Post-treatment LSM rather than change during treatment predicts decompensation in patients with cACLD after HCV cure.

BACKGROUND & AIMS: Baveno VII has defined a clinically significant (i.e., prognostically meaningful) decrease in liver stiffness measurement (LSM) in cACLD as a decrease of ≥20% associated with a final LSM <20 kPa or any decrease to <10 kPa. However, these rules have not yet been validated against direct clinical endpoints. METHODS: We retrospectively analysed patients with cACLD (LSM ≥10 kPa) with paired liver stiffness measurement (LSM) before (BL) and after (FU) HCV cure by interferon-free therapies from 15 European centres. The cumulative incidence of hepatic decompensation was compared according to these criteria, considering hepatocellular carcinoma and non-liver-related death as competing risks. RESULTS: A total of 2,335 patients followed for a median of 6 years were analysed. Median BL-LSM was 16.6 kPa with 37.1% having ≥20 kPa. After HCV cure, FU-LSM decreased to a median of 10.9 kPa (<10 kPa: 1,002 [42.9%], ≥20 kPa: 465 [19.9%]) translating into a median LSM change of -5.3 (-8.8 to -2.4) kPa corresponding to -33.9 (-48.0 to -15.9) %. Patients achieving a clinically significant decrease (65.4%) had a significantly lower risk of hepatic decompensation (subdistribution hazard ratio: 0.12, 95% CI 0.04-0.35, p <0.001). However, these risk differences were primarily driven by a negligible risk in patients with FU-LSM <10 kPa (5-year cumulative incidence: 0.3%) compared to a high risk in patients with FU-LSM ≥20 kPa (16.6%). Patients with FU-LSM 10-19.9 kPa (37.4%) also had a low risk of hepatic decompensation (5-year cumulative incidence: 1.7%), and importantly, the risk of hepatic decompensation did not differ between those with/without an LSM decrease of ≥20% (p = 0.550). CONCLUSIONS: FU-LSM is key for risk stratification after HCV cure and should guide clinical decision making. LSM dynamics do not hold significant prognostic information in patients with FU-LSM 10-19.9 kPa, and thus, their consideration is not of sufficient incremental value in the specific context of HCV cure. IMPACT AND IMPLICATIONS: Liver stiffness measurement (LSM) is increasingly applied as a prognostic biomarker and commonly decreases in patients with compensated advanced chronic liver disease achieving HCV cure. Although Baveno VII proposed criteria for a clinically significant decrease, little is known about the prognostic utility of LSM dynamics (changes through antiviral therapy). Interestingly, in those with a post-treatment LSM of 10-19.9 kPa, LSM dynamics did not provide incremental information, arguing against the consideration of LSM dynamics as prognostic criteria. Thus, post-treatment LSM should guide the management of patients with compensated advanced chronic liver disease achieving HCV cure.

Computational aptamer design for spike glycoprotein (S) (SARS CoV-2) detection with an electrochemical aptasensor.

A new bioinformatic platform (APTERION) was used to design in a short time and with high specificity an aptamer for the detection of the spike protein, a structural protein of SARS-CoV-2 virus, responsible for the COVID-19 pandemic. The aptamer concentration on the carbon electrode surface was optimized using static contact angle and fluorescence method, while specificity was tested using differential pulse voltammetry (DPV) associated to carbon screen-printed electrodes. The data obtained demonstrated the good features of the aptamer which could be used to create a rapid method for the detection of SARS-CoV-2 virus. In fact, it is specific for spike also when tested against bovine serum albumin and lysozyme, competitor proteins if saliva is used as sample to test for the virus presence. Spectrofluorometric characterization allowed to measure the amount of aptamer present on the carbon electrode surface, while DPV measurements proved the affinity of the aptamer towards the spike protein and gave quantitative results. The acquired data allowed to conclude that the APTERION bioinformatic platform is a good method for aptamer design for rapidity and specificity. KEY POINTS: • Spike protein detection using an electrochemical biosensor • Aptamer characterization by contact angle and fluorescent measurements on electrode surface • Computational design of specific aptamers to speed up the aptameric sequence time.

Comparative meta-analysis of antimicrobial resistance from different food sources along with one health approach in the Egypt and UK.

