Silafluorene as a promising core for cell-permeant, highly bright and two-photon excitable fluorescent probes for live-cell imaging

International audienceIn this study, we report the synthesis, linear and non-linear photophysical studies and live cell imaging of two two-photon activatable probes based on a silafluorene core: SiFluo-V and SiFluo-L. Thanks to their quadrupolar (A-π-D-π-A) design, these probes exhibit respectively good to impressive two-photon cross sections (from 210 GM to 2150 GM). TD-DFT calculations support the experimental evidence that SiFluo-L displays far better two-photon absorption properties than SiFluo-V. Moreover, SiFluo-L possesses all requirements for bioimaging as it is water soluble, cell-permeant and and presents a low cytotoxicity (IC80≥10 µM). It labels mitochondria in live-cell imaging at low laser power with high brightness, contrast and photostability. This study demonstrates that silafluorene is a promising core to develop new two-photon fluorophores for live cell imaging

The Listeria monocytogenes Homolog of the Escherichia coli era Gene Is Involved in Adhesion to Inert Surfaces▿

Two transposon-insertional mutants of Listeria monocytogenes showing smaller viable surface-attached cell populations after disinfection with N,N-didecyl-N,N-dimethylammonium chloride were identified. In both mutants, transposon Tn917-lac was found to be inserted into the same gene, lmo1462, which is homologous to the essential Escherichia coli era gene. Both L. monocytogenes lmo1462-disrupted mutants displayed lower growth rates, as was also shown for several E. coli era mutants, and the lmo1462 gene was able to complement the growth defect of an E. coli era mutant. We showed that the disruption of lmo1462 decreased the ability of L. monocytogenes cells to adhere to stainless steel. Our results suggest that this era-like gene is involved in adhesion and contributes to the presence of L. monocytogenes on surfaces

Role of milk and milk products in the spread of methicillin‐resistant Staphylococcus aureus in the dairy production chain

International audienceMilk and milk products can harbor a multiple varieties of microorganisms. Therefore, they can be an important source of foodborne pathogens, including multidrug-resistant bacteria. Methicillin-resistant Staphylococcus aureus (MRSA) causes a wide spectrum of infections both in animals and humans. Over the last two decades, the presence of MRSA in foods and food-producing animals, including milk and milk products, has been frequently reported worldwide, raising public health concerns. In order to monitor and prevent foodborne MRSA contamination, it is necessary to understand their sources, the pheno/genotypic characteristics of the strains, and their transmission dynamics. In this review, studies conducted worldwide were summarized in order to assess the prevalence and diversity of MRSA circulating in milk and milk products. The risk factors for the occurrence of MRSA in milk and milk products were also discussed with preventive and control measures to avoid MRSA contamination in the dairy food chain

Organization and ELISA-Based Results of the First Proficiency Testing to Evaluate the Ability of European Union Laboratories to Detect Staphylococcal Enterotoxin Type B (SEB) in Buffer and Milk

The aim of this work was to organize the first proficiency test (PT) dedicated to staphylococcal enterotoxin B (SEB) detection in milk and buffer solutions. This paper describes the organization of the PT trial according to EN ISO 17043 requirements. Characterization of the SEB stock solution was performed using SDS-PAGE and SE-specific ELISA, and amino acid analysis was used to assign its protein concentration. The solution was then used to prepare six PT materials (four milk and two buffer batches) at a ng/g toxin level, which included one blank and one SEA-containing milk as specificity control. Suitable material homogeneity and stability were assessed using screening and quantitative ELISAs. Among the methods used by the participants, ELISA-based methods demonstrated their efficiency for the detection of SEB in both simple and complex matrices. The results serve as a basis for further improving the detection capabilities in expert laboratories and can therefore be considered as a contribution to biopreparedness

Dose-response Modelling of Staphylococcal Enterotoxins Using Outbreak Data

AbstractStaphylococcal food poisoning (SFP) is one of the most common food-borne diseases and results from the ingestion of staphylococcal enterotoxins (SEs). Yet, small amount of data are available for establishing a dose response. The objective of this work was to build a dose response relation based on the systematic investigations carried out during recent years in France. Over the period 2010-2014, more than 60 SFP outbreaks involving SEs, mainly from France, were microbiologically investigated. The enterotoxins were characterized as well as quantified. Attack rates, appearance times and natures of symptoms collected during epidemiological investigations were related to microbiological data. The outbreaks collected focused on enterotoxins SEA, SEB, SEC, and SED. Distribution of appearance times of symptoms and their natures were not influenced by the type of enterotoxins. The US EPA benchmark dose (BMD) methodology was then used to establish dose response. Attack rates of SFP outbreaks were modelled as a function of ingested doses and a BMD have been estimated for SEA

Ultrabright two-photon excitable red-emissive fluorogenic probes for fast and wash-free bioorthogonal labeling in live cells

Fluorogenic bioorthogonal reactions are promising tools for tracking small molecules or biomolecules in living organisms. Two-photon excitation, by shifting absorption towards the red, significantly increases the signal-to-noise ratio and decreases photodamages, while allowing to image about 10 times deeper than with a confocal. However, efficient two-photon excitable fluorogenic probes are currently lacking. We report here the design and synthesis of fluorogenic probes based on a two-photon excitable fluorophore and a tetrazine quenching moiety. These probes react with bicyclo[6.1.0]no-4-yn-9yl)methanol (BCN) with good to impressive kinetic rate constant (up to 1.1x103 M-1.s-1) and emit in the red window with moderate to high turn-on. TDDFT allowed to rationalize both the kinetic and fluorogenic performance of the different probes. The best candidate displays a turn-on of 13.8-fold measured by fluorescence intensities quantification in living cells under one-photon excitation, whereas a value of 3 is enough for live-cell imaging with high contrast. Moreover, live-cell imaging under two-photon excitation confirmed that there was no need for washing to monitor the reaction between BCN and this probe as a turn-on of 8.0-fold was measured under two-photon excitation. Finally, the high two-photon brightness of the clicked adduct (>300 GM) allows the use of a weak laser power compatible with in vivo imaging

On the road for more efficient biocompatible two-photon excitable fluorophores

International audienceRed to NIR absorption and emission wavelengths are key requirements for intravital bioimaging. One of the way to reach such excitation wavelengths is to use two-photon excitation. Unfortunately, there is still a lack of two-photon excitable fluorophores that are both efficient and biocompatible. Thus, we design a series of biocompatible quadrupolar dyes in order to study their ability to be used for live-cell imaging, and in particular for two-photon microscopy. Hence, we report the synthesis of 5 probes based on different donor cores (phenoxazine, acridane, phenazasiline and phenothiazine) and the study of their linear and non-linear photophysical properties. TD-DFT calculations were performed and were able to highlight the structure-property relationship of this series. All these studies highlight the great potential of three of these biocompatible dyes for two-Photon microscopy, as they both exhibit high two-photon cross-sections (up to 3 650 GM) and emit orange to red light. This potential was confirmed through live-cell two-photon microscopy experiments, leading to images with very high brightness and contrast

Influence de la sarcopénie sur la survie globale des patients pris en charge par chimiothérapie pour un cancer de l’œsophage

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