Social Interactions and Biofilm Formation in Bacillus subtilis

Međustanična je komunikacija (engl. quorum sensing, QS) oblik kooperativnog socijalnog ponašanja bakterija što se oslanja na prepoznavanje izvanstaničnih signalnih molekula. Signalna se molekula veže na receptor i inducira promjenu transkripcije gena, odgovornih za stvaranje biofilma, proizvodnju izvanstaničnih enzima i druge kooperativne značajke populacije. Svrha je ovoga rada bila dati pregled objavljenih znanstvenih radova koji se bave kooperativnim socijalnim ponašanjem bakterije Bacillus subtilis, a naročito doprinosom komunikacijskog sustava ComQXPA. Sustav QS obuhvaća četiri komponente koje su u međusobnoj interakciji: izoprenil transferazu ComQ što procesira i modificira signal, peptid ComX koji ima ulogu signala, receptor ComP i transkripcijski regulator ComA. Fosforilirani ComA kontrolira transkripciju brojnih gena, uključujući i one odgovorne za proizvodnju surfaktina te izvanstaničnog matriksa, važnog za nastajanje biofilma. Sustav ComQXPA QS ima visok stupanj genetičkog polimorfizma, što je vidljivo iz činjenice da se sojevi Bacillus subtilis mogu podijeliti u četiri skupine. Sojevi jedne skupine (ferotipa) mogu razmjenjivati signale i informacije, dok to nije moguće između različitih ferotipova. Nedavno smo pokazali da je ovaj fenomen djelomično posljedica ekološke raznolikosti sojeva, ali su moguće i alternativne hipoteze, koje daju prednost socijalnoj evoluciji. Između ostalog, sustav ComQXPA kontrolira i proizvodnju izvanstaničnog matriksa, koji se sastoji od polisaharida, proteina i nukleinskih kiselina. U ovom je radu dan pregled trenutnih spoznaja o regulaciji, strukturi, kemijskom sastavu i funkciji izvanstaničnog matriksa. Usprkos mnogim važnim nedavnim otkrićima u području regulacije formiranja biofilma B. subtilis, molekularne interakcije među komponentama matriksa i njihov utjecaj na QS i stabilnost biofilma nisu još dobro poznati, pa se u ovom radu razmatraju i moguća rješenja ovih zanimljivih pitanja.Quorum sensing (QS) is a form of cooperative social behaviour which relies on extracellular signalling molecules that elicit the QS response across many cells and controls the development of many cooperative traits including biofilm formation. The main aim of this work is to review the published work on cooperative social behaviour of Bacillus subtilis and especially its QS system ComQXPA. This QS system involves four interacting components: the signal-processing enzyme ComQ, the ComX signal, the ComP receptor and the ComA transcriptional regulator. Phosphorylated ComA controls the transcription of many genes including those responsible for the production of surfactin and extracellular matrix, essential for biofilm formation. The ComQXPA QS shows a high degree of genetic polymorphism, which manifests itself in the separation of Bacillus subtilis strains into four different communication groups (pherotypes). The information exchange is possible between members of the same pherotype but not across pherotypes. We have recently suggested that this phenomenon is at least in part driven by the ecological divergence of strains, but may also be induced by frequency-dependent selection. The ComQXPA QS system controls the production of extracellular matrix (ECM) components: polysaccharides, proteins and nucleic acids. We will address the present understanding of the ECM structure-function relationships in B. subtilis biofilms and review published results on regulation, composition and distribution of ECM components. Despite many important recent discoveries on regulation of B. subtilis biofilm development, we know little about the molecular interactions in the ECM and the role they play in the QS and stability of the biofilm. Future research needs to address these questions better

Self-assembly of polysaccharides gives rise to distinct mechanical signatures in marine gels