BACKGROUND: Antimicrobial resistance (AMR) is a critical global issue that poses significant threats to human health, animal welfare, and the environment. With the increasing emergence of resistant microorganisms, the effectiveness of current antimicrobial medicines against common infections is diminishing. This study aims to conduct a competitive meta-analysis of surveillance data on resistant microorganisms and their antimicrobial resistance patterns in two countries, Egypt and the United Kingdom (UK). METHODS: Data for this study were obtained from published reports spanning the period from 2013 to 2022. In Egypt and the UK, a total of 9,751 and 10,602 food samples were analyzed, respectively. Among these samples, 3,205 (32.87%) in Egypt and 4,447 (41.94%) in the UK were found to contain AMR bacteria. RESULTS: In Egypt, the predominant resistance was observed against ß-lactam and aminoglycosides, while in the United Kingdom, most isolates exhibited resistance to tetracycline and ß-lactam. The findings from the analysis underscore the increasing prevalence of AMR in certain microorganisms, raising concerns about the development of multidrug resistance. CONCLUSION: This meta-analysis sheds light on the escalating AMR problem associated with certain microorganisms that pose a higher risk of multidrug resistance development. The significance of implementing One Health AMR surveillance is emphasized to bridge knowledge gaps and facilitate accurate AMR risk assessments, ensuring consumer safety. Urgent actions are needed on a global scale to combat AMR and preserve the effectiveness of antimicrobial treatments for the well-being of all living beings.

Light-Up Split Broccoli Aptamer as a Versatile Tool for RNA Assembly Monitoring in Cell-Free TX-TL Systems, Hybrid RNA/DNA Origami Tagging and DNA Biosensing.

Binary light-up aptamers are intriguing and emerging tools with potential in different fields. Herein, we demonstrate the versatility of a split Broccoli aptamer system able to turn on the fluorescence signal only in the presence of a complementary sequence. First, an RNA three-way junction harbouring the split system is assembled in an E. coli-based cell-free TX-TL system where the folding of the functional aptamer is demonstrated. Then, the same strategy is introduced into a 'bio-orthogonal' hybrid RNA/DNA rectangle origami characterized by atomic force microscopy: the activation of the split system through the origami self-assembly is demonstrated. Finally, our system is successfully used to detect the femtomoles of a Campylobacter spp. DNA target sequence. Potential applications of our system include the real-time monitoring of the self-assembly of nucleic-acid-based devices in vivo and of the intracellular delivery of therapeutic nanostructures, as well as the in vitro and in vivo detection of different DNA/RNA targets.

Detection of Listeria monocytogenes in foods with a textile organic electrochemical transistor biosensor.

Foods contaminated by pathogens are responsible for foodborne diseases which have socioeconomic impacts. Many approaches have been extensively investigated to obtain specific and sensitive methods to detect pathogens in food, but they are often not easy to perform and require trained personnel. This work aims to propose a textile organic electrochemical transistor-based (OECT) biosensor to detect L. monocytogenes in food samples. The analyses were performed with culture-based methods, Listeria Precis™ method, PCR, and our textile OECT biosensor which used poly(3,4-ethylenedioxythiophene) (PEDOT):polystyrene sulfonate (PSS) (PEDOT:PSS) for doping the organic channel. Atomic force microscopy (AFM) was used to obtain topographic maps of the gold gate. The electrochemical activity on gate electrodes was measured and related to the concentration of DNA extracted from samples and hybridized to the specific capture probe immobilized onto the gold surface of the gate. This assay reached a limit of detection of 1.05 ng/µL, corresponding to 0.56 pM of L. monocytogenes ATCC 7644, and allowed the specific and rapid detection of L. monocytogenes in the analyzed samples. KEYPOINTS: • Textile organic electrochemical transistors functionalized with a specific DNA probe • AFM topographic and surface potential maps of a functionalized gold gate surface • Comparison between the Listeria monocytogenes Precis™ method and an OECT biosensor.

Comparative meta-analysis of antimicrobial resistance from different food sources along with one health approach in Italy and Thailand.

