The rod‐shaped cells of Myxococcus xanthus, a Gram‐negative deltaproteobacterium, differentiate to environmentally resistant spores upon starvation or chemical stress. The environmental resistance ...depends on a spore coat polysaccharide that is synthesised by the ExoA‐I proteins, some of which are part of a Wzx/Wzy‐dependent pathway for polysaccharide synthesis and export; however, key components of this pathway have remained unidentified. Here, we identify and characterise two additional loci encoding proteins with homology to enzymes involved in polysaccharide synthesis and export, as well as sugar modification and show that six of the proteins encoded by these loci are essential for the formation of environmentally resistant spores. Our data support that MXAN_3260, renamed ExoM and MXAN_3026, renamed ExoJ, are the Wzx flippase and Wzy polymerase, respectively, responsible for translocation and polymerisation of the repeat unit of the spore coat polysaccharide. Moreover, we provide evidence that three glycosyltransferases (MXAN_3027/ExoK, MXAN_3262/ExoO and MXAN_3263/ExoP) and a polysaccharide deacetylase (MXAN_3259/ExoL) are important for formation of the intact spore coat, while ExoE is the polyisoprenyl‐phosphate hexose‐1‐phosphate transferase responsible for initiating repeat unit synthesis, likely by transferring N‐acetylgalactosamine‐1‐P to undecaprenyl‐phosphate. Together, our data generate a more complete model of the Exo pathway for spore coat polysaccharide biosynthesis and export.
Myxococcus xanthus differentiates to environmentally resistant spores in response to various stresses. Resistance is provided by the spore coat polysaccharide that is synthesised by the ExoA‐I proteins, which make up an incomplete Wzx/Wzy pathway for polysaccharide synthesis and export. Here, we identified the Wzx flippase and Wzy polymerase as well as three glycosyltransferase and a polysaccharide deacetylase that are important for formation of the intact spore coat, thus, providing a more complete Exo pathway.
Summary
Myxococcus xanthus is a model bacterium to study social behavior. At the cellular level, the different social behaviors of M. xanthus involve extensive cell–cell contacts. Here, we used ...bioinformatics, genetics, heterologous expression and biochemical experiments to identify and characterize the key enzymes in M. xanthus implicated in O‐antigen and lipopolysaccharide (LPS) biosynthesis and examined the role of LPS O‐antigen in M. xanthus social behaviors. We identified WbaPMx (MXAN_2922) as the polyisoprenyl‐phosphate hexose‐1‐phosphate transferase responsible for priming O‐antigen synthesis. In heterologous expression experiments, WbaPMx complemented a Salmonella enterica mutant lacking the endogenous WbaP that primes O‐antigen synthesis, indicating that WbaPMx transfers galactose‐1‐P to undecaprenyl‐phosphate. We also identified WaaLMx (MXAN_2919), as the O‐antigen ligase that joins O‐antigen to lipid A‐core. Our data also support the previous suggestion that WzmMx (MXAN_4622) and WztMx (MXAN_4623) form the Wzm/Wzt ABC transporter. We show that mutations that block different steps in LPS O‐antigen synthesis can cause pleiotropic phenotypes. Also, using a wbaPMx deletion mutant, we revisited the role of LPS O‐antigen and demonstrate that it is important for gliding motility, conditionally important for type IV pili‐dependent motility and required to complete the developmental program leading to the formation of spore‐filled fruiting bodies.
Myxococcus xanthus displays several social behaviors involving cell–cell contacts. To understand the role of the lipopolysaccharide (LPS) in these behaviors, we identified key enzymes involved in O‐antigen and LPS biosynthesis: the polyisoprenyl‐phosphate hexose‐1‐phosphate transferase, responsible for priming O‐antigen synthesis and the O‐antigen ligase. We show that LPS O‐antigen is important for gliding motility, conditionally important for type IV pili‐dependent motility and required to complete the developmental program resulting in fruiting body formation.
