Currently, identification of pathogenic bacteria present at very low concentration requires a preliminary culture-based enrichment step. Many research efforts focus on the possibility to shorten this ...pre-enrichment step which is needed to reach the minimal number of cells that allows efficient identification. Rapid microbiological controls are a real public health issue and are required in food processing, water quality assessment or clinical pathology. Thus, the development of new methods for faster detection and isolation of pathogenic culturable bacteria is necessary. Here we describe a specific enrichment technique for culturable Gram negative bacteria, based on non-lethal click chemistry and the use of magnetic beads that allows fast detection and isolation. The assimilation and incorporation of an analog of Kdo, an essential component of lipopolysaccharides, possessing a bio-orthogonal azido function (Kdo-N3), allow functionalization of almost all Gram negative bacteria at the membrane level. Detection can be realized through strain-promoted azide-cyclooctyne cycloaddition, an example of click chemistry, which interestingly does not affect bacterial growth. Using E. coli as an example of Gram negative bacterium, we demonstrate the excellent specificity of the technique to detect culturable E. coli among bacterial mixtures also containing either dead E. coli, or live B. subtilis (as a model of microorganism not containing Kdo). Finally, in order to specifically isolate and concentrate culturable E. coli cells, we performed separation using magnetic beads in combination with click chemistry. This work highlights the efficiency of our technique to rapidly enrich and concentrate culturable Gram negative bacteria among other microorganisms that do not possess Kdo within their cell envelope.
Angeklickte Bakterien: Metabolisch aktive Gram‐negative Bakterien können ein azidmodifiziertes Analogon von 3‐Desoxy‐D‐mannooctulosonsäure (1; siehe Schema) spezifisch in Lipopolysaccharide auf ihrer ...Membran einbauen. Dieser Prozess versieht die Zelloberfläche mit einem bioorthogonalen chemischen Reporter und ermöglicht die schnelle Fluoreszenzmarkierung lebensfähiger Zellen mithilfe der Klick‐Chemie.
Toluene degradation performances were studied in a 10 L Two-Phase Partitioning Bioreactor（TPPB）.The liquid phase consisted of a mixture of water and PDMS 50（Poly Di Methyl Siloxane,i.e.silicone ...oil,viscosity of 46 m Pa·s） in the volume ratio of 75%/25%.Two series of experiments were carried out：in the first,the reactor was sequentially supplied with toluene whereas in the second,toluene was continuously supplied.Activated sludge from the wastewater treatment plant of Beaurade（Rennes,France） was used at an initial concentration of 0.5 dry mass g·（mixture L）~（-1）.The elimination capacity（EC） was investigated as well as the change in biomass concentration over time.Toluene biodegradation was very ef ficient（removal ef ficiency,RE=100%） for toluene flows ranging from 0.2 to 1.2 ml·h~（-1）,corresponding to elimination capacities of up to 104 g·m~（-3）·h~（-1）.For a toluene flow of 1.2 ml·h~（-1）,the biomass concentration measured at the end of the experiment was 4.7 dry mass g·（mixture L）~（-1）.The oxygen concentration in the liquid phase was clearly not a limiting factor in these operating conditions.Based on these results,an extrapolation leading to the design of a large-scale pilot TPPB can now be considered to study toluene degradation performances in industrial conditions.
Salmonella enterica is an intracellular bacterial pathogen that causes gastroenteritis and typhoid fever. Inside host cells, the bacterium is enclosed in a membrane bound compartment, the ...Salmonella-containing vacuole (SCV). Intracellular replication of Salmonella requires the translocation of effector proteins into the host cytosol. The SifA effector protein is important for the membrane stability of the SCV. Recently, we have shown that the Salmonella sifA- mutant presents on its vacuole an important accumulation of kinesin-1, a molecular motor involved in the plus-end-directed transport of various organelles. Kinesin-1 is not recruited on SCVs of mutants that do not translocate effector proteins. This indicates that SifA is a negative regulator of the recruitment of this molecular motor and reveals the existence of another effector that recruits kinesin-1. This chapter describes techniques that are used to screen by immunofluorescence microscopy the accumulation of kinesin-1 on strains of Salmonella carrying multiple mutations.
Legionella pneumophila is a pathogenic bacterium involved in regular outbreaks characterized by a relatively high fatality rate and an important societal impact. Frequent monitoring of the presence ...of this bacterium in environmental water samples is necessary to prevent these epidemic events, but the traditional culture‐based detection and identification method requires up to 10 days. Reported herein is a method allowing identification of Legionella pneumophila by metabolic lipopolysaccharide labeling which targets, for the first time, a precursor to monosaccharides that are specifically present within the O‐antigen of the bacterium. This new approach allows easy detection of living Legionella pneumophila, while other Legionella species are not labeled.
Mörderjagd: Auch fast 40 Jahre nach dem ersten nachweislichen Ausbruch in Philadelphia ist Legionella pneumophila noch immer schwer zu identifizieren. Die Lipopolysaccharid‐Metabolitmarkierung mit einem spezifischen Monosaccharid macht nun die Erkennung und Identifizierung lebender Vertreter dieses gefährlichen Pathogens möglich, ohne dass andere Legionella‐Spezies markiert werden.
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