Serotype switching in Pseudomonas aeruginosa ST111 enhances adhesion and virulence.

Evolution of the highly successful and multidrug resistant clone ST111 in Pseudomonas aeruginosa involves serotype switching from O-antigen O4 to O12. How expression of a different O-antigen serotype alters pathogen physiology to enable global dissemination of this high-risk clone-type is not understood. Here, we engineered isogenic laboratory and clinical P.

Biosynthesis enhancement of tropodithietic acid (TDA) antibacterial compound through biofilm formation by marine bacteria on micro-structured polymer surfaces.

Although aquaculture is a major player in current and future food production, the routine use of antibiotics provides ample ground for development of antibiotic resistance. An alternative route to disease control is the use of probiotic bacteria such as the marine bacteria which produces tropodithietic acid (TDA) that inhibit pathogens without affecting the fish. Improving conditions for the formation of biofilm and TDA-synthesis is a promising avenue for biocontrol in aquaculture.

Microbial biofilms in biorefinery - Towards a sustainable production of low-value bulk chemicals and fuels.

Harnessing the potential of biocatalytic conversion of renewable biomass into value-added products is still hampered by unfavorable process economics. This has promoted the use of biofilms as an alternative to overcome the limitations of traditional planktonic systems. In this paper, the benefits and challenges of biofilm fermentations are reviewed with a focus on the production of low-value bulk chemicals and fuels from waste biomass.

Bacterial Cell Cultures in a Lab-on-a-Disc: A Simple and Versatile Tool for Quantification of Antibiotic Treatment Efficacy.

Pathogenic bacterial biofilms can be life-threatening, greatly decrease patient's quality of life, and are a substantial burden on the healthcare system. Current methods for evaluation of antibacterial treatments in clinics and systems used in drug development and screening either do not facilitate biofilm formation or are cumbersome to operate, need large reagent volumes, and are costly, limiting their usability. To address these issues, this work presents the development of a robust cell culture platform compatible with confocal microscopy.

Loss of AA13 LPMOs impairs degradation of resistant starch and reduces the growth of .

Background: Lytic polysaccharide monooxygenases (LPMOs) are often studied in simple models involving activity measurements of a single LPMO or a blend thereof with hydrolytic enzymes towards an insoluble substrate. However, the contribution of LPMOs to polysaccharide breakdown in complex cocktails of hydrolytic and oxidative enzymes, similar to fungal secretomes, remains elusive. Typically, two starch-specific AA13 LPMOs are encoded by mainly Ascomycota genomes.

Synthesis of carbon quantum dot-poly lactic-co-glycolic acid hybrid nanoparticles for chemo-photothermal therapy against bacterial biofilms.

Bacterial biofilm represents a protected mode of bacterial growth that significantly enhances the resistance to antibiotics. Poly lactic-co-glycolic acid (PLGA)-based nanoparticle delivery systems have been intensively investigated to combat the bacterial biofilms-associated infections. However, some drawbacks associated with current PLGA-based nanoformulations (e.

Ultrasmall TPGS-PLGA Hybrid Nanoparticles for Site-Specific Delivery of Antibiotics into Biofilms in Lungs.

Inhaled antibiotic treatment of cystic fibrosis-related bacterial biofilm infections is challenging because of the pathological environment of the lungs. Here, we present an "environment-adaptive" nanoparticle composed of a solid poly lactic--glycolic acid (PLGA) core and a mucus-inert, enzymatically cleavable shell of d-α-tocopheryl polyethylene glycol 1000 succinate (TPGS) for the site-specific delivery of antibiotics to bacterial biofilms via aerosol administration. The hybrid nanoparticles with ultrasmall size were self-assembled via a nanoprecipitation process by using a facile microfluidic method.

Utilizing nanoparticles for improving anti-biofilm effects of azithromycin: A head-to-head comparison of modified hyaluronic acid nanogels and coated poly (lactic-co-glycolic acid) nanoparticles.