Marine-gel biopolymers were recently visualized at the molecular level using atomic force microscopy (AFM) to reveal fine fibril-forming networks with low to high degrees of cross-linking. In this work, we use force spectroscopy to quantify the intra- and intermolecular forces within the marine-gel network. Combining force measurements, AFM imaging, and the known chemical composition of marine gels allows us to identify the microscopic origins of distinct mechanical responses. At the single-fibril level, we uncover force-extension curves that resemble those of individual polysaccharide fibrils. They exhibit entropic elasticity followed by extensions associated with chair-to-boat transitions specific to the type of polysaccharide at high forces. Surprisingly, a low degree of cross-linking leads to sawtooth patterns that we attribute to the unraveling of polysaccharide entanglements. At a high degree of cross-linking, we observe force plateaus that arise from unzipping, as well as unwinding, of helical bundles. Finally, the complex 3D network structure gives rise to force staircases of increasing height that correspond to the hierarchical peeling of fibrils away from the junction zones. In addition, we show that these diverse mechanical responses also arise in reconstituted polysaccharide gels, which highlights their dominant role in the mechanical architecture of marine gels

Control of the aqueous solubility of cellulose by hydroxyl group substitution and its effect on processing

Native cellulose is insoluble in water, despite the high number of hydrogen bonding sites per chain, as molecules preferably hydrogen bond to each other, preventing its use in industrial applications. The modification of cellulose has received considerable recent attention, motivated by the move away from conventional petroleum-based, water-soluble polymers, however, a systematic analysis of the effects of modification is rare. Herein a detailed study of hydroxypropyl (HP)- and (2-hydroxypropyl) trimethylammonium chloride-modified cellulose, with degrees of substitution (DS) determined by NMR, establishes modification-property relationships. TEM, small-angle X-ray scattering and rheology demonstrated that increasing DS gradually changes the aqueous solubility, resulting in the formation of different morphologies, including micron-sized aggregates, needle-like cellulose nanoparticles (CNPs) and solvated molecules. It was found that aqueous dispersions with DSHP of 50 %, assigned to a ‘sweet spot’ in cellulose modification, are suitable for the fiber formation. It is shown that this state of the material can be easily detected by rheo-optical methods based on birefringence. Using structural analysis, molecular dynamic simulation and fiber-spinning results, it is proposed that co-existing CNPs and cellulose molecules, interacting via H-bonding, form a network which orients under shear, acting as a precursor for the fiber formation from aqueous solutions

Biofilm formation by <em>Bacillus subtilis</em>:new insights into regulatory strategies and assembly mechanisms

Biofilm formation is a social behaviour that generates favourable conditions for sustained survival in the natural environment. For the Gram-positive bacterium Bacillus subtilis the process involves the differentiation of cell fate within an isogenic population and the production of communal goods that form the biofilm matrix. Here we review recent progress in understanding the regulatory pathways that control biofilm formation and highlight developments in understanding the composition, function and structure of the biofilm matrix

eDNA Provides a Scaffold for Autoaggregation of B. subtilis in Bacterioplankton Suspension

The self-binding of bacterial cells, or autoaggregation, is, together with surface colonization, one of the first steps in the formation of a mature biofilm. In this work, the autoaggregation of B. subtilis in dilute bacterial suspensions was studied. The dynamics of cell lysis, eDNA release, and bacterial autoaggregate assembly were determined and related to the spatial autocorrelation of bacterial cells in dilute planktonic bacterial suspensions. The non-random distribution of cells was associated with an eDNA network, which stabilized the initial bacterial cell-cell aggregates. Upon the addition of DNase I, the aggregates were dispersed. The release of eDNA during cell lysis allows for the entrapment of bacterial drifters at a radius several times the size of the dying bacteria. The size of bacterial aggregates increased from 2 to about 100 &mu;m in diameter in dilute bacterial suspensions. The results suggest that B. subtilis cells form previously unnoticed continuum of autoaggregate structures during planktonic growth