Antimicrobial resistance (AMR) is increasing worldwide due to overuse, misuse and incomplete treatment of antibiotics. Many countries are facing the excessive issue due to the spreading of AMR not only in humans and animals, but also in water and agri-food sector. Our main aim was to perform a competitive meta-analysis of surveillance-resistant microbes and their antimicrobial superintendence in Italy and Thailand. Data have been collected from reports published for the period 2012-2021. A total of 9507 and 11,753 food samples contained 3905 (41.07%) and 3526 (30%) AMR bacteria in Italy and Thailand, respectively. In Italy, the highest microbial prevalence was ß-lactam and tetracycline, while in Thailand mostly isolates showed resistance to cephalosporin and aminoglycoside. Our findings contribute to highlighting the increment of AMR related to different microbes with tendency to become multidrug resistant.

Specific and sensitive detection of Influenza A virus using a biotin-coated nanoparticle enhanced immunomagnetic assay.

This study reports the development of a sensitive magnetic bead-based enzyme-linked immunoassay (MELISA) for the pan-reactive detection of the Influenza A virus. The assay combines immunomagnetic beads and biotin-nanoparticle-based detection to quantify a highly conserved viral nucleoprotein in virus lysates. At the capture step, monoclonal antibody-coated magnetic microbeads were used to bind and concentrate the nucleoprotein in samples. The colorimetric detection signal was amplified using biotinylated silica nanoparticles (NP). These nanoparticles were functionalized on the surface with short DNA spacers bearing biotin groups by an automated supported synthesis method performed on nano-on-micro assemblies with a DNA/RNA synthesizer. A biotin-nanoparticle and immunomagnetic bead-based assay was developed. We succeeded in detecting Influenza A viruses directly in the lysis buffer supplemented with 10% saliva to simulate the clinical context. The biotin-nanoparticle amplification step enabled detection limits as low as 3 × 103 PFU mL-1 and 4 × 104 PFU mL-1 to be achieved for the H1N1 and H3N2 strains respectively. In contrast, a classical ELISA test based on the same antibody sandwich showed detection limit of 1.2 × 107 PFU mL-1 for H1N1. The new enhanced MELISA proved to be specific, as no cross-reactivity was found with a porcine respiratory virus (PRRSV). Graphical abstract.

Enrichment Free qPCR for Rapid Identification and Quantification of Campylobacter jejuni, C. coli, C. lari, and C. upsaliensis in Chicken Meat Samples by a New Couple of Primers.

Campylobacter is the main cause of bacterial foodborne disease and poultry meat is the principal source of human infections. Rapid methods for Campylobacter detection are urgently needed to decrease high bacterial prevalence in poultry products. In this study, we developed new primers, CampyPFw and CampyPRv, that target the 16S-23S rRNA genes of Campylobacter jejuni, C. coli, C. lari and C. upsaliensis. The primers were tested on positive and negative reference strains in pure cultures and in inoculated poultry meat samples before their application in real-time PCR (qPCR) protocol for analyzing chicken meat samples. In parallel, the samples were tested by using the ISO 10272-1:2006 method. The qPCR protocol based on CampyPFw and CampyPRv showed good sensitivity, with the limit of detection of 4.6 × 102 cells/mL in chicken samples without enrichment steps.

Highly sensitive detection of Campylobacter spp. In chicken meat using a silica nanoparticle enhanced dot blot DNA biosensor.

Paper-based DNA biosensors are powerful tools in point-of-care diagnostics since they are affordable, portable, user-friendly, rapid and robust. However, their sensitivity is not always as high as required to enable DNA quantification. To improve the response of standard dot blots, we have applied a new enhancement strategy that increases the sensitivity of assays based on the use of biotinylated silica-nanoparticles (biotin-Si-NPs). After immobilization of a genomic Campylobacter DNA onto a paper membrane, and addition of a biotinylated-DNA detection probe, hybridization was evidenced using streptavidin-conjugated to horseradish peroxidase (HRP) in the presence of luminol and H2O2. Replacement of the single biotin by the biotin-Si-NPs boosted on average a 30 fold chemiluminescent read-out of the biosensor. Characterization of biotin-Si-NPs onto a paper with immobilized DNA was done using a scanning electron microscope. A limit of detection of 3 pg/µL of DNA, similar to the available qPCR kits, is achieved, but it is cheaper, easier and avoids inhibition of DNA polymerase by molecules from the food matrices. We demonstrated that the new dot blot coupled to biotin-Si-NPs successfully detected Campylobacter from naturally contaminated chicken meat, without needing a PCR step. Hence, such an enhanced dot blot paves the path to the development of a portable and multiplex paper based platform for point-of-care screening of chicken carcasses for Campylobacter.

