Establishment of a transparent soil system to study Bacillus subtilis chemical ecology

Abstract

Bacterial secondary metabolites are structurally diverse molecules that drive microbial interaction by altering growth, cell differentiation, and signaling. Bacillus subtilis, a Gram-positive soil-dwelling bacterium, produces a wealth of secondary metabolites, among them, lipopeptides have been vastly studied by their antimicrobial, antitumor, and surfactant activities. However, the natural functions of secondary metabolites in the lifestyles of the producing organism remain less explored under natural conditions, i.e. in soil. Here, we describe a hydrogel-based transparent soil system to investigate B. subtilis chemical ecology under controllable soil-like conditions. The transparent soil matrix allows the growth of B. subtilis and other isolates gnotobiotically and under nutrient-controlled conditions. Additionally, we show that transparent soil allows the detection of lipopeptides production and dynamics by HPLC-MS and MALDI-MS imaging, along with fluorescence imaging of 3-dimensional bacterial assemblages. We anticipate that this affordable and highly controllable system will promote bacterial chemical ecology research and help to elucidate microbial interactions driven by secondary metabolites.

Construction and characterisation of a structured, tuneable, and transparent 3D culture platform for soil bacteria

“…Nafion was selected due to its commercial availability, documented use for bioimaging, stability during autoclave sterilisation, low refractive index, potential for particle size refinement, and ion exchange capacity for nutrient imbibement [15,34]. Using Nafion surmounted the optical and material complexities and costs of previously published artificial soil scaffolds such as those using cryolite [17,35], FEP [36], silica [37,38], and hydrogels [32,39]. The material properties of Nafion permitted particle size refinement from centimetre to micron-scale particles.…”

Construction and Characterisation of a Structured, Tuneable, and Transparent 3D Culture Platform for Soil Bacteria

“…Nafion™ was selected due to its commercial availability, documented use for bioimaging, stability during autoclave sterilisation, low refractive index, potential for particle size refinement, and ion exchange capacity for nutrient imbibement (18,34). Using Nafion™ surmounted the optical and material complexities and costs of previously published artificial soil scaffolds, such as those using cryolite (20,35), fluorinated ethylene propylene (36), silica (37,38), and hydrogels (39,40). Moreover, we demonstrated that our platform was compatible with conventional fluorescence microscopy.…”

Complex extracellular biology drives surface competition during colony expansion in Bacillus subtilis

“…Advances in single-cell microscopy, transcriptomics and molecule tracking will likely continue to push our understanding of surface-bound microbial communities [21,54,55]. In addition, we believe important challenges lie in developing laboratory settings that most accurately reflect the complex ecologies to which bacteria are exposed in nature [56].…”