Preconcentration and sensitive determination of the anti-inflammatory drug diclofenac on a paper-based electroanalytical platform

This work describes the development of a paper-based platform for highly sensitive detection of diclofenac. The quantification of this anti-inflammatory drug is of importance in clinical (e.g. quality and therapeutic control) and environmental (e.g. emerging contaminant determination) areas. The easy-to-handle platform here described consists of a carbon-ink paper-based working electrode and two metallic wires, provided by a gold-plated standard connector, as reference and counter electrodes. The porous paper matrix enables the preconcentration of the sample, decoupling sample and detection solutions. Thus, relatively large sample volumes can be used, which significantly improves the sensitivity of the method. A wide dynamic range of four orders of magnitude, between 0.10 and 100 μM, was obtained for diclofenac determination. Due to the predominance of adsorption at the lowest concentrations, there were two linear concentration ranges: one comprised between 0.10 and 5.0 μM (with a slope of 0.85 μA μM-1) and the other between 5.0 and 100 μM (with a slope of 0.48 μA μM-1). A limit of detection of 70 nM was achieved with this simple device that provided accurate results with an RSD of ca. 5%. The platform was applied for diclofenac quantification in spiked tap water samples. The versatility of this design enabled the fabrication of a multiplexed platform containing eight electrochemical cells that work independently. The low cost, small size and simplicity of the device allow on-site analysis, which is very useful for environmental monitoring.The authors would like to thank the EU and FCT/UEFISCDI/FORMAS for funding, in the frame of the collaborative internationalconsortium REWATERfinanced under the ERA-NET CofundWaterWorks 2015 Call. This ERA-NET is an integral part of the 2016Joint Activities developed by the Water Challenges for a ChangingWorld Joint Programme Initiative (Water JPI). This work was alsosupported by the EU (FEDER funds through COMPETE) and FCT(project FOODnanoHEALTH, Portugal2020, Norte-01-0145-FEDER-000011 and project UID/QUI/50006/2013) and by the SpanishMinistry of Economy and Competitiveness (MINECO, projectCTQ2014-58826-R). Estefanía Costa Rama thanks the Governmentof Principado de Asturias and Marie Curie-Cofund Actions for thepost-doctoral grant“Clarín-Cofund”ACA17-20.info:eu-repo/semantics/publishedVersio

Green zero-valent iron nanoparticles for the degradation of amoxicillin

In the last years, it has been proven that zerovalent iron nanoparticles, including those produced using green methods, are efficient remediation agents for a wide range of target contaminants. However, apart from the known advantages of these green nanomaterials, the knowledge of how they act on distinct contaminants is not yet fully understood and requires further investigation. The objectives of this work were to study the degradation of a common antibiotic, amoxicillin, in water and in a sandy soil using green zero-valent iron nanoparticles (gnZVIs) as reductants and as catalysts for the Fenton reaction. It represents the first study of the use of gnZVI, as alternative for the zero-valent iron nanoparticles produced with sodium borohydride, for the degradation of amoxicillin. The results of the performed tests indicate that gnZVIs have the potential to be used in remediation processes. In both chemical tests, the gnZVI was able to degrade up to 100% of amoxicillin in aqueous solutions, using an amoxicillin/ gnZVI molar ratio of 1:15 when applied as a reductant, and an amoxicillin/H2O2/gnZVI molar ratio of 1:13:1 when applied as a catalyst for the Fenton reaction. The soil tests showed that the required molar ratios for near complete degradation were higher in the reduction test (1:150) than in the gnZVI-catalyzed Fenton reaction (1:1290:73). This is possibly due to parallel reactions with the soil matrix and/or limitations of the reagents to reach the entire soil sample. The degradation efficiencies obtained in these tests were 55 and 97% for the reduction and catalyzed Fenton processes, respectively.info:eu-repo/semantics/publishedVersio

Transforming Shiga toxin-producing Escherichia coli surveillance through whole genome sequencing in food safety practices

IntroductionShiga toxin-producing Escherichia coli (STEC) is a gastrointestinal pathogen causing foodborne outbreaks. Whole Genome Sequencing (WGS) in STEC surveillance holds promise in outbreak prevention and confinement, in broadening STEC epidemiology and in contributing to risk assessment and source attribution. However, despite international recommendations, WGS is often restricted to assist outbreak investigation and is not yet fully implemented in food safety surveillance across all European countries, in contrast to for example in the United States.MethodsIn this study, WGS was retrospectively applied to isolates collected within the context of Belgian food safety surveillance and combined with data from clinical isolates to evaluate its benefits. A cross-sector WGS-based collection of 754 strains from 1998 to 2020 was analyzed.ResultsWe confirmed that WGS in food safety surveillance allows accurate detection of genomic relationships between human cases and strains isolated from food samples, including those dispersed over time and geographical locations. Identifying these links can reveal new insights into outbreaks and direct epidemiological investigations to facilitate outbreak management. Complete WGS-based isolate characterization enabled expanding epidemiological insights related to circulating serotypes, virulence genes and antimicrobial resistance across different reservoirs. Moreover, associations between virulence genes and severe disease were determined by incorporating human metadata into the data analysis. Gaps in the surveillance system were identified and suggestions for optimization related to sample centralization, harmonizing isolation methods, and expanding sampling strategies were formulated.DiscussionThis study contributes to developing a representative WGS-based collection of circulating STEC strains and by illustrating its benefits, it aims to incite policymakers to support WGS uptake in food safety surveillance

