Biofilms are microbial communities that represent a highly abundant form of microbial life on Earth. Inside biofilms, phenotypic and genotypic variations occur in three-dimensional space and time; microscopy and quantitative image analysis are therefore crucial for elucidating their functions. Here, we present BiofilmQ—a comprehensive image cytometry software tool for the automated and high-throughput quantification, analysis and visualization of numerous biofilm-internal and whole-biofilm properties in three-dimensional space and time.

Biofilms are structured communities of bacteria, which are adhered to a surface and embedded in a self-produced matrix of extracellular polymeric substances. Since biofilms are very resistant to antimicrobial agents, they are at the basis of a range of problems, including quality and safety issues in food industry. Recently, major advances have been made in elucidating the different structural components of the biofilm matrix, the regulatory pathways involved in biofilm formation, and signaling molecules involved in biofilm formation and dispersal, which provide opportunities for prevention and control of these biofilms in the food industry.

Trophic interactions play a central role in driving microbial community assembly and function. In gut or soil ecosystems, successful inoculants are always facilitated by efficient colonization; however, the metabolite exchanges between inoculants and resident bacteria are rarely studied, particularly in the rhizosphere. Here, we used bioinformatic, genetic, transcriptomic, and metabonomic analyses to uncover syntrophic cooperation between inoculant ( Bacillus velezensis SQR9) and plant-beneficial indigenous Pseudomonas stutzeri in the cucumber rhizosphere. We found that the synergistic interaction of these two species is highly environmental dependent, the emergence of syntrophic cooperation was only evident in a static nutrient-rich niche, such as pellicle biofilm in addition to the rhizosphere. Our results identified branched-chain amino acids (BCAAs) biosynthesis pathways are involved in syntrophic cooperation. Genome-scale metabolic modeling and metabolic profiling also demonstrated metabolic facilitation among the bacterial strains. In addition, biofilm matrix components from Bacillus were essential for the interaction. Importantly, the two-species consortium promoted plant growth and helped plants alleviate salt stress. In summary, we propose a mechanism in which synergic interactions between a biocontrol bacterium and a partner species promote plant health.

Organisms as simple as bacteria can engage in complex collective actions, such as group motility and 28 fruiting body formation. Some of these actions involve a division of labor, where phenotypically 29 specialized clonal subpopulations, or genetically distinct lineages cooperate with each other by 30 performing complementary tasks. Here, we combine experimental and computational approaches to 31 investigate any benefits arising from division of labor during biofilm matrix production. We show that 32 both phenotypic and genetic strategies for a division of labor can promote collective biofilm formation 33 in the soil bacterium Bacillus subtilis. In this species, biofilm matrix consists of two major components; 34 EPS and TasA. We observed that clonal groups of B. subtilis phenotypically segregate in three 35 subpopulations composed of matrix non-producers, EPS-producers, and generalists, which produce 36 both EPS and TasA. We further found that this incomplete phenotypic specialization was 37 outperformed by a genetic division of labor, where two mutants, engineered as strict specialists, 38 . CC-BY 4.0 International license not peer-reviewed) is the author/funder. It is made available under a The copyright holder for this preprint (which was . http://dx.doi.org/10.1101/237230 doi: bioRxiv preprint first posted online Dec. 21, 2017; 2 complemented each other by exchanging EPS and TasA. The relative fitness of the two mutants 39 displayed a negative frequency dependence both in vitro and on plant roots, with strain frequency 40 reaching an evolutionary stable equilibrium at 30% TasA-producers, corresponding exactly to the 41 population composition where group fitness is maximized. Using individual-based modelling, we could 42show that asymmetries in strain ratio can arise due to differences in the relative benefits that matrix 43 compounds generate for the collective; and that genetic division of labor can be favored when it 44 breaks metabolic constraints associated with the simultaneous production of two matrix components. Microbes can act collectively in groups, and thereby substantially influence their local environment in 62 their own benefit. Such beneficial collective actions include the secretion of nutrient-degrading 63 enzymes [1,2], iron-scavenging siderophores [3], biosurfactants for group motility [4,5], and structural 64 components for biofilm formation [6,7]. In certain cases, cooperation even involves a division of labor, 65where subpopulations of cells specialize to perform different tasks [8][9][10]. For instance, during sliding 66 colony expansion Bacillus subtilis cells phenotypically differentiate into surfactant producers and 67 matrix producers where the role of the first is to reduce surface tension, while the latter allows 68 expanding colony 'arms' to form and explore new territories [10]. Given the high relatedness between 69 cells, specialization is likely beneficial for the group as a whole [11,12], with individuals gaining an 70 inclusive fitness benefit from helping their clone ...