Multiscale spatial segregation analysis in digital images of biofilms

Quantifying the degree of spatial segregation of two bacterial strains in mixed biofilms is an important topic in microbiology. Spatial segregation is dependent on spatial scale as two strains may appear to be well mixed if observed from a distance, but a closer look can reveal strong separation. Typically, this information is encoded in a digital image that represents the binary system, e.g., a microscopy image of a two species biofilm. To decode spatial segregation information, we have developed quantitative measures for evaluating the degree of the spatial scale-dependent segregation of two bacterial strains in a digital image. The constructed algorithm is based on the new segregation measures and overcomes drawbacks of existing approaches for biofilm segregation analysis. The new approach is implemented in a freely available software and was successfully applied to biofilms of two strains and bacterial suspensions for detection of the different spatial scale-dependent segregation levels

eDNA Provides a Scaffold for Autoaggregation of <i>B. subtilis</i> in Bacterioplankton Suspension

The self-binding of bacterial cells, or autoaggregation, is, together with surface colonization, one of the first steps in the formation of a mature biofilm. In this work, the autoaggregation of B. subtilis in dilute bacterial suspensions was studied. The dynamics of cell lysis, eDNA release, and bacterial autoaggregate assembly were determined and related to the spatial autocorrelation of bacterial cells in dilute planktonic bacterial suspensions. The non-random distribution of cells was associated with an eDNA network, which stabilized the initial bacterial cell-cell aggregates. Upon the addition of DNase I, the aggregates were dispersed. The release of eDNA during cell lysis allows for the entrapment of bacterial drifters at a radius several times the size of the dying bacteria. The size of bacterial aggregates increased from 2 to about 100 μm in diameter in dilute bacterial suspensions. The results suggest that B. subtilis cells form previously unnoticed continuum of autoaggregate structures during planktonic growth

Physicochemical data on aqueous polymeric systems of methyl cellulose and lambda- and kappa-carrageenan: SAXS, rheological, densitometry, and sound velocity measurements

General as well as more specific physicochemical data obtained by studying the structure and various dynamical properties of aqueous polymer systems of methyl cellulose, λâcarrageenan, and κâcarrageenan are presented in graphical and numeric tabular form. The data provide basic polymer characterization info as also a specific structural and dynamical info for aqueous solutions of three industrially very important polymers (food additives) that are available commercially. The commercial availability has much bigger impact to applications, research and connected advances, when the basic substances are well characterized â a feature that is still missing for many commercially available polymers unfortunately. Keywords: Food-grade polymers, Basic polymer characterization, Methyl cellulose, kappa-Carrageenan, lambda-Carrageena

Thermal and Rheological Properties of Gluten-Free, Starch-Based Model Systems Modified by Hydrocolloids

Obtaining good-quality gluten-free products represents a technological challenge; thus, it is important to understand how and why the addition of hydrocolloids influences the properties of starch-based products. To obtain insight into the physicochemical changes imparted by hydrocolloids on gluten-free dough, we prepared several suspensions with different corn starch/potato starch/hydroxpropyl methyl cellulose/xanthan gum/water ratios. Properties of the prepared samples were determined by differential scanning calorimetry and rheometry. Samples with different corn/potato starch ratios exhibited different thermal properties. Xanthan gum and HPMC (hydroxypropyl methyl cellulose) exhibited a strong influence on the rheological properties of the mixtures since they increased the viscosity and elasticity. HPMC and xanthan gum increased the temperature of starch gelatinization, as well as they increased the viscoelasticity of the starch model system. Although the two hydrocolloids affected the properties of starch mixtures in the same direction, the magnitude of their effects was different. Our results indicate that water availability, which plays a crucial role in the starch gelatinization process, could be modified by adding hydrocolloids such as, hydroxypropyl methyl cellulose and xanthan gum. By adding comparatively small amounts of the studied hydrocolloids to starch, one can achieve similar thermo-mechanical effects by the addition of gluten. Understanding these effects of hydrocolloids could contribute to the development of better quality gluten-free bread with optimized ingredient content