Hypothesis: The widespread resistance of bacteria to traditional antibiotic treatments has expedited the search for novel therapies against these pathogens. The hypothesis of this work is that two distinctively different polymeric delivery systems, specifically D-α-tocopherol polyethylene glycol 1000 succinate (TPGS)-poly(lactic-co-glycolic acid) (PLGA) nanoparticles and octenyl succinic anhydride-modified low molecular weight hyaluronic acid (OSA-HA) nanogels may be used to substantially improve the properties of azithromycin, allowing its use for effective treatment of Pseudomonas aeruginosa biofilm infections.

Experiments: Azithromycin was encapsulated in both delivery systems and the physicochemical properties of the loaded delivery systems, including size, surface charge and drug loading were evaluated.

Differential bacterial capture and transport preferences facilitate co-growth on dietary xylan in the human gut.

Metabolism of dietary glycans is pivotal in shaping the human gut microbiota. However, the mechanisms that promote competition for glycans among gut commensals remain unclear. Roseburia intestinalis, an abundant butyrate-producing Firmicute, is a key degrader of the major dietary fibre xylan.

Application of RNA-seq and Bioimaging Methods to Study Microbe-Microbe Interactions and Their Effects on Biofilm Formation and Gene Expression.

Complex interactions between pathogenic bacteria, the microbiota, and the host can modify pathogen physiology and behavior. We describe two different experimental approaches to study microbe-microbe interactions in in vitro systems containing surface-associated microbial populations. One method is the application of RNA sequencing (RNA-seq) to determine the transcriptional changes in pathogenic bacteria in response to microbial interspecies interactions.

Critical review on biofilm methods.

Biofilms are widespread in nature and constitute an important strategy implemented by microorganisms to survive in sometimes harsh environmental conditions. They can be beneficial or have a negative impact particularly when formed in industrial settings or on medical devices. As such, research into the formation and elimination of biofilms is important for many disciplines.

Biofilm as a production platform for heterologous production of rhamnolipids by the non-pathogenic strain Pseudomonas putida KT2440.

Background: Although a transition toward sustainable production of chemicals is needed, the physiochemical properties of certain biochemicals such as biosurfactants make them challenging to produce in conventional bioreactor systems. Alternative production platforms such as surface-attached biofilm populations could potentially overcome these challenges. Rhamnolipids are a group of biosurfactants highly relevant for industrial applications.

Utilization and control of ecological interactions in polymicrobial infections and community-based microbial cell factories.

Microbial activities are most often shaped by interactions between co-existing microbes within mixed-species communities. Dissection of the molecular mechanisms of species interactions within communities is a central issue in microbial ecology, and our ability to engineer and control microbial communities depends, to a large extent, on our knowledge of these interactions. This review highlights the recent advances regarding molecular characterization of microbe-microbe interactions that modulate community structure, activity, and stability, and aims to illustrate how these findings have helped us reach an engineering-level understanding of microbial communities in relation to both human health and industrial biotechnology.

Methods for studying biofilm formation: flow cells and confocal laser scanning microscopy.

In this chapter methods for growing and analyzing biofilms under hydrodynamic conditions in flow cells are described. Use of flow cells allows for direct microscopic investigation of biofilm formation. The flow in these chambers is essentially laminar, which means that the biofilms can be grown under highly controlled conditions, and that perturbations such as addition of antibiotics or change of the growth medium can be done efficiently at a defined time point.

Methods for dynamic investigations of surface-attached in vitro bacterial and fungal biofilms.

Three dynamic models for the investigation of in vitro biofilm formation are described in this chapter. In the 6-well plate assay presented here, the placing of the plate on a rotating platform provides shear, thereby making the system dynamic with respect to the static microtiter assay.The second reported model, especially suitable for harvesting high amounts of cells for transcriptomic or proteomic investigations, is based on numerous glass beads placed in a flask incubated with shaking on a rotating platform, thus increasing the surface area for biofilm formation.

Secreted single-stranded DNA is involved in the initial phase of biofilm formation by Neisseria gonorrhoeae.