Development and Evaluation of qPCR Detection Method and Zn-MgO/Alginate Active Packaging for Controlling Listeria monocytogenes Contamination in Cold-Smoked Salmon.

To answer to food industry requests to monitor the presence of L. monocytogenes in cold-smoked salmon samples and to extend their shelf-life, a qPCR protocol for the detection of L. monocytogenes, and an antibacterial active packaging reinforced with zinc magnesium oxide nanoparticles (Zn-MgO NPs) were developed. The qPCR allowed the sensitive and easy detection of L. monocytogenes in naturally contaminated samples, with specificity in full agreement with the standard methods. The halo diffusion study indicated a high antibacterial efficiency of 1 mg/mL Zn-MgO NPs against L. monocytogenes, while the flow cytometry showed only moderate cytotoxicity of the nanoparticles towards mammalian cells at a concentration above 1 mg/mL. Thus, the novel active packaging was developed by using 1 mg/mL of Zn-MgO NPs to reinforce the alginate film. Cold-smoked salmon samples inoculated with L. monocytogenes and air-packed with the Zn-MgO NPs-alginate nanobiocomposite film showed no bacterial proliferation at 4 °C during 4 days. In the same condition, L. monocytogenes growth in control contaminated samples packed with alginate film alone. Our results suggest that Zn-MgO nanoparticles can extend the shelf-life of cold-smoked salmon samples.

Acoustic Multi-Detection of Gliadin Using QCM Crystals Patterned with Controlled Sectors of TEM Grid and Annealed Nanoislands on Gold Electrode.

Celiac diseases are a group of gluten ingestion-correlated pathologies that are widespread and, in some cases, very dangerous for human health. The only effective treatment is the elimination of gluten from the diet throughout life. Nowadays, the food industries are very interested in cheap, easy-to-handle methods for detecting gluten in food, in order to provide their consumers with safe and high-quality food. Here, for the first time, the manufacture of controlled micropatterns of annealed gold nanoislands (AuNIs) on a single QCM crystal (QCM-color) and their biofunctionalization for the specific detection of traces of gliadin is reported. In addition, the modified quartz crystal with a TEM grid and 30 nm Au (Q-TEM grid crystal) is proposed as an acoustic sensitive biosensing platform for the rapid screening of the gliadin content in real food products.

Food Sensing: Detection of Bacillus cereus Spores in Dairy Products.

Milk is a source of essential nutrients for infants and adults, and its production has increased worldwide over the past years. Despite developments in the dairy industry, premature spoilage of milk due to the contamination by Bacillus cereus continues to be a problem and causes considerable economic losses. B. cereus is ubiquitously present in nature and can contaminate milk through a variety of means from the farm to the processing plant, during transport or distribution. There is a need to detect and quantify spores directly in food samples, because B. cereus might be present in food only in the sporulated form. Traditional microbiological detection methods used in dairy industries to detect spores show limits of time (they are time consuming), efficiency and sensitivity. The low level of B. cereus spores in milk implies that highly sensitive detection methods should be applied for dairy products screening for spore contamination. This review describes the advantages and disadvantages of classical microbiological methods used to detect B. cereus spores in milk and milk products, related to novel methods based on molecular biology, biosensors and nanotechnology.

Exploring the impact of Mg-doped ZnO nanoparticles on a model soil microorganism Bacillus subtilis.