The benefits of whole genome sequencing for foodborne outbreak investigation from the perspective of a national reference laboratory in a smaller country

Gradually, conventional methods for foodborne pathogen typing are replaced by whole genome sequencing (WGS). Despite studies describing the overall benefits, National Reference Laboratories of smaller countries often show slower uptake of WGS, mainly because of significant investments required to generate and analyze data of a limited amount of samples. To facilitate this process and incite policy makers to support its implementation, a Shiga toxin-producing Escherichia coli (STEC) O157:H7 (stx1+, stx2+, eae+) outbreak (2012) and a STEC O157:H7 (stx2+, eae+) outbreak (2013) were retrospectively analyzed using WGS and compared with their conventional investigations. The corresponding results were obtained, with WGS delivering even more information, e.g., on virulence and antimicrobial resistance genotypes. Besides a universal, all-in-one workflow with less hands-on-time (five versus seven actual working days for WGS versus conventional), WGS-based cgMLST-typing demonstrated increased resolution. This enabled an accurate cluster definition, which remained unsolved for the 2013 outbreak, partly due to scarce epidemiological linking with the suspect source. Moreover, it allowed detecting two and one earlier circulating STEC O157:H7 (stx1+, stx2+, eae+) and STEC O157:H7 (stx2+, eae+) strains as closely related to the 2012 and 2013 outbreaks, respectively, which might have further directed epidemiological investigation initially. Although some bottlenecks concerning centralized data-sharing, sampling strategies, and perceived costs should be considered, we delivered a proof-of-concept that even in smaller countries, WGS offers benefits for outbreak investigation, if a sufficient budget is available to ensure its implementation in surveillance. Indeed, applying a database with background isolates is critical in interpreting isolate relationships to outbreaks, and leveraging the true benefit of WGS in outbreak investigation and/or prevention

Validation strategy of a bioinformatics whole genome sequencing workflow for Shiga toxin-producing Escherichia coli using a reference collection extensively characterized with conventional methods

Whole genome sequencing (WGS) enables complete characterization of bacterial pathogenic isolates at single nucleotide resolution, making it the ultimate tool for routine surveillance and outbreak investigation. The lack of standardization, and the variation regarding bioinformatics workflows and parameters, however, complicates interoperability among (inter)national laboratories. We present a validation strategy applied to a bioinformatics workflow for Illumina data that performs complete characterization of Shiga toxin-producing Escherichia coli (STEC) isolates including antimicrobial resistance prediction, virulence gene detection, serotype prediction, plasmid replicon detection and sequence typing. The workflow supports three commonly used bioinformatics approaches for the detection of genes and alleles: alignment with blast+, kmer-based read mapping with KMA, and direct read mapping with SRST2. A collection of 131 STEC isolates collected from food and human sources, extensively characterized with conventional molecular methods, was used as a validation dataset. Using a validation strategy specifically adopted to WGS, we demonstrated high performance with repeatability, reproducibility, accuracy, precision, sensitivity and specificity above 95 % for the majority of all assays. The WGS workflow is publicly available as a ‘push-button’ pipeline at https://galaxy.sciensano.be. Our validation strategy and accompanying reference dataset consisting of both conventional and WGS data can be used for characterizing the performance of various bioinformatics workflows and assays, facilitating interoperability between laboratories with different WGS and bioinformatics set-ups

Gut Microbiome Dysbiosis in Antibiotic-Treated COVID-19 Patients is Associated with Microbial Translocation and Bacteremia

Although microbial populations in the gut microbiome are associated with COVID-19 severity, a causal impact on patient health has not been established. Here we provide evidence that gut microbiome dysbiosis is associated with translocation of bacteria into the blood during COVID-19, causing life-threatening secondary infections. We first demonstrate SARS-CoV-2 infection induces gut microbiome dysbiosis in mice, which correlated with alterations to Paneth cells and goblet cells, and markers of barrier permeability. Samples collected from 96 COVID-19 patients at two different clinical sites also revealed substantial gut microbiome dysbiosis, including blooms of opportunistic pathogenic bacterial genera known to include antimicrobial-resistant species. Analysis of blood culture results testing for secondary microbial bloodstream infections with paired microbiome data indicates that bacteria may translocate from the gut into the systemic circulation of COVID-19 patients. These results are consistent with a direct role for gut microbiome dysbiosis in enabling dangerous secondary infections during COVID-19