Summary
Global dRNA‐seq analysis of transcription start sites combined with in silico annotation using Infernal software revealed the expression of 91 putative non‐coding sRNA in Sphingopyxis granuli ...TFA cells grown on different carbon sources. Excluding housekeeping sRNAs, only one additional sRNA, which belongs to the Rfam SuhB family (RF00519), was detected by Infernal but with an incorrect size according to the experimental results. SuhB is highly conserved across the Sphingopyxis genus. Expression data revealed that SuhB is present in rapidly growing TFA cells. A suhB deletion mutant exhibited de‐repression of tetralin degradation (thn) gene expression and higher amounts of their LysR‐type activator, ThnR, under conditions of carbon catabolite repression (CCR). Interaction between SuhB and the 5′UTR of thnR mRNA was demonstrated in vitro. Moreover, co‐immunoprecipitation experiments, combined with fluorescence measurements of gfp fusions to the 5′UTR of thnR mRNA and the phenotype of an hfq deletion mutant, suggest the involvement of Hfq in this interaction. Taken together, these data support an Hfq‐mediated repressive role for SuhB, on ThnR mRNA translation that prevents thn gene induction. SuhB, which is a highly conserved sRNA in the Sphingopyxis genus, is the first identified element directly involved in CCR of thn gene expression in S. granuli strain TFA.
•Rapeseed straw hemicellulosic hydrolysate was used for xylitol production.•Different strategies were evaluated for the hydrolysate detoxification.•C. guilliermondii exhibited higher tolerance to the ...toxic compounds than D. hansenii.•Toxic compounds and glucose concentration affected the yeast’s performance.•Partial removal of toxic compounds was enough to achieve an efficient bioconversion.
This study evaluated the possibility of using rapeseed straw hemicellulosic hydrolysate as a fermentation medium for xylitol production. Two yeast strains, namely Debaryomyces hansenii and Candida guilliermondii, were used for this bioconversion process and their performance to convert xylose into xylitol was compared. Additionally, different strategies were evaluated for the hydrolysate detoxification before its use as a fermentation medium. Assays in semi-defined media were also performed to verify the influence of hexose sugars on xylose metabolism by the yeasts. C. guilliermondii exhibited higher tolerance to toxic compounds than D. hansenii. Not only the toxic compounds present in the hydrolysate affected the yeast’s performance, but glucose also had a negative impact on their performance. It was not necessary to completely eliminate the toxic compounds to obtain an efficient conversion of xylose into xylitol, mainly by C. guilliermondii (YP/S=0.55g/g and 0.45g/g for C. guilliermondii and D. hansenii, respectively).
Current strategies to treat pelvic organ prolapse (POP) or stress urinary incontinence (SUI), include the surgical implantation of vaginal meshes. Recently, there have been multiple reports of issues ...generated by these meshes conventionally made of poly(propylene). This material is not the ideal candidate, due to its mechanical properties leading to complications such as chronic pain and infection. In the present manuscript, we propose the use of an alternative material, thermoplastic polyurethane (TPU), loaded with an antibiotic in combination with fused deposition modelling (FDM) to prepare safer vaginal meshes. For this purpose, TPU filaments containing levofloxacin (LFX) in various concentrations (e.g., 0.25%, 0.5%, and 1%) were produced by extrusion. These filaments were used to 3D print vaginal meshes. The printed meshes were fully characterized through different tests/analyses such as fracture force studies, attenuated total reflection-Fourier transform infrared, thermal analysis, scanning electron microscopy, X-ray microcomputed tomography (μCT), release studies and microbiology testing. The results showed that LFX was uniformly distributed within the TPU matrix, regardless the concentration loaded. The mechanical properties showed that poly(propylene) (PP) is a tougher material with a lower elasticity than TPU, which seemed to be a more suitable material due to its elasticity. In addition, the printed meshes showed a significant bacteriostatic activity on both
and
cultures, minimising the risk of infection after implanting them. Therefore, the incorporation of LFX to the TPU matrix can be used to prepare anti-infective vaginal meshes with enhanced mechanical properties compared with current PP vaginal meshes.
In a broad sense, all materials with a biological origin can be called biomass, which includes those materials that have been obtained from the direct conversion of solar energy, such as plants and ...crops, their residues, the materials obtained from their industrial transformation as well as subproducts and residues and the organic fraction of the municipal solid wastes ...
