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This content will become publicly available on April 22, 2027

Title: Genetic diversification of Pseudomonas fluorescens maintained by multi-niche selection within biofilms
ABSTRACT We established a research-education partnership, EvolvingSTEM, that currently provides thousands of secondary school students the opportunity to conduct authentic research experiments centered on microbial evolution each year. Providing high school students access to research experiences improves learning, can have positive and long-lasting impacts on their attitudes toward science, and gives them the opportunity to make impactful scientific contributions. Through EvolvingSTEM, students evolve populations ofPseudomonas fluorescensin a bead model that includes daily cycles of bacterial dispersal, attachment, and biofilm growth, and observe heritable changes in colony morphology. We sequenced the genomes of 69 evolved clones with unique colony morphologies and identified parallel mutations in genes known to regulate or affect biofilm growth (wsp,yfiBNR,morA,andfuzY). We also uncovered novel adaptations: loss-of-function mutations in phosphodiesterase PFLU0185 that did not alter colony morphology and mutations affecting periplasmic disulfide bond formation producing small colonies. PFLU0185 mutants rapidly and consistently reached high frequencies, and phenotyping revealed roles in cyclic di-GMP regulation, biofilm formation, and motility, prompting us to name this genebmo(biofilm andmotility regulator). Competition experiments and microscopy demonstrated thatbmomutants are ecological generalists that compete with their ancestor and specialist mutants. Consequently, phenotypic diversity is maintained, with smooth (ancestral andbmo) colonies consistently outnumbering wrinkly and fuzzy variants. This study advances our understanding of biofilm genetic architecture while demonstrating that student-led research can uncover mechanisms of microbial adaptation relevant toPseudomonasinfection biology. IMPORTANCEBacterial biofilms dominate microbial life; however, their evolutionary genetics remain incompletely understood. Extensive replication of biofilm selection experiments by secondary school students can provide valuable insights into mechanisms of adaptation. Notably, mutations in a conserved phosphodiesterase, PFLU0185/bmo, dominate evolved populations without changing colony morphology. Phenotypic assays reveal thatbmoplays a generalist role and competes with its ancestor and other diverse, less frequent biofilm-specialist mutants, resulting in populations that have differentiated to fill multiple niches within the biofilm life cycle. This work also demonstrates the power of distributed research networks for addressing complex biological questions while promoting scientific literacy to the public.  more » « less
Award ID(s):
2444414 2147075
PAR ID:
10690825
Author(s) / Creator(s):
; ; ; ; ;
Editor(s):
Rudi, Knut
Publisher / Repository:
American Society of Microbiology
Date Published:
Journal Name:
Applied and Environmental Microbiology
Volume:
92
Issue:
4
ISSN:
0099-2240
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
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