Effect of two types of biosurfactants on phenanthrene availability to the bacterial bioreporter Burkholderia sartisoli strain RP037

Biosurfactants are tensio-active agents that have often been proposed as a means to enhance the aqueous solubility of hydrophobic organic contaminants, such as polycyclic aromatic hydrocarbons (PAHs). Biosurfactant-producing bacteria such as those belonging to the genus Pseudomonas might therefore enhance PAH availability to PAH-degrading bacteria. We tested the effects of two types of biosurfactants produced by Pseudomonas sp., cyclic lipopeptides and rhamnolipids, on phenanthrene bioavailability. Bioavailability was judged from growth rates on phenanthrene and from specific induction of a phenanthrene-responsive GFP-reporter in Burkholderia sartisoli strain RP037. Co-culturing of strain RP037 with the lipopeptide-producing bacterium Pseudomonas putida strain PCL1445 enhanced GFP expression compared to a single culture, but this effect was not significantly different when strain RP037 was co-cultivated with a non-lipopeptide-producing mutant of P. putida. The addition of partially purified supernatant extracts from the P. putida lipopeptide producer equally did not unequivocally enhance phenanthrene bioavailability to strain RP037 compared to controls. In contrast, a 0.1% rhamnolipid solution strongly augmented RP037 growth rates on phenanthrene and led to a significantly larger proportion of cells in culture with high GFP expression. Our data therefore suggest that biosurfactant effects may be strongly dependent on the strain and type of biosurfactan

Cooperation controls partners spatial intermixing in a synthetic bacterial consortium

Microbes are essential actors in all environments (nutrients cycling, pollutants bioremediation, and more), and their functions depend on multispecies communities that possess intricate interspecies interactions and high degree of spatial organization. Due to the complexity of natural microbial systems, mechanistic understanding of community spatial assembly and activity remains fragmentary. Here, we used a synthetic ecology approach to ask how simple factors such as carbon utilization control spatial patterns of a two-partner bacterial consortium grown on surfaces. The experimental model consortium consisted of two mutant strains of Pseudomonas putida that cooperate to degrade and utilize toluene. Strains tagging with different autofluorescent proteins allowed for microscopic visualization and pattern quantification by image analysis. We showed that trophic cooperation (toluene degradation) led to convergence of partner abundance (1:1) regardless of the initial ratio, and to strong strain intermixing at the microscale (10-100 m). In contrast, competition for a carbon source (benzoate) degraded independently by both strains resulted in distinct segregation patterns. The consortium productivity (cell growth) on toluene was affected by the initial partner ratio on surfaces but not in liquid cultures. This study pinpoints general principles of microbial community spatial organization with potential applications for natural and engineered microbial systems

Cell distribution and habitat fragmentation affecting the spread of plasmids in soil bacterial populations

Aims and Objectives. Transfer of conjugative plasmids among soil bacteria is an important evolutionary driver for fostering adaptation to environmental stresses such as antibiotic or xenobiotic contamination. However, plasmids impose a metabolic burden on host cells and the reason for their persistence over evolutionary times remains unclear. Here, we hypothesize that soil environments favor plasmid maintenance due to habitat fragmentation and nutrient spatial heterogeneity that promote plasmid transfer rates and reduce competition in microniches. We aim to test that hypothesis and disentangle the contributions of transmission and selection. Materials and Methods. The soil bacterium Pseudomonas putida was used as donor and recipient of the conjugative plasmid pIPO2tet (conferring resistance to tetracycline). A tagging system with fluorescent proteins allowed us to visually discriminate recipients, donors and transconjugants using microscope image analysis and plating on selective media. Bacteria were grown in controlled systems of varying complexity, from agar surfaces to micromodels, and in presence or absence of tetracycline as a selective agent. Results. Experiments on homogeneous agar surfaces showed that transfer rate and the final size of the plasmid-carrying population increased with cell density, while competition (as measured by selection coefficient) tended to decrease. The presence of antibiotics at sub-inhibitory concentrations also affected plasmid transfer rate and selection. To address the role of spatial isolation of microhabitats, we have designed micromodels that allow us to observe local variations in the spread of bacterial plasmids, with results still pending. Conclusions. Local (microscale) conditions such as cell density and spatial confinement can enhance or suppress plasmid transfer among bacterial populations, as well as affect the selective effects of carrying a plasmid. The study offers new insights linking soil microhabitats to ecological and evolutionary adaptations of soil bacteria

Bacterial flagellar motility on hydrated rough surfaces controlled by aqueous film thickness and connectedness

Recent studies have shown that rates of bacterial dispersion in soils are controlled by hydration conditions that define size and connectivity of the retained aqueous phase. Despite the ecological implications of such constraints, microscale observations of this phenomenon remain scarce. Here, we quantified aqueous film characteristics and bacterial flagellated motility in response to systematic variations in microhydrological conditions on porous ceramic surfaces that mimic unsaturated soils. We directly measured aqueous film thickness and documented its microscale heterogeneity. Flagellar motility was controlled by surface hydration conditions, as cell velocity decreased and dispersion practically ceased at water potentials exceeding –2 kPa (resulting in thinner and disconnected liquid films). The fragmentation of aquatic habitats was delineated indirectly through bacterial dispersal distances within connected aqueous clusters. We documented bacterial dispersal radii ranging from 100 to 10 μm as the water potential varied from 0 to –7 kPa, respectively. The observed decrease of flagellated velocity and dispersal ranges at lower matric potentials were in good agreement with mechanistic model predictions. Hydration-restricted habitats thus play significant role in bacterial motility and dispersal, which has potentially important impact on soil microbial ecology and diversity.ISSN:2045-232