Neisseria gonorrhoeae is an obligate human pathogen that colonizes the genital tract and causes gonorrhoea. Neisseria gonorrhoeae can form biofilms during natural cervical infections, on glass and in continuous flow-chamber systems. These biofilms contain large amounts of extracellular DNA, which plays an important role in biofilm formation.

Growing and analyzing biofilms in flow chambers.

This unit describes the setup of flow chamber systems for the study of microbial biofilms, and methods for the analysis of structural biofilm formation. Use of flow chambers allows direct microscopic investigation of biofilm formation. The biofilms in flow chambers develop under hydrodynamic conditions, and the environment can be carefully controlled and easily changed.

Quantification of specific E. coli in gut mucosa from Crohn's disease patients.

We here present a method based on qRT-PCR to quantify E. coli LF82 in intestinal human samples. Two different primer-probe sets were designed to detect LF82, and a third to target total E.

Pseudomonas aeruginosa and Saccharomyces cerevisiae biofilm in flow cells.

Many microbial cells have the ability to form sessile microbial communities defined as biofilms that have altered physiological and pathological properties compared to free living microorganisms. Biofilms in nature are often difficult to investigate and reside under poorly defined conditions(1). Using a transparent substratum it is possible to device a system where simple biofilms can be examined in a non-destructive way in real-time: here we demonstrate the assembly and operation of a flow cell model system, for in vitro 3D studies of microbial biofilms generating high reproducibility under well-defined conditions(2,3).

An individual-based approach to explain plasmid invasion in bacterial populations.

We present an individual-based experimental framework to identify and estimate the main parameters governing bacterial conjugation at the individual cell scale. From this analysis, we have established that transient periods of unregulated plasmid transfer within newly formed transconjugant cells, together with contact mechanics arising from cellular growth and division, are the two main processes determining the emergent inability of the pWW0 TOL plasmid to fully invade spatially structured Pseudomonas putida populations. We have also shown that pWW0 conjugation occurs mainly at advanced stages of the growth cycle and that nongrowing cells, even when exposed to high nutrient concentrations, do not display conjugal activity.

Advanced microscopy of microbial cells.

Growing awareness of heterogeneity in cells of microbial populations has emphasized the importance of advanced microscopy for visualization and understanding of the molecular mechanisms underlying cell-to-cell variation. In this review, we highlight some of the recent advances in confocal microscopy, super-resolution optical microscopy (STED, SIM, PALM) as well as atomic force microscopy and Raman spectroscopy. Using examples of bistability in microbial populations as well as biofilm development and differentiation in bacterial and yeast consortia, we demonstrate the importance of microscopy for visualization of variation between cells in phenotypic traits such as gene expression.

Microfluidic dissolved oxygen gradient generator biochip as a useful tool in bacterial biofilm studies.

A microfluidic chip for generation of gradients of dissolved oxygen was designed, fabricated and tested. The novel way of active oxygen depletion through a gas permeable membrane was applied. Numerical simulations for generation of O(2) gradients were correlated with measured oxygen concentrations.

Evaluation of enoyl-acyl carrier protein reductase inhibitors as Pseudomonas aeruginosa quorum-quenching reagents.

Pseudomonas aeruginosa is an opportunistic pathogen which is responsible for a wide range of infections. Production of virulence factors and biofilm formation by P. aeruginosa are partly regulated by cell-to-cell communication quorum-sensing systems.

An in vitro model of bacterial infections in wounds and other soft tissues.

There is growing evidence that bacteria play a crucial role in the persistence of chronic wounds. These bacteria are most probably present in polymer-embedded aggregates that represent the biofilm mode of growth. Much work has been carried out to study the development of biofilms in vitro, in particular in attachment to solid surfaces.

Insight into the microbial multicellular lifestyle via flow-cell technology and confocal microscopy.

Biofilms are agglomerates of microorganisms surrounded by a self-produced extracellular matrix. During the last 10 years, there has been an increasing recognition of biofilms as a highly significant topic in microbiology with relevance for a variety of areas in our society including the environment, industry, and human health. Accordingly a number of biofilm model systems, molecular tools, microscopic techniques, and image analysis programs have been employed for the study of biofilms under controlled and reproducible conditions.