The environmental contamination of soil by metal oxide nanomaterials is a growing global concern because of their potential toxicity. We investigated the effects of Mg doped ZnO (Mg-nZnO) nanoparticles on a model soil microorganism Bacillus subtilis. Mg-nZnO exhibited only a moderate toxic effect on B. subtilis vegetative cells but was able to prevent biofilm formation and destroy already formed biofilms. Similarly, Mg-nZnO (≤1 mg/mL) was moderately toxic towards Listeria monocytogenes, Staphylococcus aureus, Escherichia coli, Salmonella enterica, Saccharomyces cerevisiae and murine macrophages. Engineered Mg-nZnO produced H2O2 and O2•- radicals in solutions of various salt and organic molecule compositions. A quantitative proteomic analysis of B. subtilis membrane proteins showed that Mg-nZnO increased the expression of proteins involved in detoxification of ROS, translation and biofilm formation. Overall, our results suggest that Mg-nZnO released into the environment may hinder the spreading, colonization and biofilm formation by B. subtilis but also induce a mechanism of bacterial adaptation.

Electrochemical and Optical Biosensors for the Detection of Campylobacter and Listeria: An Update Look.

Foodborne safety has aroused tremendous research interest in recent years because of a global public health problem. The rapid and precise detection of foodborne pathogens can reduce significantly infection diseases and save lives by the early initiation of an effective treatment. This review highlights current advances in the development of biosensors for detection of Campylobacter spp. and Listeria monocytogenes that are the most common causes of zoonosis. The consumption of pathogen contaminated food is responsible for humans hospitalization and death. The attention focused on the recognition elements such as antibodies (Ab), DNA probes and aptamers able to recognize cells, amplicons, and specific genes from different samples like bacteria, food, environment and clinical samples. Moreover, the review focused on two main signal-transducing mechanisms, i.e., electrochemical, measuring an amperometric, potentiometric and impedimetric signal; and optical, measuring a light signal by OLED (Organic Light Emitting Diode), SPR (Surface Plasmon Resonance), and Optical fiber. We expect that high-performance of devices being developed through basic research will find extensive applications in environmental monitoring, biomedical diagnostics, and food safety.

Robust SERS Platforms Based on Annealed Gold Nanostructures Formed on Ultrafine Glass Substrates for Various (Bio)Applications.

In this study, stable gold nanoparticles (AuNPs) are fabricated for the first time on commercial ultrafine glass coverslips coated with gold thin layers (2 nm, 4 nm, 6 nm, and 8 nm) at 25 °C and annealed at high temperatures (350 °C, 450 °C, and 550 °C) on a hot plate for different periods of time. Such gold nanostructured coverslips were systematically tested via surface enhanced Raman spectroscopy (SERS) to identify their spectral performances in the presence of different concentrations of a model molecule, namely 1,2-bis-(4-pyridyl)-ethene (BPE). By using these SERS platforms, it is possible to detect BPE traces (10-12 M) in aqueous solutions in 120 s. The stability of SERS spectra over five weeks of thiol-DNA probe (2 µL) deposited on gold nano-structured coverslip is also reported.

Point-of-Need DNA Testing for Detection of Foodborne Pathogenic Bacteria.

Foodborne pathogenic bacteria present a crucial food safety issue. Conventional diagnostic methods are time-consuming and can be only performed on previously produced food. The advancing field of point-of-need diagnostic devices integrating molecular methods, biosensors, microfluidics, and nanomaterials offers new avenues for swift, low-cost detection of pathogens with high sensitivity and specificity. These analyses and screening of food items can be performed during all phases of production. This review presents major developments achieved in recent years in point-of-need diagnostics in land-based sector and sheds light on current challenges in achieving wider acceptance of portable devices in the food industry. Particular emphasis is placed on methods for testing nucleic acids, protocols for portable nucleic acid extraction and amplification, as well as on the means for low-cost detection and read-out signal amplification.

Effects of Silicone Implants on the Mammary Gland: Ultrasonographic and 3D Study.