Biophysical processes supporting the diversity of microbial life in soil

Soil, the living terrestrial skin of the Earth, plays a central role in supporting life and is home to an unimaginable diversity of microorganisms. This review explores key drivers for microbial life in soils under different climates and land-use practices at scales ranging from soil pores to landscapes. We delineate special features of soil as a microbial habitat (focusing on bacteria) and the consequences for microbial communities. This review covers recent modeling advances that link soil physical processes with microbial life (termed biophysical processes). Readers are introduced to concepts governing water organization in soil pores and associated transport properties and microbial dispersion ranges often determined by the spatial organization of a highly dynamic soil aqueous phase. The narrow hydrological windows of wetting and aqueous phase connectedness are crucial for resource distribution and longer range transport of microorganisms. Feedbacks between microbial activity and their immediate environment are responsible for emergence and stabilization of soil structure—the scaffolding for soil ecological functioning. We synthesize insights from historical and contemporary studies to provide an outlook for the challenges and opportunities for developing a quantitative ecological framework to delineate and predict the microbial component of soil functioning.ISSN:0168-6445ISSN:1574-697

Fragmented aqueous habitats affect bacterial plasmid transfer in porous environments

Horizontal transfer of plasmids among bacterial populations is critical to genetic exchange and adaptation to environmental stresses. Studies have examined how bacterial cell density, donor-recipient ratio and nutrient availability affect plasmid transfer rates in homogeneous cultures. The insights are useful, however, they often neglect the ubiquitous physical heterogeneity of natural microbial habitats such as soil environments. Our hypothesis is that unsaturated and dynamic water conditions prevailing in most soils lead to fragmentation of the aquatic bacterial habitat, which could enhance plasmid spread through local (microscale) higher cell densities with reduced competition. However, the biophysical processes at play eschew quantitative observations at the cell level owing to soil complexity and opacity. We report and new microfluidic chip comprised of connected microhabitats that permit external control of aqueous fragmentation and observations at the single-cell level. These elements were used to quantify conjugation events in the microchip at various scales as function of fragmentation dynamics. We used the soil bacterium Pseudomonas putida as donor and recipient of a conjugative plasmid carrying a tetracycline resistance gene, while a tagging system with autofluorescent proteins allowed us to distinguish donors and transconjugants with epifluorescence microscopy. We successfully generated various patterns of aqueous fragmentation in the microchips, hence resulting in disconnected aqueous habitats for the bacteria. Plasmid transfer rate in individual microhabitats was linked to local cell densities as well as aqueous and gas phase distribution. This study highlights the importance of microhydrological processes that affect the ecology and evolution of bacteria in natural soil habitats

Symplasmata are a clonal, conditional, and reversible type of bacterial multicellularity.

Microorganisms are capable of remarkable social behaviours, such as forming transient multicellular assemblages with properties and adaptive abilities exceeding those of individual cells. Here, we report on the formation and structure of genets known as symplasmata produced by Pantoea eucalypti bacteria. Each symplasmatum develops clonally and stochastically from a single bacterium into a membrane-delimited, capsule-embedded cluster of progeny cells and with a frequency that depends on temperature, pH, and nutrient availability. Transposon mutagenesis identified several gene products required for symplasmata formation, including master regulator LrhA, replication inhibitor CspD, polysaccharide transporter RfbX3, and autoinducer synthase PhzI. We also show that bacteria inside symplasmata are shaped irregularly with punctuated cell-to-cell contacts, metabolically responsive to environmental stimuli, dispersal-ready, and transcriptionally reprogrammed to anticipate multiple alternative futures in terms of carbon source availability. The structured and conditionable nature of symplasmata offers exciting prospects towards a mechanistic understanding of multicellular behaviours and their ecological significance

Cooperation in carbon source degradation shapes spatial self-organization of microbial consortia on hydrated surfaces

Mounting evidence suggests that natural microbial communities exhibit a high level of spatial organization at the micrometric scale that facilitate ecological interactions and support biogeochemical cycles. Microbial patterns are difficult to study definitively in natural environments due to complex biodiversity, observability and variable physicochemical factors. Here, we examine how trophic dependencies give rise to self-organized spatial patterns of a well-defined bacterial consortium grown on hydrated surfaces. The model consortium consisted of two Pseudomonas putida mutant strains that can fully degrade the aromatic hydrocarbon toluene. We demonstrated that obligate cooperation in toluene degradation (cooperative mutualism) favored convergence of 1:1 partner ratio and strong intermixing at the microscale (10–100 μm). In contrast, competition for benzoate, a compound degraded independently by both strains, led to distinct segregation patterns. Emergence of a persistent spatial pattern has been predicted for surface attached microbial activity in liquid films that mediate diffusive exchanges while permitting limited cell movement (colony expansion). This study of a simple microbial consortium offers mechanistic glimpses into the rules governing the assembly and functioning of complex sessile communities, and points to general principles of spatial organization with potential applications for natural and engineered microbial systems.ISSN:2045-232