis by the melon concentrate involves antioxidant defenses, as well as relaxin and ANP pathways restoration. It is suggested that dietary SOD supplementation could be a useful additional strategy against cardiac hypertrophy and fibrosis.