This review focuses on the role of small extracellular vesicles in the pathophysiological mechanisms of retinal degenerative diseases. Many of these mechanisms are related to or modulated by the ...oxidative burden of retinal cells. It has been recently demonstrated that cellular communication in the retina involves extracellular vesicles and that their rate of release and cargo features might be affected by the cellular environment, and in some instances, they might also be mediated by autophagy. The fate of these vesicles is diverse: they could end up in circulation being used as markers, or target neighbor cells modulating gene and protein expression, or eventually, in angiogenesis. Neovascularization in the retina promotes vision loss in diseases such as diabetic retinopathy and age-related macular degeneration. The importance of micro RNAs, either as small extracellular vesicles' cargo or free circulating, in the regulation of retinal angiogenesis is also discussed.
arranges into two morphologically distinct biofilms depending on its nutritional status, i.e., coordinately spreading colonies in the presence of nutrients and spore-filled fruiting bodies in the ...absence of nutrients. A secreted polysaccharide, referred to as exopolysaccharide (EPS), is a structural component of both biofilms and is also important for type IV pilus-dependent motility and fruiting body formation. Here, we characterize the biosynthetic machinery responsible for EPS biosynthesis using bioinformatics, genetics, heterologous expression, and biochemical experiments. We show that this machinery constitutes a Wzx/Wzy-dependent pathway dedicated to EPS biosynthesis. Our data support that EpsZ (MXAN_7415) is the polyisoprenyl-phosphate hexose-1-phosphate transferase responsible for the initiation of the repeat unit synthesis. Heterologous expression experiments support that EpsZ has galactose-1-P transferase activity. Moreover, MXAN_7416, renamed Wzx
, and MXAN_7442, renamed Wzy
, are the Wzx flippase and Wzy polymerase responsible for translocation and polymerization of the EPS repeat unit, respectively. In this pathway, EpsV (MXAN_7421) also is the polysaccharide copolymerase and EpsY (MXAN_7417) the outer membrane polysaccharide export (OPX) protein. Mutants with single in-frame deletions in the five corresponding genes had defects in type IV pilus-dependent motility and a conditional defect in fruiting body formation. Furthermore, all five mutants were deficient in type IV pilus formation, and genetic analyses suggest that EPS and/or the EPS biosynthetic machinery stimulates type IV pilus extension. Additionally, we identify a polysaccharide biosynthesis gene cluster, which together with an orphan gene encoding an OPX protein make up a complete Wzx/Wzy-dependent pathway for synthesis of an unknown polysaccharide.
The secreted polysaccharide referred to as exopolysaccharide (EPS) has important functions in the social life cycle of
; however, little is known about how EPS is synthesized. Here, we characterized the EPS biosynthetic machinery and showed that it makes up a Wzx/Wzy-dependent pathway for polysaccharide biosynthesis. Mutants lacking a component of this pathway had reduced type IV pilus-dependent motility and a conditional defect in development. These analyses also suggest that EPS and/or the EPS biosynthetic machinery is important for type IV pilus formation.
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Cardiovascular diseases constitute a number of conditions which are the leading cause of death globally. To combat these diseases and improve the quality and duration of life, several ...cardiac implants have been developed, including stents, vascular grafts and valvular prostheses. The implantation of these vascular prosthesis has associated risks such as infection or blood clot formation. In order to overcome these limitations medicated vascular prosthesis have been previously used. The present paper describes a 3D printing method to develop medicated vascular prosthesis using fused deposition modelling (FDM) technology. For this purpose, rifampicin (RIF) was selected as a model molecule as it can be used to prevent vascular graft prosthesis infection. Thermoplastic polyurethane (TPU) and RIF were combined using hot melt extrusion (HME) to obtain filaments containing RIF concentrations ranging between 0 and 1% (w/w). These materials are capable of providing RIF release for periods ranging between 30 and 80 days. Moreover, TPU-based materials containing RIF were capable of inhibiting the growth of Staphylococcus aureus. This behaviour was observed even for TPU-based materials containing RIF concentrations of 0.1% (w/w). TPU containing 1% (w/w) of RIF showed antimicrobial properties even after 30 days of RIF release. Alternatively, these methods were used to prepare dipyridamole containing TPU filaments. Finally, using a dual extrusion 3D printer vascular grafts containing both drugs were prepared.
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