Interrogating laboratory-adapted strains of Bacillus subtilis to elucidate the selective pressures of laboratory conditions on multicellular bacterial behaviors

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Abstract Bacteria carry out a variety of multicellular processes that influence their pathogenesis and environmental roles in the natural environment. However, it has become apparent that when these organisms are studied in the laboratory, they undergo significant genetic modification over time. The long-term goal of this project is to understand how the laboratory environment itself selects against and/or changes the fitness effects of multicellular bacterial behaviors including biofilm formation, motility, and the secretion of pigmented secondary metabolites. These features, however, are critical due to their influence on bacterial pathogenesis and their positive or negative environmental effects. The PI will characterize laboratory adapted strains of B. subtilis isolated from populations that grew in the common laboratory medium LB for approximately 300 generations. These strains have distinctive changes in motility, biofilm formation, and pigment production. The goals of this project are to use these laboratory-adapted strains to: 1) Identify the molecular mechanism(s) that causes an unusual “social swimming” behavior in one laboratory adapted strain, as surprisingly, this strain actively forms large, multicellular aggregates in broth culture. 2) Quantify the costs and benefits of motility in laboratory culture, as preliminary data suggest many laboratory-adapted strains have reduced or altered motility. 3) Identify the pigments produced by a laboratory-adapted strain and the wildtype B. subtilis strain NCIB3610 under distinct laboratory conditions, and quantify the effects of the production of these pigments on fitness in the laboratory. 4) Identify the environmental and metabolic pathways responsible for the triggering of production of pigmented natural products by Bacillus subtilis, focusing on pulcherrimin; and determine the relationship between pigment production and biofilm formation. This research is innovative because social swimming is a novel phenotype that could inform understanding of the evolution of multicellularity. Also, preliminary data identify discrepancies between our observations and the current model explaining the role of the pigment pulcherrimin in Bacillus subtilis, suggesting that additional research on the role of this pigment in B. subtilis is needed. This is important due to the role of pulcherrimin in biofilm formation and its antimicrobial properties. Additionally, this work will provide insights to scientists studying multicellular processes like motility in the laboratory, as this work will identify probable effects of the laboratory environment itself. Furthermore, this proposed project will enhance the research environment at Siena College significantly by providing support to involve more undergraduate students in research, increasing research capacity, and due to Siena’s student population, will aid in the larger goal of increasing diversity in STEM.