BACKGROUND: Breast implants may be responsible for secondary deformities produced by parenchymal atrophy. However, few studies in the literature have reported changes in breast tissue after augmentation surgery. In this study, the breast thickness of patients undergoing breast augmentation was monitored by ultrasound, and correlations with surface, volume and projection measurements were examined. METHODS: We studied the parenchymal thickness at the lower pole of the breast with ultrasound in 36 women (72 breasts). In another group of 33 patients (66 breasts), we studied the thickness at the upper and lower poles along the meridian of each breast by ultrasound and measured the anthropometric metrics, volume and projection of the breast with a 3D camera. RESULTS: Midline measurements close to the areola showed reduced thickness at the lower pole, with 31.8% at the midpoint of the lower pole and 42% at the infra-areolar level (p < 0.0001). At the upper pole, there was a decrease of 14.6% (p < 0.001), but the thickness was increased by 6% and 38% at more cranial levels. No correlations with volume were found. Anatomical implants produced more atrophy at the lower pole, and round implants at the upper pole. More atrophy was found with subfascial than submuscular augmentation. Compared with the expected values, the final volume was very similar, but the projection was 29% less. Surface measurements changed significantly up to 4 months postoperatively and remained stable thereafter. CONCLUSIONS: Implants affect significatively the thickness of the glandular tissue. All changes occur very soon postoperatively but stabilize after 4 months. LEVEL OF EVIDENCE IV: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Rapid and label-free electrochemical DNA biosensor for detecting hepatitis A virus.

Diagnostic systems that can deliver highly specific and sensitive detection of hepatitis A virus (HAV) in food and water are of particular interest in many fields including food safety, biosecurity and control of outbreaks. Our aim was the development of an electrochemical method based on DNA hybridization to detect HAV. A ssDNA probe specific for HAV (capture probe) was designed and tested on DNAs from various viral and bacterial samples using Nested-Reverse Transcription Polymerase Chain Reaction (nRT-PCR). To develop the electrochemical device, a disposable gold electrode was functionalized with the specific capture probe and tested on complementary ssDNA and on HAV cDNA. The DNA hybridization on the electrode was measured through the monitoring of the oxidative peak potential of the indicator tripropylamine by cyclic voltammetry. To prevent non-specific binding the gold surface was treated with 3% BSA before detection. High resolution atomic force microscopy (AFM) confirmed the efficiency of electrode functionalization and on-electrode hybridization. The proposed device showed a limit of detection of 0.65pM for the complementary ssDNA and 6.94fg/µL for viral cDNA. For a comparison, nRT-PCR quantified the target HAV cDNA with a limit of detection of 6.4fg/µL. The DNA-sensor developed can be adapted to a portable format to be adopted as an easy-to- use and low cost method for screening HAV in contaminated food and water. In addition, it can be useful for rapid control of HAV infections as it takes only a few minutes to provide the results.

DNA origami nanorobot fiber optic genosensor to TMV.

In the quest of greater sensitivity and specificity of diagnostic systems, one continually searches for alternative DNA hybridization methods, enabling greater versatility and where possible field-enabled detection of target analytes. We present, herein, a hybrid molecular self-assembled scaffolded DNA origami entity, intimately immobilized via capture probes linked to aminopropyltriethoxysilane, onto a glass optical fiber end-face transducer, thus producing a novel biosensor. Immobilized DNA nanorobots with a switchable flap can then be actuated by a specific target DNA present in a sample, by exposing a hemin/G-quadruplex DNAzyme, which then catalyzes the generation of chemiluminescence, once the specific fiber probes are immersed in a luminol-based solution. Integrating organic nanorobots to inorganic fiber optics creates a hybrid system that we demonstrate as a proof-of-principle can be utilized in specific DNA sequence detection. This system has potential applications in a wide range of fields, including point-of-care diagnostics or cellular in vivo biosensing when using ultrathin fiber optic probes for research purposes.

Development of a chemiluminescent DNA fibre optic genosensor to Hepatitis A Virus (HAV).

Hepatitis A virus (HAV) infection has caused substantial morbidity and economic losses to human society, presenting a major public health problem in many parts of the world. Despite the capability for low-concentration detection, current PCR-based techniques are limited by the requirement of specialized lab equipment, trained personnel and a relatively large time-commitment. The need for a prompt in-field quantitative identification of HAV in real samples has led us to develop a chemiluminescent fibre optic genosensor system. In this study, a two-probe sandwich-type hybridization process was implemented on the tip of a fibre optic with an area of 0.12mm2. After optimization of the probes and the working conditions, we showed that the biosensor was able to work for both cDNA and RNA with a relatively large signal/noise ratio and a good sensitivity. An excellent specificity was also confirmed by screening with a broad range of other pathogen samples. The nucleic acid probes method was validated by optimized PCR and qPCR, and may thus be used when field testing would be required.